间充质干细胞对衰老大鼠睾丸的影响.docVIP

间充质干细胞对衰老大鼠睾丸的影响 李艳菊，胡亮杉，郭坤元（南方医科大学珠江医院血液科，广东 广州 510282） [摘要] 目的：研究间充质干细胞(MSCs)延缓睾丸衰老的机制。方法：通过皮下注射D-半乳糖建立衰老大鼠模型的同时，大鼠尾静脉注射MSCs进行生物治疗；采用硫代巴比妥酸和黄嘌呤氧化法分别检测血清和睾丸中丙二醛(MDA)含量和超氧化物歧化酶(SOD)活性；应用Real-time PCR和免疫印记法检测各组睾丸组织中VEGF的表达。结果：MSCs治疗组与模型组相比，血清与睾丸中SOD的活性均明显增高、MDA含量也显著降低(p＜0.05)；与模型组相比，MSCs治疗组能够使衰老大鼠睾丸内的VEGF mRNA和VEGF蛋白表达升高(p＜0.05)。结论：MSCs上调VEGF的表达、抑制MDA生成并增强SOD活性，可能是其改善衰老睾丸功能的机制之一。 【关键词】间充质干细胞；衰老；睾丸；VEGF； The effect of mesenchymal stem cells on the spermary of aging rats AIM: To study the mechanism of mesenchymal stem cells on the spermary of aging rats. METHODS: While rats were injected with D-galactose daily,rats were injected with MSCs by vena caudalis; The content of superoxide dismutase (SOD) and malondialdehyde (MDA) in serum and spermary were measured by hydroxylamine and chromatometry methods; The expression of VEGF mRNA and protein in rats’spermary are detected by method of Real-time PCR and Western-blotting.RESULTS: In the group used MSCs,the activity of SOD was higher significantly and the content of MDA was lower significantly in serum and spermary than that in the model control group(p＜0.05);The expression of VEGF mRNA and protein in the spermary of MSCs group increased significantly compared with model group (P0.05).CONCLUSION: MSCs could increase the expression of VEGF,could suppress the production of MDA and reinforce the activity of SOD ,That maybe one of mechanism what it has the ability of anti-aging. Key words:MSCs;aging; spermary;VEGF; 0 引言 骨髓间充质干细胞(MSCs)是中胚层来源的具有多向分化能力的干细胞, 随着对MSCs的深入研究，目前认为MSCs除了具有多向分化潜能，还能在损伤局部反应性分泌各种各样的营养因子，可能使MSCs具有抗大鼠睾丸衰老的作用。本研究通过皮下注射D-半乳糖建立大鼠衰老模型，采用硫代巴比妥酸和黄嘌呤氧化法分别检测血清和睾丸中丙二醛(MDA)含量和超氧化物歧化酶(SOD)活性；应用real-time PCR和免疫印记法检测睾丸组织中VEGFmRNA和VEGF蛋白的表达水平，从而探讨MSCs对衰老睾丸功能影响的可能机制。 1 材料和方法 1.1 材料 D-半乳糖(Sigma公司)；SOD测定试剂盒、MDA试剂盒(南京建成公司)； RT-PCR试剂盒（大连宝生物公司）；引物序列由primer 3设计,上海英俊公司合成；Western Blot 所用抗体为Santa Cruz公司；流式细胞仪(美国COULTER公司)、721紫外-可见分光光度计(上海光电研究所)。 1.2 方法 1.2.1 大鼠MSCs的体外分离培养和鉴定 ①细胞的培养 采用贴壁法培养MSCs，倒置显微镜下观察MSCs形态并作HE染色；将细胞接种于6孔板内