大鼠肌肉卫星细胞的分离培养鉴定与诱导分化

发布时间：2017-12-30 20:22

本文关键词：大鼠肌肉卫星细胞的分离培养鉴定与诱导分化　出处：《内蒙古大学》2010年硕士论文　论文类型：学位论文

【摘要】： 骨骼肌肌肉控制着脊椎动物的精确的运动。成体骨骼肌的组成单元是肌纤维。肌纤维是由一群处于有丝分裂间期的成肌细胞组成的合胞体。而未分化的肌肉卫星细胞参与了合胞体的修复和维持。肌肉卫星细胞是未分化的单核肌源性细胞群落,在哺乳动物、爬行动物、鸟类、两栖动物的骨骼肌内都能发现。1961年在蛙的肌肉中首先发现了卫星细胞的存在。电镜观察提示卫星细胞位于骨骼肌纤维的肌细胞膜与基底膜之间,其核较大、胞浆少,细胞器数量少。这些特征说明卫星细胞处于有丝分裂静止期,其转录活性低于肌纤维。由于能够从病人骨骼肌中获得并在体外大量扩增,肌肉卫星细胞可以作为细胞移植的资源,并且已经应用于骨骼肌和心肌的重建。尽管发现卫星细胞的存在已经有50多年的历史了,但分离鉴定肌肉卫星细胞的方法仍然具有不稳定性。本研究使用两步酶消化和差速贴壁相结合的方法分离大鼠肌肉卫星细胞。Desmin免疫细胞化学染色、Myf5及Myodl的RT-PCR法证明了所分离的细胞是肌肉卫星细胞。成肌和成脂诱导试验证明了所分离的肌肉卫星细胞具有多分化潜能。主要研究结果如下： 1.通过两步酶消化和差速贴壁相结合的方法分离得到了肌肉细胞系。Desmin免疫细胞化学染色、Myf5及Myodl的RT-PCR法证明了所分离的细胞是肌肉卫星细胞。 2.分离得到的肌肉卫星细胞能够向成肌细胞和脂肪细胞分化的潜能,具有一定的多能性。 3. DCA处理促进了肌肉卫星细胞的增殖并激活了WNT/beta-Catenin信号通路。
[Abstract]:Skeletal muscle muscles control the precise movement of vertebrates. Adult skeletal muscles are composed of muscle fibers. Muscle fibers are syncytial cells consisting of a group of myoblasts in the mitotic phase. Undifferentiated muscle satellites. Cells are involved in the repair and maintenance of syncytial bodies. Muscle satellite cells are undifferentiated mononuclear myogenic cell communities in mammals, reptiles, and birds. In 1961, satellite cells were first found in frog muscle. Electron microscopic observation indicated that satellite cells were located between muscle cell membrane and basement membrane of skeletal muscle fiber. These characteristics indicate that the satellite cells are in mitotic quiescent phase and their transcriptional activity is lower than that of muscle fibers. Muscle satellite cells can be used as a resource for cell transplantation because they can be obtained from patients' skeletal muscles and expanded in vitro. Although the existence of satellite cells has been discovered for more than 50 years, the method of isolation and identification of muscle satellite cells is still unstable. In this study, two steps of enzyme digestion and differential adhesion were used to isolate rat muscle satellite cells. Desmin immunocytochemical staining was used. The RT-PCR method of Myf5 and Myodl proved that the isolated cells were muscle satellite cells, and the myoblast and adipogenic induction tests showed that the isolated muscle satellite cells had multi-differentiation potential. The main findings are as follows: 1. The muscle cell line. Desmin immunocytochemical staining was obtained by two-step enzyme digestion and differential adhesion. The RT-PCR method of Myf5 and Myodl proved that the isolated cells were muscle satellite cells. 2. The isolated muscle satellite cells have the potential to differentiate into myoblast and adipocytes, and have certain pluripotency. 3. DCA treatment promoted the proliferation of muscle satellite cells and activated the WNT/beta-Catenin signaling pathway.
【学位授予单位】：内蒙古大学
【学位级别】：硕士
【学位授予年份】：2010
【分类号】：R329

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