Isolation, culture and identification of sheep skeletal muscle satellite cells

Skeletal muscle satellite cells (SMSCs) play an important role in muscle growth, regeneration and maintenance. This study aimed to isolate SMSCs from sheep, establish a system for isolation, culture and identification of SMSCs in vitro, and provide seed cells for subsequent studies. SMSCs were isolated and purified from newborn 2-day-old healthy sheep by collagenase type I and trypsin two-step digestion and pre-plating method. The results indicated that the isolated and purified SMSCs showed full spindle shape and strong refractive index. The cell growth curve detected by CCK-8 kit was typical “S” type. Immunofluorescence analysis showed that the isolated cells expressed SMSC marker proteins Pax7 and MyoD1. After induction of myogenic differentiation, the cells fused with each other to form multinucleated myotubes and expressed the myoblast specific marker MHC. RT-PCR results showed that the cells expressed SMSC marker gene Pax 7. This experiment established an in vitro isolation, purification and identification system for sheep skeletal muscle satellite cells, which provided a good cell model for studying the biological mechanism of sheep skeletal muscle cells, optimizing sheep breeds, and cell transplantation repair.