摘要
目的在用血小板源性生长因子-BB(PDGF-BB)联合20%胎牛血清(FBS)诱导骨髓间充质干细胞(BM-SC)向平滑肌细胞(SMC)分化过程中,通过对促血管SMC分化因子myocardin和平滑肌分化标志物基因α-actin,SM-22等表达的研究,探讨myocardin在促血管SMC分化中的重要作用。方法采用骨髓干细胞培养基和贴壁法从小鼠骨髓中分离BMSC,然后细胞分为两组,诱导其向SMC分化:(1)20%FBS单独诱导;(2)PDGF-BB(50ng/ml)联合20%FBS诱导BMSC。分别于诱导0d,4d,8d,12d后用逆转录-聚合酶链反应法(reverse transcription polymerase chain reaction,RT-PCR)和免疫组化法检测细胞myocardin,α-actin,SM-22的mRNA表达和蛋白表达情况.结果RT-PCR和免疫组化结果显示在对小鼠BMSC的诱导过程中,检测出了myocardin,同时还有平滑肌分化标志物基因α-actin,SM-22等的表达。0d,4d,8d两诱导组细胞的myocardin,α-actin,SM-22的mRNA和蛋白表达水平随着诱导时间延长而增强(P<0.05),而诱导8d及12d时表达基本趋于稳定,与SMC相比无明显统计学差异(P>0.05);并且各项指标的表达水平在PDGF-BB与20%FBS联合诱导时比20%FBS单独诱导表达增强。不同诱导组之间统计学比较有差异(P<0.05)。结论PDGF-BB联合20%FBS可有效诱导BMSC向SMC分化,myocardin在此过程中起着关键性作用,且PDGF-BB,20%FBS两者联合诱导比20%FBS单独诱导作用要强。
Objective To investigate the importance of myocardin in the differentiation of bone marrow mesenchymal stem cells （BMSC） into smooth muscle cells （SMC） in vitro. Methods Mouse BMSCs were isolated and purified from bone marrow, and then treated in two groups： （1） by FBS （20%） only; （2） only by 50ng/ml PDGF-BB and FBS （20%） . The cells of each group were cultured for 0, 4, 8, 12 days respectively. At the end, RT-PCR and Immunohistochemistry assay were used to detect the expression of the mRNA and protein of myocardin, α-actin, and SM-22. Results 1. The expressing of the mRNA and protein of myocardin, α-actin, and SM-22 increased gradually and remaind stabilization since it had achieved the highest point at 8days after induction 2. The expression of mRNA of myocardin, α-actin, and SM-22 are higher in the second group than the first one. 3. Statistically there was significance in different induction times and groups （P〈0.05） . Conclusion 1 . BMSC could be effectively induced into smooth muscle-like cells by PDGF-BB with 20% FBS. 2. Myocardin plays an important role in differentiation of BMSC into SMC. 3. The combination effect of PDGF-BB （50ng/ml） and FBS （20%） was stronger than that of FBS （20M） alone.
出处
《中国组织化学与细胞化学杂志》
CAS
CSCD
2008年第5期401-406,共6页
Chinese Journal of Histochemistry and Cytochemistry
基金
国家自然科学基金资助项目(30570725)
