摘要
目的:探讨骨形态发生蛋白(bone morphogenetic protein,BMPs)信号通路在压力刺激兔骨髓间充质干细胞(bone marrow-derived mesenchymal stem cells,BMSCs)成软骨响应中的作用。方法:体外分离、培养兔BMSCs,经鉴定后随机分为4组:空白对照组、压力刺激组、BMPs拮抗剂Noggin刺激组、压力+BMPs拮抗剂Noggin刺激组,其中压力刺激组和压力联合Noggin刺激组置于静态液压细胞加载装置中培养,并给予120 kPa、每天1 h、连续4 d的力学刺激。Real-time PCR和Western Blot方法检测压力刺激前后细胞中BMPs信号相关分子BMP2、Smad1、Smad5表达的变化;Western Blot方法检测4个实验组中P-Smad1/Smad5和软骨特异性基因Col-II、Aggrecan的蛋白表达。结果:兔BMSCs传代后细胞生长状态稳定,细胞表面标记物CD29阳性率97.2%、CD44阳性率93.2%、CD45阳性率0.8%;成骨诱导3周后,茜素红染色可见矿化结节形成;成脂诱导2周后,油红O染色可见红色脂滴;压力刺激后BMP2、Smad1、Smad5的基因与蛋白表达均明显增加(P<0.05);Western Blot检测显示,压力加载方式作用于兔BMSCs后,软骨细胞特异性指标Col-Ⅱ、Aggrecan的表达明显上调(P<0.05);BMPs特异性拮抗剂Noggin能明显抑制压力刺激兔BMSCs中P-Smad1/Smad5、Col-Ⅱ、Aggrecan的蛋白表达。结论:BMP/Smad信号通路介导了压力刺激兔BMSCs成软骨响应的过程。
AIM： To investigate the role of bone morphogenetic proteins （BMPs） pathway in chondrogenic mechanotransduction in bone marrow mesenchymal stem ceils （BMSCs） under mechanical pressure. METHODS ： BMSCs were cultured and identified by flow cytometry. BMSCs at passage 3 were randomly divided into blank control group, mechanical pressure treantment group, BMPs inhibitor Noggin treatment group, mechanical pressure and Noggin treatment group. Hydrostatic pressure at 120 kPa was adopted for 1 hour per day for 4 consecutive days as the mechani- cal pressure treantment. , The expressions of BMP2, Smadl and Smad5 were examined by Real-time PCR and Western Blot. Western Blot was also adopted to assay the protein expression of P-smadl/Smad5 and collagen II and aggrecan. RESULTS： The subcultured BMSCs grew well with positive expression of CD29 of 97.2%, CD44 of 93.2% and CD45 of 0.8%. Osteogenic and adipogenic differentiations of BMSCs were confirmed by induction assays. Hydrostatic pressure up - regulated both the mRNA and protein expressions of BMP2, Smadl and SmadS. The mechanical pressuresignificantly increased, collagen II and aggrecan expression in BMSCs. Noggin inhibited the phosphorylation of smadl/ Smad5 and chondrogenic differentiation of BMSCs. CONCLUSION： BMPs pathway participates the mechanotranduc- tion process of BMSCs and paly an important role in chondrogenic differentiation of BMSCs under the mechanical pres- sure.
出处
《牙体牙髓牙周病学杂志》
CAS
北大核心
2013年第4期218-224,共7页
Chinese Journal of Conservative Dentistry
基金
国家自然科学基金资助项目(31170888)
关键词
骨髓间充质干细胞
压力
软骨形成
BMPS
bone marrow mesenchymal stem cells （BMSCs）
mechanical pressure
chondrogenesis
bone mor-phogenetic protein （BMP）
