| 摘要 |
[Objectives] To characterize the expression pattern of Fibroblast growth factor 10 (FGF10) during the differentiation of rat L6 myoblasts and to identify potential key transcription factors (TFs) regulating its expression through bioinformatics approaches. [Methods] Rat L6 myoblasts were induced to differentiate by culturing them in DMEM supplemented with 2% donor horse serum (DHS). Morphological changes were observed using an inverted microscope. Cell samples were collected prior to induction (day 0) and on days 1, 3, 5, and 7 post-induction. The relative expression levels of FGF10 mRNA and protein at each time point were quantified using RT-qPCR and Western blot analysis, respectively. Furthermore, a 2 000 bp sequence upstream of the transcription start site of the rat Fgf10 gene was extracted as the promoter region. Putative TF binding sites were predicted using four databases (TRANSFAC, JASPAR, HOCOMOCO, and CISBP), and high-confidence candidates were screened to construct a regulatory network. [Results] Morphological observations confirmed successful differentiation, as evidenced by the appearance of binucleated myotubes on day 3 and the formation of numerous thick, multinucleated myotubes by day 7. Both RT-qPCR and Western blot analysis demonstrated a significant dynamic expression pattern of FGF10. Expression levels were markedly upregulated during the early phase (days 1-3), reaching a peak on day 3 (P<0.01), followed by a decline to basal levels during the late phase (days 5-7). Cross-validation across multiple databases identified 48 high-confidence TFs, among which Elf5, Tcf3, Nkx3-2, Zic2, Tcf7, and Egr1 were consistently predicted by all four databases. [Conclusions] FGF10 exhibits high expression levels during the early stage of differentiation, indicating its crucial role in the initiation of myogenesis. The six identified TFs serve as core candidate regulators of Fgf10 expression, offering novel insights into the molecular mechanisms underlying muscle development. |
