人脐带间充质干细胞外泌体对多柔比星致扩张型心肌病大鼠心肌纤维化的影响

Effect of human umbilical cord mesenchymal stem cell exosomes on myocardial fibrosis in rats with Adriamycin-induced dilated cardiomyopathy

目的:探讨大鼠尾静脉注射人脐带间充质干细胞来源外泌体(hUCMSCs-ex)对多柔比星致扩张型心肌病(DCM)大鼠心功能与心肌纤维化的影响。方法:将100只SD雄性大鼠按随机数字表法分为正常组(20只)与DCM组(80只)。采用腹腔注射多柔比星进行造模。造模成功后大鼠按随机数表法分为DCM组、低剂量组、中剂量组和高剂量组,分别经大鼠尾静脉注射1 mL/kg氨基酸葡萄糖培养基(DMEM),外泌体20μg/kg、100μg/kg和250μg/kg。造模结束后按随机数字法表随机选取正常组10只、DCM组30只大鼠行心脏超声评估大鼠心功能。外泌体治疗后按随机数字表法随机选取各组大鼠各10只行心脏超声评估大鼠心功能。大鼠心室肌Masson染色,观察各组大鼠心肌病理学的改变;Western blot检测各组大鼠心肌组织Ⅰ型胶原蛋白(COLⅠ)、Smad2、α平滑肌肌动蛋白(α-SMA)的表达情况。结果:DCM组大鼠的左心室射血分数(LVEF)和左心室短轴缩短率(LVFS)[(64.30±3.51)%和(38.70±2.85)%]较正常组[(78.80±1.52)%和(50.60±1.50)%]显著降低,差异均有统计学意义(t=20.518、22.311,均P<0.01)。DCM组大鼠的左心室舒张末期内径(LVEDD)和左心室收缩末期内径(LVESD)[(4.62±0.13)mm和(3.40±0.12)mm]较正常组[(3.29±0.24)mm和(3.16±0.33)mm]明显增加,差异均有统计学意义(t=2.854、3.800,均P<0.01)。外泌体治疗后,中剂量组和高剂量外泌体组LVEF[(84.3±2.6)%和(83.4±3.2)%]明显高于DCM组[(79.2±2.4)%],差异均有统计学意义(均P<0.01)。Masson染色发现外泌体治疗组蓝染的胶原纤维较DCM组有所改善。Western blot结果显示,高剂量外泌体可显著降低α-SMA和Smad2的表达(t=2.828、2.972,均P<0.05),高剂量及低剂量外泌体均可降低COLⅠ的表达(t=4.276、2.972,均P<0.05)。结论:大鼠尾静脉注射hUCMSCs-ex能显著降低多柔比星诱导大鼠DCM所致的心肌纤维化程度,改善大鼠心功能。

Objective To evaluate the effects of human umbilical cord mesenchymal stem cells-derived exosomes(hUCMSCs-ex)injection on cardiac function and myocardial fibrosis in dilated cardiomyopathy(DCM)rats induced by Adriamycin(ADR).Methods One hundred male SD rats were randomly divided into the normal group(20 rats)and the DCM group(80 rats).The rats in DCM group were treated with ADR by intravenous injection to induce DCM.DCM rats were randomly divided equally into DCM group,low-dose group,medium-dose group and high-dose group which were received intravenous injection 1 mL/kg Dulbecco′s modified eagle medium(DMEM),20μg/kg,100μg/kg and 250μg/kg exosomes.After modeling,10 rats in normal group and 30 rats in DCM group were randomly selected to receive echocardiography to evaluate the cardiac function.After exosomes treatment,10 rats were randomly selected form each group for echocardiography to evaluate the cardiac function.The morphological changes in myocardial cells were observed by using Masson staining in each group;Western blot detection between groups of rats was used to analyze the expression of myocardial collagenⅠtype(COLⅠ),Smad2 and alpha smooth muscle actin(α-SMA).Results Left ventricular ejection fraction(LVEF)and left ventricular fraction shortening(LVFS)in the DCM group[(64.30±3.51)%and(38.70±2.85)%]were significantly lower than those of the normal group[(78.80±1.52)%and(50.60±1.50)%],and the differences were statistically significant(t=20.518,22.311,all P<0.01).The left ventricular end-diastolic diameter(LVEDD)and left ventricular end-systolic diameter(LVESD)[(4.62±0.13)mm and(3.40±0.12)mm]of the DCM group were significantly higher than those of the normal group[(3.29±0.24)mm and(3.16±0.33)mm],and the differences were statistically significant(t=2.854,3.800,all P<0.01).After exosomes treatment,LVEF[(84.3±2.6)%and(83.4±3.2)%]in the medium-dose and high-dose groups were significantly higher than that in the DCM group[(79.2±2.4)%],and the diffe-rences were statistically significant(all P<0.01).Masson staining found that collagen fibers were less in exosomes treating group than those in the DCM group;Western blot test showed that high-dose exosomes can reduce the expression ofα-SMA and Smad2,high-dose and low-dose exosomes can both significantly reduce the expression of COLⅠ.Conclusions It suggests that exosomes intravenous injection from hUCMSCs-ex can significantly improve myocardial fibrosis in DCM rats induced by ADR and cardiac function.