Identification of key pathways and hub genes in the myogenic differentiation of pluripotent stem cell: A bioinformatics and experimental study

Abstract

Background: The regeneration of muscle cells from stem cells is an intricate process and various genes are included in the process as myoD, mf5, mf6 etc. The key genes and pathways in the differentiating stages is various. Therefore, the differential expression of key genes after 4weeks of differentiation were investigated in our study. Method: Three published gene expression profiles, GSE131125, GSE148994, GSE149055, about the comparisons of pluripotent stem cells to differentiated cells after 4 weeks were obtained from the Gene Expression Omnibus (GEO) database. Common differentially expressed genes (DEGs) were obtained for further analysis as proteinprotein interaction (PPI) network, Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) and GSEA analysis. After hub genes and key pathways were obtained, we manipulated in vitro cell research for substantiation as immunohistochemical staining and semi-quantitative analysis and quantitative real-time PCR. Results: A total of 824 DEGs including 350 up-regulated genes and 474 down-regulated genes were identified in the three GSEs. 19 hub genes were identified from the PPI network. The GO and KEGG pathway analysis confirmed that myogenic differentiation at 4 weeks was strongly associated with pathway in cancer, PI3K pathway, actin cytoskeleton regulation and metabolic pathway, biosynthesis of antibodies and cell cycle. GSEA analysis indicated the differentiated cells were enriched in muscle cell development and myogenesis. Meanwhile, the core genes in each pathway were identified from the GSEA analysis. The in vitro cell research revealed that actin cytoskeleton and myoD were up-regulated after 4 weeks differentiation. Conclusions: The research revealed the potential hub genes and key pathways after 4 weeks differentiation of stem cells which contribute to further study about the molecular mechanism of myogenesis regeneration. Paving a way for more accurate treatment for muscle dysfunction.