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In vitro culture and induced differentiation of sheep skeletal muscle satellite cells

Authors

  • Haiqing Wu,

    1. Key Laboratory of Mammalian Reproductive Biology and Biotechnology Ministry of Education, Inner Mongolia University, Inner Mongolia, Hohhot 010021, Peoples Republic of China
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  • Yu Ren,

    1. Key Laboratory of Mammalian Reproductive Biology and Biotechnology Ministry of Education, Inner Mongolia University, Inner Mongolia, Hohhot 010021, Peoples Republic of China
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  • Shuo Li,

    1. Key Laboratory of Mammalian Reproductive Biology and Biotechnology Ministry of Education, Inner Mongolia University, Inner Mongolia, Hohhot 010021, Peoples Republic of China
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  • Wei Wang,

    1. Key Laboratory of Mammalian Reproductive Biology and Biotechnology Ministry of Education, Inner Mongolia University, Inner Mongolia, Hohhot 010021, Peoples Republic of China
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  • Jianlong Yuan,

    1. Key Laboratory of Mammalian Reproductive Biology and Biotechnology Ministry of Education, Inner Mongolia University, Inner Mongolia, Hohhot 010021, Peoples Republic of China
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  • Xudong Guo,

    1. Key Laboratory of Mammalian Reproductive Biology and Biotechnology Ministry of Education, Inner Mongolia University, Inner Mongolia, Hohhot 010021, Peoples Republic of China
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  • Dongjun Liu,

    1. Key Laboratory of Mammalian Reproductive Biology and Biotechnology Ministry of Education, Inner Mongolia University, Inner Mongolia, Hohhot 010021, Peoples Republic of China
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  • Ming Cang

    Corresponding author
    1. Key Laboratory of Mammalian Reproductive Biology and Biotechnology Ministry of Education, Inner Mongolia University, Inner Mongolia, Hohhot 010021, Peoples Republic of China
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To whom correspondence should be addressed (email cangming1970@yahoo.com.cn).

Abstract

Skeletal muscle satellite cells are adult muscle-derived stem cells receiving increasing attention. Sheep satellite cells have a greater similarity to human satellite cells with regard to metabolism, life span, proliferation and differentiation, than satellite cells of the rat and mouse. We have used 2-step enzymatic digestion and differential adhesion methods to isolate and purify sheep skeletal muscle satellite cells, identified the cells and induced differentiation to examine their pluripotency. The most efficient method for the isolation of sheep skeletal muscle satellite cells was the type I collagenase and trypsin 2-step digestion method, with the best conditions for in vitro culture being in medium containing 20% FBS+10% horse serum. Immunofluorescence staining showed that satellite cells expressed Desmin, α-Sarcomeric Actinin, MyoD1, Myf5 and PAX7. After myogenic induction, multinucleated myotubes formed, as indicated by the expression of MyoG and fast muscle myosin. After osteogenic induction, cells expressed Osteocalcin, with Alizarin Red and ALP (alkaline phosphatase) staining results both being positive. After adipogenic induction, cells expressed PPARα2 (peroxisome-proliferator-activated receptor α2) and clear lipid droplets were present around the cells, with Oil Red-O staining giving a positive result. In summary, a successful system has been established for the isolation, purification and identification of sheep skeletal muscle satellite cells.

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