诱导人脐带间充质干细胞向心肌细胞分化及对小鼠心肌梗死的治疗作用

目的研究脐带间充质干细胞(UC-MSC)体外分化为心肌细胞的可行性以及观察UC-MSC体内移植对心肌梗死模型小鼠的治疗效果。方法 10μmol/L 5-氮胞苷(5-aza)体外诱导UC-MSC 14 d,通过RT-PCR、免疫荧光染色鉴定其分化效果;采用腹腔注射盐酸异丙肾上腺素(ISO)每只3.0 mg/(kg/d),制作心肌梗死模型鼠;在注射ISO 48 h后,实验组将DAPI标记的UC-MSC经两次尾静脉移植给心肌梗死模型鼠,移植后第4周和第8周,分别采集实验小鼠的心脏、脾脏,以未移植细胞组的小鼠心肌损伤模型作为对照,通过心脏指数和脾脏指数测量,免疫荧光和碱性复红-苦味酸(HBFP)染色鉴定其体内分化和修复作用。结果 RT-PCR分析表明诱导的UC-MSC表达心肌特异性基因:心肌α-actin、TBX5、GATA4和NKx2.5,免疫荧光染色显示诱导细胞呈心肌α-actin和NKx2.5阳性,且呈双核现象。尾静脉移植后第4周和第8周,模型受体鼠心脏均发现有DAPI阳性细胞迁移至心肌组织且呈现心肌α-actin阳性,HBFP染色及心脏和脾脏指数结果显示移植UC-MSC对心肌损伤的模型鼠有明显的修复和治疗效果。结论 UC-MSC在体外经5-aza诱导可定向分化为心肌细胞,尾静脉体内移植UC-MSC对心肌损伤小鼠有明显的治疗效果。

Induction of human umbilical cord-derived mesenchymal stem cells to differentiate into cardiomyocytes and the therapeutic effect of UC-MCSs transplantation on mouse models of myocardial infarction

Objective To investigate the feasibility of differentiation of umbilical cord-derived mesenchymal stem cells（UC-MSCs） into cardiomyocytes in vitro and the effects of UC-MSCs transplantation on the mouse models of myocardial infarction.Methods UC-MSCs were induced by 10 μmol/L 5-aza for 14 days in vitro and then identified by RT-PCR and immunofluorescence staining.The mouse models of myocardial infarction were developed by intraperitoneal injection of isoproterenol hydrochloride（ISO） 3.0 mg/（kg/d）.After 48 h,DAPI-labeled UC-MSCs were transplanted via the tail vein twice within 48 to 72 hours in the experimental group.At 4 and 8 weeks after transplantation,the mouse heart and spleen of both groups were taken out.The cardiac index and spleen index of the mouse models were measured,and the differentiation in vivo and the repair of cardiomyocyte injury were evaluated histologically using immunofluorescence and basic fuchsin-picric acid（HBFP） staining,with the mouse models of myocardial injury without UC-MSCs transplantation as control.Results RT-PCR analysis showed that the induced cells expressed cardiac-specific genes α-actin,TBX5,GATA4 and NKx2.5.In addition,immunofluorescence staining showed that the induced cells were positive for α-actin and NKx2.5,and some cells showed double nuclei.The models with UC-MSC transplantion were found to have DAPI-positive cardiac cells migrated to the myocardium and showed cardiac α-actin positive at 4 weeks and 8 weeks after transplantation.Obvious therapeutic effect was shown by HBFP staining and by the heart and spleen indexes after UC-MSC transplantation in the myocardial injury mouse models.Conclusions 5-azainduced UC-MSCs can be directed to differentiate into cardiomyocytes in vitro,and significant therapeutic effect on the mouse myocardial injury in vivo can be obtained through UC-MSCs transplantation via the tail vein.