ANALYSIS OF HUMAN MESENCHYMAL STEM CELL TRANSCRIPTOME DURING OSTEOGENESIS
RNAseq, stem cell, Osteoblast, ncRNA.
Human mesenchymal stem cells (hMSC) differentiation into osteoblast follows a specific gene expression program, committing itself initially under Wnt and BMPs pathways influence then differentiating into osteoblasts. However, specifically activated pathways during the first three days of differentiation are still poorly studied. Considering next generation sequencing technologies efficiency, and the lack of characterization in early hMSC differentiation process, in this work we used Illumina RNA-Seq to investigate the changes in these cells transcriptome. We used ex-vivo cultures of two human umbilical cord veins. Data from the complete transcriptome were analyzed in Transcriptogramer for the production of an ordered list of functionally associated genes, obtained by the mean of the gene expression taken on neighboring genes in this list, thus facilitating their biological interpretation. To study gene ontology and gene expression profile design during osteogenesis, the metabolic processes and molecular functions significantly altered during the course of the differentiation process were analyzed in several tools (REVIGO, GOrila, PANTHER, LNCipedia and NONCODE) and properly described. During hMSC differentiation, an increase in expression of osteoblastic phenotype genes was observed as soon as the first day of differentiation started. Noncoding RNAs were also identified, depending on the evolution of the differentiation process, as well as genes involved in the formation of membrane rafts, on the third day of differentiation. During the third day of induction, cell differentiation regulation genes and other important genes on biological processes that precede differentiation, such as cell adhesion, signaling and external chemical factors response, have already increased expression. The study of gene expression during these first three days also revealed the decrease in the expression of groups of genes crucial to basal metabolism maintenance, RNA degradation and cytoskeleton organization, thus indicating that the cellular changes that lead the cell into differentiation may originate during the first three days of osteogenic treatment.