5-氮杂胞苷诱导骨髓源性心肌干细胞向心肌分化过程中心肌肌钙蛋白T的表达变化

CHANGES OF cTnT EXPRESSION OF THE CARDIOMYOCYTES DIFFERENTIATED FROM 5-AZACYTIDINE-INDUCED MCSCs

目的研究5_氮杂胞苷诱导骨髓源性心肌干细胞(MCSC)向心肌分化过程中,心肌肌钙蛋白T(cTnT)的表达变化和超微结构。方法从SD大鼠骨髓间充质干细胞(MMSC)中筛选MCSC,用5_氮杂胞苷诱导向心肌定向分化。取分化后不同时间的细胞作cTnT免疫细胞化学染色,并对结果进行图像分析。透射电镜下观察分化细胞的超微结构。取不同发育期和成熟期的SD大鼠心肌作对比研究。结果用5_氮杂胞苷诱导后2周,细胞内开始表达cTnT;诱导后3周,cTnT表达增强,可见横纹样结构;诱导后4周,cTnT表达明显增强,横纹增多,但排列不规则。诱导后4周的细胞内出现心房粒,可见丰富的粗面内质网和糖原。胚胎11d大鼠的心肌内,横纹样结构较少且排列不规则。新生大鼠心肌内横纹增多,排列较规则。1月龄和3月龄心肌的横纹样结构发达,排列规则。诱导分化后4周的细胞cTnT表达特征与胚胎11d的心肌相似。结论MCSC经5_氮杂胞苷诱导后可分化为心肌细胞,表现为未成熟心肌的结构和功能特征。

Objective To study the changes of expression of cardiac troponin T(cTnT) and the ultrastructures of the cardiomyocytes differentiated from 5_azacytidine_induced marrow_derived cardiac stem cells(MCSCs). Methods MCSCs were isolated from marrow mesenchymal stem cells(MMSCs) of SD rats and then were induced with 5_azacytidine to differentiate into cardiomyocytes.Expression of cTnT of the cells was determined 2 weeks,3 weeks and 4 weeks after induction with 5_azacytidine.Mean optical density of cTnT was analyzed with an image analysis system.The ultrastructures of the cells were viewed with a transmission electron microscope.Expression of cTnT in the cardiomyocytes of the embryonic and adult rats was examined. Results Expression of cTnT of the cells could be observed 2 weeks after induction with 5-azacytidine.In 3 weeks after induction, expressin of cTnT increased,striation-like structures occurred.In 4 weeks after induction,expression of cTnT and the striation-like structures increased,but they were not irregular-organized.There were atrial granules,rough endoplasmic reticulum and glycogen.There were a few irregular-organized striation-like structures in the cardiomyocytes of the embryonic rats in day eleven.In the cardiomyocytes of the neonatal rats,the striation-like structures were increased and arranged regularly.In the cardiomyocytes of the one month-old and three month-old rats,striation-like structures were well developed and organized regulatlly.The characteristics of cTnT expression of the cells 4 weeks after induction were similar to that of the cardiomyocytes of the embryonic rats in day eleven.Conclusion MCSCs may differentiate into cardiomyocytes after induction with 5-azacytidine,the differentiated cells demonstrate the structural and functional characteristics of the immature cardiomyocytes.;