心肌细胞裂解成分诱导骨髓间充质干细胞分化的超微结构观察

目的:观察骨髓间充质干细胞(MSCs)在心肌细胞裂解成分作用下超微结构改变,了解心肌细胞裂解成分对MSCs分化的诱导作用。 方法:分离并裂解新生大鼠的心肌细胞;自成年大鼠骨髓中分离MSCs;将分离的MSCs分3组培养:仅用普通培养基培养(对照组);5-氮杂胞苷(5-aza)诱导后用普通培养基培养(5-aza诱导组);含有心肌细胞裂解成分的培养基培养(心肌细胞裂解成分培养组)。培养1周,观察细胞形态及超微结构的改变,对培养的细胞进行抗心脏特异性肌钙蛋白T(cTnT)及抗分化决定簇31(CD31)细胞免疫化学染色,并分析各组细胞的增殖情况。 结果:对照组MSCs无明显的肌样细胞或内皮样细胞形成,抗cTnT和抗CD31染色阴性。5-aza诱导组部分MSCs分化为肌样细胞,电镜下可见大量细胞器空化,抗cTnT染色阳性,但细胞增殖缓慢。心肌细胞裂解成分培养组的MSCs分化为肌样细胞,电镜下可见肌丝样结构,抗cTnT染色阳性,细胞增殖旺盛,另见部分MSCs分化为内皮样细胞,形成内皮细胞特有的胞质突起和质膜小泡等超微结构,且抗CD31染色阳性。 结论:含有心肌细胞裂解成分的培养基可以诱导MSCs向心肌样细胞和内皮样细胞方向分化。

Ultrastructure of Differentiated Bone Mesenchymal Stem Cells Induced by Cardiomyocyte Lysate

Objective: To observe the ultrastrutural changes of bone mesenchymal stem cells ( MSCs) cultured with cardiomyocyte lysate , and to demonstrate the effects of cardiomyocyte lysate on MSCs differentiation.Methods: Cardiomyocytes isolated from newborn rats were lysed by repeated freezing and defrosting. MSCs isolated from adult rats were cultured in 3 different systems for 1 week: medium alone (control group) ,medium alone after 5-aza inducement (5-aza group)and medium containing cardiomyocyte lysate (cardiomyocyte lysate group). The changes of MSC morphology and ultrastructure were observed. At the end of culture, immunostaining against cTnT and CD31 was performed, and cell proliferation of MSCs in the 3 groups was analyzed.Results: MSCs in control group did not express cTnT or CD31. Myogenic cells or endotheloid cells were not found. MSCs in 5-aza group were differentiated into myogenic cells and expressed cTnT, but the cells grew slowly and cell organellae became empty. In cardiomyocyte lysate group, a great number of cells were differentiated into myogenic cells and expressed cTnT, a lot of filaments were observed in these cells by electron microscopy; some other cells differentiated into endotheloid cells also expressed CD31, and had endothelial specific ultrastructures such as plasmal projections and plasmalemmal vesicles.Conclusions: The methods of adding cardiomyocyte lysate into the culture system can be used to induce MSCs differentiated into cardiomyogenic cells and endotheloid cells.