Myostatin (), a member of the transforming growth factor-β superfamily, inhibits the activation of muscle satellite cells. However, the role and regulatory network of in equine muscle cells are not well understood yet. We discovered that knockdown significantly reduces the proliferation rate of equine muscle satellite cells. In addition, after the RNA sequencing of equine satellite cells transfected with -interference plasmid and control plasmid, an analysis of the differentially expressed genes was carried out. It was revealed that regulatory networks mainly involve genes related to muscle function and cell-cycle regulation, and signaling pathways, such as Notch, MAPK, and WNT. Subsequent real-time PCR in equine satellite cells and immunohistochemistry on newborn and adult muscle also verified the regulatory network found in RNA sequencing analysis. The results of this study provide new insight into the regulatory mechanism of equine .