骨髓间充质干细胞在mdx鼠体内分化为骨骼肌细胞及其dystrophin/utrophin蛋白的表达  

Mesenchymal stem cells transplanted in mdx mice differentiate into myocytes and express dystrophin/utrophin

作　　者：冯善伟[1,2] 张成[1,2] 卢锡林[1] 柳太云[1] 李才明[1] 姚晓黎[1] 于美娟[3] 

机构地区：[1]中山大学附属第一医院神经科,广东广州510080 [2]中山大学干细胞与组织工程中心,广东广州510080 [3]广州医学院附属第二医院神经病学研究所,广东广州510260

出　　处：《南方医科大学学报》2009年第5期974-978,共5页Journal of Southern Medical University

基　　金：国家自然科学基金(30370510,30170337);广东省自然科学基金(31693);CMB基金(982667);中国博士后基金(20080430799)

摘　　要：目的研究骨髓间充质干细胞(MSCs)移植入放疗的mdx鼠后dystrophin/utrophin的表达变化。方法将BrdU标记的大鼠P5代MSCs移植入放疗处理的mdx鼠,分别于移植后4、8、12和16周断头处死取其腓肠肌,冰冻切片应用免疫荧光、RT-PCR及Western blotting检测BrdU/dystrophin及utrophin的表达变化。另分别取5只正常C57BL/6鼠和mdx鼠作阳性和阴性对照。结果经γ射线放疗预处理的mdx鼠,在尾静脉移植MSCs后4周时肌纤维抗肌萎缩蛋白的表达少于1%,随移植时间延长增加,16周时为15%,dystrophin/BrdU激光共聚焦免疫荧光显示肌细胞核中移现象较同龄mdx鼠对照组减少,边居增多。RT-PCR及Western blotting检测到dystrophin表达也随移植时间延长增多。移植后mdx鼠肌细胞膜utrophin表达较对照组mdx鼠减少,RT-PCR结果显示mRNA在移植前后没有明显改变。结论MSCs移植后mdx鼠的肌细胞膜有dystrophin表达,激光共聚焦技术表明dystrophin阳性肌纤维部分来自于移植的MSCs。utrophin在mdx鼠肌膜上表达上调,MSCs移植后dystrophin表达对utrophin有一定的抑制作用,两者存在互补关系。Objective To investigate the differentiation of rat bone marrow mesenchymal stem cells （MSCs） into myocytes and their expression of dystrophin/utrophin after transplantation in mdx mice. Methods BrdU-labeled fifth-passage rat MSCs were transplanted in mdx mice with previous total body γ irradiation （7 Gy）. At 4, 8, 12 and 16 weeks after the transplantation, the mice were sacrificed to detect dystrophin/BrdU and utrophin expressions in the gastrocnemius muscle using immunofluorescence assay, RT-PCR and Western blotting. Five normal C57 BL/6 mice and 5 mdx mice served as the positive and negative controls, respectively. Results Four weeks after MSC transplantation, less than 1% of the muscle fibers of the mdx mice expressed dystrophin, which increased to 15% at 16 weeks. Donor-derived nuclei were detected in both single and clusters of dystrophin-positive fibers. Some BrdU-positive nuclei were centrally located, and some peripherally within myofibers. Utrophin expression decreased over time after transplantation. Conclusion The myofibers of mdx mice with MSC transplantation express dystrophin, which is derived partially from the transplanted MSCs. Dystrophin expression from the transplanted MSCs partially inhibits the upregulation of utrophin in mdx mouse muscle, showing a complementary relation between them.

关 键 词：骨髓间充质干细胞 MDX鼠 细胞移植 5-溴脱氧尿核苷 抗肌萎缩蛋白 UTROPHIN 

分 类 号：R746.2[医药卫生—神经病学与精神病学]