右归饮含药血清对TGF-β1诱导骨髓间充质干细胞向软骨细胞分化的影响  

Effects of Yougui Drink Medicated Serum on Chondrogenic Differentiation of Bone Marrow Mesenchymal Stem Cells Induced by Tansforming Growth Factor Beta_1

作　　者：陈祁青[1] 金红婷[1,2] 吴承亮[1,2] 王萍儿[1,2] 应俊[1] 肖鲁伟[1,2] 童培建[1,3] 

机构地区：[1]浙江中医药大学,杭州310053 [2]浙江省骨伤研究所 [3]浙江中医药大学附属第一医院

出　　处：《浙江中医药大学学报》2015年第4期304-311,共8页Journal of Zhejiang Chinese Medical University

基　　金：国家自然科学基金资助项目(81373669);浙江省自然基金重点项目(LZ12H270001);浙江省重点科技创新团队计划资助项目(2011R50022-01);浙江省重点实验室计划资助项目(2013E10024)~~

摘　　要：[目的]观察不同浓度右归饮含药血清对转化生长因子β1(tansforming growth factor,TGF-β1)诱导大鼠骨髓间充质干细胞(bone marrow mesenchymal stem cells,BMSCs)向软骨细胞分化的影响。[方法]密度梯度离心法分离培养大鼠BMSCs,取第3代,分为正常对照组、诱导对照组、右归饮高剂量组、右归饮中剂量组、右归饮低剂量组。正常对照组加入大鼠空白血清,诱导对照组加入诱导液(10 g·L-1 TGF-β1、10-7mol·L-1地塞米松、50mg·L-1维生素C)及大鼠空白血清,右归饮各组加入诱导液及相应的含药血清。14d后,进行甲苯胺蓝染色及Ⅱ型胶原免疫细胞化学染色,采用Real time PCR检测Ⅱ型胶原、聚集蛋白聚糖m RNA的表达。[结果]甲苯胺蓝染色及免疫细胞化学染色显示,正常对照组未见阳性细胞,诱导对照组及右归饮各组细胞均呈阳性。与诱导对照组比较,右归饮各组平均累积光密度值均增高(P<0.01),右归饮低剂量组高于高、中剂量组(P<0.01)。Real time PCR检测结果显示,与正常对照组相比,诱导对照组及右归饮各组Ⅱ型胶原、聚集蛋白聚糖m RNA的表达均增高(P<0.01);与诱导对照组比较,右归饮各组Ⅱ型胶原、聚集蛋白聚糖m RNA的表达均增高(P<0.01),其中右归饮低剂量组显著高于高、中剂量组(P<0.01)。[结论]右归饮含药血清能在一定程度上促进TGF-β1诱导的BMSCs成软骨细胞分化,低剂量组效果较显著。[Objective] To investigate the effects of Yougui Drink（YGD） medicated serum on chondrogenic differentiation of rat bone marrow mesenchymal stem cells（BMSCs） induced by tansforming growth factor betal. [Methods] Rat BMSCs were isolated and cultured by density gradient centrifugation method. The 3rd-passage BMSCs were divided into the normal control group, the induction control group, and the high-dose, middle-dose,low-dose YGD medicated serum groups. Serum of the blank group rats was added to the normal control group, induction medium（10g ＂L-1 TGF-β1,10-Tmol·L^-1 dexamethasone, 50mg ·L^-1 ascorbic acid） and serum of the blank group rats was added to the induction control group, the induction fluid and YGD medicated serum was added to the corresponding YGD medicated serum group. 21 days later, the expressions of type I1 collagen and aggrecan were detected using toludine blue staining, immunocytochemical staining and Real-time PCR. [Results] The results of Toluidine blue staining and immunocytochemistry staining were positive in the induction control group and all YGD medicated serum group while negative in the normal control group. Compared with the induction control group, integrated optical density value（IOD value） increased in all YGD medicated serum group（P〈0.01） and IOD value of low-dose YGD medicated serum group was highest（P〈0.01）. Real-time PCR showed that the mRNA relative expression of type 11 collagen and aggrecan in the induction control group and all YGD medicated serum group was significantly higher than that in the normal control group. Compared with the induction control group, the mRNA relative expression of type Ⅱ collagen and aggrecan in increased in all YGD medicated serum group（P〈0.01）, and it was highest in the low-dose YGD medicated serum group（P〈0.01）. [Conclusion] YGD medicated serum can effectively facilitate the differentiation of BMSCs into chondrocytes induced by TGF-β1, and the effect of low-dose YGD medicated serum is significant relativ

关 键 词：骨髓间充质干细胞 软骨细胞 诱导分化 右归饮 转化生长因子β1 

分 类 号：R331[医药卫生—人体生理学]