丹酚酸B对大鼠骨髓间充质干细胞分化过程中Desmin、α-actinmRNA表达的影响  

Effect of SalB on expression of desmin and a-actin mRNA in differentiation process of bone marrow mesenchymal stem cells into cardiomyocytes in rats

作　　者：华声瑜[1] 范英昌[1] 马轶文[1] 王强[1] 

机构地区：[1]天津中医药大学基础医学院,天津300193

出　　处：《天津中医药》2009年第2期145-148,共4页Tianjin Journal of Traditional Chinese Medicine

基　　金：天津市社会发展科技计划项目(043111211)

摘　　要：[目的]探讨丹参单体丹酚酸B(SalB)在体外诱导大鼠骨髓间充质干细胞(MSCs)分化为心肌样细胞的情况及Desmin和α-actin mRNA的表达,确定大鼠MSCs体外诱导为心肌样细胞的条件、规律及转化细胞的特点。[方法]选用成年近交系Wistar大鼠,利用percoll分离液进行密度梯度离心法和直接贴壁法进行分离、提纯MSCs,并进行培养扩增。免疫组化方法对第9代MSCs表面抗原进行鉴定。分别应用10μmol/L5-氮胞苷(5-azacytidine,5-aza)及250μg/LSalB联合5-aza(5-aza+SalB)对第9代的MSCs进行联合诱导24h,于诱导后第4周,用实时荧光定量逆转录-聚合酶链反应(RT-PCR)法检测Desmin、α-actin mRNA的表达。[结果]1)第9代MSCs免疫组化染色CD44呈阳性表达,CD34呈阴性表达。2)诱导4周后,细胞体积变小,可见由几个细胞连接形成的多核肌管样结构。5-aza组、5-aza+SalB组均出现明确的心肌早期分化基因与5-aza组相比,5-aza+SalB组的Desmin和α-actin mRNA表达明显增强。[结论]SalB具有促进5-aza诱导的MSCs向心肌样细胞分化的作用。[Objective] To study the differentiation of bone marrow mesenchymal stem cells （MSCs） into cardiomyocytes induced by 5- azacytidine and SalB in vitro and the expressions of Desmin and α-actin to decide the requirements, rules and characteristics of MSCs differentiated into cardiomyocytes. [Methods] The density gradient centrifugation and adherent filtration were used to isolate and purify MSCs. The MSCs were cultured and amplified. MSCs surface antigen profiles were obtained by immunity tissue chemistry. The MSCs of the ninth passage were induced with 5-azacytidine at 10 μmol/L and SalB at 250 mg/L for 24 hours, and the expressions of Desmin and aactin were tested by RT-PCR at the 4th week, respectively after treating MSCs with 5-azacytidine and SaIB. [Results] The samples of MSCs of ninth passage were tested repetitively. The MSCs were positive for CD44 and negative for CD34. After the MSCs treated with 5- azacytidine and SalB for 24 hours, their morphology and arrangement changed obviously. Four weeks later, the cells became small, and the muhinuclear myotube-shape constructions were formed by a few cells. The Desmin and a-actin were positive in the MSCs treated group, and negative in control group. With the culture time passed, the levels of Desmin and a-actin in MSCs increased.

关 键 词：丹酚酸B 骨髓间充质干细胞 5-氮胞苷 心肌样细胞 基因 

分 类 号：R285.5[医药卫生—中药学]