摘要
为研究绵羊胚胎肌源性干细胞的生物学特性,取30日龄的绵羊胚胎,采用联合酶消化法及差速贴壁法分离培养获得均质的肌源性干细胞,并对其进行一系列生物学特性检测。结果显示,绵羊肌源性干细胞能传代培养至35代以上;免疫荧光染色和RT-PCR鉴定肌源性干细胞表面标记物Sca-1、CD34、CD144和Desmin呈阳性表达;生长曲线和细胞周期结果显示绵羊肌源性干细胞具有较强自我更新能力;通过不同的诱导方法可以将肌源性干细胞成功地诱导为肝样细胞和胰岛样细胞,Schiff染色和双硫腙染色均呈阳性,RT-PCR检测肝样细胞特异性标记物AFP和ALB以及胰岛样细胞特异性标记物Insulin均呈阳性表达,进一步证明了其诱导分化潜能。结果表明,本试验成功地分离培养获得了均质并具有较强自我更新潜能的绵羊肌源性干细胞,这种干细胞具有向肝样细胞和胰岛样细胞分化的潜能,这为临床治疗提供了潜在的理论基础和大量的种子细胞。
The 30-day-old sheep embryo limb skeletal muscle were taken to isolate and culture the homogeneous muscle-derived stem cells by enzymatic digestion and differential adherence method. A series of biological characteristics were studied. The sheep muscle-derived stem ceils could be subcultured to more than 35 generations ; the surface marker Sca-1,CD34,CD144 and Desmin of muscle-derived stem cells were identified to be positive by means of immunofluorescence staining and RT-PCR;the growth curve and cell cycle analysis showed that the muscle-derived stem cells had a strong cell proliferation ability;the muscle-derived stem cells were induced into hepatic-like cells and islet-like cells successfully by different induce treatment,and Schiff staining and dithizone staining were positive, meanwhile, RT-PCR detection showed that hepatic-like cell specific markers AFP and ALB and islet-like cell specific marker insulin were positive expressed. The above identi- fication further demonstrated its induction differentiation potential. This test results showed that the sheep muscle-derived stem cells were isolated successfully,and had the potential of self-renewal and induction differentiation of hepatic-like cells and islet-like cells. This provides potential theoretical basis and great quantity of seed ceils for clinical therapy.
作者
张萍
马月辉
关伟军
ZHANG Ping MA Yue-hui GUAN Wei-jun(Institute of Animal Sciences, Chinese Academy of Agricultural Sciences, Beij ing 100193, China)
出处
《中国兽医学报》
CAS
CSCD
北大核心
2017年第10期1973-1979,共7页
Chinese Journal of Veterinary Science
基金
中国农业科学院科技创新工程基金资助项目(ASTIP-IAS01)
国家绒毛用羊产业体系基金资助项目(nycytx-40-01)
国家家养动物种质资源平台基金资助项目(2015year)
关键词
绵羊
肌源性干细胞
分离培养
诱导分化
细胞周期
sheep
muscle-derived stem ceils
isolation and culture
induction differentiation
cell cycle
