Banca de QUALIFICAÇÃO: RAPHAEL PASCHOAL SERQUIZ

Uma banca de QUALIFICAÇÃO de MESTRADO foi cadastrada pelo programa.
DISCENTE : RAPHAEL PASCHOAL SERQUIZ
DATA : 29/09/2017
HORA: 13:00
LOCAL: Auditório do Museu de Ciências Morfológicas
TÍTULO:

 

A TRYPSIN INHIBITOR WITH STIMULATING HEPARIN ACTIVITY AND ANTI-INFLAMMATORY POTENTIAL: PARTIAL ISOLATION AND CHARACTERIZATION

PALAVRAS-CHAVES:

Mimosa regnellii; Protease; Anticoagulant; Inflammation; Neutrophil; Macrophage;

PÁGINAS: 68
GRANDE ÁREA: Ciências Biológicas
ÁREA: Bioquímica
RESUMO:

 

Failures in the hemostatic mechanisms of mammals contribute to development of thrombotic diseases. The principal drug for the treatment of thromboses is heparin, which has several adverse effects when used for long periods. Some patients have demonstrated resistance to the anticoagulant effects of heparin, commonly caused by a lack of antithrombin, an endogenous coagulation inhibitor that is potentiated by heparin. Faced with this, the researches for new molecules with potential for substitution or alternation of established clinical treatments are necessary. Protease inhibitors have been shown to be effective in control of protease-dependent metabolic processes, such as clot formation, inflammation and mechanisms of death programmed by various cells. In view of this, the present work provides a partial isolation of a novel trypsin inhibitor (ITJ) from the Mimosa regnellii Benth. seed and characterize its inhibitory activity for different proteases, as well as to detect its ability to interfere in the time of clot formation in the presence and absence of heparin or to block inflammatory activities of neutrophils and macrophages. ITJ was partially isolated by affinity and ion exchange (DEAE) chromatography, presenting only two protein bands of 11.9 and 19.2 kDa in polyacrylamide electrophoresis, and was able to inhibit trypsin and chymotrypsin, but did not show activity against elastase. In coagulation, ITJ extended the clot formation time, at concentrations greater than 2μg for each 100μL of human plasma in the APTT test. The same effect was not observed in the PT test. When preincubated with heparin the inhibitor was able to duplicate the anticoagulant effect of heparin on both APTT and PT. ITJ induced release of elastase by human neutrophils in a discrete manner, but when cells were activated with PAF, the inhibitor reduced more than 40% of elastase release levels. The same effect was not observed when neutrophils were activated with fMLP. In contact with RAW 264.7 macrophages and human erythrocytes, ITJ did not reduce cell viability and stimulated the production of TNF by macrophages without stimulating the production of nitric oxide (NO), IL-1, IL-6 or IL-10. LPS-activated macrophages had a reduction in NO, IL-6 and TNF levels when treated with ITJ, with normalization of cytokine levels at levels between 20-160 μg/mL of inhibitor. The trypsin inhibitor of M. regnellii has been shown to be effective as adjuvant to the anticoagulant effect of heparin and has promoted the reduction of inflammatory stimuli of neutrophils and macrophages. More efficient purification techniques should be employed in the future for total inhibitor isolation and structural characterization, as well as interaction studies of the inhibitor to elucidate the mechanism of heparin supplementation.

MEMBROS DA BANCA:
Presidente - 2578619 - ANA HELONEIDA DE ARAUJO MORAIS
Externo à Instituição - ARIANE FERREIRA LACERDA - UFRN
Externo à Instituição - PAULA IVANI MEDEIROS DOS SANTOS - IFRN