牛骨骼肌卫星细胞的分离鉴定和诱导分化

doi:10.16431/j.cnki.1671-7236.2020.10.024

Abstract

Abstract: In order to study the muscle growth and development process of Simmental cattle in vitro and establish a primary cell model of bovine skeletal muscle satellite cells.Bovine skeletal satellite cells (BSSC) were obtained by digestion with 0.2% type Ⅱ collagenase and then digested with 0.25% trypsin-EDTA,and using the differential adhesion method to separate and obtain purified BSSC,reverse transcription PCR (RT-PCR),immunofluorescence staining and Western blotting were used to identify BSSC.BSSC was induced differentiation into adipocytes by adipogenic inducers.Oil red O staining was used to identify the adipogenic capacity of differentiated cells.It was also induced differentiation into myocytes by 2% horse serum,and RT-PCR was used to identify the expression of PAX7 and MyoG marker genes in resting phase before and after induction.The results showed that the isolated and purified BSSC was fusiform or spindle-shaped,with full cell morphology and strong refraction.With the extension of the culture time,the cells changed from disordered growth to ordered growth.It was found that BSSC surface marker genes Desmin,c-Met,Myf5 and specific marker Paired box 7 (PAX7) were all positively expressed by RT-PCR detection.Immunofluorescence staining and Western blotting showed that PAX7 and MyoD gene were both positively expressed.Adipocyte induction after differentiation,it was heavily stained with oil red O and a large number of lipid droplets were observed.The expression of PAX7 gene in the resting period after induction of myoblasts was higher than that before induction,but the expression of myogenin marker gene MyoG was opposite.The above results indicated that this study successfully isolated BSSC and proved that BSSC had the ability to differentiate into adipocytes and myocytes,which provided a primary cell model for the in vitro study of the regulation mechanism of Simmental beef products.

Key words: bovine skeletal muscle satellite cells; PAX7; induced differentiation; myoblasts; adipocytes

CLC Number:

Q291

ZHAO Xinyan, GUO Yanting, CHEN Junzhen, LI Zeyu, SHI Huijun, FU Qiang. Isolation,Identification and Differentiation of Bovine Skeletal Muscle Satellite Cells[J]. China Animal Husbandry and Veterinary Medicine, 2020, 47(10): 3249-3258.

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Isolation,Identification and Differentiation of Bovine Skeletal Muscle Satellite Cells

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