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目的 通过向骨髓间充质干细胞 (MSCs)培养体系中添加心肌细胞裂解液的方法,体外模拟心肌微环境,观察MSCs向心肌细胞分化的诱导作用,并与诱导分化剂 5 氮杂胞苷 ( 5 aza)比较。方法 分离新生乳鼠的心肌细胞并制成心肌细胞裂解液,自成年大鼠骨髓中分离MSCs,用含有心肌细胞裂解液的培养基(A组)、含有 5 aza的培养基 (B组 )、含有 5 aza和心肌细胞裂解液的培养基(C组)以及普通培养基(对照组)培养。观察细胞形态的改变,并通过免疫组化分析分化后细胞表达α 肌动蛋白、心脏特异性肌钙蛋白T(cTnT)、连接蛋白 43及CD31的情况。结果 A、B组的MSCs在培养 1周后均形成肌细胞形态,并且均表达α 肌动蛋白和cTnT;A组MSCs分化的肌样细胞所含的肌纤维较B组更丰实,细胞生长趋势也优于B组,并且可以表达CD31;B组MSCs分化的肌样细胞不表达CD31;对照组细胞仅表达α 肌动蛋白。结论 心肌细胞裂解液是体外诱导MSCs分化为心肌样细胞的理想条件,优于传统的 5 aza,在心肌细胞移植技术中可以用于体外模拟心肌细胞微环境。
OBJECTIVE: To observe the induction of MSCs to cardiomyocytes by simulating the myocardial microenvironment in vitro by adding cardiomyocyte lysate to the culture system of MSCs and to observe the effect of inducing differentiation of 5-azacytidine (5 aza) comparison. Methods Cardiomyocytes from neonatal neonatal rats were isolated and cardiomyocytes lysates were prepared. MSCs were isolated from bone marrow of adult rats and cultured in medium containing cardiomyocyte lysate (group A), medium containing 5 aza (group B) , Medium containing 5 aza and cardiomyocyte lysate (group C), and normal medium (control group). The morphological changes of the cells were observed. The expressions of α-actin, cardiac troponin T (cTnT), connexin 43 and CD31 were analyzed by immunohistochemistry. Results MSCs in groups A and B all formed myoblast morphology after 1 week of culture, and all expressed α-actin and cTnT. Muscle fibers contained in group A and group B were more abundant than those in group B, and their cell growth tended to be better In group B, and could express CD31; in group B, the differentiated myogenic cells of group B did not express CD31; in control group, the cells expressed only α-actin. Conclusions Cardiomyocytes lysate is an ideal condition for inducing MSCs to differentiate into cardiomyocytes in vitro. Compared with the traditional 5 aza, cardiomyocyte lysate can be used to simulate cardiomyocyte microenvironment in vitro.