电针和bmscs移植联合应用对大鼠脑出血病灶周围微环境影响的实验研究

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1、中图分类号：R89；R36；R39硕士学位论文电针和BMSCs移植联合应用对大鼠脑出血病灶周围微环境影响的实验研究院（系）、所基础医学院研究生姓名李振学科、专业人体解剖与组织胚胎学导师姓名萧洪文教授二Ο一二年四月1目录1电针和BMSCs移植联合应用对大鼠脑出血神经元修复的机制研究„„„„„„„„„„„„„„„„„„„„„„„„11.1中文摘要„„„„„„„„„„„„„„„„„„„„„11.2英文摘要„„„„„„„„„„„„„„„„„„„„„41.3前言„„„„„„„„„„„„„„„„„„„„„„„81.4材料与方法„„„„„„„„„„„„„„„„„„„„101.5结果„„„„„„„„„„„„„

2、„„„„„„„„„„231.6讨论„„„„„„„„„„„„„„„„„„„„„„„291.7结论„„„„„„„„„„„„„„„„„„„„„„„381.8参考文献„„„„„„„„„„„„„„„„„„„„„391.9英汉缩略词对照表„„„„„„„„„„„„„„„„„442致谢„„„„„„„„„„„„„„„„„„„„„„„463可溶性环氧化物酶与缺血性脑卒中的防治（综述）„„„„„„„„„„„„„„„„„„„„„„„„„472电针和BMSCs移植联合应用对大鼠脑出血神经元修复的机制研究摘要目的：观察电针和骨髓间充质干细胞（bonemarrowmesenchymalstemcells,BMSCs）移植联

3、合应用对脑出血(intracerebralhemorrhage,ICH)病灶及周围微环境的影响，探讨神经元修复的机制。方法：1.将复苏的BMSCs培养贴壁达80%时，将培养液更换为预诱导液培养24小时后，再更换为无血清诱导液诱导6h。收集细胞备用。2.采用胶原酶及肝素注入尾状核制备大鼠ICH模型。3.术后分别在2h、24h、48h时间点对SD大鼠进行神经功能评分(NeurologicalSeverityScore,NSS)评分，评分大于9分者则纳入实验。并随机分为：对照组（control组）、细胞移植组（BMSCs组）、电针与细胞移植联合组（electro-acupuncturecombine

4、dwithBMSCstransplantation，EaBMSCs组）。4.在术后第3天时通过脑立体定位仪导航，将20l0.9%生理盐（Normalsaline，NS）水和BMSCs悬液移植入ICH病灶中。根据移植后喂养天数每组再分为：1d、3d、5d、7d、14d五个亚组。移植2h后，对EaBMSCs组取大椎穴和百会穴，进行电针治疗，每天一次，直至处死。5.对各组SD大鼠每天行NSS评分一次，直至处死取材。6.通过蛋白免疫印迹(WeternBlotting,WB)方法检测各组不同时间点神经生长因子（nervegrowthfactor，NGF）、脑源性神经营养因子(brainderivedn

5、europhicfactor,BDNF)、β-微管蛋白（tubulin）Ⅲ及甘油醛-3-磷酸脱氢酶（glyceraldehyde-3-phosphate1dehydrogenase，GAPDH）蛋白表达变化情况。结果：1.制备大鼠ICH模型成功后，移植细胞前对control组、BMSCs组、EaBMSCs组进行NSS评分，各组之间评分无显著性差异；移植后第5天时，移植后第5天，BMSCs组评分显著低于control组（P0.05）；EaBMSCs组（除1d外）明显低于control组，（P0.05）。EaBMSCs组和control组相比较（除1d外）均有显著差异，有统计学意义（P0.0

6、5）。改善神经功能缺失症状更明显。2.EaBMSCs组和BMSCs组的NGF蛋白表达明显增加；于移植后7天、14天，BMSCs组较control组增加显著，有统计学意义（P0.05）；EaBMSCs组与BMSCs组和control组比较（除1d外）增加更明显，差异有统计学（P0.05）。EaBMSCs组大鼠ICH病灶周围NGF蛋白表达上调更明显。3.EaBMSCs组和BMSCs组BDNF蛋白表达明显增加，移植后5天、7天、14天时，较control组差异显著有统计学意义（P0.05）；在移植后第3天、5天、14天，EaBMSCs组较BMSCs组表达增加显著，有统计学意义（P0.05）。

7、EaBMSCs组较单纯BMSCs组和control组大鼠ICH病灶周围BDNF蛋白表达上调更明显。4.在同一时间点，EaBMSCs组和BMSCs组与control组比较β-tubulinⅢ蛋白表达明显增加；在移植后5天、7天、14天，BMSCs组较control组增加显著，有统计学意义（P0.05）；EaBMSCs组（除移植1天）均较BMSCs组明显增加，有统计学意义P0.05）。电针和BMS