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SIGAA - Sistema Integrado de Gestão de Atividades Acadêmicas

PPGBqBM PROGRAMA DE PÓS-GRADUAÇÃO EM BIOQUÍMICA E BIOLOGIA MOLECULAR ADMINISTRAÇÃO DO CB Phone: Not available E-mail: ppgbioquimica@gmail.com https://posgraduacao.ufrn.br/ppgbqbm

Dissertations/Thesis

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2025

	Dissertations
1	MARIA SANTANA DA SILVA EVALUATION OF TRAZODONE EFFECTS ON REPRODUCTIVE PARAMETERS AND EPIDIDYMAL DUCT CONTRACTION IN RATS Advisor : EDILSON DANTAS DA SILVA JUNIOR COMMITTEE MEMBERS : EDILSON DANTAS DA SILVA JUNIOR ELAINE CRISTINA GAVIOLI MIRIAM DAS DORES MENDES FONSECA Data: Jan 30, 2025 Show Abstract Trazodone is an antidepressant widely used by the male population. However, the effects of this drug on male reproductive health are not yet fully understood. Therefore, this study aims to evaluate the effects of trazodone on epididymal duct contraction and various reproductive parameters in rats. Adult Wistar rats were orally treated for 21 days with saline (control) or trazodone at doses of 10 mg/kg and 20 mg/kg. After the treatment period, the animals were euthanized, reproductive organs were weighed, and the testis and epididymis were isolated for sperm parameter analysis and assessment of the contraction of the distal cauda epididymal duct. In silico studies were also conducted to predict molecular targets related to the effects of trazodone on the male reproductive tract. We observed that trazodone has anti-reproductive effects in rats, characterized by damage to testicular tissue, reduced sperm production, as well as decreased quantity and quality of sperm collected from the cauda epididymis. The contraction of the isolated distal cauda epididymal duct was also affected by in vivo and in vitro trazodone treatment, indicating possible alterations during the emission phase of ejaculation. Additionally, both trazodone and its active metabolite m-CPP showed in silico predictors suggesting nonspecific interactions with pharmacological targets, which may contribute to an unfavorable reproductive toxicological profile. In conclusion, trazodone exhibited significant anti-reproductive effects that may impair male fertility.
2	MARINA ROCHA DO NASCIMENTO DANTAS Evaluation of osteogenic and genotoxic potentials of nanostructured hydroxyapatite-based microspheres containing zinc Advisor : SUSANA MARGARIDA GOMES MOREIRA COMMITTEE MEMBERS : CARLOS AUGUSTO GALVAO BARBOZA GILDACIO PEREIRA CHAVES FILHO SUSANA MARGARIDA GOMES MOREIRA Data: Feb 14, 2025 Show Abstract A wide range of biomaterials is being studied as new alternatives in the field of bone regeneration. Among these, hydroxyapatite (HA) is a mineral component present in the inorganic phase of bones. HA is not only biocompatiblebut also possesses suitable physicochemical properties, allowing for ionic substitutions that enhance its performance for various applications. The REGENERA Institute has developed nanostructured hydroxyapatite (nHA) microspheres with different compositions, including zinc ions (Zn²⁺) incorporation,which are notable for their ability to increase cell viability and promote bone mineralization. However, any modification to HA results in the production of a newbiomaterial that, in addition to confirming its osteogenic capabilities, needs to have its biosafety tested. The osteogenic differentiation capacity of nHA-based microspheres containing zinc ions using stroma stem cells isolated from the Wharton's jelly of human umbilical cords (hWJ-MSCs) was evaluated in this study. Cytotoxicity results, assessed by the MTT assay (3-(4,5-dimethylthiazol-2- yl)-2,5-diphenyltetrazolium bromide), indicated that the samples did not significantly reduce the ability of hWJ-MSCs cells to reduce MTT, suggesting thatthe samples are not cytotoxic under the tested conditions. The bone mineralization results, alkaline phosphatase (ALP) activity, and gene expression quantification assay by qRT-PCR indicated that the samples, under the tested conditions, exhibited osteogenic activity by increasing extracellular matrix mineralization, ALP activity, and the expression of genes related to osteogenesis in hWJ-MSCs. Considering clinical applications, the genotoxic potential of these materials was also assessed. The genotoxicity results, evaluated by the Comet assay and the CBMN assay (cytokinesis-blocked micronucleus assay), showed that the samples do not exhibit genotoxic potential under the tested conditions. Therefore, it is expected that this biomaterial may have applications in bone regeneration.
3	PEDRO VITOR VALE BEZERRA Extracts from Amburana cearensis and Erythrina velutina seeds: impacts on survival, intestinal protease activity and morphology of Aedes aegypti and Aedes albopictus larvae Advisor : ADRIANA FERREIRA UCHOA COMMITTEE MEMBERS : ADRIANA FERREIRA UCHOA FELIPE ARLEY COSTA PESSOA GIULIAN CÉSAR DA SILVA SÁ PATRICIA BATISTA BARRA MEDEIROS BARBOSA VALTER FERREIRA DE ANDRADE NETO Data: Feb 17, 2025 Show Abstract Arboviruses constitute a significant group of viral diseases primarily transmitted by hematophagous mosquitoes, notably Aedes aegypti (Linnaeus, 1762) and A. albopictus (Skuse, 1894). Conventional management strategies for these diseases predominantly rely on synthetic chemical insecticides as the primary control measure. However, this approach raises concerns regarding environmental toxicity and the potential development of resistance among mosquito populations. In light of these challenges, botanical insecticides, including plant extracts, represent a compelling alternative due to their biocompatibility and diverse modes of action, minimizing environmental impacts and insect resistance. This study investigates the insecticidal properties of extracts from the seeds of two legumes indigenous to the Caatinga biome, specifically Amburana cearensis ((Allem.) A. C. Smith) and Erythrina velutina (Willd.), against A. aegypti and A. albopictus larvae (L4). The extraction process involved solubilizing the seed powder in distilled water (1:10, w/v) at a temperature of 80 °C. The lethal concentrations for 50% (LC50) and 90% (LC90) of the larvae were assessed over 24 and 48 hours. Morphological alterations were examined using a stereomicroscope equipped with a camera, and larval development was monitored for 15 days. Furthermore, zymographic analyses were conducted to assess the inhibition of digestive proteases in the treated larvae. The results indicated that both extracts exhibited larvicidal activity, with A. cearensis extract demonstrating particularly notable efficacy. Specifically, the extract recorded LC50 values of 0,45 mg/mL and 0,23 mg/mL for 24 and 48 hours in A. aegypti, respectively, and LC50 values of 0,34 mg/mL and 0,11 mg/mL in A. albopictus. Additionally, A. cearensis extract reduced the enzymatic activity of intestinal proteases of both species by more than 70%, while E. velutina extract had no significant effect. Delays in larval development in A. albopictus suggest a pronounced influence of protease inhibition resulting from A. cearensis extract. Morphological changes suggest the presence of other deleterious effects of the extracts: acetylcholinesterase inhibition (A. cearensis and E. velutina), disruption of chitin synthesis, and internal tissue damage or infection leading to melanization (E. velutina). This study highlights the promising potential of A. cearensis and E. velutina extracts for larval control of A. aegypti and A. albopictus, with A. cearensis extract emerging as a particularly viable candidate for the development of botanical larvicides. The findings underscore the importance of plant biodiversity within the Caatinga biome in the creation of bioproducts, thereby enhancing its cultural, biotechnological, and economic significance while contributing to integrated management strategy for arboviral diseases.
4	LILIANE SOARES DE CASTRO BEZERRA Characterization of EXO-3/APE1 and XPA-1/XPA proteins in development of neurodegenerative phenotypes in Caenorhabditis elegans Advisor : RIVA DE PAULA OLIVEIRA COMMITTEE MEMBERS : JULLIANE TAMARA ARAUJO DE MELO CAMPOS RIVA DE PAULA OLIVEIRA EVANDRO ARAUJO DE SOUZA Data: Feb 24, 2025 Show Abstract The BER and NER pathways are well preserved in C. elegans. In this organism, the endonuclease EXO-3, homologous to APE1 of the BER pathway, and the ortholog XPA-1 of the NER pathway play key roles. exo-3 mutants present reduced progeny, increased production of reactive oxygen species (ROS) and larger mitochondrial DNA (mtDNA) deletions. On the other hand, xpa-1 mutants are sensitive to UV radiation and present deficiencies in the ubiquitin-proteasome system (UPS) and mitochondrial metabolism.Thus, we evaluated how deficiencies of EXO-3 and XPA-1 influence mitochondrial integrity and the development of neurodegenerative phenotypes in transgenic models of C. elegans. For this purpose, we analyzed exo-3 and/or xpa-1 knockdown animals from strains that emit GFP in intestinal and muscle mitochondria. In addition to observing the effect of BER and NER pathway deficiency on apoptosis, we also evaluated cholinergic neuronal integrity in animals deficient for exo-3 or xpa-1. The results indicated evidence of neurodegeneration and signs of mitochondrial dysfunction in animals deficient in both repair pathways, reinforcing the overlap of these two pathways. In addition, they are in line with what was observed in previous studies on increased mitochondrial content in exo-3;xpa-1 knockdown animals. Thus, the focus is on the development of exo-3 and xpa-1 deletion mutant animals for a better characterization of mitochondrial function and neuronal integrity. This work is expected to contribute to a better understanding of neurodegenerative diseases and the role of EXO-3, a central endonuclease in BER repair, in the emergence of neurodegenerative phenotypes and their association with mitochondrial dysfunctions.
5	MATHEUS OLIVEIRA DE SENA Multiple Symmetric Lipomatosis: Genetic Diagnosis and Genotype/Phenotype Relationship of MFN2 Gene Pathogenic Variants Advisor : JULLIANE TAMARA ARAUJO DE MELO CAMPOS COMMITTEE MEMBERS : JULLIANE TAMARA ARAUJO DE MELO CAMPOS LEONARDO CAPISTRANO FERREIRA RENAN MAGALHAES MONTENEGRO JUNIOR Data: Mar 10, 2025 Show Abstract Multiple Symmetric Lipomatosis (MSL), also known as Madelung's disease or Launois Bensaude disease, consists of a rare genetic disease with an autosomal recessive or compound heterozygous character that affects adipose tissue and is also characterized as a rare mitochondrial syndrome. MSL, genetically caused by a pathogenic variant in the MFN2 gene (c.2119 C>T; p.Arg707Trp), has already been phenotypically characterized by the development of non-encapsulated lipomas, mainly in the upper region of the trunk, highlighting the neck, supraclavicular and interscapular regions, commonly associated with axonal neuropathy, metabolic complications such as hypertriglyceridemia, decreased HDL-c, hypoadiponectinemia and hypoleptinemia in addition to mitochondrial disorders. Patients' history of excessive alcohol consumption has been associated with the accelerated development of LSM. It is worth mentioning that other variants were found in patients with LSM, such as the pathogenic variants c.1027_1029delAGG (p.Arg343del), c.600_816del (exons 7 and 8 deletion) and c.1496-2A>G, all found in compound heterozygosity together with pathogenic variant c.2119 C>T (p.Arg707Trp). Furthermore, several other pathogenic variants have already been described in the MFN2 gene. Still, only c.2119 C>T (p.Arg707Trp) in homozygosity or compound heterozygosity (along with the variants c.1027_1029delAGG, c.600_816del and c.1496-2A>G) were reported to determine the development of LSM, while the vast majority of other variants described in the gene are known to cause the axonal neuropathy called Charcot-Marie-Tooth type 2 (CMT2). The clinical manifestations of LSM present a significant heterogeneity, which is reflected in the chronology of their appearance and severity. In this study, we performed the 16th diagnosis of MFN2-associated MSL worldwide, the first in Brazil, in which the c.2119C>T variant was identified in homozygosis. The patient had low serum levels of adiponectin, leptin and HDL-c, in addition to hyperglycemia. The genotype x phenotype relationship of this patient with other already described in the literature was established. Using bioinformatic tools, simulations of three-dimensional structure, prediction of membrane topology and homotypic and heterotypic protein-protein interactions were performed for the wild type and mutants of mitofusin 2. In addition, an interaction network of MFN2 gene was made, in which the most enriched biological processes involving this gene were observed.
6	ANGELIS MARIA ALVES FALCÃO Interactions between vector and hosts in the transmission dynamics of Leishmania in- fantum in endemic areas Advisor : SELMA MARIA BEZERRA JERONIMO COMMITTEE MEMBERS : SELMA MARIA BEZERRA JERONIMO FABIANO OLIVEIRA JOANNA GARDEL VALVERDE GALVÃO LUCAS PEDREIRA DE CARVALHO Data: Mar 10, 2025 Show Abstract Rio Grande do Norte state (RN) is an endemic area for human and canine visceral leishmaniasis (VL) in Brazil. Leishmania infantum is the major etiologic agent for VL in Latin America. The transmission cycle of this pathogen is anthropozoonotic, and Lutzomyia longipalpis is the most important epidemiological vector in the Americas. During a blood meal, the vector releases chemical factors contained in its saliva, which have anti-hemostatic activity, facilitating blood suction, but have also been shown to facilitate the chemotaxis process of macrophages and neutrophils. Studies have shown that re-exposure to the vector insect can increase the transmissibility of Leishmania from the infected mammalian host to the vector. This study aims to determine the main blood sources, the vector infection rate by L. infantum in sandflies from an endemic VL area, and the role of Lu. longipalpis saliva in dogs infected with L. infantum. A total of 248 female sandflies were collected using light traps in the peridomestic area of residences with the following characteristics: (1) households with a recent history of human VL, (2) neighboring households of a home with human VL, and (3) households without a recent history of human VL. DNA was extracted individually from the sandflies' intestines, and infection with L. infantum and blood sources were identified by qPCR. Additionally, dogs with VL, selected through the surveillance survey of canine visceral leishmaniasis by the Zoonoses Surveillance Unit (UVZ), were studied. Peripheral blood from the dogs was collected before intradermal inoculation of a Lu. longipalpis salivary glands homogenate or saline into the dog's ear. After 24 hours, the dog's blood were collected again to evaluate parasitemia and protein profile in canine plasma.. Molecular analysis detected L. infantum DNA in 56% of the sandflies captured. Households with a history of human VL were 3.73 (95% CI = 1.64 to 8.48; p = 4.92e-04) times more likely to have L. infantum DNA in the intestines of sandflies compared to those captured in households without a recent history of VL. Multiple blood sources were detected, with dogs being the primary blood source in all three studied areas. A significant increase in L. infantum was observed in peripheral blood after inoculation with Lu. longipalpis saliva (p=0.0034), unlike dogs inoculated with saline. The concentration of Leishmania in the DNA of skin tissue, including ear and contralateral axillary regions, was higher in both groups compared to the amount of Leishmania present in the left ear, which was used for inoculation. Proliferation of Leishmania in the amastigote form occurred with the presence of Lu. longipalpis saliva. The analysis of differential expression of plasma proteins revealed overexpression of proteins such as coagulation factor V and ApoE, 24 hours after exposure to the vector's saliva. Based on these findings, we conclude that there is an important reservoir of L. infantum near the residences of human VL cases and that dogs are the primary blood source for sandflies in the studied areas. Additionally, we conclude that the intradermal administration of Lu. longipalpis saliva increases parasitemia in dogs naturally infected with L. infantum. Therefore, daily re-exposure to the vector in endemic areas may increase the infectivity of the hosts. These findings contribute to a better understanding of transmission in endemic areas of visceral leishmaniasis and the discussion of possible control measures for VL.
7	RAYSSA THAINNA SILVA LOPES NUT carcinoma: case report and tumor microenvironment analysis using VEGFA, STAT3 and CD163 markers Advisor : JULLIANE TAMARA ARAUJO DE MELO CAMPOS COMMITTEE MEMBERS : JULLIANE TAMARA ARAUJO DE MELO CAMPOS NAISANDRA BEZERRA DA SILVA FARIAS KARUZA MARIA ALVES PEREIRA Data: Apr 10, 2025 Show Abstract NUT carcinoma (NC) is a rare neoplasm with rapidly progressing affecting both sexes across all age groups, with primary tumors most commonly located in the mediastinum and genetically characterized by a translocation involving the NUTM1 gene and one of the following genes: BRD4, BRD3, BRD2, NSD3, ZNF532, or ZNF592. Generally, resistance to conventional therapies and high toxicity of available treatments are the main challenges associated with this disease. In this context, the present study aimed to characterize a case of NC diagnosed in 2019 in Natal, Rio Grande do Norte, Brazil, and to analyze the tumor microenvironment using VEGFA, STAT3, and CD163 markers through immunohistochemistry (IHC). For these purposes, the patient’s medical record and imaging exams were used for clinical characterization, hematoxylin and eosin-stained histopathological slides were obtained for capturing histopathological images, and paraffin-embedded tumor tissue samples were acquired for IHC testing. The clinical case involved a 28- year-old man with a primary tumor located in the nasal cavity and metastasis in the T11 thoracic vertebra. He underwent conventional therapies, including surgical resection, chemotherapy, and radiotherapy; however, the tumor showed resistance to these treatments, progressed rapidly, and the patient died 21 months after diagnosis. Histopathological analyses revealed a monomorphic population of round tumor cells, PMN infiltration, hemorrhagic foci, and the presence of macrophages, findings that guided the choice of three additional IHC markers, all of which showed positivity in the tumor sample. In this context, VEGFA overexpression suggests intense angiogenic activity, STAT3 presence indicates signaling that promotes resistance to apoptosis and immunosuppression, and CD163+ macrophages suggest a strong infiltration of immunosuppressive cells in the tumor microenvironment. Thus, these markers may assist in monitoring NC, providing information on metastatic potential, inflammatory activity, and tumor resistance to treatments, as well as promoting the development of immunotherapybased approaches.
	Thesis
1	ÉRIKA GEICIANNY DE CARVALHO MATIAS QUANTUM BIOCHEMISTRY APPROACH APPLIED TO THE MODULATION OF SIRT6 AND PIM1: UNVEILING NEW THERAPEUTIC STRATEGIES WITH FLAVONOIDS Advisor : UMBERTO LAINO FULCO COMMITTEE MEMBERS : UMBERTO LAINO FULCO EDILSON DANTAS DA SILVA JUNIOR ANTONIO DE MACEDO FILHO KATYANNA SALES BEZERRA LUIZ ANTONIO RIBEIRO JUNIOR Data: Jan 24, 2025 Show Abstract Metabolic diseases and cancers affect millions of people worldwide, representing a health and well-being problem for society. In this scenario, two proteins, SIRT6 and PIM1, emerge. These macromolecules have stood out as potential therapeutic targets for these pathologies since their regulatory functions are directly associated with preventing these diseases. SIRT6 is an NAD+-dependent deacylase, and its activation has potential against metabolic and aging-related diseases. In contrast, its inhibition is considered promising in cancer therapy. The investigation of the modulation of this protein by specific molecules stands out as a promising area. PIM1 is a proto-oncogene overexpressed in several types of human cancer, and its inhibition is considered a promising target for cancer therapy. Here, we explore the modulation of SIRT6 with the flavonoids quercetin (QUE) and its derivative isoquercetin (ISO), in addition to catechin gallate (GC) and trichostanin-A (TSA), as well as the PIM1 protein with the flavonoids quercetin (QUE), myricetin (MYC) and pentahydroxyflavone (PTH), providing a detailed view of the molecular interactions that lead to the activation and/or inhibition of these proteins. For this, we used computational methods from the perspective of Molecular Modeling through the Molecular Fractionation with Conjugated Caps (MFCC) technique in accordance with the calculation parameters of the Density Functional Theory (DFT). These evaluations were carried out within a radius of up to 10.0 Å away from the modulators. The energy analysis of the SIRT6-modulator complex demonstrated that the activating substances (GC) and (TSA) were more expressive compared to the inhibitory molecules (QUE) and (ISO). In contrast, for the PIM1-inhibitor complex, the flavonoid that presented the most expressive interaction was (PTH) followed by (QUE) and (MYC). Our results provide relevant evidence for the development of therapeutic strategies targeting SIRT6 and PIM1, which have the potential to impact the regulation of these proteins and open new perspectives in the research of treatments for metabolic diseases and cancer.

2024

	Dissertations
1	MONIQUE ALVARES DA SILVA GENETIC DIAGNOSIS AND GENOTYPE/PHENOTYPE CHARACTERIZATION OF VARIANTS OF THE PPARG GENE, PRECURSOR OF FAMILIAR PARTIAL LIPODYSTROPHY TYPE 3 IN PATIENTS FROM NORTHEAST BRAZIL Advisor : JULLIANE TAMARA ARAUJO DE MELO CAMPOS COMMITTEE MEMBERS : VIRGINIA OLIVEIRA FERNANDES CORTEZ DANIEL CARLOS FERREIRA LANZA JULLIANE TAMARA ARAUJO DE MELO CAMPOS NAISANDRA BEZERRA DA SILVA FARIAS Data: Feb 8, 2024 Show Abstract Lipodystrophy type 3 (FPLD3) has been described as molecularly associated with the PPARG gene, with predominantly missense variants on chromosome 3. The syndrome causes loss of adipose tissue in the upper and lower limbs, hips, and face. It is generally associated with metabolic disorders such as type 2 diabetes, insulin resistance, hypertriglyceridemia, and liver dysfunction. Therefore, the objective of this work was to molecularly diagnose and characterize the genotype/phenotype of patients with a clinical diagnosis of LPF3 recruited by the endocrinology clinic of the Hospital Universitário Onofre Lopes.To this end, a clinical evaluation was carried out, oral swabs and blood samples were collected. Biochemical analyses and dosages of the hormones adiponectin and leptin were performed using the ELISA technique to evaluate the metabolic disorders inherent to FPLD3. The next-generation sequencing (NGS) technique was chosen to diagnose the variants through a panel of genes related to lipodystrophies including LMNA and PPARG. To confirm the NGS data, amplification of genetic material was carried out by conventional PCR and Sanger sequencing using primers specifically designed for this work. The tools used for bioinformatic analysis of the variants were Poly-Phen2, TCoffee, and Mutation Taster. It was observed that the patients presented clinical characteristics compatible with the presence of lipodystrophy, such as loss of adipose tissue in the extremities, hypertriglyceridemia, and steatosis, in addition to a reduced adiponectin and leptin dosage within normal limits concerning the reference value for the index of corresponding body mass. Molecularly, new heterozygous variants located at positions c.533T>C promoting the replacement of the amino acid leucine by proline at 178 (p.Leu178Pro) and c.641C>T promoting the replacement of the amino acid proline by leucine at 214 were identified and confirmed in three patients. (p.Pro214Leu). The variants respectively affect the DBD domain of the protein, responsible for binding PPAR to the promoter region of target genes, and the Hinge region, involved in the interaction with coactivators and corepressors. Through bioinformatic analysis of each identified variant and its comparative alignment with the wild type, it was observed that the scores corroborated what was observed clinically, classifying the variants as harmful to their function. This was the first study to molecularly characterize patients with familial partial lipodystrophy type 3 in Brazil and the first to report the c.533T>C and c.641C>T variants in the global literature.
2	MARIA EDUARDA CARDOSO DE MELO Analysis of pathogenic variants in AGPAT2 and biochemical-molecular evaluation of individuals with congenital generalized lipodystrophy types 1 and 2 Advisor : JULLIANE TAMARA ARAUJO DE MELO CAMPOS COMMITTEE MEMBERS : JULLIANE TAMARA ARAUJO DE MELO CAMPOS LEONARDO CAPISTRANO FERREIRA VIRGINIA OLIVEIRA FERNANDES CORTEZ Data: Mar 28, 2024 Show Abstract Congenital Generalized Lipodystrophy (CGL) is a rare disease that causes the loss of adipose tissue (AT) throughout the body from birth. Patients with CGL have low levels of leptin and adiponectin (APN), important adipokines produced by AT. There are four types of CGL, with types 1 and 2 being responsible for more than 80% of the cases described in the literature and with patients already reported in northeastern Brazil. The molecular etiology for LGC 1 and LGC 2 is the presence of pathogenic variants (PVs) in the AGPAT2 and BSCL2 genes, respectively. This study carried out the molecular analysis of the main VPs described for AGPAT2 (c.366-588del, c.589-2A>G, c.646A>T, c.570C>A, c.369-372delGCTC, c.202C>T, c.514G>A, c.144C>A) and an analysis of the effects on their transmembrane and functional domains. The genotype vs phenotype relationship of the first patients described in the literature for the PVs in question demonstrates the relationship between molecular alterations and clinical symptoms, so that patients with PVs that more significantly affect the proteins show a higher rate of clinical symptoms. In addition, we performed the molecular diagnosis of two sisters with VPs in compound heterozygosity for c.299G>A and c.493-1G>C and a new patient described for the c.366-588del variant. Using next-generation sequencing (NGS), a panel of genes associated with adipose tissue-related diseases was analyzed in LGC patients, which showed the presence of different VPs for the BSCL2 gene. The comparative study of leptin and APN dosages between the LGC 1 and LGC 2 groups showed that LGC type 2 patients had lower leptin levels, while no differences were observed between APN dosages in these two groups. At the level of expression of enzymes involved in repairing oxidized DNA damage (APE-1, OGG1 and PPARG), no significant differences were observed between the LGC 1 and LGC 2 groups, although both types had higher levels of expression of these genes when compared to that observed in control individuals. This suggests that the different molecular etiology of the two types of LGC is not a significant parameter for altering redox homeostasis between these two groups. Further studies are needed to better understand the relationship between LGC and oxidative stress, DNA repair and cellular response.
3	GEORGGIA FÁTIMA SILVA NALIATO Tenebrio molitor as an experimental model for comparing virulence profiles and immune response to pathogenic fungi of the genus Sporothrix Advisor : RAQUEL CORDEIRO THEODORO COMMITTEE MEMBERS : RAQUEL CORDEIRO THEODORO DANIEL CARLOS FERREIRA LANZA RAFAEL WESLEY BASTOS ÉVERTON KORT KAMP FERNANDES Data: Apr 5, 2024 Show Abstract Diseases caused by pathogenic fungi have a substantial impact on both immunocompromised and immunocompetent patients, requiring lengthy treatments that, in some cases, demand hospitalization. In this regard, species and/or genotype-specific diagnosis can enhance disease prognosis, enabling a more targeted treatment depending on the etiological agent. Nevertheless, comparative studies on parasitism relationships involving closely related fungal species, often cryptic within the same genus, are still scarce. Species-specific virulence categorization directly impacts therapeutic direction and clinical prognosis, thereby optimizing case resolution. Consequently, the gold standard model for assessing the virulence of these pathogens is the murine model. However, currently, due to increased concerns about bioethical and financial issues, invertebrate animals have been proposed as attractive alternatives for infection models, given their innate immunity evolutionarily close to mammals, infrastructural ease, and robust data reproducibility. In this study, the virulence of different species of medical importance, from both clinical and environmental clades of the dimorphic fungus genus Sporothrix, as well as aspects of the innate immune response, were assessed in the invertebrate model Tenebrio molitor, in both the mycelial and yeast phases of the fungus. Thus, it was possible to compare virulence profiles and the host-parasite relationship among isolates of species within the genus. Significant differences were observed between the mycelial and yeast phases of pathogenic species from both Sporothrix clades. The species S. chilensis and S. pallida (environmental clade) demonstrated higher virulence in the mycelial morphology, whereas the species S. schenckii and S. brasiliensis (clinical clade) showed higher virulence in the yeast morphology. The expression of antimicrobial peptides (AMPs) from the invertebrate model was quantified, showing a significant difference in expression between groups infected with S. pallida and S. schenckii, indicating alterations in the pattern of the innate immune response concerning the Sporothrix species inoculated into the model.
4	MARYANA THALYTA FERREIRA CÂMARA DE OLIVEIRA MALE FERTILITY: MARKET IN LATIN AMERICA AND DEVELOPMENT OF SYSTEMS FOR DETECTING PATHOGENS IN HUMAN SEMEN Advisor : DANIEL CARLOS FERREIRA LANZA COMMITTEE MEMBERS : DANIEL CARLOS FERREIRA LANZA EDILSON DANTAS DA SILVA JUNIOR DANIELLE BARBOSA MORAIS JANAINA FERREIRA ADERALDO KYVIA BEZERRA MOTA Data: Apr 15, 2024 Show Abstract According to the World Health Organization (WHO), infertility is a disease of the reproductive system that results in limited fertility in men and women. On a global scale, more than 186 million individuals face infertility challenges, with the majority of them residing in developing countries. This has driven the growth of the infertility treatment industry, including the male fertility market, since around 15% of men of reproductive age face infertility problems. However, diagnosing infertility in men still faces challenges, with semen infections being one of the potential causes. Recent studies have shown that different microorganisms, such as viruses and bacteria, can impact on semen quality, causing damage to organs and cells through inflammatory mediators. In this context, this dissertation aimed to characterize the main topics related to the fertility and assisted reproduction market in Latin America and Brazil, to emphasize the growth of the male fertility market and the challenges faced in diagnosing infertility in men, as well as to develop systems for detecting the main pathogens that infect semen. Three studies were conducted to achieve these objectives: two literature reviews to describe the markets and the last study developed primers and validated in silico three multiplex PCR panels for the detection of the main seminal pathogens of clinical importance already described. The main conclusions of the studies include: 1) Fertility services could earn up to $31.59 billion by 2029 and the use of assisted reproductive technologies (ART) has become a thriving commercial business within fertility services. It was noted that the fertility market has seen significant growth in both Brazil and Latin America. Specifically in Brazil, there has been growth over the last few years, with approximately 40% of assisted reproduction clinics in Latin America and revenues of around R$1.3 billion. It is estimated that between 2021 and 2025, around 77,588 patients up to the age of 35 will be treated via ART in the country. At the same time, both public policies and private sector initiatives aimed at fertility are increasing, underlining the importance of fertility in people's lives (Chapter I). 2) The male fertility market has also shown significant growth as the fertility and assisted reproduction market expands. According to market reports, global trade related to male fertility is estimated to be worth billions of dollars and is expected to reach more than $6 billion by 2027 (Chapter II). 3) Based on the data collected, three panels were developed in silico that focus on seminal quality, covering HPV-16, HPV18, HBV, HHV-5, HSV-1, HSV-2, HIV-1, HIV-2, ZIKV, HCV, Treponema pallidum, Neisseria gonorrhoeae and Chlamydia trachomatis. The implementation of these panels represents a significant advance in seminal quality control, providing more accurate diagnoses for semen banks and guidance for couples facing infertility. The proposed assay promises to meet the needs of the male fertility market, as well as simplifying workflow, allowing it to be used in routine diagnostic laboratories with basic molecular facilities (Chapter III). Overall, all the information gathered in this dissertation contributes to new areas of research in the areas of the market, improving techniques and diagnosing conditions related to male infertility and global reproductive health.
5	RAFAELA ALVES DE LIMA Comparative analysis of the effects of microgravity, heat, and hypoxia on the gene expression pattern of plants: a bioinformatic approach Advisor : KATIA CASTANHO SCORTECCI COMMITTEE MEMBERS : ALESSANDRO DE MELLO VARANI KATIA CASTANHO SCORTECCI LEONARDO CAPISTRANO FERREIRA Data: Apr 24, 2024 Show Abstract Plants are subjected to daily environmental changes and must respond optimally to tolerate these modifications. Alterations in gravity, hypoxia, and heat are examples of environmental changes that can induce abiotic stress, primarily triggering oxidative signaling. Since microgravity is not a condition naturally occurring on Earth, some exposure methods are required to place plants in environments with gravity approaching zero. The use of international space stations and space flights are ways to expose plants to this type of environment. However, there is still no consensus on all the effects that microgravity may trigger. Thus, comparing microgravity studies with other environmental factors can enhance understanding of plant responses to microgravity. In this context, the integration of transcriptomic data from studies available on open-access platforms was performed, investigating the impacts of microgravity, hypoxia, and thermal stress on plants. Since these environmental conditions may share oxidative signaling pathways, the objective of this integration was to enrich the understanding of plant responses to microgravity, heat, and hypoxia in a comparative manner, as well as to explore the differential and correlated effects of microgravity in international space station and space flight settings. Therefore, bioinformatics tools were employed to identify significantly activated central genes, ontologies, and metabolic pathways. For data integration, a selection was made according to inclusion and exclusion criteria. For example, studies should clearly state experimental conditions and compare stressed and non-stressed plants. Consequently, 16 microgravity studies were selected, including 11 from international space stations and 6 from space flights, along with 10 thermal stress studies and 9 hypoxia studies. This integration revealed significant regulation of genes related to chlorophyll biosynthesis. These results suggest that photooxidation may be an important effect under microgravity, hypoxia, and heat conditions. This effect appears to be independent of microgravity exposure type. There was also an overlap in the activation of antioxidant metabolic pathways: glutathione metabolism and phenylpropanoid biosynthesis, demonstrating that plants employ redox homeostasis pathways using different strategies. Primary metabolism, specifically through the production of sugars such as starch and fructose, is significantly affected under hypoxia, heat, and microgravity conditions. These sugars are essential for energy production, carbon metabolism, and gravity signaling. Therefore, oxidative signaling triggered by microgravity, heat, and hypoxia significantly affected chlorophyll production and primary metabolism; however, this impact may be mitigated by intense antioxidant activity. Comparing these responses to those induced by other environmental stresses provides important insights into the adaptive capacity of plants in environments with different gravitational forces. Such understanding is crucial for developing strategies to enhance crop resilience in terrestrial and space agriculture contexts.
6	OHANA LETÍCIA TAVARES DA SILVA Exploring the pharmacological potential of carrageenan disaccharides as antitumor agents: an in silico approach Advisor : HUGO ALEXANDRE DE OLIVEIRA ROCHA COMMITTEE MEMBERS : HUGO ALEXANDRE DE OLIVEIRA ROCHA LEANDRO SILVA COSTA ÉDER GALINARI FERREIRA Data: Apr 30, 2024 Show Abstract Carrageenans, sulfated galactans, have antitumor activity that can be enhanced by their depolymerization. This means that their oligosaccharides, including disaccharides, being less structurally complex, may have pharmacological potential in the treatment of various tumors. However, it is still unknown whether these disaccharides/oligosaccharides are pharmacologically viable and by what mechanisms they act to inhibit tumor growth. Therefore, the objective of this study was to utilize bioinformatics tools to investigate the pharmacological properties and potential molecular targets of disaccharides most representative of iota, kappa, and lambda carrageenans. To achieve this, the pharmacokinetic profile and drug-like physicochemical properties of disaccharides were initially predicted. Subsequently, the molecular targets of these disaccharides were predicted, and among them, those that are already targets of oncological drugs were selected. Molecular docking was then conducted to evaluate the binding capacity and affinity between the selected targets and disaccharides. For comparative analysis of binding energies, oncological drugs acting on these targets were also subjected to docking. In general, disaccharides exhibited favorable metabolization, excretion, and toxicity profiles, but they face permeability challenges in biological membranes, affecting their absorption and distribution. Furthermore, they met most of the physicochemical criteria proposed in drug similarity tests, except for polarity and lipophilicity, which were below and above the desirable limits, respectively. In the selection of molecular targets, five common targets were predicted: carbonic anhydrases (CAs) I, II, IX, XII, and XIV, which are already targeted by several oncological drugs. In docking, the binding energies were similar or superior to those of drugs already used for these targets. Regarding the interactions between disaccharides and CAs, those involving the zinc cofactor stood out, as it is the primary mechanism for inhibiting these enzymes. These findings indicate a promising avenue for carrageenan disaccharides as cancer therapeutics.
7	MELISSA FARIAS ALVES DA SILVA PROSPECTION OF ANTIFUNGAL ACTIVITY AND MODULATORY POTENTIAL OF EXTRACTS AND FRACTIONS OF Tephrosia toxicaria (Sw.) Pers in REFERENCE ANTIMICROBIALS Advisor : ADRIANA FERREIRA UCHOA COMMITTEE MEMBERS : GABRIEL BONAN TAVEIRA ADRIANA FERREIRA UCHOA KATIA CASTANHO SCORTECCI THALES DOMINGOS ARANTES Data: May 9, 2024 Show Abstract Fungal infections represent a major contributor to the increasing global mortality rate associated with infectious diseases. This increase is largely attributed to the indiscriminate use of antibiotics and antifungals agents, whose effectiveness is compromised by the development of resistance in pathogens and the adverse effects observed in non-target organisms. Botanical extracts and their fractions have been highlighted as promising sources for the development of antifungals and therapeutic adjuvants for reference drugs as they are derived from renewable sources and present toxic selectivity. Tephrosia toxicaria (Sw.) Pers, a legume well adapted to the northeastern semi-arid region, has stood out for its biopotential. The present research aimed to investigate the antifungal potential of extracts and protein fractions obtained from T. toxicaria seeds against species of the Candida and Cryptococcus genera and their modulating activity for reference antimicrobials. The extracts and fractions obtained by the Scopes and Osborne methods were partially biochemically characterized and evaluated for their antifungal activity by determining the MICs (Minimum Inhibitory Concentration). Subsequently, the most active fraction and its combinations were also investigated regarding their possible mode of action. The extracts and fractions showed antifungal activity to different degrees for at least one of the yeasts tested, in which the T 50-75 fraction was the most active against Cryptococcus gattii, Cryptococcus neoformans and Candida parapsilosis and the GLB fraction against Candida albicans. The lowest MIC100 values (MIC, capable of inhibiting 100% of growth) found were 32 µg/mL for the T 50-75 fraction against Cryptococcus gattii (R-265), 128 µg/mL for the ST extract and 256 µg/mL for fractions T 50-75 and T 75-100, all against C. gattii (FC1). The ST extract and the fractions T 50-75 and T 75-100, as they presented the lowest MIC100 values, were selected to investigate their modulating potential in combination with Amphotericin B, Fluconazole and 5-Flucytosine, using the association method (checkerboard) by FICI (Fractional Inhibitory Concentration Index) and by mathematical models (Blis, HSA, Loewe and ZIP). All tested combinations were able to reduce the MIC100 for C. gattii and after data harmonization, the combinations between the T 50-75 and T 75-100 fractions with Amphotericin B and Fluconazole were classified as synergistic, however the combinations with the fraction T 50-75 showed greater sensitivity (93%) and degree of synergistic intensity. The combination of the fraction T 50-75 with the drugs caused greater interference in the growth curve, melanization and a significant reduction in capsule size. However, exposure of C. gattii to treatment with only the T 50-75 fraction also caused morphological changes in the cell wall and capsule, indicating that the fraction contributed strongly to the result observed in combination with the drug. Therefore, the T 50-75 fraction presents itself as a promising candidate for the development of adjuvants and/or new drugs for antifungal chemotherapy. Analysis of the results suggests that the fungistatic action of the T 50-75 fraction can contribute to minimizing the emergence of resistance in the pathogen, acting in redundant pathways and reducing the effective concentrations of clinical reference drugs, thus improving treatment and minimizing toxicity generated.
8	JOHN WESLLEY LIRA SANTOS Effect of simulated microgravity using clinostat 2D in Lemna aequinoctialis Welw. Advisor : KATIA CASTANHO SCORTECCI COMMITTEE MEMBERS : KATIA CASTANHO SCORTECCI RAQUEL CORDEIRO THEODORO TERCILIO CALSA JUNIOR Data: Jun 21, 2024 Show Abstract Understanding the effects of microgravity on plants is still limited, it has been observed that the response changes according to tissue, plant and microgravity conditions. This study evaluated the effects of simulated microgravity on Lemna aequinoctialis, a plant from the subfamily Lemnoideae, important for bioremediation and animal feeding. Using a 2D clinostat machine to simulate microgravity, the plants were exposed to different rotations (15, 30, 45 rpm) for 2 and 4 hours, along with a control (0 rpm). Plant development was monitored at 0, 5, and 10 days after treatment. Biochemical analyses were conducted to measure total antioxidant capacity (CAT), reducing power, phenolic compounds, protein content, and catalase enzyme activity. The results showed that simulated microgravity significantly increased antioxidant markers and protein content. It has been observed that CAT and reducing power increased up to three times, in special after 4 hours of treatment. The 45 rpm rotation for 2 hours resulted it was verified an increase in phenolic compounds on days 5 and 10, while for the 30 rpm rotation for 4 hours treatment is as verified an important increase on day 0. For the catalase activity, it was higher with 30 rpm in both time treatment on day 10. In addition, protein content increased up to seven times with 45 rpm for 2 hours. These results suggest that simulated microgravity can enhance the nutritional profile of L. aequinoctialis, boosting its biotechnological applications in sustainable food production and bioremediation. This study advances the understanding of the mechanisms involved in L. aequinoctialis response to microgravity.
9	DYONATAN FONSECA SILVA EVALUATION OF THE METALLOPROTEINASE PROFILE IN TISSUE SAMPLES FROM ANIMALS EXPERIMENTALLY INFECTED WITH Trypanosoma cruzi Advisor : MARCELO DE SOUSA DA SILVA COMMITTEE MEMBERS : MARCELO DE SOUSA DA SILVA ALINE RIMOLDI RIBEIRO CLÁUDIA JASSICA GONÇALVES MORENO TAFFAREL MELO TORRES Data: Jun 26, 2024 Show Abstract Chagas disease (DCh) is caused by the parasite Trypanosoma cruzi (T. cruzi) and is characterized by two clinical phases: acute and chronic. During the acute phase, the parasite targets specific tissues and dysregulates cytokines and matrix metalloproteinases (MMPs), which are major risk factors for the development of cardiopathies, common complications in both the acute and chronic phases of the disease. This study aimed to optimize gel-based methods, such as SDS-PAGE and zymography, to evaluate whether T. cruzi infection could alter the metalloproteinase profile of key organs during the acute phase of DCh. For this purpose, total protein extract from epimastigote forms of T. cruzi Y strain from axenic culture was obtained, and experimental infection with blood trypomastigote forms was performed in female mice, divided into three groups: uninfected (CN, negative control, n=3) and infected, euthanized at 17 (D17, n=3) and 32 (D32, n=3) days post-infection. Control animals were used as a baseline for comparison with infected animals. Heart, spleen, liver, and blood from all groups were collected to produce total extract and blood plasma. Optimization allowed the determination of the protein profile of T. cruzi, revealing bands with approximate molecular weights of 95, 85, 63, 57, 48-52, 43-45, 40-41, 29, and 18-20 kDa, which were discussed in terms of their possible identification and function in the infection. The results also demonstrated the protein profiles of cardiac, splenic, hepatic tissues, and plasma in the three analyzed groups. The T. cruzi zymogram revealed three activity bands with approximate molecular weights of 66 kDa, 55 kDa, and 35 kDa, along with a faint activity band between 66 and 35 kDa. In the infected animal group, clinical signs of hepatomegaly and splenomegaly characteristic of the acute phase of DCh were observed. Several gelatin-degrading metalloproteinases were detected in the zymogram of tissues and plasma, especially in splenic tissue. Differences in their expression during acute infection were observed only in splenic and cardiac tissues, with the alteration in cardiac tissue related to the increased expression of a 93 kDa protease, likely proMMP-9, and in the spleen, a 146 kDa unknown protease. In summary, the findings provide information on the protein and proteolytic expression patterns for T. cruzi and the analyzed organs plus plasma, revealing abundant proteins as potential immunogenic targets against T. cruzi. These data serve for comparison of the genetic and functional diversity of the parasite along with other studies, besides confirming changes in protease expression during the acute phase of DCh, opening avenues for diagnostics and treatment based on the necessity and feasibility of inhibiting these proteins.
10	PABLO FELIPE FERREIRA FARIAS SULFATED POLYSACCHARIDES FROM THE SEAWEED CAULERPA CUPRESSOIDES VAR. FLABELLATA WITH ANTIOXIDANT ACTIVITY: EVALUATION OF BIOSAFETY IN NEURO-2A CELLS AND ZEBRAFISH MODELS Advisor : SUSANA MARGARIDA GOMES MOREIRA COMMITTEE MEMBERS : SUSANA MARGARIDA GOMES MOREIRA KATIA CASTANHO SCORTECCI RAFAEL BARROS GOMES DA CAMARA DOUGLAS DOURADO OLIVEIRA Data: Sep 25, 2024 Show Abstract Neurodegenerative diseases (NDs), such as Alzheimer's and Parkinson's, are global health issues affecting millions of people. Oxidative stress is believed to be a key factor in the cause and/or progression of NDs, damaging neuronal cells and leading to symptoms such as memory loss and motor problems. Therefore, searching for new antioxidant agents to mitigate those damages is crucial. Among the potential antioxidant agents, sulfated polysaccharides (SPs) obtained from seaweeds stand out. Beyond their antioxidant properties, SPs also exhibit various bioactivity properties, including anticoagulant, antiproliferative, and anti-inflammatory effects. Thus, this study evaluated the antioxidant potential of SP-rich fractions of the green seaweed Caulerpa cupressoides var. flabellata against H2O2 in murine neuroblastoma cells (Neuro-2A). Four SP-rich samples (CCB-F0.3, CCB-F0.5, CCB-F1.0, and CCB-F2.0) were obtained from polysaccharide-crude extract of Caulerpa cupressoides (PCE). The cytotoxicity and protective effect of the samples against H2O2 in Neuro-2A cells were evaluated, using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) reduction method. No cytotoxic effects were observed, except for the PCE and CCB-F0.5 samples at higher concentrations. The CCB-F2.0 sample presented antioxidant potential and protective effects against H2O2 under the tested conditions. Additionally, the CCB-F2.0 sample reduced the reactive oxygen species (ROS), evaluated by the 2,7-dichlorofluorescein diacetate (DCFH-DA) fluorescent probe. This sample did not exhibit genotoxicity or mutagenicity, as determined by single-cell gel electrophoresis assays (comet assay) and the Salmonella microsuspension reversion assay (Kado test), respectively. Furthermore, embryotoxicity and optomotor response were evaluated using the zebrafish model. Thus, CCB-F2.0 sample showed no embryotoxic or neurotoxic effects in the zebrafish embryos at different concentrations, since no changes in mortality, malformations, and optomotor response were observed in the larvae. Therefore, it is expected that the results of this study may contribute to the development of an alternative treatment for NDs.
11	GABRIEL CHRISTIAN DE FARIAS MORAIS ADMET and Quantum Biochemistry Predictions of Bioactives with Antimicrobial or Neuroactive Potential Advisor : JONAS IVAN NOBRE OLIVEIRA COMMITTEE MEMBERS : JONAS IVAN NOBRE OLIVEIRA EDILSON DANTAS DA SILVA JUNIOR JÉSSICA DE FÁTIMA VIANNA Data: Nov 26, 2024 Show Abstract The increasing demand for novel neuroactive and antimicrobial drugs motivated this study to conduct ADMET (Absorption, Distribution, Metabolism, Excretion, and Toxicity) predictions and quantum biochemical analyses of bioactive compounds with eugeroic (modafinil), noradrenergic (atomoxetine), antiviral (tecovirimat), and antiparasitic (BZTS) activities. Advanced computational tools such as ADMETlab 2.0, admetSAR, FAFDrugs4, MolInspiration, SwissADME, ADMET-AI, pkCSM, and PRED-HERG were utilized to evaluate the pharmacokinetic and toxicological profiles of Modafinil, Atomoxetine, and Tecovirimat, identifying risks such as hepatotoxicity, cardiotoxicity, and mutagenic potential. The focus then shifted to Chagas disease, selecting six compounds with antichagasic potential, with BZTS emerging as a standout due to its favorable ADMET profile and low toxicity risk, meeting all medicinal chemistry guidelines. Molecular modeling studies and Density Functional Theory (DFT) calculations revealed robust and specific interactions between BZTS and cruzain, a key Trypanosoma cruzi enzyme, indicating stability and high affinity within the ligand-receptor complex through interactions with amino acids GLU208 (-10.2 kcal/mol), MET68 (-4.75 kcal/mol), LEU67 (-4.08 kcal/mol), ASN69 (-3.68 kcal/mol), LEU160 (-3.38 kcal/mol), ALA138 (-1.98 kcal/mol), GLU117 (-1.97 kcal/mol), and ASP161 (-1.68 kcal/mol). Additionally, frontier orbital analyses (HOMO -5.85 eV, LUMO -3.37 eV) and quantum chemical descriptors—ionization potential (5.85 eV), electron affinity (3.37 eV), chemical hardness (1.24 eV), softness (0.81 eV), chemical potential (-4.61 eV), electronegativity (4.61 eV), and electrophilicity (13.19 eV)—confirmed the BZTS compound’s potential for effective biological target interactions, reinforcing its therapeutic potential. This study underscores the effectiveness of in silico methodologies in identifying and characterizing promising bioactive compounds, with BZTS emerging as a promising candidate for Chagas disease treatment. Integrating computational predictions in drug development proves crucial for accelerating the discovery and optimization of safer, more effective drugs, significantly contributing to advances in pharmacology and public health.
	Thesis
1	LÍDIA LEONIZE RODRIGUES MATIAS New alternatives for treatment against bacterial infections: evaluation of the potential of Trypsin Inhibitor isolated from Tamarind seed (TTI) as an anti-infective agent Advisor : ANA HELONEIDA DE ARAUJO MORAIS COMMITTEE MEMBERS : ANA HELONEIDA DE ARAUJO MORAIS FRANCISCO CANINDE DE SOUSA JUNIOR KARLA SUZANNE FLORENTINO DA SILVA CHAVES DAMASCENO MAYARA SANTA ROSA LIMA PATRICIA SANTOS LOPES RICHELE JANAINA ARAUJO MACHADO Data: Feb 23, 2024 Show Abstract Infections have become a worrying threat to public health, especially when caused by bacteria resistant to currently available antimicrobials. This thesis aimed to study new alternatives for treatments against bacterial infections. For this purpose, a narrative review and an in vitro and in vivo study were performed. Thus, the first chapter of this thesis presents a narrative review of new biomedical alternatives in antibacterial treatments. As a result, it was found that some areas of research aiming to control bacterial infections have been highlighted, namely bioinformatics, nanotechnology, and genomics, in addition to studies developed with natural molecules such as antimicrobial peptides. Furthermore, the articles in this narrative review presented promising research in developing antimicrobial agents, and their mechanisms of action and strategies to minimize the consequences of bacterial resistance. The review will present current alternatives may contribute to future studies in the area. In the second chapter, the effect of trypsin inhibitor isolated from tamarind seeds (TTI) on the survival of Caenorhabditis elegans (C. elegans) (wild strain) under conditions of bacterial infection was investigated. The TTI was isolated in Trypsin-Sepharose CNBr-4B affinity chromatography. Its protein quantification was performed using the Bradford method, and an antitryptic assay monitored its specific activity. The animals were maintained in Nematode Growth Medium (NGM) and Escherichia coli bacteria (OP50) as a food source at 20 ⸰C and, for the analysescarried out, they were previously exposed to different concentrations of TTI, from stage L1 to L4, in addition to a group control without treatment. Besides, the toxicity test was performed by evaluatingoviposition, progeny, locomotion, and body size. Subsequently, the bacterial infection test was carried out with Staphylococcus aureus in C. elegans previously subjected to concentrations of 0.1 and 1.0 mg/mL of TTI. Nematodes were exposed to the stressor tert-butyl hydroperoxide (TBOOH) in an oxidative stress test. Then, bacterial growth curve and antioxidant and antibacterial activity tests were carried out in vitro. In the toxicity test, no statistical difference was found between the control group and the other concentrations tested (p> 0.05). Regarding body size, the tested TTI concentrations showed a statistical difference (p<0.05) and presented higher length than the untreated group. Regarding bacterial infection analyses, animals subjected to 0.1 mg/mL and 1.0 mg/mL of TTI showed survival of 16.10% and 30.32%, respectively, higher than the group infected without treatment (8.08%). To understand the role of TTI on animal survival, in the oxidative stress test, at all tested TTI concentrations, greater survival was observed about the control at 12h and 18h (p< 0.05). In in vitro studies about the bacterial growth curve, no inhibitory activity of TTI was observed about Escherichia coli (OP50) (p>0.05) at the concentrations tested, nor was antibacterial and antioxidant activity observed. Given the results, a greater survival of C. elegans was found when previously exposed to TTI; however, in a different antioxidant and antibacterial pathway. In this way, the study contributed to consolidating theoretical and scientific understanding through narrative review and ITT tests, respectively, on new alternatives for controlling bacterial infection.
2	MARYELLE DE CÁSSIA ALBINO Effects of sucrose and ethanol withdrawal on behaviors related toanxiety and depression: investigation of the role of the KynureninePathway Advisor : VANESSA DE PAULA SOARES RACHETTI COMMITTEE MEMBERS : VANESSA DE PAULA SOARES RACHETTI EDILSON DANTAS DA SILVA JUNIOR ELAINE CRISTINA GAVIOLI ISABELLA MARIA DE OLIVEIRA PONTES FERNANDES JANAINA MENEZES ZANOVELI Data: Feb 27, 2024 Show Abstract In disorders related to substance use/abuse, the withdrawal phase favors negative reinforcement and continued consumption despite physical and mental harm. Women constitute a risk group for these disorders and few studies investigate changes in neural substrates in females. The enzyme indoleamine 2,3-dioxygenase (IDO), responsible for the metabolism of tryptophan into kynurenines, was seen hyperactivated in males undergoing alcohol withdrawal and associated with emotional disorders. Thus, the present work investigated the effects of sucrose and ethanol withdrawal on behaviors related to anxiety and depression and on IDO activity in brain areas that participate in the modulation of these emotions in Wistar rats. In experiment 1, the rats were subjected to a liquid sucrose diet for 16 days and to behavioral tests on days 3, 5, 22 and 24 after sucrose withdrawal. In experiment 2, females were subjected to an exclusive liquid ethanol diet in increasing concentrations (2, 4 and 6%) for 21 days and to a battery ofbehavioral tests on days 22, 23, 24, 25 and 26 after withdrawal of ethanol. The rats that received ethanol were treated with inulin (10 or 30 mg/kg orally, gavage) or mineral water. After the behavioral assessment, the animals were euthanized and had blood and frontal cortex, striatum and hippocampus samples collected. In rats exposed to sucrose there was a reduction in immobility time in the forced swimming test and an increase in plasma triglyceride concentration. No biochemical and behavioral changes were seen in animals on sucrose withdrawal. Rats withdrawing from ethanol exhibited less exploration of open arms in the elevated plus maze, greater grooming in the sucrose spray test, greater burying of glass balls in the marble burying test and greater immobility time in the forced swimming test. There was a trend towards an increase in kynurenine concentration in the cortex. Inulin at doses of 10 and 30 mg/Kg attenuated this increase, and the dose of 30 mg/Kg mitigated the behavioral effects on animals in the elevated plus maze, sucrose spray, marble burying and forced swimming tests. In conclusion, females were not sensitive to sucrose withdrawal. Prolonged consumption of sucrose has demonstrated an antidepressant effect. Ethanol withdrawal promoted anxious and depressive-like behaviors and increased the activity of the kynurenine pathway in females, such changes were reversed by treatment with inulin. The prebiotic demonstrated a potential candidate for the treatment of alcohol use disorder.
3	MYCHELLE KYTCHIA RODRIGUES NUNES DUARTE NUTRIGENETIC TESTS: POSSIBLE APPLICATIONS IN THE PREVENTION AND TRE-ATMENT OF OBESITY Advisor : LUCYMARA FASSARELLA AGNEZ LIMA COMMITTEE MEMBERS : LUCYMARA FASSARELLA AGNEZ LIMA HUGO ALEXANDRE DE OLIVEIRA ROCHA VIVIANE SOUZA DO AMARAL ANNETE BRESSAN RENTE FERREIRA MARUM TATIANE MIEKO DE MENESES FUJII Data: Jun 26, 2024 Show Abstract Obesity is a complex disease and mostly polygenic disease. In this context, several trea-tments are listed to mitigate it and nutritional genomics has been gaining prominence withpossible tools to assist in the prevention and/or treatment of obesity with nutrigenetic testing(NT) . Therefore, this thesis is divided into two chapters, aiming to evaluate whether NT canbe used to prevent and/or treat obesity. In the first chapter, a narrative review was carriedout to understand the current panorama of obesity and NT, covering the themes of precisionnutrition, exposome, genetic testing sold directly to the consumer. The benefits and nega-tive aspects of using NT as a tool to help prevent or treat overweight and obesity wereanalyzed. Some entities, such as the Brazilian Association for the Study of Obesity andMetabolic Syndrome (ABESO) are against the use of NT for obesity. The Academy of Nu-trition and Dietetics recommends its use, however not in isolation, but within a context ofwell-conducted anamnesis. Therefore, NT should not be used alone in the treatment ofobesity, but always associated with a good anamnesis for the individualized treatment ofobesity or at a public health level. In the second chapter, a retrospective study was carriedout, evaluating the relationship of 29 candidate gene SNP related to obesity. Thus, 112individuals were divided into two groups: 64 eutrophic (BMI< 25 kg/m²) and 48 overweightand obese (BMI≥ 25 g/m²). It was verified that the population was in Hardy-Weinberg equi-librium and a multivariate logistic analysis was performed to verify whether the SNP wereassociated with overweight and obesity. In this analysis, it was found that of the 29 SNPfound in NT marketed in Brazil, only ADIPOQ rs17300539, PPARG rs1801282 were asso-ciated with overweight and obesity, while SOD rs4880 was related to protection againstoverweight and obesity. Thus, it was shown that more studies are needed to validate whichSNP are related to overweight and obesity in the Brazilian population. Furthermore, lifestyleanalysis must be added, as these factors can epigenetically alter gene expression and im-pact the phenotype.
4	JOELTON IGOR OLIVEIRA DA CRUZ ANTIBACTERIAL, ANTIBIOTIC MODULATING AND ANTIPROTEASIC ACTIONS OF PROTEIN FRACTIONS FROM CATINGUEIRA SEEDS (Poincianella pyramidalis Tul.) Advisor : ELIZEU ANTUNES DOS SANTOS COMMITTEE MEMBERS : ELIZEU ANTUNES DOS SANTOS RAFAEL BARROS GOMES DA CAMARA BRUNO OLIVEIRA DE VERAS MARIA TATIANA ALVES OLIVEIRA PAULA IVANI MEDEIROS DOS SANTOS Data: Aug 30, 2024 Show Abstract Bacterial resistance, as well as inflammatory disorders, are significant public health problems that take thousands of people to hospitals every year, often requiring several days of hospitalization for treatment, generating excessive costs and, sometimes, without solving the problem. Considering the importance of discovering new substances that mitigate these problems, this study aimed to evaluate the crude extract and protein fractions of the seed of Poincianella pyramidalis (Tul.) for antibacterial, antibiotic-modulating, and protease-inhibiting activities. The protein extracts were obtained from extraction with 0.05M Tris-HCL buffer pH 7.5 and subsequent fractionation with ammonium sulfate in different saturation ranges 0-30%, 30-60% and 60-90% of sodium sulfate ammonium. The electrophoretic profile revealed several protein bands between 12 KDa and 225 KDa. The inhibitor-enriched fraction (FE) was obtained using only a Trypsin-Sepharose affinity chromatography after extraction. The antimicrobial activity test was performed through the microdilution assay and the modulating action of the antibiotics ampicillin, norfloxacin and gentamicin, using a subinhibitory concentration MIC/8. Although the samples did not show direct antimicrobial action against the ATCC strains Escherichia coli, Staphylococcus aureus and methicillin-resistant S. aureus (MRSA), it was observed that, at a concentration of 128 µg/mL, the samples associated with the different antibiotics (aminoglycoside, β-lactam and fluoroquinolone) had a synergistic action against the multiresistant strains of E. coli, S. aureus and Pseudomonas aeruginosa. While the other samples inhibited the action of the serine proteases tested in percentages above 80%, FE reduced 50% of the activities of the enzymes trypsin, human neutrophil elastase (HNE) and chymotrypsin, at concentrations of 0.017, 0.023 and 0.156 µg/µL, respectively, and its antitrypsin activity was maintained at high temperatures (80 ºC) and a wide pH range (2 to 12). No hemolytic activity was observed at concentrations up to 10x higher than the IC50, thus enabling its use in possible pharmacological preparations. In this way, we were able to separate, in a relatively simple way, products derived from P. pyramidalis seeds with relevant activities in the clinical context, considering their potential application in combating infections caused by resistant bacteria, as well as in inflammatory protease disorders, and with reduced treatment costs.
5	DANIEL MELO DE OLIVEIRA CAMPOS Development of Multi-Epitope Vaccine and Potential Inhibitors of Sars-Cov-2 Mpro Protease: Reverse Vaccinology and Computational Chemistry Approaches Advisor : JONAS IVAN NOBRE OLIVEIRA COMMITTEE MEMBERS : JONAS IVAN NOBRE OLIVEIRA UMBERTO LAINO FULCO JOAO FIRMINO RODRIGUES NETO JÉSSICA DE FÁTIMA VIANNA KATYANNA SALES BEZERRA Data: Sep 9, 2024 Show Abstract Infections caused by SARS-CoV-2 have posed a significant threat to global public health and continue to be a relevant and complex issue. This thesis aimed to explore new approaches to combating the virus, focusing on the development of a multi-epitope vaccine and the identification of potential Mpro protease inhibitors. Two in silico studies were conducted: one dedicated to the design of a multi-epitope vaccine using reverse vaccinology, and the other focused on developing non-covalent inhibitors through computational chemistry. Additionally, the thesis includes a critical review of emerging antiviral strategies. The first chapter of this thesis presents a critical commentary on promising targets and pharmacological strategies adopted during the pandemic. Among the main targets were Mpro, the S glycoprotein, and TMPRSS2. It concludes that, despite advancements, definitive evidence on the efficacy of these treatments was still necessary. Meanwhile, repurposing existing drugs and using convalescent plasma remained the best alternatives until an effective vaccine was developed. In the second chapter, we conducted an in silico study to develop a new multi-epitope vaccine against SARS-CoV-2, including the Alpha, Beta, Gamma, Delta, and Omicron variants. We used an immunoinformatics approach to identify the best immunogenic epitopes from the four structural proteins of the virus (S, M, N, and E) from 475 genomes sequenced from regions with high disease incidence. The vaccine was modeled, refined, validated, and its molecular docking with the TLR3 receptor was evaluated. The results suggest that the candidate vaccine increases antibody response and should be tested in clinical trials. In the third chapter, we evaluated and compared two potential non-covalent inhibitors of SARS-CoV-2 Mpro, WU-04 and ML188, using computational chemistry approaches, including quantum calculations and ADMET analysis. Our results revealed that residues Met165, Asn142, Glu166, His41, and Leu141 are critical as they interact with high affinity with both inhibitors, suggesting that these residues are essential for complex stabilization. The total energy calculated for the Mpro-WU-04 complex was -52.21 kcal/mol, while for Mpro-ML188, it was -40.47 kcal/mol, indicating that the WU-04 inhibitor has greater energetic affinity.
6	CYNTHIA HAYNARA FERREIRA DA SILVA Development of Blends Containing Fucoidan and Evaluation of Antioxidant Activity In Vitro and In Vivo in Caenorhabditis elegans and Zebrafish (Danio rerio) Models Advisor : HUGO ALEXANDRE DE OLIVEIRA ROCHA COMMITTEE MEMBERS : DAYANNE LOPES GOMES HUGO ALEXANDRE DE OLIVEIRA ROCHA LEANDRO SILVA COSTA MARÍLIA MEDEIROS FERNANDES DE NEGREIROS ÉDER GALINARI FERREIRA Data: Sep 30, 2024 Show Abstract Oxidative stress is a crucial physiological process involved in regulating and maintaining several fundamental processes in organisms, as well as in diseases such as cardiovascular diseases, neurodegenerative diseases, and cancer. There is growing interest in identifying new molecules, particularly from natural sources, that possess antioxidant properties. Marine macroalgae have emerged as functional foods and sources of active molecules with various properties, including sulfated polysaccharides (PS). One example is PS fucoidan A (FucA), extracted from the algae Spatoglossum schroederi, which has been studied for its antithrombotic, anti-inflammatory, and non-genotoxic potential. However, FucA does not exhibit significant antioxidant activity. The presence of antioxidant properties could enhance the effects of the biological activities already identified. One strategy to enhance the effects of a molecule is through chemical modification by adding functional groups and developing blends that combine the properties of two components. To improve the antioxidant activity of FucA, factorial design and response surface plots were employed to develop blends containing FucA and dextran modified with gallic acid (Dex-Gal). Chemical and structural analyses confirmed that Dex-Gal was modified with the addition of 3% GA and exhibited a total antioxidant capacity (TAC) 3.2 times higher than that of unmodified Dex. Dex-Gal and FucA were subjected to factorial design to create different proportions and develop five blends (BLD1, BLD2, BLD3, BLD4, and BLD5). Two blends, BLD1 and BLD5, demonstrated low activity in the antioxidant tests performed. In the TAC test, three blends stood out: BLD4 (22 Eq. AA), BLD3 (17 Eq. AA), and BLD2 (13 Eq. AA). In the reducing power test, BLD4 showed a reduction of 28%, compared to 17% for BLD2 and 15% for BLD3. Notably, BLD4 also excelled in the hydroxyl radical scavenging test, demonstrating approximately 100% activity. Analysis of the surface graphs confirmed that the ideal proportion of the FucA and Dex-Gal blend is that formulated in BLD4, with a 1:1 ratio of each component. BLD4 was selected for antioxidant tests in the 3T3 cell line, as well as in biological models of Caenorhabditis elegans and zebrafish (Danio rerio). BLD4 did not exhibit cytotoxicity in 3T3 cells and provided protective effects against oxidative stress induced by H2O2; it enhanced the survival of C. elegans larvae under oxidative stress with T-BOOH and reduced intracellular ROS by 30%; it also protected zebrafish embryos from oxidative stress caused by H2O2. In both cellular and C. elegans models, BLD4 performed better compared to its isolated components.
7	ELLYNES AMANCIO CORREIA NUNES PURIFICATION, CHARACTERIZATION AND BACTERIOSTATIC ACTIVITY OF A MANGANESE-DEPENDENT LECTIN FROM THE VENOM OF Bothrops moojeni Advisor : LUDOVICO MIGLIOLO COMMITTEE MEMBERS : LUDOVICO MIGLIOLO ADRIANA FERREIRA UCHOA ELIZEU ANTUNES DOS SANTOS JULIANA PAVAN ZULIANI OCTÁVIO LUIZ FRANCO Data: Oct 15, 2024 Show Abstract Bioactive molecules isolated from venomous animals constitute a variety of components with biological activities that can be directed to the treatment of different diseases. Snake venoms are composed of approximately 90% proteins and peptides. These toxins perform different biochemical mechanisms and are therefore responsible for the clinical response observed in snakebites. Knowledge of snake proteins allows the study of heterologous activities related to these toxins, such as antibacterial and antitumor activity. In this context, the objective of this study was to conduct a bibliographic review on the protein group of lectins isolated from snake venoms and to isolate, characterize and understand the biological activities of a lectin from the venom of the snake Bothrops moojeni. A literature review was conducted on lectins isolated and characterized from snake venoms, focusing on their main characteristics and classification, with an emphasis on studies from the last decade. It was found that, to date, snake venom lectins are predominantly classified as C-type lectins and C-type lectin-like proteins, exhibiting diverse biological activities. These include potential applications in the treatment of neoplastic and hematologic diseases, as well as demonstrated antibacterial properties. In addition, the identification of a protein with lectin activity was performed using Size Exclusion Chromatography (SEM) and Reverse-Phase High Performance Liquid Chromatography (RP-HPLC). The following procedures included analysis by Polyacrylamide Gel (SDS-PAGE), as well as hemagglutination assays, inhibition of hemagglutinating activity, and evaluation of ion, pH and temperature dependence. Additionally, hemolytic activity, antibacterial activity against the bacterial strains Acinetobacter baumannii and Staphylococcus aureus, and antibiofilm activity was evaluated. Molecular exclusion generated a chromatographic profile with 6 peaks that were analyzed by polyacrylamide gel electrophoresis and subjected to hemagglutination assays. Hemagglutinating activity was observed in fractions 5 and 6, with the best result for fraction 5, which was named BmoojL. It was also confirmed that BmoojL is manganese-dependent. Furthermore, BmoojL was inhibited in the presence of the carbohydrate’s glucose, fructose, mannose, fucose and N-acetylgalactosamine, and was active at temperatures up to 80 °C and in pH ranges from 5 to 9. The hemolytic assay showed that the molecule did not promote hemagglutination at the concentrations tested (31.25 - 500 µg. mL-1), with a hemolysis percentage below 10%. The antibacterial assays showed that despite not reaching the minimum inhibitory concentration, BmoojL reduced the bacterial growth of A. baumannii from the concentration of 64 µg. mL-1. The antibiofilm assay showed that for S. aureus, BmoojL was effective, considerably reducing biofilm growth at all concentrations tested (16 - 512 µg. mL-1) by approximately 50%. It was also shown that the ion, in isolation and associated with another protein, did not perform significant activity, as observed with BmoojL. Furthermore, it was found that BmoojL inhibited bacterial growth, indicating that the molecule acts bacteriostatically. Thus, the study allowed the isolation of a molecule with manganese-dependent lectin activity with the potential to exert antibacterial and antibiofilm activity, contributing to the research of bioactive molecules derived from venomous snakes. Although the literature recommends some studies involving such molecules and their heterologous activity, when compared to other molecular groups, the study involving lectins from venomous animals remain an underexplored area of research. These findings show what there is still to know about these molecules and to direct and propose new activities that may add to biotechnological research involving these animal groups.
8	GEOVANNA MARIA DE MEDEIROS MOURA Unveiling the Antibiotic Modulating and Antioxidant Properties of the Wild Mushroom Langermannia bicolor Advisor : ELIZEU ANTUNES DOS SANTOS COMMITTEE MEMBERS : ELIZEU ANTUNES DOS SANTOS BRUNO OLIVEIRA DE VERAS MARIA TATIANA ALVES OLIVEIRA PAULA IVANI MEDEIROS DOS SANTOS WESLLEY DE SOUZA PAIVA Data: Dec 4, 2024 Show Abstract The emergence of multidrug-resistant bacteria has driven the search for new therapeutic agents and adjuvants. In this study, we investigated the properties of an extract from the mushroom Langermannia bicolor as a modulator of antibiotics (gentamicin, norfloxacin, and ampicillin) against multidrug-resistant strains of Staphylococcus aureus, Escherichia coli, and Pseudomonas aeruginosa, as well as its antioxidant activity and toxicity. The crude aqueous extract of L. bicolor was fractionated using increasing volumes of acetone, resulting in three fractions (F0.5, F1, and F2), which exhibited different biochemical profiles. Tests with human erythrocytes revealed that the extract and its fractions did not exhibit hemolytic activity, while MTT reduction assays in fibroblast cells (NHI/3T3) showed that the crude extract reduced viability by 50%. In vivo tests with zebrafish embryos, up to young larvae (Danio rerio), confirmed the absence of toxicity of the fractions through embryotoxic analyses, heart rate measurements, and behavioral tests. Although none of the fractions demonstrated direct antibacterial activity, all exhibited significant modulatory effects when combined with conventional antibiotics against the tested multidrug-resistant bacteria. Notably, fraction F2, rich in proteins with higher proteolytic and hemagglutinating activities, reduced the Minimum Inhibitory Concentration (MIC) of gentamicin across all tested strains. The fractions also displayed potent antioxidant properties both in vitro and in vivo. Fraction F0.5, enriched with phenolic compounds and flavonoids, exhibited the broadest spectrum of antioxidant activities, including metal chelation (Fe²⁺ and Cu²⁺), free radical scavenging (•OH and O₂⁻•), and both concurrent and preventive cellular protection against hydrogen peroxide (H₂O₂) induced oxidative stress in 3T3 cells. Meanwhile, fraction F1, which was carbohydrate-rich, was able to repair oxidative damage following H₂O₂ stress. In vivo, all fractions protected zebrafish embryos exposed to H₂O₂. When quantifying reactive oxygen species (ROS) using a fluorescent probe (DFCH), zebrafish larvae treated with F0.5 followed by H₂O₂ significantly reduced ROS production in their cells, yielding values even lower than those of larvae not exposed to the oxidizing agent. To a lesser extent, F1 and F2 also protected zebrafish cells by reducing ROS production. By pioneering the therapeutic potential of L. bicolor, our study paves the way for future investigations into the bioactive composition of this mushroom and its respective molecular mechanisms involved in the observed properties: antibiotic potentiation and protection against oxidative stress.
9	ALESSANDRA MARINHO MIRANDA LUCENA ANTIOXIDANT ACTION AND NEUROPROTECTIVE EFFECT OF SULFATED POLYSACCHARIDES Advisor : RAFAEL BARROS GOMES DA CAMARA COMMITTEE MEMBERS : RAFAEL BARROS GOMES DA CAMARA HUGO ALEXANDRE DE OLIVEIRA ROCHA KATIA CASTANHO SCORTECCI FAUSTO PIERDONA GUZEN PABLO DE CASTRO SANTOS ÉDER GALINARI FERREIRA Data: Dec 4, 2024 Show Abstract Neurodegenerative diseases are the result of multifactorial biological mechanisms, involving both exogenous and endogenous causes. Due to the increase in life expectancy and the predicted growth of dementia conditions, there is a growing concern in the search for more effective prevention and treatment for neurodegeneration. Oxidative stress has been shown to be an important factor in the development and progression of these diseases, due to intrinsic characteristics of nervous tissue (e.g., low regenerative capacity and high energy demand). Seaweeds are sources of various bioactive molecules, especially sulfated polysaccharides (SP). These compounds are important for the maintenance and survival of sea weeds and have significant biological activities, such as antioxidant and anti-inflammatory. These properties can be useful in the treatment of neurodegenerative diseases. Seaweeds SP have shown promise as an alternative for neuroprotection. Nine species of seaweed have been identified whose SPs exhibit antioxidant action under different conditions. Forty-four sulfated polysaccharides rich fractions (PRF) were obtained. These PRF were evaluated for their cytotoxicity in Neuro-2A cells at different concentrations (0.1, 0.25, 0.5, or 1 mg/mL). From each PRF, two concentrations were selected, one higher and one lower, which showed MTT reduction values equal to or greater than 80%. They were analyzed for their ability to protect against oxidative damage caused by hydrogen peroxide. To select the PRF that would be evaluated for the possible protection mechanisms employed, the results of yield parameters, cytotoxic effect, and neuroprotective potential in cell culture were considered to construct a Score. The possible mechanisms involved were analyzed by determining the hydrogen peroxide scavenging potential and the effect on the expression of genes in the antioxidant pathway (Keap1, Nrf2, Sod1, Cat, Gpx, Hmox1, Gclc, Gclm) and the subunits of the NFκB transcription factor (Nfkb1 and Rela). Additionally, the antioxidant protection capacity was evaluated in an in vivo model (Zebrafish). Forty-two PRF did not show cytotoxic effects against Neuro-2A cells at at least one of the evaluated concentrations. All evaluated seaweeds had PRF that protected Neuro-2A cells from oxidative damage induced by hydrogen peroxide. Furthermore, through the Score, it was possible to observe that brown seaweed had the best results. Based on these data, four PRF (DJ 1.2, SF 0.5, SS 1.3, and DM 1.0) were selected for the evaluation of the possible mechanisms involved. The data obtained here demonstrate that the mechanisms used by these samples range from direct action on oxidant molecules to the modulation of endogenous antioxidant mechanisms, promoting an increase in the expression of genes related to the translation of antioxidant enzymes and a reduction in the expression of genes related to a pro-inflammatory and pro-oxidant response. Additionally, these polysaccharides were able to promote protection against oxidative stress in an in vivo model. These data are important for the development of future evaluations of these molecules, as they provided knowledge about their potential neuroprotective effect and antioxidant behavior in more complex organisms, thus guiding the possible mechanisms employed.
10	ISAIANE MEDEIROS STUDY OF THE EFFECT OF NATURAL BIOACTIVE PROTEINS AND PEPTIDES IN THE TREATMENT OF DIABETES MELLITUS Advisor : ANA HELONEIDA DE ARAUJO MORAIS COMMITTEE MEMBERS : ADRIANA AUGUSTO DE REZENDE ANA HELONEIDA DE ARAUJO MORAIS ANA LUISA PIRES MOREIRA AUGUSTO MONTEIRO DE SOUZA EDUARDO BORGES DE MELO Data: Dec 27, 2024 Show Abstract Diabetes mellitus (DM) is a public health problem characterized by hyperglycemia. The aim of this study was to evaluate the effect of natural bioactive proteins and peptides in the treatment of DM. Thus, this thesis is divided into three chapters. The first chapter describes the protocol for the systematic review (SR), which was registered in the International Register of Prospective Systematic Reviews (PROSPERO, number: CRD42022355540) and guided the construction of the SR presented in the second chapter. In the second chapter, the SR was conducted with the objective of answering the starting question: which peptides or proteins have been studied in silico for the treatment of diabetes mellitus? The articles were selected based on PECOs (Population, Exposure, and Context) from the databases PubMed, ScienceDirect, Scopus, Web of Science, Virtual Health Library, and EMBASE. Based on eligibility criteria, five articles were included, and the risk of bias was assessed using the adapted Strengthening the Reporting of Empirical Simulation Studies (STRESS) tool. The results showed that proteins and/or peptides extracted from natural sources interacted in silico with therapeutic targets involved in DM management, such as α-amylase, dipeptidyl peptidase IV (DPP-IV), α-glucosidase, insulin receptor (IR), glucose transporter type 2 (GLUT-2), and sodium-glucose co-transporter protein (SGLT1). When re-evaluated in vivo, these interactions promoted a reduction in fasting blood glucose, improvement in pancreatic morphology, positive regulation of insulin secretion and expression, reduction or maintenance of plasma insulin levels, a decrease in HOMA-IR, an increase in HOMA-β, and maintenance of GLP-1. This demonstrated that in silico studies provided crucial insights into therapeutic strategies for DM. In the third chapter, a preclinical study was conducted to evaluate the effect of trypsin inhibitor isolated from tamarind seed (ITT) in zebra fish with increased fasting blood glucose developed by overfeeding with Artemia sp., aiming to determine the mechanism of action of this inhibitor through glycation or insulin-like pathways. Biochemical parameters, relative expression of the IR gene, and morphological changes in organs and tissues relevant to type 2 diabetes (T2DM) and protein glycation (in vitro and in vivo) were evaluated. The diagnosis of T2DM was made using Accu-Chek® for fasting blood glucose measurement before treatment began. The animals (n = 140), of both sexes, were divided into four groups (n = 35): 1) healthy, untreated, and normo-fed animals, 2) T2DM animals without treatment and overfed, 3) T2DM animals treated with 25 mg of ITT/L and overfed, and 4) T2DM animals treated with 25 mg of ITT/L and normo-fed for 10 days. Fasting blood glucose was 62.33 mg/dL (2.52) for normo-fed animals and 104.70 mg/dL (4.16) for overfed animals pre-treatment (p = 0.008). After treatment, a significant reduction (p < 0.01) in fasting blood glucose, HOMA-IR, and QUICKI index, and a significant increase (p < 0.01) in HOMA-β were observed in the animals treated with ITT and overfeeding compared to the untreated T2DM group. However, these values showed no significant difference (p > 0.05) compared to the healthy animals, except for the QUICKI index. A significant increase (p < 0.01) in insulin was observed in all groups compared to the healthy control. No significant change (p > 0.05) in in vitro glycation of bovine serum albumin was observed for ITT concentrations of 1.4 and 5 mg/mL, while the concentration of 25 mg/mL of ITT significantly increased (p < 0.01). This finding did not persist when checking the formation of advanced glycation end products (AGEs) in vivo (p > 0.05) among the groups treated with 25 mg of ITT/L. Additionally, ITT promoted negative regulation of relative IR expression in adipose tissue and skeletal muscle. Histopathological findings showed reduced visceral adiposity, pancreatic and hepatic steatosis, and renal degeneration in animals treated with ITT and overfeeding. However, further studies are still needed to elucidate the mechanisms of fasting blood glucose reduction.

2023

	Dissertations
1	MARIA KAROLAYNNE DA SILVA Development of a Multi-Epitope Subunit Vaccine Against a Neglected Arbovirus of the Americas: an Immunoinformatics and Molecular Modeling Approach. Advisor : JONAS IVAN NOBRE OLIVEIRA COMMITTEE MEMBERS : JONAS IVAN NOBRE OLIVEIRA JOAO FIRMINO RODRIGUES NETO CLAUDIO BRUNO SILVA DE OLIVEIRA Data: Mar 30, 2023 Show Abstract The Mayaro virus (MAYV) is an emerging arbovirus in the Americas that can cause debilitating arthritogenic diseases. Although the biology of MAYV is not fully understood, it is known to be closely related to arthritogenic alphaviruses such as chikungunya, Ross River, and O’nyong nyong viruses. Effective control of the spread of these diseases requires identification of infected individuals and the development of preventive and prophylactic therapies. However, currently, the only available approach for controlling MAYV is vector control, as there are no licensed vaccines to prevent MAYV infection or therapies to treat it. In this study, we used immunoinformatics and molecular modeling approaches to identify potential T cell epitopes for vaccination against the Mayaro virus. To do this, we analyzed 127 MAYV genomes sequenced in the Americas (08 Bolivia, 72 Brazil, 04 French Guiana, 05 Haiti, 20 Peru, 04 Trinidad and Tobago, and 14 Venezuela) and identified short protein sequences that can bind to MHC class I and class II alleles. These promiscuous epitopes were selected based on their conservation and immunogenicity. Through immunoinformatics and molecular modeling analyses, we identified 56 potential MHCI/TCD8+ and 29 potential MHC-II/TCD4+ epitopes, with only specific protein sequences (nsP1191-199, nsP1501-509, nsP1498-506, nsP3348-356, nsP4305-314 and nsP4212-221(nsP157-71 , nsP116-30, nsP1182-196, nsP1180-194, nsP115-29, nsP2640-654, nsP2639-653, nsP2679-693, nsP2677-691 , nsP2680-694, nsP3127-141, nsP362-76, nsP4414-428, nsP4413-427, nsP4412-426 and nsP4372-386) TCD8+ (TCD4+) exhibiting high antigenicity, conservation, non-allergenicity, non-toxicity, and excellent population coverage. Based on these results, we developed a multiepitope vaccine coupled to the TLR3 receptor and improved its quality through quantum mechanical calculations. These results have important implications for advancing diagnosis, vaccine development, and immunotherapeutic interventions against the Mayaro virus.
2	RONALD MURYELLISON OLIVEIRA DA SILVA GOMES Application of group I introns from the mitogenome of Cryptococcus neoformans and Cryptococcus gattii for species identification and their association with antifungal susceptibility Advisor : RAQUEL CORDEIRO THEODORO COMMITTEE MEMBERS : RAQUEL CORDEIRO THEODORO SILVIA REGINA BATISTUZZO DE MEDEIROS VANIA SOUSA ANDRADE MARCUS DE MELO TEIXEIRA Data: Apr 17, 2023 Show Abstract The species complexes of Cryptococcus neoformans and Cryptococcus gattii are composed of pathogenic fungi that kill over 180,000 people annually worldwide, having meningoencephalitis as the main symptom of cryptococcosis. Characteristics such as virulence and antifungal susceptibility can vary within each species according to fungal genotype: C. neoformans is divided into molecular types VNI, VNII, VNIII, and VNIV, and C. gattii into VGI, VGII, VGIII, and VGIV. Therefore, specific molecular markers are necessary for differential diagnosis. Autocatalytic group I introns can be a potential target for distinguishing between molecular types of these yeasts, as they have polymorphisms regarding their presence and base pair sequences. Furthermore, since the auto-splicing of these elements is vital for the fungal cell, they are considered important therapeutic targets since they are absent from the human genome. Therefore, this study aimed to evaluate the presence of group I introns in the mitochondrial cob and cox1 genes of Cryptococcus fungal isolates, search for ORFs with endonuclease genes in intronic sequences, investigate the potential use of these genetic elements as molecular markers for differentiation of genotypes and/or species, and analyze their relationship with antifungal susceptibility andin addition to study their origin, distribution, and evolution through phylogenetic analyses. The data obtained from intron amplification of cob and cox1 genes showed that the C. neoformans complex has, on average, fewer introns in its mitogenome, and there is a great polymorphism of presence and size of these elements among and within genotypes, making it impossible to use a single intronic marker to differentiate genotype and/or species in the C. neoformans and C. gattii complexes. However, differentiation between species is possible with combinations of PCRs of mtLSU and cox1 introns for C. neoformans species and mtLSU and cob introns for C. gattii. Approximately 80.5% of the sequenced introns had homing endonucleases, and phylogenetic analyses showed that introns occupying the same insertion site form monophyletic clades and probably have a common ancestor that invaded this site before the species diverged. There was only one case of heterologous invasion, probably originating from horizontal transfer between a VGIV genotype isolate and the lichenized fungus Arthonia susa. Regarding susceptibility assays to antifungal agents, it was observed that the minimum inhibitory concentration (MIC) values of Fluconazole, 5-Flucytosine, and Pentamidine were statistically associated with species complexes, with C. gattii having higher MICs for Fluconazole and C. neoformans having higher MICs for 5-Flucytosine and Pentamidine. The presence of introns was related to susceptibility to Amphotericin B, for which a higher number of introns was associated with lower MIC values, and to 5-Flucytosine and Pentamidine, for which the influence of the presence of introns varied between complexes: regarding 5-Flucytosine, the presence of introns was related to lower susceptibility in C. neoformans and higher in C. gattii, and the opposite scenario was observed for Pentamidine, for which introns were associated with higher susceptibility in C. neoformans and lower in C. gattii.
3	MARIANNA BARROS SILVA Characterization of Chondroitin-4-Sulfate Isolated from Tilapia Viscera (Oreochromis Niloticus) and Its Effect on Modulating Calcium Oxalate Crystallization Advisor : RAFAEL BARROS GOMES DA CAMARA COMMITTEE MEMBERS : RAFAEL BARROS GOMES DA CAMARA MOACIR FERNANDES DE QUEIROZ NETO MOACIR FRANCO DE OLIVEIRA Data: May 15, 2023 Show Abstract Glycosaminoglycans (GAGs) are sulfated polysaccharides that are naturally present in urine and have been identified as modulators of urinary stone formation. Among GAGs, chondroitin sulfate (CS) has shown promise as a possible modulator of crystal growth. Studies have revealed that aquatic organisms are alternative sources of CS with therapeutic potential. The fish Oreochromis niloticus, also known as Nile tilapia, was chosen for this study due to its high production and potential for environmental damage. The objective of this study was to isolate CS from O. niloticus viscera and evaluate its modulating effect on calcium oxalate (OxCa) crystal growth. The tilapia culture residues were subjected to an enzymatic proteolysis process, followed by complexation and decomplexation with Lewatit cationic resin to obtain a mixture of GAGs. The GAGs were then purified using fractionation with acetone and ion exchange chromatography. The purified CS was identified as chondroitin-4-sulfate (CSA) and named tilapia chondroitin sulfate (CST). CST (0.01 mg/mL) was found to decrease the size of OxCa crystals and increase the number of crystals formed by 15 times. Fluorescence microscopy assays revealed that CST interacts with the faces of the CaOx monohydrate (COM) crystal, but not with dihydrate crystals. Scanning electron microscopy indicated that CST alters the morphology of COM crystals, making them assume a more elongated shape. Additionally, the study showed that CST does not present cytotoxicity under the evaluated conditions (0.01 – 0.2 mg/mL) in Madin-Darby canine renal cell (MDCK) cells. These findings suggest that CST has potential as an anti-kidney stone agent, but further in vivo studies are needed to confirm its efficacy.
4	TATIANA DOS SANTOS PAIS TERATOGENIC SCREENING IN ZEBRAFISH (Danio rerio) OF THE CRUDE EXTRACT RICH IN CAROTENOIDS OF CANTALOUPE MELON (Cucumis melo L.) AND NANONOENCAPSULATED IN GELATIN Advisor : ANA HELONEIDA DE ARAUJO MORAIS COMMITTEE MEMBERS : ANA HELONEIDA DE ARAUJO MORAIS CHRISTINA DA SILVA CAMILLO THAIS SOUZA PASSOS KELLY ALENCAR SILVA Data: Sep 14, 2023 Show Abstract Cantaloupe melon has an orange pulp rich in carotenoids, which can promote several beneficial effects on human health. However, carotenoids are extremely unstable in the presence of oxygen, light, and heat, which may reduce their bioactive properties. In this context, nanoencapsulation is an excellent strategy to ensure the preservation and enhancement of bioactive effects. On the other hand, one of the major current concerns is related to the toxicity effects of these nanoparticles. Therefore, this study aimed to evaluate the teratogenicity of the crude extract rich in carotenoids from Cantaloupe melon (CE) and of the porcine gelatin-based nanoparticles containing CE (EPG) in an animal model of zebrafish (Danio rerio). The CE was obtained from Cantaloupe melon (Cucumis melo L.) in processes that involved drying the melon pulp (55 ̊C/24 h) to obtain flour, maceration in ethanol (1:4 w/v) and partition in hexane (1:1 v/v). The nanoparticles were obtained using the oil-in-water (O/W) emulsification technique. For the characterization of nanoparticles in terms of morphology, chemical interactions, particle size and encapsulation efficiency, Scanning Electron Microscopy (SEM), Fourier Transform Infrared Spectroscopy (FTIR), Laser Diffraction and Incorporation Efficiency (IE) analyses were carried out (IE), respectively. In the experiments in an animal model with zebrafish, for the embryotoxicity test, the 308 embryos, divided into seven groups, were exposed to concentrations of 12.5 mg/L and 50.0 mg/L of CE and EPG during 96 hours post fertilization (hpf) and seven days post fertilization (dpf) the optomotor test was performed. In the toxicity tests with adult zebrafish, the 60 fish were divided into six groups exposed to 12.5 mg/L and 50.0 mg/L of CE and EPG for 96 hours, and then submitted to light/dark neurotoxicity, new and social tank. Based on the characterization results, EPG showed an IE of 94% (4.04), a smooth surface without depressions, a diameter of 88.7 nm (7.02), and a polydispersion index of 0.41 (0.03). FTIR evidenced the emergence of new vibrational bands in EPG compared to CE, demonstrating new chemical interactions. In animal model experiments, no anomalies were observed after exposure to 96 hpf, in the groups treated with CE and EPG, and heartbeats remained between 132 and 138 rpm within the expected range for embryos, similar to the negative control and DMSO groups. The groups that received CE and EPG did not show significant morphological alterations. The mortality level was below 20%, revealing neither an embryotoxic nor teratogenic character for CE and EPG in the concentrations used. There was a significant improvement in the visual motor response of larvae exposed to CE and EPG, which can be attributed to the antioxidant power and vitamin A precursor of carotenoids. No adverse effects were identified on the behavior of adults, all were similar to the behavior of the control group, without signs of anxiety, stress, or changes in swimming, speed or sociability that would indicate any toxicity. Therefore, it was found that CE and EPG maintained characteristics found in previous studies, proving the method's reproducibility. There were no signs of teratogenicity, cardiotoxicity, or neurotoxicity in the larvae, and both CE and EPG improved cognitive function and response in the evaluated model. Adult fish showed no signs of anxiety or any other evidence of toxicity, showing that CE and EPG are potentially safe.
5	ELISÂNGELA DA COSTA SILVA TOXICITY ANALYSIS OF THE BIOMATERIAL HYDROXYAPATITE FUNCTIONALIZED WITH STRONTIUMION (Sr-HA) IN ZEBRAFISH (Danio rerio) Advisor : SILVIA REGINA BATISTUZZO DE MEDEIROS COMMITTEE MEMBERS : SILVIA REGINA BATISTUZZO DE MEDEIROS VANESSA DE PAULA SOARES RACHETTI CÉSAR KOPPE GRISÓLIA Data: Oct 27, 2023 Show Abstract Hydroxyapatite (HA) is an essential inorganic component found in human bone tissue, and its compatibility with biological systems makesitwidelyapplicableinvariousbiomedicalfields.Theabilityto incorporate different ions into the HA structure allows the modification of its properties, making them more similar to those of bone tissue itself. One of these ions that can be incorporated is strontium, known for its ability to stimulate osteogenic activity. However, the biological effects of strontium-functionalized HA nanoparticles(SrHA)arenotyetcompletelyunderstood.Toaddress this gap, an embryotoxicity study was performed using zebrafish (Danio rerio), following OECD 236 guidelines. Zebrafish embryos were exposed to microspheres containing synthesized nSrHA nanoparticles at 5 °C and 90 °C for 120 hours. The mortality rate, development parameters, and production of reactive oxygen species (ROS) were evaluated during the test. At 168 hours post fertilization (hpf), behavioral parameters were assessed through motor responses and anxiety-like behavior to verify whether the biomaterialscausedneurotoxiceffects.Theresultsrevealedthatthe survival rate decreased in the SrHA group synthesized at 5 °C, and there was an increase in ROS production in this group. However, none of the biomaterials caused morphological changes that would suggest toxicity during larval development. Furthermore,behavioral tests showed no significant differences in any experimental groups, suggesting that exposure to the tested biomaterials did not have neurotoxic effects. These findings provide valuable information on the safety of HA-based nanostructured biomaterials and highlight theirpromisingapplicabilityinbonetissuerepair.Withtheincreasing use of hydroxyapatite-based biomaterials, investigations into the safety of these materials areessential.
6	NATHÁLIA CRISTINA LOPES DE JORGE Chemical, antioxidant and toxicological prospecting of Marine Sponges Suberites aurantiacus, Mycale angulosa and Halichondrida from the Potiguar Basin - RN. Advisor : ELIZEU ANTUNES DOS SANTOS COMMITTEE MEMBERS : ELIZEU ANTUNES DOS SANTOS HUGO ALEXANDRE DE OLIVEIRA ROCHA DAYANNE LOPES GOMES Data: Nov 10, 2023 Show Abstract Marine sponges are very promising organisms in the search for bioactive molecules with a range of biological activities already reported in the scientific literature, with an emphasis on antioxidant agents. Knowing that oxidative stress is associated with various health conditions, such as cancer, Parkinson's, Alzheimer's, among others, the search for compounds that act on oxidative control becomes a key element in the quest for new treatments. The Potiguar Basin of RN has a vast coastline with a rich diversity of marine sponge species, but, on the other hand, it is little explored in terms of its biotechnological potential. In this work, we conducted chemical, antioxidant, and toxicological prospecting of three species of sponges collected on the coast of the Potiguar Basin: Suberites aurantiacus, Mycale (Zygomycale) angulosa, and Halichondrida. Sequential extractions were carried out with n-hexane (HE) and chloroform/methanol, and the extracts obtained only with HE were named HESUB, HEMYC, and HEHALI, while the extracts obtained only with CM were named CMSUB, CMMYC, and CMHALI for S. aurantiacus, M. angulosa, and Halichondrida, respectively. The yield of the extracts was the determining factor for choosing the CM extracts that would be chemically characterized by GC/MS and evaluated for their antioxidant activity using the following methods: Total Antioxidant Capacity (CAT), Reducing power, ion sequestration (superoxide, DPPH, Hydroxyl Radical (OH)) and Ion Chelating activity (Ferrous (Fe2+) and Cuprous (Cu2+)). CM extracts from the three sponges were also subjected to toxicological tests in vitro with human erythrocytes and in vivo with Tenebrio molitor larvae. Chemical characterization by GC/MS revealed the presence of alcohols, alkaloids, phenolic compounds, and lipids (mainly sterol classes). In in vitro antioxidant tests, the extracts showed the ability to scavenge DPPH radicals (CMHALI 60.91% ± 3.05; CMSUB 59.32% ± 3.71 and CMMYC 53.78% ± 6.96 at 1000 µg/mL) and hydroxyl radicals (CMSUB 96% ± 0.04 at a concentration of 2000 µg/mL; CMMYC 37% ± 0.17 and CMHALI 54% ± 0.086 with 1000 µg/mL of extract) and chelating copper ions (CMHALI 113.5% ± 1.93; CMSUB 112.1% ± 5.81 and CMMYC with 110.6% ± 3.19 activity with 25 µg/mL of the sample), with a particular emphasis on the copper chelating activity for three extracts. In vitro and in vivo toxicological tests revealed that, in both tests, at different concentrations, the samples did not exhibit toxic effects. Furthermore, antibacterial tests were conducted against the pathogenic bacteria Staphylococcus aureus and Escherichia coli, but no such activity was observed. The results presented here suggest that marine sponges from this coast have pharmacological properties that can be investigated with a view to isolating the metabolites responsible for the antioxidant activities presented. Furthermore, there is the possibility of investigating the potential of these metabolites in relation to other biological activities.
7	BEATRIZ HELENA DANTAS RODRIGUES DE ALBUQUERQUE CHARACTERIZATION OF MOLECULAR MARKERS WITH POTENTIAL FOR SEMINAL SCREENING: VIRUSES AND METABOLIC DISEASES Advisor : DANIEL CARLOS FERREIRA LANZA COMMITTEE MEMBERS : DANIEL CARLOS FERREIRA LANZA DANIELLE BARBOSA MORAIS JULLIANE TAMARA ARAUJO DE MELO CAMPOS KYVIA BEZERRA MOTA Data: Nov 10, 2023 Show Abstract Infertility is a condition that affects millions of couples of reproductive age worldwide, involving both the male and female reproductive systems. While male factors contribute to half of infertility cases, a significant portion of these cases remains without an identified cause. This suggests the possibility of unidentified etiological factors that are not adequately assessed by current diagnostic investigations. In this context, the conducted study aimed to characterize molecular markers and viruses associated with male health and semen quality. The first chapter provides a comprehensive review of the human seminal virome. It was observed that the number of studies investigating viruses occurring in human semen has increased, and thus far, these studies have mostly been prospective or related to specific clinical findings. Through the combined analysis of all these articles, viruses related to deteriorating seminal parameters were listed, and a new panel with the main viruses already described that possibly affect fertility and male health was proposed. This panel can assist in evaluating semen quality and serve as a research tool in cases of infertility. The second chapter aimed to identify genes relevant to the diagnosis and early detection of hereditary metabolic disorders, using renowned databases such as OMIM and Orphanet. A total of 228 genes associated with metabolic disorders, referred to as GADM here, were identified from the 372 records extracted from OMIM. Notably, these genes are distributed across nearly all chromosomes, with a notable absence on the Y chromosome. In terms of genetic variants, the APOB gene stood out with the highest number recorded. Delving into phenotype prevalence, amino acid metabolism disorders emerged as the most prevalent category. It was also observed that autosomal recessive inheritance was dominant. Through a meticulous analysis of protein-protein interactions, an intricate network linking genes with highly prevalent phenotypes to less frequent ones was revealed. This mapping has significant implications as it indicates key genes for future investigations. From this research, targeted panels of primary genes for screening and diagnosing metabolic diseases in various contexts were established. This study underscores the need to integrate protein-protein interaction data to illuminate the underlying mechanisms of metabolic phenotypes and potentially discover new therapeutic targets or biomarkers for these conditions. In summary, the findings of this dissertation contribute to advancing diagnostic strategies for male factor infertility and to a more comprehensive understanding of genetic and reproductive counseling for metabolic diseases.
8	BELICIA SANTANA DA SILVA CHARACTERIZATION AND EXPRESSION OF GENE AND ENVIRONMENTAL STRAINS OF THE GENUS ACINETOBACTER SP. ON OIL BIODEGRADATION Advisor : LUCYMARA FASSARELLA AGNEZ LIMA COMMITTEE MEMBERS : LUCYMARA FASSARELLA AGNEZ LIMA THIAGO BRUCE RODRIGUES VÂNIA MARIA MACIEL MELO Data: Dec 1, 2023 Show Abstract Pollution from oil can have significant and long-lasting effects on the environment. Bioremediation can be an alternative to this problem. Several studies report the biotechnological potential for bioremediation by environmental Acinetobacter baumannii. However, it is still necessary to expand the understanding of the genetic mechanisms of this species to maximize its biotechnological potential. Therefore, this work aims to characterize strains of Acinetobacter baumannii oleumficedula, identify genes related to the biodegradation of hydrocarbons, as well as their presence in degradation pathways and analyze the expression of target genes. To this end, analyzes of biosurfactant production, estimation of hydrocarbon degradation, gene expression analysis and evaluation of the strains' genome were carried out. From these analyses, it was possible to verify the efficiency of all strains in the production of biosurfactant, the strains presented an estimate of hydrocarbon degradation that varied from 7.94 to 69.82%. Approximately 81 different genes associated with bioremediation were identified in 16 distinct metabolic pathways. The expression of 12 genes was observed, all of which were upregulated. By observing the gnomic context, the genetic similarity of the strains was verified. However, even with genetic similarities, the strains show significant phenotypic differences, which can be attributed to the presence of a master regulator or regulon. Finally, it can be inferred that all strains are promising for application in hydrocarbon degradation.
9	ARIANA PEREIRA DA SILVA PROSPECTION OF THE ANTIOXIDANT ACTIVITY IN VITRO AND IN VIVO OF CHAÑAR SEED (GEOFFROEA DECORTICANS) EXTRACTS (GILL. EX HOOK. ET ARN.) BURKART (FABACEAE). Advisor : KATIA CASTANHO SCORTECCI COMMITTEE MEMBERS : KATIA CASTANHO SCORTECCI RAFAEL BARROS GOMES DA CAMARA CARLOS HENRIQUE SALVINO GADELHA MENESES Data: Dec 22, 2023 Show Abstract The use of plants for medicinal purposes has been practiced since time immemorial. Various parts of plants are used for this purpose, and their use is as a complementary therapeutic source Geoffroea decorticans, popularly known as Chañar, is a plant native to Chile, All parts of this plant are used in folk medicine, ranging from expectorant activity, including bronchopulmonary disorders and pain relief, as well as antioxidant and antinociceptive activity. However, few studies are reporting such activities with Chañar seed extracts. That said, our work aimed to evaluate and characterize the antioxidant, cytotoxic and protective potential against oxidative stress induced by copper sulfate (CuSO4) of the ethanolic (EE) and aqueous (EA) extracts of Chañar seed using in vitro and in vivo methodologies. The two extracts produced were phytochemically characterized using HPLC - GC-MS/MS, as well as spectrophotometric tests such as total phenolic compounds and total flavonoids. Subsequently, biochemical tests were carried out to assess the in vitro antioxidant potential using five spectrophotometric tests (total antioxidant capacity-CAT, reducing power; 2,2-diphenyl-1-picrylhydrazyl-DPPH; copper and iron chelation) aimed at verifying the possible modes of action by which these bioactive compounds could be contributing to the maintenance of cellular redox homeostasis. The 3T3 ATCCCCL-92 fibroblast cell line was also used to assess the cytotoxicity of EE and EA, as well as the protective role of the extracts when this cell line was subjected to the stress agent copper sulfate (CuSO4), in addition to the potential to induce cell migration or not. After carrying out the in vivo tests, the extracts were tested on an invertebrate animal model (Tenebrio molitor). Survival tests were carried out to assess the toxicity of the extracts and their protective capacity against oxidative stress caused by copper sulfate. Melanisation parameters were also assessed. The results obtained by chromatography detected the presence of secondary metabolites such as phytol, alpha-tocopherol, vitexin and rutin. The in vitro spectrophotometric tests demonstrated the antioxidant capacity of the extracts to act by different mechanisms of action (inhibiting, sequestering and chelating). The tests on the 3T3 cell model demonstrated the absence of EE and EA cytotoxicity, also conferring protection against (CuSO4) stress and cell migration capacity. In the animal model, the extracts once again showed their excellent protective capacity against oxidative stress, capable of combating the free radicals generated, mitigating the melanization exacerbated by stress and the lack of modulation in the insect's metamorphosis process. Taken together (in vitro and in vivo), the EE and EA extracts of Chañar seeds are a source of bioactive compounds with excellent antioxidant properties.
	Thesis
1	ANNA BEATRIZ SANTANA LUZ Experimental models that simulate human proteolytic digestion and in silico prospecting of peptides derived from purified trypsin inhibitor from tamarind seeds with anti-SARS-CoV-2 potential Advisor : ANA HELONEIDA DE ARAUJO MORAIS COMMITTEE MEMBERS : ADRIANA FERREIRA UCHOA ALEXANDRE COELHO SERQUIZ ANA HELONEIDA DE ARAUJO MORAIS DAVI SERRADELLA VIEIRA NORBERTO DE KASSIO VIEIRA MONTEIRO RICHELE JANAINA ARAUJO MACHADO Data: Jan 23, 2023 Show Abstract Proteins are multifunctional macromolecules and also act as sources of amino acids and energy. Protein hydrolysis also generates numerous bioactive peptides with different functions in the body. In this context, proteins are sources of biological products that can be used for the control and treatment of various human diseases, and may be candidates for the management and treatment of diseases such as COVID-19. To investigate experimental models that mimic human digestion and prospect peptides from trypsin inhibitor purified from tamarind (TTIp) seeds. The first two chapters of this study are referring to a systematic review (SR), and are organized as follows: the first chapter presents the SR protocol registered in the International Prospective Register of Systematic Reviews (under number CRD42020198709); and the second chapter is the SR of in vitro studies that used methodological procedures to mimic the gastrointestinal digestion of proteins. Articles were selected according to the study population, interventions, control, results and type of study strategy. The searches were carried out in Pubmed, Web of Science, Science direct, Embase, Virtual Health Library and Scopus databases. The methodological quality assessment was performed using the Office of Health Assessment and Translation tool. A total of 1.986 articles were selected, resulting in 20 eligible studies. In the results section, we describe the methodological procedures used in vitro to simulate the digestion of animal or vegetable proteins. The third chapter refers to an in silico prospecting study of native peptides and analogues with anti-SARS-Cov-2 potential, derived from TTIp. Native peptides were obtained from enzymatic cleavages of TTIp in silico and analogues were generated by replacing amino acids in the primary sequences of native peptides. The anti-SARS-CoV-2 potential was investigated by simulating molecular dynamics between the peptides and the binding sites of transmembrane serine protease 2 (TMPRSS2), necessary for the entry of SARS-CoV-2 into the host cell, and generated a patent application: BR 10 2022 020330 0. The best interaction potential energy occurred between TMPRSS2 and one of the native peptides obtained by cleavage with trypsin (-287.48 kJ.mol-1) and its analogue peptide (-293.49 kJ.mol-1). Thus, both peptides had many hydrophobic residues, a common physical-chemical property among peptides that inhibit the entry of enveloped viruses, such as SARS-Cov-2, present in drugs used to treat of diseases related to these enveloped viruses. Therefore, this research presents an expressive scientific contribution, since the SR grouped important data that will provide subsidies for development of studies related to the clinical application of bioactive peptides, concluding that, although in vitro dynamic experimental models are more accurate, static models can be used to simulate human proteolytic digestion, provided that the physiological conditions are controlled and there is a comparison with in vivo studies to ensure greater accuracy of the results. In addition, the in silico study resulted, from the cleavage of TTIp 56/287, in potential candidates for agents that inhibit SARS-CoV-2 infection, through interaction with TMPRSS2, encouraging future in vitro and in vivo investigations that aim to use the peptides prospected here as specific medicines for the treatment of COVID-19 and other diseases.
2	FRANCISCO CARLOS DA SILVA JUNIOR A LOOK BEYOND THE PARTICULATE MATTER AND PRIORITY POLYCYCLIC AROMATIC HYDROCARBONS (PAHs): A COMPREHENSIVE INVESTIGATION OF THE TOXICITY OF RETENE Advisor : SILVIA REGINA BATISTUZZO DE MEDEIROS COMMITTEE MEMBERS : SILVIA REGINA BATISTUZZO DE MEDEIROS VIVIANE SOUZA DO AMARAL FABIANO PERES MENEZES JULIANA DA SILVA RENATA CANALLE Data: Feb 7, 2023 Show Abstract Particulate matter (PM) is considered one of the greatest threats to human health worldwide, there is still significant uncertainty and knowledge gaps with regard to many of the chemicals responsible for these effects such as PAHs. Polycyclic Aromatic Hydrocarbons (PAHs) are a wide class of chemical compounds with significant mutagenic and carcinogenic potentials, thereby harming human well-being. The United States – Environmental Protection Agency (US-EPA) includes 16 priority PAHs in risk assessment or routine environmental analyses. Retene (1-methyl-7-isopropylphenanthrene; RET), a non-priority PAH, is one of the most widely produced PAHs following forest fires. At present, the toxic endpoints of RET remain unknown, especially in human health. Therefore, divided into five chapters, this work comprehensively investigated the toxic endpoints of RET. In the first chapter, through a systematic review and meta-analysis, it was demonstrated the presence of an association between exposure to PM and genetic instability in different human population, which this early damage may lead to susceptibility to complex diseases, including lung cancer. The second chapter demonstrated that much of the knowledge on the PAHs is restricted to the priority ones; however, there are other non-priority PAHs in the environment, whose mutagenic and carcinogenic potentials are underestimated in risk assessments and routine environmental analysis, especially RET. In the third chapter, using human lung cells (A549), the results revealed that RET can significantly decrease cell viability, increase oxidative stress, mitochondrial membrane potential, and mitochondrial contents, leading to an increased reactive oxygen species (ROS) production. Besides, RET led to a significant increase in chromosomal mutations such as micronuclei (MN), nucleoplasmic bridges (NPBs), and nuclear buds (NBUDs) frequency but not mutagenicity in Salmonella strains, as well as cell death, mainly due to necrosis. The fourth chapter showed, using in-silico analysis of differentially expressed genes (DEGs), interaction networks, and transcriptional profiles in A549 cells, that RET induced variations in several genes related to metabolism, transcriptional and translational control, oxidative stress, cell cycle, DNA replication, and repair. Genes involved in these processes may explain the toxic phenotypes demonstrated using the in-silico analyses such as genotoxicity, mutagenicity, and carcinogenicity. In the fifth chapter, using zebrafish (Danio rerio) as an experimental model, RET affected DNA generating micronuclei in erythrocytes and provided new evidence suggesting behavioral alterations due to changes in redox status and the mRNA expression of the neurotransmitter systems in zebrafish brains. Overall, these results reinforce the need to look beyond the priority in toxicological research on PAHs, especially those whose toxic potentials remain underestimated such as RET, highlighting the importance of including this PAH in risk assessments and routine environmental analysis in the future especially due its toxic effects in humans.
3	LUCIANA FENTANES MOURA DE MELO EXTRACTS AND FRACTIONS OF Coccoloba alnifolia: ANTIOXIDANT, IMMUNOMODULATION AND HEALING POTENTIAL in vitro Advisor : KATIA CASTANHO SCORTECCI COMMITTEE MEMBERS : KATIA CASTANHO SCORTECCI RAFAEL BARROS GOMES DA CAMARA RIVA DE PAULA OLIVEIRA CARLOS HENRIQUE SALVINO GADELHA MENESES DEBORAH YARA ALVES CURSINO DOS SANTOS Data: Mar 29, 2023 Show Abstract Oxidative stress is an imbalance between the production of reactive species and antioxidants. The accumulation of reactive species can result in damage to tissues and organs, and it can induce various physiological changes that compromise cellular homeostasis. This imbalance can be associated with diseases such as cardiac, neurological, and cancer. It has been observed that plants' phenolic compounds can scavenge and neutralize the excess of free radicals. C. alnifolia, the plant that is the focus of our research, is a tree from the Atlantic Forest and is known to have few ethnobotanical studies. Previous studies by our group have shown that extracts obtained from the leaves of this plant have antioxidant potential. Thus, in this work, the aim was to evaluate the antioxidant and immunomodulatory potential of the ethanolic extract (EE), aqueous extract (AE), and two fractions obtained from the aqueous extract – the ethyl acetate (AF) and butanolic (BF). These two fractions were chosen based on the profile obtained after analysis in high-performance liquid chromatography coupled to a diode detector (HPLC-DAD). In addition, these fractions were characterized here for their antioxidant potential by using in vitro assays. It was observed that both have antioxidant potential. The FB, however, displayed higher antioxidant potential compared to the FA. Within the antioxidant activity, the results obtained pointed out the chelation of copper, which had not been observed previously in the crude extract (EA). From these results, the immunomodulatory potential was evaluated through assays with the RAW 267.4 cell. First, it was verified that both the extracts and the two fractions were not cytotoxic. Then, considering the complexity of the inflammatory process, the effect of EE and EA on the production of ROS after treatment with lipopolysaccharide (LPS) was analyzed. For this, three concentrations of extracts were used (100, 250, and 500 µg/mL). Both extracts were found to be able to reduce ROS production when compared to the positive control (treated with LPS). Furthermore, the ROS reduction achieved with EE was similar to the negative control (without LPS treatment). Next, the ability of the extracts and fractions to inhibit nitric oxide production was verified. Both the extracts and the fractions were able to inhibit the production and release of NO at the three concentrations tested, in a dose-dependent manner, with a reduction of more than 50%. Moreover, the phagocytic activity was evaluated only for EE and AE. This analysis was done only at a concentration of 500 μg/mL, and a reduction in phagocytosis of 30% for EE and 50% for EA was observed. Considering that cytokines are important mediators for the immune and inflammatory response, they were evaluated using two methods such as the measurement of cytokines by flow cytometry, where there was observed an increase in IL-17, and by qRT-PCR, where there was a reduction in iNOS and IL -10 mRNA in the treatment with EE. Based on these results, the potential of extracts and fractions in cell migration was also evaluated using the NHI/3T3 cell line (fibroblasts) in cultures submitted to the wound assay. The results obtained showed that both extracts and fractions in 24 h promoted cell migration, with 60% wound closure for the extracts, and 75% for the FB fraction, while the control (no extract) migration percentage was around 40%. Therefore, the data presented here show that the EE and EA and the FA and FB fractions of C. alnifolia have antioxidant and immunomodulatory potential, as well as the ability to induce cell migration involved in the in vitro wound healing process. Besides, it was evidenced in this study important properties of extracts and fractions that have the potential to act on the dynamics of the inflammatory process and tissue damage. Consequently, both extracts and fractions have potential applications for health biotechnology.
4	ANA CAMILA CAMPELO DE ALBUQUERQUE NUNES BIOQUÍMICA QUÂNTICA DOS ESTEROIDES ANABOLIZANTES AGONISTAS E SEUS MECANISMOS NO SÍTIO DE LIGAÇÃO DO RECEPTOR ANDROGÊNICO Advisor : UMBERTO LAINO FULCO COMMITTEE MEMBERS : UMBERTO LAINO FULCO EDILSON DANTAS DA SILVA JUNIOR EUDENILSON LINS DE ALBUQUERQUE JONAS IVAN NOBRE OLIVEIRA LUIZ ANTONIO RIBEIRO JUNIOR VALDER NOGUEIRA FREIRE Data: May 25, 2023 Show Abstract Anabolic androgenic steroids (AAS), natural and synthetic derivatives of testosterone, are substances with both androgenic and anabolic characteristics, which are used both clinically and also by athletes for their anabolic properties. However, the anabolic effects of testosterone and its derivatives are questioned, as what is known is that they are effective in increasing performance and improving physical appearance, even with side effects that can impact health and well-being. This thesis presents two researches carried out, one related to a narrative review and the second in the field of ab initio simulation, based on principles of Quantum Mechanics. The first study exposes a narrative review through experimental studies pointing out the main effects of anabolic androgenic steroids on the cardiovascular system. The articles used for this review were published from 2010 to 2019 in the Pubmed and Scielo databases and in independent journals, after screening by title and abstract, 26 articles were selected. The results of this review confirmed that the use of anabolic steroids can cause structural and functional alterations in the cardiac musculature due to their use, through direct and/or indirect effects, which can cause and perpetuate cardiovascular diseases. The second study characterizes the binding site of AAS Testosterone, DHT and THG in complex with the androgen receptor, identifying the chemical interactions responsible for maintaining the drugreceptor complementarity and evaluating the changes that occur in these interactions, using the MFCC strategy in the light of the DFT and dielectric constant parameterizations. Our results confirm the importance of amino acid residues Asn 705, Leu707, Leu704, Leu701, Leu873, Met742, Met780, Phe764, Met745 and Thr877 for the DHT steroid. When we analyzed THG, the important residues were Arg752, Asn705, Leu873, Leu707, Leu704, Met745, Met742 and Phe764. As for the steroid Testosterone, the results confirm the importance of the amino acid residues Asn705, Gly708, Leu704, Leu701, Leu873, Met745, Met742 Met895, Met780, Phe764, Phe876, Thr877. 17 The computational method used in the second study emerges as an efficient alternative for drug development and can aid in the discovery and design of improved antiandrogen ligands with enhanced action.
5	RAFAEL OLIVEIRA DE ARAÚJO COSTA Encapsulation of proteins and peptides with anti-obesity properties: study of the effect of trypsin inhibitor isolated from tamarind seeds on weight change and satiety Advisor : ANA HELONEIDA DE ARAUJO MORAIS COMMITTEE MEMBERS : ANA HELONEIDA DE ARAUJO MORAIS ARNOBIO ANTONIO DA SILVA JUNIOR Catarina Gonçalves PRISCILLA VANESSA FINOTELLI SEVERINA CARLA VIEIRA CUNHA LIMA THAIS SOUZA PASSOS Data: May 30, 2023 Show Abstract Obesity drives the search for new strategies to combat it. In this sense, several peptides and proteins with satietogenic activity have been studied because they regulate dietary intake and body weight. Thus, this thesis is divided into two chapters. First, we sought to understand how peptides and proteins, when encapsulated, acted on satiety, impacting weight gain and the state of obesity. For this, an integrative review was carried out by searching databases, using the keywords: peptide, protein, obesity, encapsulation, and anti-obesity agents, and selecting preclinical studies that compared the effect of peptides or proteins, encapsulated or not, on satiety. A total of 836 studies were selected, of which only four articles met the review's focus under the selection criteria. Although it has been a limited number of studies, among the effects observed with the encapsulation of these molecules are the greater efficiency in reducing weight gain, changes in the function of adipose tissue, in addition to the reduction of hormone levels that modulate appetite and body weight, promoting the potentiation of the effects of these peptides and proteins when encapsulated. In the second chapter, the impact of the trypsin inhibitor isolated from tamarind seeds (TTI) was investigated compared to the same one, nanoencapsulated in chitosan and isolated whey protein (EQPI). Thus, the ECW was analyzed and exposed to different pH conditions (7.0, 3.0, and 7.0) and temperatures (37°C, 7°C and -18°C) to evaluate the interaction between the TTI and its encapsulating agents monitored by antitrypsin activity. Subsequently, a preclinical study was carried out in which Wistar rats (n = 25) with obesity induced by a high glycemic index and high glycemic load (HGLI) diet for 17 weeks were divided into five groups and evaluated for ten days, with them: no treatment (HGLI + water), treatment 1 (nutritionally adequate diet), treatment 2 (with nutritionally adequate diet and ECW/12.5 mg/kg), treatment 3 (diet HGLI and ECW/12.5 mg/kg ) and treatment 4 (diet HGLI and TTI/25 mg/kg). These groups were evaluated for dietary intake, zoometric, biochemical, and inflammatory parameters. The ECW preserved the TTI, showing no inhibition for trypsin efficiently. When subjected to the optimal condition (constant stirring at 300 rpm, for approximately 12 h, at 20°C) for the release of the TTI, it showed 100% antitrypsin activity. However, exposing ECW to different pHs, significant anti-trips activity was observed only at gastric pH for 2 h. And regarding exposure to temperatures, ECW showed high antitrypsin activity, similar to TTI (37°C for 4 h, 7°C for three days, and 7°C for 7 days), demonstrating the complete release of TTI contained in ECW. In the preclinical study, obese Wistar rats treated with ECW significantly reduced body weight change (p < 0.05) compared to treatment with TTI. It was observed that only TTI treatment led to significant changes in inflammatory parameters (p < 0.05). The effect of ECW on weight indicates that encapsulation promoted an increase in the impact of TTI with sustained release. Therefore, the encapsulation of peptides and proteins is a promising strategy for the applicability of these molecules in the treatment of diseases such as obesity.
6	CASSIO RICARDO DE MEDEIROS SOUZA ANTI-FLAVIVIRIDAE ANTIBODIES IN GUILLAIN-BARRÉ AND CONGENITAL ZIKA SYNDROMES Advisor : SELMA MARIA BEZERRA JERONIMO COMMITTEE MEMBERS : ELIARDO GUIMARAES DA COSTA ERYVALDO SOCRATES TABOSA DO EGITO LUCAS PEDREIRA DE CARVALHO LUIZ BEZERRA DE CARVALHO JR SELMA MARIA BEZERRA JERONIMO Data: Oct 9, 2023 Show Abstract Between 2015 and 2016, Brazil has experienced the major ZIKV epidemics registered until now, which became a public health problem not only because the great number of cases, but mainly by the onset of critical infection-associated clinical conditions, like Guillain-Barré (GBS) and Congenital Zika Syndromes (CZS). The evaluation of serological diagnosis efficacy and antibody response profile are two great issues in ZIKV infection context. Serological diagnosis is mainly performed by detection of anti-NS1 antibodies, but there is evidence about cross-reactivity with antibodies against other flaviviruses, which impairs the ZIKV infection specific diagnosis. Also, urge the importance of study anti-ZIKV antibodies response and kinetics post-infection, to determine the existence of long-term antibodies and its protective capacity. So, the present work focuses in these two issues. First, efficacy NS1-based serology to detection of IgM, total IgG and IgG3 anti-ZIKV and anti-DENV and the application in differential diagnosis of ZIKV infection in SGB was evaluated. The presence of anti-ZIKV and anti-DENV antibodies in patients who developed GBS between 2009 and 2018, residents of DENV endemic area, was registered. The presence of these antibodies in individuals who developed the syndrome before ZIKV introduction indicates the occurrence of cross-reactivity between anti-flaviviridae antibodies. Data indicates IgM and total IgG serology, besides considerable sensitivity, do not present specificity enough to supports differential diagnosis of ZIKV infection. However, anti-NS1 IgG3 serology shows better performance, making possible the identification of ZIKV-associated GBS cases with more efficacy. Next, the presence of anti-NS1, anti-E and neutralizing antibodies against ZIKV and/or DENV, was determined in woman who gave birth children with CZS, evaluated in two distinct periods, 8-24 months post-partum and 25-44 months post-partum, to identify the occurrence of long-term protective antibody response. The results indicate significant presence of anti-NS1, anti-E and neutralizing antibodies in woman evaluated between 8- and 24-months post-partum, as an important sign of previous ZIKV exposition. IN the second group, it was not possible to identify significant presence of anti-NS1 and anti-E antibodies, suggesting time-associated decay of the response. However, the presence of neutralizing antibodies was registered. Anti-ZIKV neutralizing response are related to presence of anti-ZIKV antibodies in first group and, int the second one, this phenomenon was associated with the occurrence of anti-DENV antibodies, suggesting cross-protective activity between anti-ZIKV and anti-DENV antibodies. It is necessary more studies to a better characterization of IgG3 role as specific markers of ZIKV infection, and long-term anti-NS1 and anti-E as anti-ZIKV neutralizing antibodies, to improve serological diagnosis strategies and enforces the knowledge about kinetics and protective role of anti-ZIKV antibodies.
7	JOHNY WYSLLAS DE FREITAS OLIVEIRA CHARACTERIZATION OF SODIUM DIETYLDITHIOCARBAMATE AS A POTENTIAL DRUG FOR THE TREATMENT OF CHAGAS DISEASE Advisor : MARCELO DE SOUSA DA SILVA COMMITTEE MEMBERS : ALINE RIMOLDI RIBEIRO ARNOBIO ANTONIO DA SILVA JUNIOR CLAUDIA JASSICA GONÇALVES MORENO HUGO ALEXANDRE DE OLIVEIRA ROCHA MARCELO DE SOUSA DA SILVA Data: Oct 19, 2023 Show Abstract Chagas disease is an endemic zoonosis in Latin America, mainly affecting the poorer populations. The available drugs for treating both diseases have low effectiveness against intracellular forms, high toxicity, low bioavailability, and end up being unsafe for patients' health. Therefore, for several decades, efforts have been made to evaluate the possibility of using other drugs to combat this disease. One promising candidate is sodium diethyldithiocarbamate (DETC), previously described as a highly effective compound against Leishmania sp. and Trypanosoma cruzi in in vitro and in vivo models. However, in an attempt to enhance its bioavailability and reduce potential toxicity, this study evaluated the use of a nanosystem in combination with DETC through in vitro, in silico, and in vivo approaches against Trypanosoma cruzi. n this study, the nanoparticle was nanoformulated through nanoprecipitation using PLA as a polymer and tested in vitro for physicochemical characterization, cytotoxicity, cellular penetration, and antiparasitic activity. The nanoformulated PLA-DETC was found to be stable, with an average size of <200 nm and with IPD <0.3 and a zeta potential of -20 mV. It exhibited low cellular toxicity, causing no reduction of more than 20% in cell viability, had the ability to penetrate 3T3 and RAW cells within a 24-hour period, and also inhibited almost 70% of the Dm28c strain of T. cruzi within 24 hours. Additionally, it was observed that the nanosystem was capable of inducing an almost 70% increase in reactive oxygen species inside the parasites, which is a crucial factor that can lead to the parasite's death.Furthermore, an in silico analysis revealed that DETC had a more suitable ADMET profile based on physicochemical and medicinal chemistry properties compared to traditionally used drugs against Chagas disease, indicating lower toxicity. In vivo toxicity of both PLA-DETC nanoparticles and free drug at concentrations of 300 mg/kg and 1000 mg/kg was evaluated, considering hematological, biochemical, histopathological, and physical parameters following the acute toxicity characterization recommended by OECD in test 423. The results showed minimal alterations compared to the control group. A 10% reduction in platelet levels was observed, along with a slight increase in urea and AST, and a decrease in creatinine levels. Moreover, there were no significant changes in the animals' weight, behavior, or physical parameters, aligning with the control group. Finally, infection and treatment protocols were evaluated using DETC (300 mg/kg, 100 mg/kg, 0.36 mg, and 0.18 mg) and PLA-DETC nanoparticles (0.36 mg and 0.18 mg). A statistically significant reduction was observed for DETC, which exhibited a profile similar to benznidazole at a concentration of 100 mg/kg. Additionally, during the 7-day treatment, few alterations were observed, indicating low compound toxicity.
8	MARIA LÚCIA DA SILVA CORDEIRO EXTRACTS OF Talisia esculenta PRESENT COMPOUNDS BIOACTIVES WITH ANTIOXIDANT ACTIVITY AND POTENTIAL ANTI-INFLAMMATORY IN IN VITRO AND IN VIVO MODELS Advisor : KATIA CASTANHO SCORTECCI COMMITTEE MEMBERS : ADRIANA FERREIRA UCHOA DAYANNE LOPES GOMES DEBORAH YARA ALVES CURSINO DOS SANTOS KATIA CASTANHO SCORTECCI RAFAEL BARROS GOMES DA CAMARA Data: Oct 24, 2023 Show Abstract Commonly referred to as Pitombeira, Talisia esculenta (Sapindaceae) is an indigenous species endemic to Northeastern Brazil with significant economic and medicinal promise. While the fruit of this plant serves as a food source, its rind remains largely underutilized in pharmacological applications and is often discarded without further biotechnological exploration. Traditional folk medicine employs leaf infusions from T. esculenta as remedies for hip pain, rheumatism, and hypertension. However, comprehensive studies evaluating its pharmacological efficacy, toxicological profile, and phytochemical constituents are notably limited. In this context, the aim of this study was to obtain hydroethanolic extracts and infusions of the leaves and fruit rinds of T. esculenta. Subsequently, it aims to perform its phytochemical characterization, measure its antioxidant and anti-inflammatory potential in vitro and its toxicity and antioxidant capacity in vivo. For this purpose, two animal models will be used, Tenebrio molitor and Zebrafish. The hydroethanolic extract (70%) and the aqueous extract (infusion) were prepared from the fresh leaves and fruit peels of the species, obtaining the hydroethanolic extract of the leaves (HF), infusion of the leaves (IF), hydroethanolic extract of the fruit peels (HC), and infusion of the fruit peels (IC). Firstly, the antioxidant capacity was assessed using various assays: Total Antioxidant Capacity (CAT), DPPH radical scavenging, reducing power, and copper chelation. Cytotoxicity was evaluated through the application of MTT and wound healing assays, while the antioxidant potential was quantified in NIH/3T3 cells. As the extracts IF and HC were selected for further investigation on their effects on T. molitor and phytochemical characterization by quantifying the content of phenolics and flavonoids and analyzing them by CLAE-DAD (High-Performance Liquid Chromatography with Diode-Array Detection). This work also analysed the antioxidant and anti-inflammatory potential of all the T. esculenta extracts on the RAW 264.7 (macrophage strain). In addition, the protective effects of T. esculenta extracts were investigated using models of oxidative stress induced by H2O2 and CuSO4 and ascorbate in zebrafish, based on the analysis of levels of reactive oxygen species (ROS). The assays indicated that both extract types exhibited effective radical scavenging capabilities and a high capacity for copper ion chelation. It was observed that T. esculenta extracts had no effects on MTT reduction or cell migration. Furthermore, these extracts protected cells from oxidative stress induced by both CuSO4 and ascorbate. Survival analysis revealed that IF and HC exhibited no significant toxicity at the evaluated concentrations. Moreover, these extracts mitigated the effects of CuSO4 exposure on T. molitor larvae. Specifically, an increase in larval survival was observed when treated with these extracts compared to the positive control. Additionally, a reduction in melanization was verified in the animals. The CLAE-DAD analyse revealed the presence of compounds mostly from the flavonoid class, and the presence of gallic acid, quercitrin and rutin was identified In RAW 264.7 cell assays, the extracts demonstrated no cytotoxicity as per MTT tests and effectively mitigated oxidative damage induced by H2O2 in macrophages. However, only HC was able to reduce nitric oxide (NO) production in LPS-stimulated macrophages. Moreover, the protective effects of T. esculenta extracts on H2O2 and CuSO4 and ascorbate-induced oxidative stress model in zebrafish were also investigated based on the analysis of ROS levels by fluorescence microscopy. All extracts reduced ROS levels in larvae exposed to H2O2, however a significant reduction was verified for HF, IF and HC, which were statistically equal to the negative control. All samples reduced ROS levels in the CuSO4 and ascorbate-induced stress model. Collectively, these findings suggest that T. esculenta is a promising source of bioactive compounds with potent antioxidant and anti-inflammatory properties, warranting further research for potential biotechnological applications. .
9	ARANTHYA HEVELLY DE LIMA COSTA Quantum Biochemistry of Ligand-Protein Interactions Between DHODH and TyrRS Enzymes of Plasmodium falciparum. Advisor : UMBERTO LAINO FULCO COMMITTEE MEMBERS : UMBERTO LAINO FULCO EUDENILSON LINS DE ALBUQUERQUE JONAS IVAN NOBRE OLIVEIRA VANESSA DE PAULA SOARES RACHETTI EVELINE MATIAS BEZERRA RONER FERREIRA DA COSTA Data: Nov 10, 2023 Show Abstract Malaria is a parasitic disease caused by unicellular protozoa of the genus Plasmodium and occurs in more than 90 countries, with the African and Asian continents predominating. The use of antimalarial drugs is critical for both treatment and prevention of the disease. However, the parasites have begun to develop resistance to antimalarials, making current therapies ineffective in controlling the parasite. Therefore, new therapeutic approaches are needed to overcome the drug resistance of parasites and increase the efficacy of drug therapy against the disease. The aim of this research is to characterize the relationships that affect the behavior of protein-ligand complexes of enzymes from Plasmodium falciparum. The first study aims to characterize dihydroorotate dehydrogenase (DHODH) and to present the energy levels of the enzyme in the complexes with wild-type PfDHODH, with ligands DSM483, DSM557, and DSM1, and with PfDHODH with the C276F mutation together with DMS1. In the second study, tyrosine RNAt synthetase (TyrRS) is examined, both in its wild-type PfTyrRS linked to ML901-Tyr and AMS-Tyr, and with the S234C mutation, again in complex with ML901-Tyr. Using the method of molecular fractionation with conjugated layers (MFCC), amino acid residues were partitioned to calculate individual interactions using the formalism DFT (Functional Density Theory). The study with PfDHODH enzyme revealed that amino acid residues Arg265, Cys184 and Phe188 were crucial for the interactions and exhibited significant interaction energies with the four complexes studied. Moreover, the energy value of the DSM1 inhibitor was not affected by the structural changes caused by the C276F mutation, demonstrating its ability to bind to the enzyme. Working with PfTyrRS, six important residues were found to be common to the three complexes. These residues include Asp61, Gln73, Gln192, Gln210, Met237, and Phe63, most of which play essential roles in ATP binding. This discovery provides a comprehensive understanding of the interaction between PfDHODH and PfTyrRS enzymes and their inhibitors. The information gathered in this study is proving to be relevant to the development of new drug therapies and could become a tool in exploring the design of new, more sophisticated and effective antimalarial drugs.
10	PAULO EDUARDO TOSCANO SOARES Application of Next Generation Sequencing in Penaeus vannamei shrimp cultives from Rio Grande do Norte Advisor : DANIEL CARLOS FERREIRA LANZA COMMITTEE MEMBERS : DANIEL CARLOS FERREIRA LANZA RODRIGO JULIANI SIQUEIRA DALMOLIN TETSU SAKAMOTO JOSÉ MIGUEL ORTEGA SÁVIO TORRES DE FARIAS Data: Dec 1, 2023 Show Abstract Over the last two decades, short-read sequencing has become a central tool in genomicstudies allowing the rapid and accurate discovery of high-quantity DNA sequences. Thismade it possible to apply sequencing in activities of economic interest such as shrimpfarming, allowing, for example, the identification of pathogens and genotyping of hundreds tothousands of shrimps simultaneously and the detection of genetic variants of these pathogens.This sequencing can also help in the discovery of genetic markers, such as microsatellites andSNPs, which can be brought together in a genotyping panel, making it scalable and reducingthe cost per sample of its application.Specifically in shrimp farming, this technology has proven to be extremely valuable,especially for studying the shrimp genome. Nuclear and mitochondrial genome analyzesprovide crucial information about origin, adaptability and other evolutionary aspects that arevital for optimizing shrimp farming. Due to the high depth of coverage of these sequences, itis possible to capture genetic diversity in the samples, allowing the discovery of geneticvariations in mitochondrial populations (heteroplasmy) occurring in shrimp and in pathogens,such as viruses that affect them. This presents a unique opportunity to study the impacts ofthis genetic diversity on the use of mitochondrial markers and the discovery of viral variantsand quasispecies. This translates into more informed management practices, minimizingoutbreaks and optimizing shrimp health. The implementation of panels based on SNPs is aclear example of the impact of genotyping, demonstrating how the technology can be used toimprove genetic selection practices in shrimp farming.In this work, the impact of mitochondrial genetic diversity was evaluated in a virusthat affects shrimp farming and shrimp using bioinformatics approaches. First, the occurrenceof heteroplasmy was assessed by analyzing muscle sequencing data from a single shrimp,detecting patterns of variability and conservation in the mitogenomes of that individual, inaddition to comparing this internal variability with that observed among other mitogenomesand observing the impact on markers in the control region, widely used in population studies.Second, the genetic variability of the shrimp infectious myonecrosis virus obtained from tanksin a viral outbreak situation was evaluated, obtaining the genotype of the most prevalentvariant for phylogenetic analyses, revealing its possible origin and relationship with otherexisting lineages, and secondary variants, evaluating the occurrence of viral quasispecies.Finally, based on previously existing data, a panel of 25 SNP markers (15 genomic and 10mitochondrial) was developed aiming at low-cost genotyping of the shrimp Penaeusvannamei by multiplex PCR amplification followed by silico parentage analysis. This lastproject is currently ongoing, and the primers have already been evaluated, and now it remainsto continue with sequencing and bioinformatics analyses. In short, this work showed thatshort-read sequencing is capable of capturing genetic diversity, bringing new insights toshrimp farming by exposing the impact on markers, viral variability and, in the future, theimpact of this variability on genotyping panels.
11	BRUNO AMORIM DO CARMO IN VITRO AND IN VIVO ACTIVITY OF ANTIMICROBIAL PEP-TIDES ANALOGS INCORPORATED IN TO PLGA NANOPARTICLES AGAINSTStaphylococcusaureus Advisor : MATHEUS DE FREITAS FERNANDES PEDROSA COMMITTEE MEMBERS : MATHEUS DE FREITAS FERNANDES PEDROSA MARCELO DE SOUSA DA SILVA ARNOBIO ANTONIO DA SILVA JUNIOR ENEAS DE CARVALHO MARIANA DE SOUZA CASTRO Data: Dec 7, 2023 Show Abstract The emergence of microorganisms resistant to conventional drugs consti-tutesaseriousglobalpublichealthissuethatlimitstherapeuticoptionsand underscores the urgent need for the discovery of new antimicrobialagents.ThevenomofthescorpionTityusstigmurusservesasarichsource of biologically active components, including antimicrobial peptideswith high therapeutic potential. Stigmurin, an antimicrobial peptide withantibiotic activity against Gram-positive bacterial strains, was used as aprototype for structural modifications, resulting in the generation of 31analogs (patent BR102015029044-6). In this study, the three-dimensionalconformation was elucidated using nuclear magnetic resonance (NMR),stability was investigated through circular dichroism, and the in vitro andin vivo antibacterial action of two analog peptides of Stigmurin, namedStigA25 and StigA31, was assessed. In addition to the remarkable antimi-crobial activity described in a previous study, StigA25 and StigA31 exhib-ited concentration-dependent hemolytic activity compared to the proto-type peptide. To mitigate cytotoxicity, enhance stability, and optimizetherapeutic efficiency of these peptides, the effect of associating thesebioactive components with poly(lactic-co-glycolic acid) (PLGA) nanopar-ticles was investigated. StigA25 and StigA31 displayed a predominant α-helical conformation by NMR, with high structural stability under varioustemperature, pH, and salt conditions as determined by circular dichroism.The incorporation of these components into PLGA nanoparticles resultedinastablenanosystemwithantimicrobialactivityequivalenttothatoffree peptides and a significant reduction in toxicity. Scanning and trans-mission electron microscopy demonstrated that both free and nanoparticulated peptides exerted their action on the bacterial cell membrane, causingmembrane disruption. Therefore, this study reveals the high therapeuticpotential of StigA25 and StigA31 when associated with PLGA nanosystems, suggesting the use of this nanocarrier as a promising tool for thetherapeutic application of these bioactive molecules in the development ofnewanti-infectious agents.

2022

	Dissertations
1	LUCAS DOS SANTOS LISBOA EVALUATION OF THE ANTIOXIDANT ACTIVITY OF ALGINATE CONJUGATED WITH GALLIC ACID AS ANTIOXIDANT AND MODULATOR OF CALCIUM OXALATE CRYSTAL FORMATION Advisor : HUGO ALEXANDRE DE OLIVEIRA ROCHA COMMITTEE MEMBERS : HUGO ALEXANDRE DE OLIVEIRA ROCHA PABLO DE CASTRO SANTOS TALITA KATIANE DE BRITO PINTO Data: May 16, 2022 Show Abstract Alginate or alginic acid is a carboxylated polysaccharide that can be extracted from marine seaweeds (Phaeophyceae) or bacteria. It is composed by two monosaccharides, manuronic acids and guluronic acid. The alginate has already been widely used in the food, textile, and pharmaceutical industries such as biofilm constituent for food coating, as an emulsifier, in the composition of paint and medication coatings. Therefore, among other properties, it is non-toxic compound and presents good bioavailability. However, the alginate does not present other desired characteristics, for example, is not a good antioxidant agent. Another commercial interesting molecule is gallic acid. This is a phenolic compound with low toxicity, easy to obtain, can be found in various food sources, and presents various biological activities, including antioxidant. However, the antioxidant activity of in vivo gallic acid is not as prominent due its low bioavailability. Since when it is ingested, it needs a large amount for its performance and even in large quantity, it has an easy degradation, which makes it difficult its action. It was observed that this compound can combine some other molecules, which is interesting, since this conjugation can improve its activity/bioavailability. The idea of the present Thesis was to link gallic acid to alginate, thus favoring a better antioxidant activity of the alginate, potentiating it and improving the bioavailability of gallic acid, since the alginate aid in the availability of gallic acid in a higher period. In view of this objective, it was carried out the combination of gallic acid to alginate, a compound called alg-GA. To verify the effectiveness of this conjugation tests such as high efficiency liquid chromatography (HPLC) and infrared spectroscopy were performed. With respect to the apparent molecular mass, it was verified that the mass of the the alg-GA was around 33.28 kDa. In vitro antioxidant tests alg-GA has exhibited high activity as iron and cooper chelating, and as hydrogen peroxide scavenger. Cytotoxicity of these compounds (300 μm) is not detected into African green monkey cells (Vero CCL-81). When these cells were exposed, the oxidative stress induced by H2O2 (2 mm) it was observed that alg-GA (300 μm) protected cells (~ 70%) of cell damage. About modulation of calcium oxalate crystal formation, alginate, at low concentrations (100 and 300 μm) induces the highest formation of crystals and at high concentrations (600 μm) induces the formation of COD crystals. alg-GA has induced only the formation of COD crystals independent of the concentrations evaluated (from 300 μm to 1.5 mm). Regarding the size of the crystals, a proportional decrease in the size of the COD was observed with the increase in the concentration of the alg-GA used, which was not observed with the alg. The data obtained indicated that the conjugation of the alginate with gallic acid promoted the synthesis of a compound with greater antioxidant activity than the alginate and that it promotes the formation of COD crystals, which are related to half people. In future studies it is intended to evaluate the activity of alg-GA in vivo to indicate its possible use in various applicability.
2	HELOÍSA SILVA SARAIVA GOMES Computational Biochemistry of Congo Red Dye Biodegradation by Laccases Advisor : JONAS IVAN NOBRE OLIVEIRA COMMITTEE MEMBERS : JONAS IVAN NOBRE OLIVEIRA EDILSON DANTAS DA SILVA JUNIOR CLAUDIO BRUNO SILVA DE OLIVEIRA Data: Jul 27, 2022 Show Abstract The worldwide increase in urbanization and industrial activities has led to the production and introduction of contaminating molecules into ecosystems. Pollution of water bodies by untreated sewage causes numerous damages to species living in the aquatic environment. Biotechnological processes and physical, chemical, and biological strategies have been developed to remove these contaminants from anthropogenically polluted waters. From the perspective of bioremediation, lacases are enzymes capable of degrading phenolic, aromatic, and non-aromatic compounds, including synthetic dyes from the textile industry. In this context, the present work proposes to analyze the intermolecular contacts of Congo red dye coupled to lacases from Tratametes versicolor (CR -LacTv) and Pycnoporus sanguineus (CR -LacPs) qualitatively (type of chemical bonding), quantitatively (interaction energy), and comparatively, including the identification of interaction patterns. To describe the individual ligand-receptor type binding energies present in these biocomplexes, we used the molecular fractionation scheme with conjugated caps (MFCC) in the framework of density functional theory (DFT). The results indicate a higher affinity of the Congo red dye for the lacase of Pycnoporus sanguineus. In this case, the dye interacts with LacPs (LacTv) with a binding energy of -38.26 kcal/mol (-19.26 kcal/mol), with 63.3% (63.6%) of the intermolecular contacts occurring with the amino acids of the receptor from the iii (i) region of this ligand. The major residues of the CR -LacPs (CR -LacTv) complex are ARG161, PHE162, GLY392, GNL160, SER393, PRO391, PHE265, and GLY266 (PHE162, ALA161, GLY266, ALA393, PHE 265, PRO391, PRO160, GLY392), in which we highlight the existence of six (five) large hydrogen bonds and two (three) hydrophobic contacts in CR -LACPs (LacTv). The structural and energetic description of the interactions of the biocomplexes in question helps to understand the differences in terms of stability and efficacy of enzymes of the same family but different organisms. Consequently, it will allow studies to increase the effectiveness of bioremediation promoted by such enzymes following targeted point changes.
3	FRANCISCO MATEUS GONCALVES TRAJANO Evaluation of histamine effects on contractility of rat cauda epididymis duct Advisor : EDILSON DANTAS DA SILVA JUNIOR COMMITTEE MEMBERS : EDILSON DANTAS DA SILVA JUNIOR JONAS IVAN NOBRE OLIVEIRA MANUELA DOS SANTOS CARVALHO SCHIAVON Data: Oct 21, 2022 Show Abstract The motor activity of the epididymal duct is an essential process for male fertility, as it results in the correct transport, maturation, storage and emission of spermatozoa. This process is regulated by hormonal (testosterone, vasopressin, oxytocin), neuronal (adrenergic and cholinergic autonomic innervation), and epithelial (endothelin-1, NO, serotonin, prostaglandins, angiotensin, etc.) mechanisms. However, although there is evidence for the presence of histamine in the epididymis, its modulatory effects on the motor activity of the epididymal duct are not yet known. Thus, the objective of this work was to evaluate the effects of histamine on contractions of the rat distal cauda epididymis duct by identifying histaminergic receptors and the participation of autonomic neurotransmitters. To achieve these objectives, segments of the epididymal duct from the distal cauda of the epididymis of Wistar rats were isolated and used in functional experiments in an isolated organ bath to evaluate the direct effects of histamine on epididymal duct or adrenergic agonist-induced contractions. These experiments were also performed in the presence of selective antagonists for histaminergic receptors (H1, H2 and H3), α1-adrenoceptors and muscarinic receptors. We found that histamine produced contractions of a phasic nature. The frequency and amplitude of histamine-induced phasic contractions in the epididymal duct were inhibited by promethazine (H1 receptor antagonist), ranitidine (H2 receptor antagonist), atropine (muscarinic receptor antagonist) or prazosin (α1-adrenoceptor antagonist). Histamine potentiated the contractile effects of noradrenaline and reduced its maximal effect, and these effects were prevented by ranitidine and promethazine, respectively. We conclude that histamine is able to promote phasic contractions with the participation of H2 receptors and release of autonomic neurotransmitters. Furthermore, this biogenic amine modified noradrenaline-induced epididymal duct contractions with the possible participation of H1 and H2 receptors.
4	LEIDIANE BARBOZA DA SILVA EFICÁCIA E SEGURANÇA DO EXTRATO DE SEMENTES DE ANGICO (Anadenanthera colubrina) CONTRA Aedes aegypti E Aedes albopictus, EM CONDIÇÕES DE LABORATÓRIO E DE CAMPO Advisor : ADRIANA FERREIRA UCHOA COMMITTEE MEMBERS : ADRIANA FERREIRA UCHOA VALTER FERREIRA DE ANDRADE NETO JOSÉ ROBERTO DA SILVA PATRICIA BATISTA BARRA MEDEIROS BARBOSA Data: Dec 22, 2022 Show Abstract Aedes aegypti (L., 1762) and Aedes albopictus (S., 1894) are vectors of the etiological agents of numerous arboviruses of medical importance. Limitations in the use of chemical insecticides have stimulated the search for effective and ecologically friendly options, such as botanical insecticides. The present study describes the aedicidal potential of Anadenanthera colubrina (Vell.) seed extract at different stages of the vectors' life cycle, its attractive/repellent action on oviposition, partial biochemical characterization and evaluation of cyto- and ecotoxicity. The hot aqueous extract of A. colubrina seeds (EAQA) was obtained by extraction in distilled water (1:10, m/v) for 7 h at 80 °C. The larvicidal effects of EAQA were evaluated in serial dilutions and the lethal LC50 and LC90 concentrations (mg/mL) determined after 24 h and 48 h of exposure. The larvicidal effect was observed for A. aegypti and A. albopictus, recording LC50 and LC90 at 24 h (0.16 and 0.94 mg/mL; 0.36 and 1.28 mg/mL) and 48 h (0. 02 and 0.26 mg/mL; 0.06 and 0.62 mg/mL), respectively. EAQA was pupicid for both insects at concentrations higher than larvicidal concentrations (24 h LC50 ≥ 0.85 mg/mL and 48 h LC50 ≥ 0.53 mg/mL). 0.26 mg/mL was defined as the diagnostic concentration for ovicidal activity and oviposition, showing ovicidal activity for both insects, compromising larval hatching by 76% and reducing adult emergence of surviving larvae by 25%. Under field conditions, EAQA exerts an attractive effect on oviposition. EAQA showed a higher concentration of total carbohydrates (15.5 mg/mL) compared to total soluble protein (2.0 mg/mL) and total phenolic compounds (0.1 mg/mL). EAQA presents bromelain, papain, trypsin and chymotrypsin inhibitors, with enzyme inhibition activity in 90.94%, 68.93%, 68.75% and 9.32%, respectively. No cytotoxicity was observed for tested cell lines (3T3) and (HepG2), nor ecotoxicity for Ceriodaphnia dubia (LC50 = 0.72 mg/mL for 24 h and 0.34 mg/mL for 48 h) and Mysidopsis juniae (LC50 = 0 .87 mg/mL for 24 h and 0.37 mg/mL for 48 h). EAQA has the potential to be an adjuvant in Aedes integrated management strategies due to the versatility of action against different stages of the vectors' life cycle, as well as their cyto- and eco-compatibility.
	Thesis
1	GIULIAN CÉSAR DA SILVA SÁ Tephrosia toxicaria (Sw.) Pers. EXTRACTS: PROSPECTION OF ANTI-Aedes, ANTIOXIDANT AND ANTIMICROBIAL ACTIVITIES Advisor : ADRIANA FERREIRA UCHOA COMMITTEE MEMBERS : ADRIANA FERREIRA UCHOA KATIA CASTANHO SCORTECCI MARIA DE FATIMA FREIRE DE MELO XIMENES PATRÍCIA MARIA GUEDES PAIVA SELENE MAIA DE MORAIS Data: Mar 25, 2022 Show Abstract The search for new natural adjuvant formulations for the management of diseases, vectors and pathogens shows that plant extracts are still promising, especially considering the great Brazilian biodiversity, they are obtained from renewable sources, their toxic selectivity to target organisms and environmental compatibility. The development of the present research was stimulated by the possibility of expanding the repertoire of options for natural adjuvants, which aimed to investigate the bioactive potential and biosecurity of aqueous and hydroethanolic extracts from seeds, roots, stems and leaves of Tephrosia toxicaria (Sw.) Pers. This legume, occurring in the Northeast region of Brazil, attracts the attention of researchers by acting as a matrix for obtaining broad spectrum bioactivity formulations. This thesis was structured in two chapters; the first chapter deals with a bibliographic review of the current state of art on the use of botanical insecticides in the integrated management of arbovirus vectors, with emphasis on control of species belonging to the genera Aedes, Culex and Anopheles. Numerous botanical insecticides formulations are presented throughout the review and its likely modes of action for different stages of insects. The second chapter describes the versatility of T. toxicaria extracts in the Aedes control, highlighting the hydroethanolic root extract (RHA) as the most promising larvicidal extract, demonstrating the lowest lethal concentration for 50% (300 µg/mL) and 90% (840 µg/mL) of Aedes larvae, observed at 24 h post-exposure. In field tests, RHA (840 µg/mL) is a promising oviposition deterrent agent, reducing Aedes egg laying by approximately 90%. In addition to manifesting insecticidal properties, RHA manifests antioxidant activity, mainly in control of metal ion homeostasis and restoration of the metabolic activity of cells under oxidative stress conditions, under low concentrations (100 µg/mL). Although leishmanicidal molecules are also expressed in RHA, its solubilization was less pronounced, requiring a higher concentration of extract to inhibit the growth of Leishmania amazonensis promastigotes (IC50 = 3.53 mg/mL). The extraction conditions did not favor the concentration of antimicrobial molecules against the bacterial and fungal strains investigated, in assays employing serial dilutions of RHA (1024 – 32 μg/mL). RHA concentrates carbohydrates, phenolic compounds and proteins, with estimated molecular masses between 10 and 24 kDa. Protein analyzes suggest the expression of protease inhibitors and lectins in RHA. The bioinsecticidal, antioxidant and leishmanicidal effects, associated with the absence of toxicity events against non-target organisms (Lactuca sativa) and cell lines (HepG2 and 3T3), suggest T. toxicaria as a natural source to be sustainably exploited to obtain extracts with wide biological and pharmacological use.
2	RONY LUCAS DA SILVA VIANA Silver nanoparticles containing xylans: synthesis, characterization and evaluation of their immunomodulatory and antibacterial effect Advisor : HUGO ALEXANDRE DE OLIVEIRA ROCHA COMMITTEE MEMBERS : HUGO ALEXANDRE DE OLIVEIRA ROCHA MONIQUE GABRIELA DAS CHAGAS FAUSTINO ALVES KAROLINE RACHEL TEODOSIO DE MELO RANIERE FAGUNDES DE MELO SILVEIRA VALQUIRIA PEREIRA DE MEDEIROS Data: May 6, 2022 Show Abstract In recent decades, the increase in bacterial resistance to conventional treatments has led to an incessant search for new effective alternatives against diseases caused by these microorganisms. Within this context, significant progress has been achieved in the development of drugs based on nanotechnology in order to combat/attenuate bacterial resistance. And among the nanoparticles, those made of silver (NANO) are pointed out as a promising option in this sense, including being already applied in different uses, such as in the textile industry. However, their use in clinical medicine has a smaller dimension, and this has been driving the development of different mixed NANO, which contain, in addition to silver, an organic portion in their constitution, since this can enhance or provide new applicability to NANO. This has even allowed the use of techniques less aggressive to the environment (green synthesis) for the synthesis of NANO. Polysaccharides are one of the organic molecules that have been used in this sense. Xylan is a hemi-cellulose found in different plants and, therefore, very abundant in nature. However, there are less than a dozen papers with NANO containing xylans, and none have evaluated their antimicrobial effect. Therefore, in this work NANO was synthesized with different xylans extracted from corn cob (Zea mays). For this, corn cob, in 0.1M NaOH, was subjected to ultrasound waves to obtain a xylan-rich extract (EBX), which was subsequently fractionated with the addition of increasing volumes of ethanol. Thus, six ethanolic fractions (EF's) were obtained, which after being characterized (sugar content, proteins, phenolic compounds, and monosaccharide composition), were used for the green synthesis of NANO. These NANO were characterized by UV-visible spectroscopy, scanning electron microscopy, dynamic light scattering, energy scattering spectroscopy and infrared spectroscopy. All NANO (NANO EBX, NANO E0.3, NANO E0.4, NANO E0.8, NANO E1.4 and NANO E2.2) were essentially composed of silver and xylans, presented round shape, average size ranging from 105, 0 and 79.7 nm and stability of 24 months. These NANO were not cytotoxic against fibroblast (3T3) and macrophages (RAW). Furthermore, they promoted the reduction of nitric oxide (NO) production in macrophages activated by bacterial lipopolysaccharide. The different NANOs, synthesized here, also showed antibacterial activity. All were effective against Gram-negative bacteria tested (Escherichia coli; carbapenemase-producing Klebsiella pneumoniae – KPPC) with emphasis on NANO EBX and NANO E1.4, which showed a minimum inhibitory concentration (MIC) of 62.5 µg/mL against E. coli. For KPPC, NANO EBX, NANO E0.4, NANO E1.4 and NANO E2.2 stand out, which presented a MIC of 250 µg/mL. The data presented here show the biotechnological potential of the different xylans in the form of nanoparticles and future tests, including in vivo, should be carried out to confirm the antibacterial potential of the different NANO synthesized with EBX and FE's.
3	LUÍZA ARAÚJO DA COSTA XAVIER Influence of radon and alpha particles in genome oxidation and LINE-1 methylation Advisor : VIVIANE SOUZA DO AMARAL COMMITTEE MEMBERS : VIVIANE SOUZA DO AMARAL SILVIA REGINA BATISTUZZO DE MEDEIROS CLÁUDIA ROHDE JULIO ALEJANDRO NAVONI MARIALVA SINIGAGLIA Data: May 10, 2022 Show Abstract The present work aims to evaluate the mutagenic and epigenetic effects induced by natural ionizing radiation in individuals exposed to high concentrations of radon (Rn). Furthermore, epigenetic effects were also investigated in assays with cell cultures exposed to different doses of alpha particles. For that purpose, an observational case-control study was conducted with 224 individuals from Lajes Pintadas (LP group) and 66 individuals from Natal city (control group). Blood and urine samples were collected from participants of both cities to perform the following tests: (a) biochemical clinical analysis tests; (b) quantification of 8-hydroxy-2’- deoxyguanosine (8-OHdG) in urine; (c) determination of the Ser326Cys polymorphism in hOGG1 gene; (d) quantification of lead in blood; (e) quantification of methylated cytosines (mC) in LINE-1 sequences as bioindicators of global genome methylation of the participant’s blood cells. In addition, all recruited individuals were interviewed with a questionnaire to collect sociodemographic information, environmental and occupational exposures, lifestyle and general health history. About the in vitro experiments, human lung fibroblasts were exposed to 0 (control), 0.25, 0.5 and 1 Gy of alpha particles and their %mC in the promoter region of the LINE-1 sequences were determined. This assay was repeated for the progeny of irradiated cells. It was observed that individuals exposed to high indoor Rn levels had 8-OHdG concentrations approximately 1.5 times higher than those exposed to low levels of this gas (p-value < 0.01). People heterozygous for the hOGG1 polymorphism had significantly lower concentrations of 8-OHdG compared to those homozygous for the wild-type allele, considering exposure to the highest Rn levels (from 145 Bq/m3 upwards; p-value < 0.05). The mean concentrations of blood lead between the control and LP groups showed no significant difference: 1.15 ± 0.91 µg/dL and 1.66 ± 1.55 µg/dL, respectively, and these values were below the limit established by the Center Disease Prevention Control (5 µg/dL). In epigenetics, there was a significant difference between the means of %mC in LINE-1 of the groups – 50.23% ± 3.44 for control and 51.87% ± 2.96 for the LP group (p-value < 0.001). Indoor Rn levels, blood lead concentrations and the individuals’ sex were factors that significantly influenced the mean %mC of LINE-1 sequences of the participants. For the in vitro assays, there was no significant difference between the means of %mC among the different doses of alpha particles applied to the cells. It can be concluded that high levels of natural ionizing radiation can cause genome instability through oxidative and epigenetic pathways even in passenger non-irradiated cells from the human body.
4	AIRTON ARAÚJO DE SOUZA JÚNIOR GAMIFIED BLENDED LEARNING OF BIOCHEMISTRY CURRICULAR COMPONENTS Advisor : ELIZEU ANTUNES DOS SANTOS COMMITTEE MEMBERS : EDUARDO GALEMBECK ELIZEU ANTUNES DOS SANTOS LEANDRO SILVA COSTA MANUEL JOÃO COSTA MONIQUE GABRIELA DAS CHAGAS FAUSTINO ALVES RAFAEL BARROS GOMES DA CAMARA Data: May 12, 2022 Show Abstract In the face of the reality of teaching biochemistry, which is permeated by the densification of the contents, the interdisciplinary complexity, and aggravated by an exclusive or excessively expository approach, it becomes increasingly necessary to evaluate methodological proposals that mitigate this scenario. Thus, in this dissertation, we developed a work proposal for a Blended Learning Gamified, based on the Flipped Classroom, in the Biochemistry classes; academic performance assessment; the interest and involvement of students in the subject; the methodological elements of the proposal, gamification, and, finally, gains in cognitive, social and socio-emotional skills. Therefore, a type of exploratory research was carried out, through a pedagogical intervention with qualitative and qualitative analysis. The proposal was applied in two biochemistry disciplines at the Federal University of Rio Grande do Norte (UFRN) for the biological sciences, in the periods of 2016.2 and 2017.1, and in medicine courses, in the periods of 2018 and 2019, a total of 179 students participated in this study. The research analyzed data from the design of two classes: face-to-face and blended classes, which were divided into online moments and in-person moments, with educational objectives combined in both moments. For the online moments, were used video lessons and the face-to-face moments, educational objects elaborated by Bloom's Taxonomy were applied. The entire process was gamified, with a scoring system (ranking), team building in the cooperative logic and obtaining benefits (reward system). The entire process was gamified, with a scoring system (ranking), team building in the cooperative logic and obtaining benefits (reward system). The results show that most students had better academic performance, and reported increased interest in the discipline, stating that learning was better in blended classes than in face-to-face classes. Furthermore, gamification had a positive evaluation and ended up promoting increased student engagement. Besides, it was shown that the method contributed to collaborative and personalization learning. Finally, it was observed that the pedagogical intervention contributed to the development of specific aspects for the discipline of biochemistry, for example, the understanding of concepts, relationships between structures and functions, and proposing solutions and arguments for solving problems, among others. After all, the results described here provide data for a discussion about the importance of Blended Learning Gamified as a promising alternative to the disciplines of biochemistry. Certainly, this requires a broad reflection on the approaches and processes involved in this model, because studies on the effectiveness of Hybrid Teaching for the Teaching of Biochemistry are still very scarce.
5	ANA BEATRIZ MEDEIROS LINS DE ALBUQUERQUE TAVARES Quantum Computational Simulation of Immuno-Oncological Drugs Advisor : EUDENILSON LINS DE ALBUQUERQUE COMMITTEE MEMBERS : EUDENILSON LINS DE ALBUQUERQUE UMBERTO LAINO FULCO LUCIANO RODRIGUES DA SILVA MANOEL SILVA DE VASCONCELOS JOSE ALZAMIR PEREIRA DA COSTA JOSE DE MIRANDA HENRIQUES NETO Data: Jun 10, 2022 Show Abstract Much of the recent excitement in the cancer immunotherapy approach has been generated by the recognition that immune checkpoint proteins, like the receptors PD-1 (Programmed cell Death-Ligand 1)and CTLA-4 (Cytotoxic T-Lymphocyte-Associated Protein 4), can be blocked by antibody-based drugs, with deeper effects. Promising clinical data have already been released, pointing out to the efficiency of the monoclonal antibody immune-oncological drugs pembrolizumab (PEM), nivolumab (NIV), and ipilimumab (IPI) to block these immune checkpoint proteins pathway, triggering the T-lymphocytes against a wide range of cancers, as well as being an important issue for clinical research. Their use was already approved by the US-FDA (United States Food and Drug Administration) office since the last decade. To date, although many structural properties of these drugs have been unveiled, binding energy features of both checkpoint proteins, PD-1 and CTLA-4, based on crystallographic X-Ray data, need a deeper understanding. In this context, by employing quantum chemistry methods based on the Density Functional Theory (DFT) and the molecular fractionation with conjugate caps (MFCC) scheme, we investigate in silico the binding energy features of the receptors PD-1 and CTLA-4 in complex with their drugs inhibitor (PEM, NIV, and IPI), highlighting the most relevant residue-residue interactions, looking for new insights into the mechanisms of the pathway blockade to further engineer their affinity and selectivity. Our computational results are not only in good agreement with the experimental binding affinity order, but also give a better understanding of the binding mechanisms, pointing out to an efficient alternative towards the development of antibody-based drugs. Besides, they lead to new treatments for cancer therapy based upon immunotherapy, unleashing the immune surveillance to destroy the cancer cells by decreasing their immune evasion. They are also an efficient alternative towards the development of new small-molecules and antibody-based drugs, unveiling new treatments for cancer therapy. On the other hand, although many structural properties of these immune-oncological drugs have been unveiled, only few studies were focused on their vibrational features. To fill this gap, quantum chemistry calculations were also employed here to depict the binding energetic modes of the PEM and NIV antibody drugs, in order to obtain their vibrational properties through their optical absorption spectra and Raman scattering spectroscopy. Detailed interpretation of their harmonic vibrational frequencies is also presented. Finally, the binding of three different anticancer drugs, Cu(BpT)Br, NAMI-A, and DOX (doxorubicin), widely used in the breast cancer treatment, to the Human Serum Albumin (HSA), were also investigated here by means of a dispersion corrected exchange-correlation functional within a fragmentation strategy. As a consequence, it is possible to identify the magnitude of the most relevant quantum binding interactions of these supramolecular complexes, and thus guide their molecular modification process. The data obtained in this work highlight the power of quantum calculations as an important tool for the drug design process, and pave the way for the use of HSA-ligand interactions during the rational design of new anticancer compounds. More important, our results show that HSA/[Cu(BpT)Br]–(NAMI-A)-(DOX) multi-drug complex increases the targeting ability compared with the isolated use of the three anticancer-drugs, in agreement with in vivo predictions. All in all, the quantum chemistry computational methods used in this PhD thesis emerged as a simple and efficient alternative to unveil the drug’s amino-acids residues that play the most important role on the binding affinity of the receptor-ligand complex. Indeed, taking into account the cost/benefit of the operation, in silico approach is becoming an important initial step not only in clinical oncology, but also in defining the research frontier in the biological, physical and chemical science.
6	SARAH DE SOUSA FERREIRA Development and evaluation of antidotes for Bothrops brazili snake envenomation: investigation of the antivenom potential of chlorogenic and rosmarinic acids and antiserum production in chitosan nanoparticulate system Advisor : MATHEUS DE FREITAS FERNANDES PEDROSA COMMITTEE MEMBERS : MATHEUS DE FREITAS FERNANDES PEDROSA HUGO ALEXANDRE DE OLIVEIRA ROCHA ARNOBIO ANTONIO DA SILVA JUNIOR DANIELA PRISCILA MARCHI SALVADOR GISELLE PIDDE QUEIROZ Data: Jun 22, 2022 Show Abstract Ophidism causes high mortality and morbidity in many regions of the world. Currently, the only treatment is serum therapy, which has some limitations, such as: high cost for its production, low effectiveness in neutralizing local effects, difficult access in some regions and adverse reactions. Considering these factors, this study has the general objective of presenting new alternatives to improve the treatment of snakebite. For this purpose, the venom of the Bothrops brazili snake was used, a little studied species that is found mainly in the Amazon region. Thus, the present study has as specific objectives (1) to analyze the inhibitory potential of chlorogenic and rosmarinic acids against the local and systemic effects induced by B. brazili envenomation, (2) to obtain and characterize chitosan nanoparticles with the venom of this species with two modes of association (incorporation and adsorption), in order to be evaluated in relation to the immunoadjuvant potential for the production of a new serum. The evaluation of the antiophidic potential of chlorogenic and rosmarinic acids was carried out through in vitro, in vivo and in silico assays. In vitro, the acids were able to inhibit proteases and phospholipases activities of the venom, in addition to reducing the pro-coagulant effect in human plasma caused by the venom. In vivo, in a mouse model, these compounds inhibited local effects such as edema, hemorrhage, increased myeloperoxidase enzyme and myotoxicity. In silico the acids were able to interact with venom phospholipases. In addition, the compounds mitigated the systemic effects resulting from the envenomation, which were: kidney, liver, muscle damage, hemostatic changes (partially activated thromboplastin time, prothrombin time and platelet count), hematological (erythrogram and leukogram) and lipid peroxidation. For the production of chitosan nanoparticles, the ionic gelation technique was used. The nanoparticles had a size between 150 and 190 nm, a potential of approximately +30 mV and a polydispersity index (PDI) of approximately 0.400. These parameters were evaluated by the Dynamic Light Scattering (DLS) technique. The nanoparticles showed 97% efficiency to incorporate and adsorb the venom and the analysis of scanning electron microscopy with field emission source (MEVFEG) and atomic force microscopy (MFA) showed that they had homogeneous shape and size. Fourier transform infrared spectroscopy (FTIR) analysis revealed that the venom showed bands at 1543 cm-1 and 1651 cm-1 referring to amide groups. The nanoparticles showed bands at 916, 1087, 1259 and 1340 cm-1 that underwent deviation after incorporation and adsorption of the venom. In conclusion, these results demonstrate that chlorogenic and rosmarinic acids have the potential to inhibit the local and systemic effects caused by B. brazili envenomation, which makes them possible alternatives to complement serum therapy. Furthermore, chitosan nanoparticles were efficient in incorporating and adsorbing the venom, showing great potential to be used as an adjuvant in the production of a new antivenom.
7	MAYARA SANTA ROSA LIMA ANTI-INFLAMMATORY EFFECT OF PEPTIDES AND PROTEINS ON INTEGRITY AND FUNCTIONALITY OF THE INTESTINAL BARRIER Advisor : ANA HELONEIDA DE ARAUJO MORAIS COMMITTEE MEMBERS : ANA HELONEIDA DE ARAUJO MORAIS BRUNA LEAL LIMA MACIEL DALINE FERNANDES DE SOUZA ARAUJO JULIANA KELLY DA SILVA MAIA FRANCISCO ADELVANE DE PAULO RODRIGUES ANA VLADIA BANDEIRA MOREIRA Data: Jul 28, 2022 Show Abstract A healthy intestinal barrier protects the individual against antigen translocation, ensuring a controlled inflammatory environment. The first two chapters of this thesis refer to a systematic review (SR), which aimed to understand the mechanisms of action of anti-inflammatory molecules that reduce TNF-α and their effects on the intestinal barrier in animal models. The protocol of the first chapter was registered in PROSPERO and guided the elaboration of the RS, presented in the second chapter. The articles were selected according to the PICO strategy (Population, Intervention, Comparison/Control and Outcomes), from PubMed, Scopus, Web of Science, EMBASE and ScienceDirect databases. Twenty-five articles were selected and the risk of bias assessment was performed using the SYRCLE tool. The results show that the anti-inflammatory molecules that acted by reducing TNF-α acted mainly on the TNF-TNFR1/TNFR2 and TLR4/MD2 complex signaling pathways, and consequently on the NF-κB pathway, which reduced the symptoms of inflammatory diseases and the macroscopic, histological and intestinal permeability aspects. In the third chapter, the proposal and methodological validation of an inflammation model with TNF-α in co-culture of Caco-2:HT29-MTX intestinal cells is presented. In the fourth chapter, the trypsin inhibitor isolated from tamarind seeds (TTI) was evaluated in vitro regarding its interaction with bacterial lipopolysaccharide (LPS) and its action against human neutrophil elastase. In addition, the effects on the integrity and functionality of the intestinal barrier in cell culture and in an experimental obesity model were evaluated, including hematological, biochemical and inflammatory parameters. The cell study was performed in differentiated Caco-2:HT29-MTX co-cultures and evaluated cell viability and production of reactive oxygen species during contact of cell monolayers with ITT. In addition, transepithelial electrical resistance and permeability were evaluated in monolayers previously inflamed with TNF-α (50 ng/mL) and treated with TTI (1.0 mg/mL, after or during the inflammatory stimulus). For the animal study, obese Wistar rats (n=15) were divided into three groups: no treatment group (n=5), group treated with a nutritionally adequate diet (n=5) and group treated with TTI (25 mg/ kg/day, n=5) for ten days. TTI did not interact with LPS, but showed inhibition against human neutrophil elastase (93%). In the cell study, ITT did not alter cell viability and did not present pro- or antioxidant properties, as well as it did not prevent damage or restore the monolayers integrity. In the in vivo experiment, TTI-treated rats had significantly lower plasma concentrations of inflammatory cytokines, and of other obesity-related parameters, such as total leukocyte count, fasting glucose, and LDL-c, compared to animals treated with a nutritionally adequate diet. They also showed better histopathological and histomorphometric results in the small intestine, although there were no significant differences between the groups regarding most semiquantitative parameters, intestinal permeability and concentration of inflammatory cytokines in the intestine. Thus, it is concluded that TTI, at the concentrations tested, was safe for cell cultures and reduced systemic inflammation in Wistar rats, which possibly reflected in the improvement of the morphology of the intestinal epithelium in the treated animals. In view of the results, it is believed that TTI has another type of action on the intestinal epithelium, probably related to the inhibitory activity against neutrophil elastase.
8	DIANA PONTES DA SILVA Analysis of the Inhibitory Potential of Chlorogenic Acid and Rosmarinic Acid against the Effects of Bothrops leucurus WAGLER (1824) snake venom Advisor : MATHEUS DE FREITAS FERNANDES PEDROSA COMMITTEE MEMBERS : FRANCISCO CANINDE DE SOUSA JUNIOR KARLA PATRÍCIA DE OLIVEIRA LUNA LUIZ ALBERTO LIRA SOARES MAIRA CONCEIÇÃO JERONIMO DE SOUZA LIMA MATHEUS DE FREITAS FERNANDES PEDROSA Data: Jul 28, 2022 Show Abstract Snakebites represent a serious public health problem in tropical countries. Bothrops leucurus snake is one of the main responsible for snakebites in Northeast Brazil, but this species is not included in the pool used in the production of antiophidic serum. Considering the limitations of the antiophidic serum, several studies have been carried out in order to develop complementary treatments for snakebite and, in this context, plants are an important source of bioactive compounds. Chlorogenic acid and rosmarinic acid are molecules derived from the secondary metabolism of plants, with several biological activities described and antiophidic potential still little explored. Thus, the aim of the study was to evaluate the inhibitory potential of chlorogenic and rosmarinic acids against the effects of B. leucurus venom. Initially, in vitro neutralization capacity of the enzymatic activities of the venom was evaluated. Then, in vivo tests were performed to verify the inhibition of acids against hemorrhagic, edematogenic, myotoxic activities and quantification of inflammatory markers, as well as the analysis of biochemical, hemostatic, hematological markers, and verification of the degree of lipid peroxidation in renal and liver tissues. Finally, the ability of the inhibitors to interact in silico with two classes of toxins was evaluated. The results showed that chlorogenic and rosmarinic acids were effective in inhibiting phospholipase A2 (21% and 60%), proteolytic (39% and 60%), hyaluronidase (59% and 75%) and coagulant (p < 0.0001) activities, respectively, as well as were effective in preventing fibrinogen degradation. In in vivo assays, the inhibitors attenuated edema (p < 0.01), reducing the levels of myeloperoxidase (p < 0.001), IL-6 (p < 0.0001) and IL-1β (p < 0.001) interleukins. The compounds also inhibited the hemorrhagic (p < 0.0001) and myotoxic (p < 0.001 – chlorogenic acid, p < 0.0001 – rosmarinic acid) actions of the venom. In serum biochemical markers, the inhibitors attenuated changes in urea levels (p < 0.01 - chlorogenic acid and p < 0.05 - rosmarinic acid), creatinine (p < 0.0001), uric acid (p < 0.0001 ), alanine aminotransferase (p < 0.05), aspartate aminotransferase (p < 0.001- chlorogenic acid and p < 0.05 - rosmarinic acid), amylase (p < 0.01), total proteins (p < 0.01), creatine kinase (p < 0.01) and lactate dehydrogenase (p < 0.0001). Furthermore, both compounds decreased the levels of lipid peroxidation in liver (p < 0.001) and renal (p < 0.0001) tissues. When evaluating hemostatic parameters, only rosmarinic acid was able to reduce the effects on circulating fibrinogen (p < 0.0001) and platelet count (p < 0.0001). Computational studies indicated that the inhibitors have the ability to interact with toxins of the phospholipase A2 and metalloprotease class. Taken together, the results obtained evidenced the antiophidic potential of chlorogenic and rosmarinic acids, which presented, in general, greater inhibition efficiency than the antiophidic serum. Thus, both inhibitors are promising candidates for the development of future adjuvants to be used to complement the currently available antivenom sorotherapy.
9	FLÁVIA ROBERTA MONTEIRO DE SOUZA SULFATED POLYSACCHARIDES OF Gracilaria birdiae, EXTRACT OF LEAVES of Baccharis trimera AND BLENDS CONSTITUTED BY THESE AGENTS: EVALUATION OF ANTIOXIDANT ACTIVITY AND THE CAPACITY OF INHIBITING LIPID ACCUMULATION Advisor : HUGO ALEXANDRE DE OLIVEIRA ROCHA COMMITTEE MEMBERS : HUGO ALEXANDRE DE OLIVEIRA ROCHA RAFAEL BARROS GOMES DA CAMARA DÁRLIO INÁCIO ALVES TEIXEIRA ALEXANDRE COELHO SERQUIZ HERYKA MYRNA MAIA RAMALHO Data: Aug 23, 2022 Show Abstract Obesity, characterized by excess body fat, is an inflammatory disease. Several factors seen in obese people, including chronic inflammation, can induce oxidative stress in these people. Within this context, antioxidants of natural origin are pointed out as promising agents to be incorporated in the treatment of obesity. The edible red algae Gracilaria birdiae synthesizes antioxidant sulfated polysaccharides. The Baccharis trimera plant is used in folk medicine for the treatment of obesity. Thus, the present study aimed to evaluate sulfated polysaccharides from the red alga G. birdiae (GB) and extracts from the leaves of B. trimera (BT), as well as the blends made from these products as antioxidants and inhibitors of lipid accumulation. In the case of blends, the objective was also to verify whether the blends made from these products could optimize such biological activities in vitro and in vivo. GB was obtained by proteolysis of the alga. BT was obtained by decoction. GB showed in vitro antioxidant capacity in five assays. GB was not cytotoxic and inhibited the differentiation of preadipocytes (3T3-L1), as well as decreased the accumulation of lipids in these cells. In studies with Caenorhabditis elegans, GB reduced the levels of reactive oxygen species (ROS) and increased the longevity of animals under stress conditions. In addition, it decreases the content of triglycerides. Phytochemical analysis of BT showed rutin, apigenin and 5-caffeoylquinic acid as the main components of BT. The data showed that BT is not toxic to 3T3 fibroblasts, nor to C. elegans. BT reduced the accumulation of lipids in worms submitted to diets rich or not in glucose. Furthermore, using RNA interference (RNAi), it was observed that BT depends on the transcription factor NHR-49 to exert its anti-obesity effect on worms. Through the formulation of blends, compounds with different concentrations of GB and BT were obtained. In most in vitro antioxidant tests, the blends GB:BT (50:50%) called GB50-BT50 and GB:BT (75:25%) (GB25-BT75) were more potent than GB and BT. ROS production in C. elegans was reduced mainly with GB50-BT50 and GB25-BT75 (5,0 mg/mL). The animals that showed the most significant reduction in fat inhibition were those treated with BT and GB25-BT75. Specifically for triglycerides, the greatest reduction was observed in animals treated with the GB25-BT75 blend. ROS quantification assays were also performed in knockdown animals for SKN-1 and DAF-16 transcription factors using RNAi and survival assay under oxidative stress conditions in skn-1(zu67) mutant animals. Still using RNAi, the mechanisms involved in the accumulation of lipids for the TUB-1 and NHR-49 transcription factors were evaluated. The results showed that the effects provided by the treatment with the blends in reducing oxidative stress were dependent on the transcription factor SKN-1. Regarding the inhibition of lipid accumulation, the main transcription factor involved was NHR-49. In view of the results presented, the potential antioxidant activity and inhibitor of the accumulation of lipids of GB and BT, separately, and the potential of the therapeutic action of these components when combined together in the form of blends were proved.
10	JALUZA LUANA CARVALHO DE QUEIROZ Carotenoid encapsulation: study of the effect on antioxidant and anti-inflammatory activity on adipose tissue Advisor : ANA HELONEIDA DE ARAUJO MORAIS COMMITTEE MEMBERS : ANA HELONEIDA DE ARAUJO MORAIS CRISTIANE FERNANDES DE ASSIS ISABEL RODRIGUEZ AMADO PRISCILLA VANESSA FINOTELLI SERGIO ADRIANE BEZERRA DE MOURA THAIS SOUZA PASSOS Data: Oct 27, 2022 Show Abstract Carotenoids act by reducing oxidative stress and, therefore, act on the inflammatory process. Encapsulation can preserve and/or enhance the functional properties of these natural pigments, such as antioxidant activity. In the present study, divided into chapters, we sought to evaluate how encapsulation acted to improve the antioxidant and anti-inflammatory activity of carotenoids. The first and second chapters, referring to the systematic review (SR), are organized as follows: the first presents the SR protocol with an international prospective registry of systematic reviews (PROSPERO, under number CRD42020142065); and the second, RS, which brought together and systematized studies in vitro, ex vivo, in vivo, which evaluated the effect of different carotenoid encapsulation techniques on antioxidant activity. The articles were selected according to the PICOS strategy (population, interventions, control, results and type of study) and searches were performed in PubMed, Embase, Virtual Health Library, Scopus, ScienceDirect and Web of Science databases. The methodological quality assessment was performed using the OHAT (Office of Health Assessment and Translation) tool. A total of 1577 articles were selected, resulting in 20 eligible studies. The results showed that encapsulation promoted changes in the chemical structure of carotenoids and the emergence of new chemical interactions, in addition to increasing the surface area, resulting in the preservation and enhancement of antioxidant activity. The third chapter refers to the experimental study, which evaluated the effect of the crude extract rich in carotenoids obtained from the Cantaloupe melon (Cucumis melo L. var cantalupensis) (EB) nanoencapsulated (EGS) in Wistar rats with inflammation induced by the consumption of high glycemic index and high glycemic load (HGLI) diet on systemic and adipose tissue inflammatory response. The EB was characterized by spectrophotometric and chromatographic methods, and the EGS was characterized by physical and chemical methods, as well as the evaluation of the incorporation efficiency. The in vivo experiment was carried out for 11 days with male Wistar rats (n = 20) with diet-induced inflammation, subdivided into four groups: no treatment (HGLI diet + water), conventional treatment (HGLI diet + nutritionally adequate diet), test 1 [HGLI and EB diet (12.5 mg/kg)] and test treatment 2 [HGLI and EGS (50 mg/kg)]. The groups were investigated in relation to food consumption, caloric intake, caloric efficiency and weight. Inflammatory cytokines were measured in plasma (TNF-α, IL-6 and leptin) and in adipose tissue (TNF-α and IL-6). In addition, the weights of visceral adipose tissue (retroperitoneal, epididymal and perirenal) were recorded, histopathological analyzes were performed, and investigation of the antioxidant activity in plasma and visceral adipose tissue through the quantification of malondialdehyde, sulfhydryls and superoxide dismutase (SOD) activity). For the in vivo experiment, no changes were observed in food consumption, intake, caloric efficiency and weight (p > 0.05). However, the animals treated with EGS had a lower mean variation in dietary intake and were the only group that showed only weight loss. This group also showed improvement (p < 0.05) in plasma concentrations of inflammatory cytokines (IL-6 and leptin), however, there was no difference in the dosage in adipose tissue. In visceral adipose tissue, it was found that the largest focal area of multilocular adipocytes was observed for the EGS-treated group. In addition, EGS may be acting in a mimetic way to SOD and, therefore, reducing oxidative stress. Thus, the present study presents a relevant scientific contribution, as RS generated information regarding the effect of nanoencapsulation on antioxidant activity and, in the pre-clinical study, on inflammation.
11	YARA CAMPANELLI DE MORAIS Blends of sulfated polysaccharides and chromium picolinate: production and evaluation of their antioxidant activity Advisor : HUGO ALEXANDRE DE OLIVEIRA ROCHA COMMITTEE MEMBERS : HUGO ALEXANDRE DE OLIVEIRA ROCHA CARLOS EDUARDO BEZERRA DE MOURA GABRIEL PEREIRA FIDELIS MARINA DE OLIVEIRA CARDOSO MACEDO PABLO DE CASTRO SANTOS Data: Nov 30, 2022 Show Abstract The red algae Gracilaria birdiae (GB) is cultivated and used as food in Northeastern Brazil. However, the economic potential of this alga has been little explored. One of its main components, a sulfated agaran, here called SPGb, is indicated as an agent with several properties, including as an antioxidant agent. Chromium picolinate (ChrPic) is a bioinorganic compound with recently described anti-inflammatory capacity. Blends are mixtures of two or more agents whose product tends to have greater activity/efficiency than the products that originated it. There is no record of blends containing SPGb and ChrPic.Therefore, the objective was to produce blends containing SPGb and ChrPic and to evaluate them as antioxidant agents. ChrPic was purchased commercially. SPGb was extracted from the alga after it was exposed to ultrasound waves and proteolysis and its identity was confirmed by chemical analysis and nuclear magnetic resonance (1H1D and HSQC). ChrPic (from 0.5 to 2.0 mg/mL) did not show copper or iron chelating activity, whereas SPGb (2.0 mg/mL) showed activity of 70 and 73% for these tests, respectively. On the other hand, ChrPic (1.0 mg/mL) showed about 80% and 100% scavenging activity for superoxide and hydroxyl radicals, respectively. While SPGb (2.0 mg/mL) showed no activity. Five blends were produced (B1; B2; B3; B4; B5) whose antioxidant activity was evaluated and data indicated B5 (SPGb:ChrPic; 1:1) as the most potent blend.Micronucleus test (CBMN) showed that B5 (0.1 to 0.4 mg/mL) has no genotoxic activity. B5 (0.1 to 0.4 mg/mL) and SPGb (0.02 to 0.05 mg/mL) were not cytotoxic to murine fibroblasts (3T3) and Chinese hamster ovary cells (CHO-K1). ChrPic (0.2 mg/mL) decreased by 30% the ability of cells to reduce MTT. When 3T3 cells were exposed to H2O2 (0,6 mM) they showed about 50% of the ability to reduce MTT compared to cells not exposed to peroxide. The presence of H2O2 together with SPGb or ChrPic decreased the cells' ability to reduce MTT. On the other hand, the presence of B5 (0.05 mg/mL) protected the cells from the action of H2O2, since the MTT reduction by these cells was ~100%. When cells were exposed to SPGb, ChrPic or B5 for 24 h, and subsequently to H2O2 (0,6 mM), it was found that the presence of SPGb was unable to prevent the decrease in the ability of cells to reduce MTT, while cells exposed to ChrPic (0.025 mg/mL) and B5 (0.05 mg/mL) reduced MTT by 100%. In another experiment, cells were exposed to peroxide and then exposed to SPGb, ChrPic or B5 for 24h. In this case, again SPGb was not effective in protecting cells from peroxide, while cells exposed to ChrPic (0.025 mg/mL) and B5 (0.05 mg/mL) reduced MTT by 100%. The data showed that B5 has antioxidant activity greater than SPGb and similar activity to ChrPic, except in the test of cell exposure to peroxide concomitantly with B5, whereas the activity of B5 was 3 times higher than activity of ChrPic. Furthermore, in the presence of B5, no signs of cytotoxicity were identified, while in the presence of ChrPic it was identified. These data point to B5 as a product to be used in future in vivo tests to confirm its antioxidant action. This blend may also be a possible nutraceutical.
12	ADRIANA MARINA E SILVA PARENTE Structural evaluation and antimicrobial activity of analog peptides from Stigmurin Advisor : MATHEUS DE FREITAS FERNANDES PEDROSA COMMITTEE MEMBERS : HUGO ALEXANDRE DE OLIVEIRA ROCHA LUDOVICO MIGLIOLO MATHEUS DE FREITAS FERNANDES PEDROSA RICHELE JANAINA ARAUJO MACHADO SYLVIE CHEVALIER LAURENCE Data: Dec 16, 2022 Show Abstract Antimicrobial peptides are molecules considered one of the first defense lines against microorganisms. From the transcriptome of a scorpion venom gland, our research group identified the antimicrobial peptide Stigmurin, from which amino acid residues were changed and two analog peptides denominated StigA6 and StigA16 were designed and studied. These peptides presented in vitro activity against Gram-positive and Gram-negative bacteria higher than the native peptide. To deepen the knowledge on these peptides here we studied their antibacterial activity against multidrug-resistant bacteria in vitro and in vivo assays of larval infection and mice polymicrobial sepsis. The structures of these peptides were evaluated when interacting with lipid vesicles by circular dichroism and by liquid-state nuclear magnetic resonance (NMR). Regarding their in vitro antibacterial activity, StigA6 and StigA16 showed high activity against multidrug-resistant S. aureus and P. aeruginosa, and antibiofilm effect on multidrug-resistant S. aureus. Both peptides were able to control infections in Galleria mellonella larvae, increasing survival and reducing colony-forming units, hemocytes and melanization. In mice polymicrobial sepsis, the analog peptides were able to reduce viable microorganisms, leukocytes migration and myeloperoxidase activity. As for their interaction with lipid vesicles both peptides showed higher interaction with bacterial membrane-mimetic vesicles than with eukaryotic membrane-mimetic vesicles. NMR data showed that both analog peptides presented high amphipathicity and longer helical structure than the native peptide, Stigmurin, suggesting that lysine placement is important to helix determination. Therefore, these peptides are interesting targets in the study of new anti-infective molecules.

2021

	Dissertations
1	MARIA FERNANDA BEZERRA DE SOUZA MOLECULAR MARKERS DEVELOPED FOR IDENTIFICATION OF Plasmodium sp. Advisor : DANIEL CARLOS FERREIRA LANZA COMMITTEE MEMBERS : DANIEL CARLOS FERREIRA LANZA PAULO MARCOS DA MATTA GUEDES PATRICIA DE ABREU MOREIRA Data: Aug 9, 2021 Show Abstract Early, the Plasmodium genus had only 4 species related to human malaria: P. vivax, P. falciparum, P. malariae and P. ovale. With the advancement of molecular tools and bioinformatics, there has been an increase in the number of species that cause this disease in humans such as P. knowlessi, P. cynolmolgi and P. simiun. The polymerase chain reaction (PCR) has been considered the most effective among the detection techniques. Most of the PCR protocols already described for the identification of Plasmodium sp. targets the ribosomal RNA 18S subunit gene.However, primers for the identification of P. vivax, P. falciparum, P. malariae and P. ovale generally cross-react with other species, species that can also cause malaria in humans. The pattern of evolution of the 18S gene in the apicomplex group is marked by copies that resemble each other by species, by genus (Plasmodium) and no apicomplex group. Despite the great efficiency of Primers for 18S rDNA, this target does not enable differentiation of Plasmodium species with great efficiency.Thus, a complementary system was developed, based on another molecular marker, for the identification of Plasmodium species. Available sequences were selected for six species that cause malaria in humans. Thus, the sequences of the several of genes were investigated. The validation of the molecular marker for Plasmodium was based on the specificity and the number of sequences available for all species. Among them, the gene chosen for the design of the primers was MSP1 (merozoite surface protein). The Plasmodium identification system was built of way to produce amplicons of different size for each species, making it possible to identify the species without the need for sequencing the amplicon. Then, due to the particularity of the 18S, a nested-PCR already proposed for the detection of apicomplex was validated for an initial screening. And a nested-PCR system was developed for P. vivax and P. falciparum, the two most recurrent species in Brazil, that were tested in vitro. Through verification of amplification, both from samples extracted from cell culture and from DNAs from patient samples. The specificity in gender was confirmed through verifying the non-amplification from samples of Plasmodium berghei, a rodent plasmid. Furthermore, it was observed that there is no cross-amplification between P. vivax and P. falciparum samples.
2	NADJA MARIA DA COSTA MELO CHEMICAL CHARACTERIZATION AND CHEMOPREVENTIVE EFFECT OF LYOPHILLED HYDROETHANOLIC EXTRACT FROM Cereus jamacaru P.DC. (MANDACARU) IN EXPERIMENTAL MODEL OF INFLAMMATORY BOWEL DISEASE Advisor : GERLANE COELHO BERNARDO GUERRA COMMITTEE MEMBERS : GERLANE COELHO BERNARDO GUERRA ANA HELONEIDA DE ARAUJO MORAIS RENAN OLIVEIRA SILVA DAMASCENO Data: Sep 28, 2021 Show Abstract Inflammatory bowel diseases (IBD) are chronic inflammatory pathologies associated with genetic alterations and environmental factors. The drugs available for treatment can cause potentially serious adverse effects, showing the need to search for new therapeutic strategies. Foods rich in bioactive compounds with anti-inflammatory activity stand out, such as Cereus jamacaru P. DC. This research aimed to chemically characterize and evaluate the anti-inflammatory effect of the Lyophilized Hydroethanolic Extract of Cereus jamacaru P.DC. in an experimental model of inflammatory bowel disease induced by 2,4-dinitrobenzene sulfonic acid (DNBS). The mandacaru cladodes were collected in the city of Jucurutu/RN, the thorns were removed, cut and dried, then they were ground, going through maceration with ethanol and water, obtaining the Lyophilized Hydroethanolic Extract of Cereus jamacaru (EHCJ). The phytochemical prospection of EHCJ was carried out by liquid chromatography coupled to a mass spectrometer to assess the presence of phenolic compounds. In the study of the intestinal anti-inflammatory activity were used Rattus norvegicus (Wistar), adult females (n = 8/group), weight 230 ± 20 g, randomly distributed in the groups: normal control, DNBS collitic control, sulfasalazine 250 mg/Kg and EHCJ (50, 100 and 200 mg/kg/day). Intestinal, macroscopic and microscopic clinical parameters, volatile compounds and fatty acids present in feces, inflammatory markers such as myeloperoxidase enzyme (MPO) dosage, gene expression were evaluated mitogen activated protein kinase (MAPK) and metalloproteinase 9 (MMP-9), expression of the kappa beta factor gene and protein (NF-κB p65); cytokine dosages (TNF-α, IL-10 and IL-1β; evaluation of oxidative stress by measuring malondialdehyde (MDA) and reduced glutathione (GSH); integrity of the intestinal barrier through the expression of the occludin gene (OCL) and expression of the mucin 2 (MUC-2) and occlusion zone (ZO-1) proteins. The results of the phytochemical analysis identified the compounds derived from benzoic acid, saccharide, Kampferol-3-O-dirhamnosyl-hexoside, Isorhamnetin-3-O-dirhamnosyl-glucoside, Kaempferol-3-O-rutinoside, Isorhamnetin-3-O-rutinoside and oxo-dihydroxy-octadecenoic acid. The GC-MS of feces identified 18 compounds, which were fatty acids, hydrocarbons, sterols and phenol according to the fragmentation patterns. In the colitis model, pretreatment with EHCJ at doses of 50, 100 and 200 mg/kg was able to cause chemoprevention observed through the Disease Activity Index (IAD) (p < 0.05). Furthermore, it reduced the colonic damage induced by DNBS with a reduction in the macroscopic damage score (p<0.05). The treatment reduced MPO enzyme activity (p<0.05) and colonic MDA levels (p<0.01) and increased GSH levels (p<0.05) for all doses tested. The extract also reduced tissue levels of the cytokines TNF-α (p<0.0001) and IL-1β (p<0.05) and increased IL-10 at a dose of 200 mg/Kg (p<0.01 ). The preventive effect was also demonstrated by the increased expression of the OCL gene, involved in the integrity of the intestinal barrier and decreased expression of the markers of the MAPK (p<0.01) and NF-κB p65 (p<0.0001) pathways. Histopathological analyzes show that EHCJ significantly reduced the infiltrate density (p<0.001) and immunohistochemistry exhibited increased immunoreactivity for MUC-2 (p<0.01) and ZO-1 (p<0.05) and a reduction of this immunostaining for NF-κB proteins p65 (p<0.05) and MMP-9 (p<0.05). Thus, preclinical results indicate that EHCJ is a product rich in bioactive compounds, beneficial in the chemoprevention of colitis, and may, in the future, become an alternative in the treatment of human IBD.
3	MELINE GOMES GONÇALVES EVALUATION OF THE EFFECT OF CREATINE SUPPLEMENTATION: A PRECLINICAL STUDY WITH A MODEL OF STREPTOZOTOCIN-INDUCED TYPE I DIABETES Advisor : JOAO PAULO MATOS SANTOS LIMA COMMITTEE MEMBERS : JOAO PAULO MATOS SANTOS LIMA BENTO JOAO DA GRACA AZEVEDO ABREU DIEGO NEVES ARAUJO Data: Oct 25, 2021 Show Abstract Diabetes Mellitus (DM) is a chronic metabolic disease growing worldwide. Researches are constantly conducted in an attempt to alleviate the complications of DM and increase the quality of life of patients. Creatine is a nutritional supplement that has been studied for this purpose due to its antioxidant and hypoglycemic features. However, there are no studies reporting the effect of creatine on both metabolism and tissues that may be affected in type I DM. Thus, this study aimed to evaluate the effects of creatine supplementation on symptoms, biochemical and histopathological parameters of the pancreas and kidneys in diabetic rats induced by streptozotocin (STZ). 32 Wistar rats were randomly divided into 4 groups, containing 8 animals each (n=8): (C) normoglycemic animals without creatine supplementation, (CCr) normoglycemic animals with creatine supplementation, (D) diabetic animals induced with STZ without creatine supplementation, (DCr) diabetic animals induced with STZ and supplemented with creatine. Groups D and DCr received a single dose of STZ (40 mg/kg i.p.) for diabetes induction. The CCr and DCr groups received creatine supplementation through isocaloric ration in two phases (saturation, five days before DMI induction, and maintenance, during the 35 days of the experiment, with 13% and 2% creatine, respectively). In order to assess the symptomatology of the pathological condition for all experimental groups, daily water and feed consumption as well as weekly body weight were measured. After anesthesia and euthanasia of the animals, blood and organs were collected and stored for analysis of biochemical parameters [fasting glucose, creatinine, serum urea, aspartate transaminase (AST) and aspartate aminotransferase (ALT)] and histopathological parameters of pancreatic tissues and renal in the animals. In addition to hyperglycemia throughout the experiment, polydipsia, polyphagia and decreased body weight were observed in STZ-induced diabetic animals. Creatine supplementation was able to reduce in animals with DMI and glycemia (p<0,05), the same serum urea and ALT in the DCr group compared to the D group (p<0,01). Histopathologically the pancreatic tissue, where it was observed that the animals in the DCr group did not differ statistically from the animals in the C and CCr groups (p>0,05). In this same group (DCr), the animals dissipated from the islet area compared to group D (p<0,01). For renal tissue the animals in the DCr group reduced reduction in renal glomerulus count compared to the other experimental groups (p<0,05). As for the glomerular areas, the animals in the DCr group were similar to the C and CCr groups, while the animals in the D group had smaller areas of the renal glomeruli (p<0,05). Creatine supplementation in STZ-induced diabetic rats, which induces the DMI experimental model, despite having attenuated the biochemical parameters of glycemia, urea and AST, as well as showing a maintenance and protection of pancreatic histomorphometry, was not able to cause beneficial effects to the DCr group regarding the tissue consequences of DM. On the contrary, CCr has histopathological characteristics referring to tissue damage from both pancreatitis and renal tubular necrosis. Therefore, a creatine supplementation is not feasible in this unpublished experimental archetype. It is evident that more investigations are needed for evidence to add as to its morphological and metabolic effects.
4	JULIETE TAVARES DE ARRUDA OLIVEIRA Effects of sucrose exposure and withdrawal on depression-related behavior in adolescent rats Advisor : VANESSA DE PAULA SOARES RACHETTI COMMITTEE MEMBERS : BRUNO LOBAO SOARES VALQUÍRIA CAMIN DE BORTOLI VANESSA DE PAULA SOARES RACHETTI Data: Nov 11, 2021 Show Abstract Refined sugar (sucrose) is very versatile and cheap and can be found in many recipes. A diet high in sugar can have negative consequences, such as obesity and diabetes, and its withdrawal can promote behavioral changes, such as binge eating. The aim of this research was to test the hypothesis that sucrose withdrawal favors depression-related behaviors. For this purpose, 30-day Wistar rats from the vivarium of the Centro de Biosciences-UFRN were used, receiving water and feed ad libitum and were submitted to the paradigm of choosing two bottles with 250 ml of water and/or 5% sucrose (group sucrose) for 16 days, followed by its withdrawal from 72h to 96h (short-term withdrawal group) or 22 to 23 days (long-term withdrawal group). Animals in the control group, sucrose, short-term withdrawal and long-term withdrawal were submitted to the open field test 72h and 22 days after the replacement of sucrose by water for the short-term and long-term withdrawal groups, respectively. Twenty-four hours after the open field test, all groups were submitted to the forced swim test. The data obtained here showed no behavioral effects of exposure or withdrawal in the short and long term of sucrose in the forced swimming test, as well as no statistical differences in the results of the locomotion parameter in the open field test. Thus, we conclude that short- and long-term exposure and withdrawal of sucrose were not able to generate depressive-like behavior in adolescent rats.
5	RODRIGO THIAGO DA SILVA EFFECTS OF SUCROSE EXPOSURE AND WITHDRAWAL ON ANXIETY-RELATED BEHAVIORS IN ADOLESCENT RATS Advisor : VANESSA DE PAULA SOARES RACHETTI COMMITTEE MEMBERS : MANUELA DOS SANTOS CARVALHO SCHIAVON RAMÓN HYPOLITO LIMA VANESSA DE PAULA SOARES RACHETTI Data: Nov 12, 2021 Show Abstract Adolescence is a critical period for the development of the central nervous system (CNS) and neuronal plasticity processes may be sensitive to environmental challenges. Evidence shows that exposure to sucrose affects brain development, even promoting behavioral changes. The aim of this study was to test the hypothesis that sucrose exposure and withdrawal alter anxiety-related behaviors in adolescent rats. Thirty-days-old Wistar rats obtained from the Animal Facility of the Biosciences Center – UFRN received water and feed ad libitum throughout the experimental period. Animals divided in the following groups were submitted to the elevated plus maze test (EPM) for 5 minutes, individually: control group (animals that received water as a source of liquid throughout the experiment), sucrose group (animals that received sucrose 5% as an additional source of liquid for 16 days, with access to sucrose until the day of the EPM test), short-term withdrawal group (animals that received sucrose 5% as an additional source of liquid for 16 days, with sucrose replaced by water 48 hours prior to the EPM test) and the long-term withdrawal group (animals that received sucrose 5% as an additional source of liquid for 16 days, with sucrose replaced by water 21 days prior to testing in the EPM). Twenty-four hours after the test in the EPM, the animals of all experimental groups were submitted to the open field test. The results showed that the exposure as well as the removal of sucrose in adolescent rats did not alter anxiety-related behaviors in the EPM and in the open field tests.
6	EMANOELLY ROBERTA DE CARVALHO MORAIS Flowering in sugarcane – identifying the role of HINT1 and SUBITILASE Advisor : KATIA CASTANHO SCORTECCI COMMITTEE MEMBERS : CARLOS HENRIQUE SALVINO GADELHA MENESES DAIANE CRISTINA FERREIRA GOLBERT KATIA CASTANHO SCORTECCI Data: Dec 21, 2021 Show Abstract Flowering is a critical stage that promotes the transition of shoot apical meristem from the vegetative to reproductive stage. This transition is orchestrated by different signal (external and internal) that have been characterized in different models as Arabidopsis thaliana, Oryza sativa and Zea mays. For sugarcane this process is not well-known in molecular way. However, it has an important impact in yield and production. Considering this, the work present here has the aim to characterize a sequence that had homology to HINT1 protein by different approaches. This sequence was previously identified using subtractive libraries from shoot apical meristem during different stages of sugarcane development. It was verified that HINT sequences were conserved in plants, and it was separate into two large branches: monocotyledons and dicots. The data obtained from the 3D modeling showed that ScHINT may be a functional protein, as it presents the important binding sites for its catalytic activity. A second protein that was worked was ScSUBTILASE. This protein was characterized had been previously identified in a protein-protein assay using the two-hybrid methods. Here, it was attempted to understand the role of SUBTILASE and HINT1. It was verified that the ScSUBTILASE protein was also conserved in plants, and the modeling showed that it might also be functional due to the 3D structure and the presence of catalytic sites. In the two-hybrid assays, a third protein was identified with homology to PHENYLALANINE AMMONIA-LYASE. The modeling was carried out and it was observed that this too must be functional. The other approached used was interactome using STRING. The results obtained for the proteins HINT, SUBTILASE and PHENYLALANINE AMMONIA-LYASE in the Arabidopsis model showed that these sequences are associated with the response of the production of ROS. Based on the results presented and on the literature data, it may be proposed that ScHINT and ScSUBTILASE may have the role of maintaining the level of ROS at the shoot apical meristematic during the transition process. The results obtained with transgenic Nicotiana tabacum plants containing over-expression cassettes in sense and antisense orientations for ScHINT1 reinforce the proposal idea that ScHINT may be associated with the flowering process.
7	GABRIELLA SILVA CAMPOS CARELLI INFLUENCE OF CAFFEINE ON PROTEIN PROFILE AND FEASIBILITY Escherichia coli CELL PHONE AND ADJUVANT POTENTIAL FOR ANTIBIOTICS Advisor : ELIZEU ANTUNES DOS SANTOS COMMITTEE MEMBERS : ANDERSON FELIPE JÁCOME DE FRANÇA ELIZEU ANTUNES DOS SANTOS VINÍCIUS CAMPELO SOEIRO Data: Dec 24, 2021 Show Abstract Caffeine is a substance found in over 100 species of plants. In addition to its neurostimulant effects, its antimicrobial activity has already been proven by science. In this context, the aim of this work was to test the influence of caffeine on the protein profile and cell viability of Escherichia coli and its adjuvant potential for antibiotics in vitro. Commercially acquired caffeine was solubilized in Mueller Hinton broth and distilled water, 10 mL of this solution were placed in seven (7) test tubes at different final concentrations of caffeine. After performing a concentration curve, it was evidenced that with the use of a concentration of only 0.20 mg/mL, caffeine allowed a partial bacterial growth. To perform the extraction of proteins from E. coli, two different methods were compared, one with acetone pa and the other with glass beads. By analyzing the SDS-PAGE and the densitometry of the extraction, it was concluded that the method with acetone was the most satisfactory. For the other tests, three generations of the bacteria were cultivated. Bacterial strains cultivated with caffeine showed alterations in the protein profile when compared to those cultivated in the absence of this substance, this fact was verified through SDS-PAGE and optical densitometry. After concluding that caffeine had an influence on the protein profile of the bacteria studied, its potential as an adjuvant to the antibiotics ampicillin and streptomycin was tested, the results of the adjuvant trial showed that caffeine was more effective when combined with ampicillin, reducing the inhibitory concentration minimum of this antibiotic from 0.250 mg/mL to 0.125 mg/mL. The combinations of caffeine and streptomycin were not statistically significant. Taken together, the data found suggest that caffeine, alone or in combination with ampicillin, is a potential candidate for applications in antibacterial therapies.
	Thesis
1	AMANDA FERNANDES DE MEDEIROS Functional aspects of vegetable peptides with anti-inflammatory effect on obesity and structural evaluation of the kunitz tamarindo seeds inhibitor Advisor : ANA HELONEIDA DE ARAUJO MORAIS COMMITTEE MEMBERS : ANA HELONEIDA DE ARAUJO MORAIS BRUNA LEAL LIMA MACIEL HERMÓGENES DAVID DE OLIVEIRA MATHEUS DE FREITAS FERNANDES PEDROSA RICHELE JANAINA ARAUJO MACHADO SEVERINA CARLA VIEIRA CUNHA LIMA Data: Oct 25, 2021 Show Abstract Subclinical inflammation in overweight and obesity alters several metabolic pathways, with positive reinforcement for the accumulation of more adipose tissue and alterations in energy metabolism. Several studies have evaluated the anti-inflammatory and immunomodulatory effects of hydrolyzed proteins and plant peptides. However, it is still necessary to explore the mechanisms of action. Thus, in the first chapter of this thesis, a narrative review is presented, aiming at understanding the mechanisms of action of proteins and peptides on inflammation, present in the accumulation of adipose tissue. Thus, a search was carried out in databases, selecting studies that involved target metabolic pathways and responses of proteins or peptides, which favored the reduction of inflammatory cytokines and adipokines, as well as the polarization of macrophages to the M2 phenotype. In the second chapter, the in vitro and computer simulation studies (in silico) study with the purified trypsin inhibitor from tamarind seeds (pTTI) is presented. This protein has been studied in vitro and preclinical studies for the treatment of obesity, its complications, and associated comorbidities. The pTTI was sequenced again by MALDI-TOF/TOF, its homology modeling was obtained, and the interaction with the trypsin enzyme was evaluated through molecular dynamics (MD) simulation under physiological conditions. A further 75 amino acid residues have been identified. Homology modeling was performed by CONCOORD and validated by MolProbity and the four best conformations of the modeling were submitted to DM. Conformation No. 287 of model No. 56 was selected, considering the RMSD analysis and the interaction energy (-301.0128 kcal.mol-1). Residues Ile (54), Pro (57), Arg (59), Arg (63) and Glu (78) of the pTTI presented lower interaction energy, which reflects the residues with greater interaction with the enzyme trypsin and the residues of Arg are mainly involved in its electrostatic binding mechanism. Once aware of potential molecular targets of plant peptides for the treatment of obesity revealed through the narrative review, in addition to the structure and function of the pTTI through the in vitro and in vivo study, this in silico study demonstrated optimization of bioprospecting of pTTI through bioinformatics. Finally, the results found and the paths addressed favor the continuity of studies of this protein for applications in pre-clinical and clinical studies in the health area, aiming to contribute to the control of obesity, a serious public health problem.
2	JÉSSICA TEIXEIRA JALES Terbufos (Organophosphate) in Diptera of forensic importance: a biological and biospectroscopic approach Advisor : RENATA ANTONACI GAMA COMMITTEE MEMBERS : BRUNO CAVALCANTE BELLINI CARINA MARA DE SOUZA MARGARETH MARIA DE CARVALHO QUEIROZ RENATA ANTONACI GAMA VALTER FERREIRA DE ANDRADE NETO VANESSA DE PAULA SOARES RACHETTI Data: Dec 15, 2021 Show Abstract Entomotoxicology studies the effect and detection of chemical substances in scavenger insects from crime scenes, being crucial for the correct estimation of the interval and causes death of decaying carcasses and cadavers. Among substances of forensic importance, Terbufos (Organophosphate) is frequently associated with cases of intoxication in Brazil and worldwide. Thus, this work aims to understand the effect of Terbufos: i. in the carrion decomposition and assemblage of sarcosaprophagous dipterans associated with carcasses intoxicated with different doses of the compound; ii. in the carcass colonization process and its implications for the calculation of the postmortem interval (PMI); iii. in the post-feeding larval dispersal behavior of scavengers and its implications as entomological evidence, as well as iv. the use of medium infrared spectroscopy (ATR-FTIR) to detect Terbufos in larvae and puparium from intoxicated rats. For this, female Wistar rats received, via gavage, 200uL of Terbufos (5mg/kg, 10mg/kg or 20mg/kg) or distilled water (control) and, after 30 minutes, were euthanized and distributed in suspended traps to decompose into environmental conditions. The decomposition was monitored daily, with photographic record and collection of visiting and colonizing dipterofauna, until the dry phase. The dispersing larvae were also used for behavior analysis and spectra collection. The data collected showed that Terbufos accelerates the decomposition of carcasses intoxicated with 10mg/kg in 24h, and changes the composition and structure of the visiting and colonizing dipterofauna, and the behavior of immature intoxicated with high doses of Terbufos. In addition, biospectroscopy has been shown to be a promising tool for toxicological analysis of intoxicated insects. Through a multidisciplinary approach, this work strengthens the importance of entomotoxicological studies to evaluate entomological evidence at crime scenes and can assist in investigative processes with suspected organophosphate poisoning.
3	JENIELLY DE NORONHA FERREIRA DE CASTRO BIOTECHNOLOGICAL POTENTIAL OF BACTERIA ISOLATED FROM CONSORTIUMS FROM PRODUCTION WATER IN OIL RESERVOIRS IN BRAZIL Advisor : LUCYMARA FASSARELLA AGNEZ LIMA COMMITTEE MEMBERS : LUCYMARA FASSARELLA AGNEZ LIMA ADRIANA FERREIRA UCHOA FRANCISCO CANINDE DE SOUSA JUNIOR LUCIANE MARIA PEREIRA PASSAGLIA MARILENE HENNING VAINSTEIN Data: Dec 22, 2021 Show Abstract During the extractive processes of oil and gas, production water is also formed, which is composed of or ery). ganic and inorganic matter, in addition to microorganisms. The microbiota of this environment, adapted to extreme conditions, has great potential for application in biotechnological processes, such as bioremediation and/or MEOR (Microbial Enhance Oil Recov Within this context, this study aimed at the isolation, identification and characterization of bacteria from three consortia, previously obtained from production water. A total of 16 bacteria were isolated, fifteen of them selected for metabolic char acterization from functional assays. Isolates were selected according to their ability to use hydrocarbons as a carbon source. Considering the evaluation of hydrocarbon degradation, the test with redox indicator 2,6 DCPIP (2,6Dichlorophenolindophenol) w as used and tests such as Emulsification Index, drop collapse and oil dispersion were applied to evaluate the production of biosurfactants / bioemulsifiers. The results obtained from the degradation tests confirmed the capacity of fifteen isolates to use p etroleum and other hydrocarbons as the only source of carbon. The tests carried out for the production of biosurfactants confirmed the capacity of 15, among the sixteen isolates, to produce biosurfactants and/or to form an emulsion. Based on the 16S gene sequence, the isolates were identified as Bacillus, Acinetobact er, Staphylococcus, Ochrobactrum and Citrobacter . Among the isolates, four were selected and had their genomes sequenced, being identified as Ochrobactrum (1) and Acinetobacter baumanii Bacillus safensis (1). Isolate AP1BH01(2), 1, classified as a member of the genus Ochrobactrum , was designated as a new species, since it does not group with another species of the genus. The genomes of the sequenced isolates showed a great diversity of genes involved in the degradation of aliphatic and aromatic hydrocarbon s, as well as genes for the synthesis of biosurfactants and bioemulsifiers. Several genes related to resistance to heavy metals have also been identified. The data obtained through functional assays and genome mining of the isolates revealed promising prof iles in hydrocarbon degradation processes and production of biosurfactants for application in bioremediation of environments contaminated with hydrocarbons and/or MEOR.

2015

	Dissertations
1	JOÃO PAULO DE FREITAS NUNES Evolução e especialização funcional da Acil-CoA oxidase 4 em Viridiplantae Advisor : JOAO PAULO MATOS SANTOS LIMA COMMITTEE MEMBERS : CÍNTIA RENATA COSTA ROCHA DANIEL CARLOS FERREIRA LANZA JOAO PAULO MATOS SANTOS LIMA Data: Jul 28, 2015 Show Abstract Stored oils are essential to the germination and initial seedling establishment of oilseeds plants. After mobilization, these oils are subjected to β-oxidation in seed’s glyoxysomes, resulting Acetyl-CoA, which in turn feeds the Glyoxylate cycle. This cycle is a variation of the tricarboxylic acid cycle that uses acetyl-CoA to produce succinate or oxaloacetate molecules that can be later utilized in carbohydrate biosynthesis to support embryo development. The first step of glyoxysomal β-oxidation is catalyzed by a specific isoform of Acyl-CoA oxidase enzyme (ACX4), a member of the protein superfamily ACAD, which occurs more commonly in germinating seeds. However, five other ACX isoforms are found in plant genomes to date, each one related to specialized functions or fatty acid chain’s sizes. In order to understand the molecular evolutionary events underlying the functional specialization of these enzymes, we analyzed DNA, mRNA and protein sequences obtained in databases (NCBI, UNIPROT, TAIR, CDD, etc.) by bioinformatic inferences, to recognize and compare the introns-exons regions, as well as protein domains of different ACX isoforms from Viridiplantae species. Then the sequences were aligned, the Cis elements of the genes and their exon/domain, secondary structures were compared, submitted to structural modeling and maximum likelihood phylogenetic inferences. It was shown that the ACX4 enzymes are more closer related to other members of the ACAD superfamily, one of them the Acyl-CoA dehydrogenase enzyme (ACDH). Since fatty acids are not commonly used in other plants tissues to energy production and also β-oxidation by ACDH in mitochondria is more related to ATP synthesis, we hypothesized that ACX4 isoform may have been subjected to specific selective pressures, which stabilized its role during seed oil metabolism.
	Thesis
1	RALFO GOES PACCHIONI Requerimentos genéticos da síntese translesão dos adutos de Benzo[a]pireno Advisor : LUCYMARA FASSARELLA AGNEZ LIMA COMMITTEE MEMBERS : CLAUDIA A S LAGE KATIA CASTANHO SCORTECCI LUCYMARA FASSARELLA AGNEZ LIMA RODRIGO DA SILVA GALHARDO TIRZAH BRAZ PETTA Data: Nov 27, 2015 Show Abstract A busca pela manutenção da integridade genômica é uma tarefa constante nas células de todos os organismos vivos. DNA polimerases replicativas atuam de forma rápida e fiel durante a replicação para que a cópia da informação genética passada de uma geração a outra seja fidedigna. Além disso, devido ao genoma ser constantemente agredido por agentes endógenos e exógenos capazes de lesionar a molécula de DNA, são acionados na célula sistemas de reparo de DNA capazes de prevenir os efeitos prejudiciais dessas lesões, uma vez que elas podem levar à mutações. Embora esses mecanismos sejam eficientes na restauração da molécula de DNA, muitas lesões escapam da correção e acabam sendo encontradas pela polimerase replicativa – enzima nem sempre capaz de sintetizar moldes de DNA danificados. Dependendo do tipo de lesão, esse encontro pode resultar no colapso da forquilha de replicação e, consequentemente, mecanismos capazes de burlar esses obstáculos são acionados. Uma das alternativas é a utilização de polimerases de síntese translesão especializadas capazes de replicar as lesões mesmo com o custo da mutagênese, em um processo chamado de Síntese Translesão. Dentre os mais diversos agentes danosos ao DNA está o poluente Benzo[a]pireno, um potente carcinógeno onipresente no ambiente. Esse químico, ao ser metabolicamente ativado na célula, gera os subprodutos BaP+ e BaP- que se ligam como adutos ao DNA de forma covalente, sendo potentes bloqueadores da forquilha de replicação. Nesse trabalho são apresentados alguns dos requerimentos para a ocorrência em E. coli da síntese translesão desses adutos estereoisômeros BaP+ e BaP-, inseridos no contexto de sequência CTGBaP+/-CAG. Os resultados mostram que enquanto o principal aduto BaP+ requer as duas enzimas de síntese translesão cataliticamente ativas Pol IV e Pol V para ser replicado, BaP- exige apenas Pol IV. Dados demonstram que a taxa de síntese translesão desses adutos possui relação positiva com a concentração de polimerases especializadas no meio celular, as quais possuem níveis aumentados pela indução da resposta SOS. Além disso, é sugerido um desbalanço do pool de nucleotídeos que desloca a reação de síntese translesão no sentido da síntese e um modelo de replicação desses adutos.