【作者】 孙树洋；

【导师】 张风河；

【作者基本信息】 山东大学 ， 口腔临床医学， 2006， 硕士

【摘要】 目的： 1．探讨骨骼肌卫星细胞体外分离培养、鉴定的方法，研究骨骼肌卫星细胞的生物学特性，为后续的实验奠定基础。 2．评价腺病毒介导的人骨形态发生蛋白2(Ad-hBMP-2)基因体外转染骨骼肌卫星细胞后，肌卫星细胞分化为成骨样细胞的能力；并测定基因转染后目的基因体外表达的情况。应用Ad-GFP基因转染肌卫星细胞，测定腺病毒转染的效率。为应用骨骼肌卫星细胞修复骨缺损提供实验依据。 方法： 1．采用Ⅰ型胶原酶和胰蛋白酶二步消化法获取大鼠骨骼肌卫星细胞，进行体外原代和传代培养。观察细胞的形态，通过生长曲线、细胞融合率等指标研究骨骼肌卫星细胞的增殖与分化能力，利用免疫细胞化学染色对所获得的细胞进行鉴定。 2．Ad-hBMP-2基因体外转染骨骼肌卫星细胞后，通过形态学观察、生长曲线的测定、碱性磷酸酶(ALP)细胞化学染色及活性检测、骨钙素(OC)及Ⅰ型胶原的免疫细胞化学染色、矿化结节形成实验和Western—blot，观察Ad-hBMP-2基因转染后骨骼肌卫星细胞生物学行为的变化和目的基因表达的情况。Ad-GFP基因转染骨骼肌卫星细胞后2天，用荧光显微镜测定腺病毒转染效率，并观测Ad-GFP基因转染的骨骼肌卫星细胞传代后绿色荧光蛋白的表达情况。更多还原

【Abstract】 Objective:1. To investigate the methods about the cultivation and identification of Skeletal Muscle Satellite Cells(SMSCs) in vitro, especially, to study the cells’ biological characters. These studies are preparations for the succeeded experiments.2. To assess the ability of inducing Skeletal Muscle Satellite Cells(SMSCs) into osteoblast-like cells, and to examine the expression of target gene in SMSCs after being transfected. To determine the adenovirus infection rate by using Ad-GFP as a contral gene vector. According to those , we can provide experimental bases for the repair of bone defect by utilizing SMSCs.Method:1. The rats’ SMSCs were acquired by enzyme digestion. Cells are cultured and subcultured in vitro. To study the proliferation and differentiation characteristics of SMSCs by cell morphological observation , cell growth curve and rate of myotube formation. Cells were identified with immunocytochemical stain.2. After being transfected by Ad-BMP2 gene, the biological characteristics of SMSCs were examined by cell morphological observation, cell growth curve ,更多还原