人脐带间质干细胞来源的外切体对大鼠心肌细胞缺血/再灌注损伤的作用

Exosome derived from human umbilical cord mesenchymal stem cell alleviates myocardial ischemia/reperfusion injury

目的:研究人脐带间质干细胞来源的外切体(hucMSC-exosome)对大鼠缺血/再灌注(I/R)损伤心肌细胞的修复作用。方法:在体内,建立大鼠I/R模型:经冠状动脉左前降支(LAD)结扎缺血30 min后取消结扎恢复灌注,与此同时,尾静脉注射hucMSC-exosome。在体外,建立大鼠H9C2(2-1)心肌细胞缺氧/复氧(H/R)模型:细胞于无血清低糖培养基中缺氧(5%CO2/93%N2)24 h后,用含hucMSC-exosome的高糖10%FBS培养液复氧24 h。采用MTT法、TUNEL法、心肌酶谱以及线粒体膜电位等方法检测大鼠心肌细胞的增殖和凋亡状况。结果:在体内,心肌细胞经I/R损伤后,hucMSC-exosome与hucMSC处理组较PBS处理组血清乳酸脱氢酶(LDH)、肌酸激酶同工酶(CK-MB)值均下降;TUNEL结果显示,前两组较PBS处理组心肌细胞凋亡明显减少;在体外,心肌细胞经H/R损伤后,MTT实验表明细胞增殖显著减弱,hucMSC-exosome作用后,TUNEL及线粒体膜电位(ΔΨm)结果分别显示受损心肌细胞凋亡以及膜电位下降明显减少。结论:hucMSC-exosome能有效抵抗心肌细胞因缺血/再灌注损伤而引起的细胞凋亡。

Objective： To investigate the repair effect of exosome derived from human umbilical cord mesenchymal stem cells（hucMSC-exosome） to ischemia/reperfusion（I/R） myocardial cells.Methods： In vivo,Female Sprague-Dawley rats′ left anterior descending coronary artery（LAD） was occluded to induce regional myocardial ischemia,reperfusion was achieved by releasing the clamp.After 30 min of ischemia,hucMSC-exosome was injected via tail vein.In vitro,I/R of H9C2（2-1） rat embryonic cardiac myoblasts was achieved by culturing the cells in low glucose DMEM without FBS in a hypoxia chamber,saturated with 5%CO2/93%N2 for 24 h and following reoxygenation using 10%FBS high glucose DMEM and hucMSC-exosome in the normal incubating condition for 24 h.Using MTT,TUNEL,myocardial enzyme spectrum and mitochondrial membrane potential and other methods to detect the proliferation and apoptosis of myocardial cell.Results： In vivo,after the injury of I/R,lower level of LDH and CK activity was observed in the hucMSC-exosome or hucMSC pretreated cells as compared with PBS control cells.TUNEL staining showed that there was a decrease in apoptosis in the injured cells after pretreated with hucMSC-exosomes or hucMSC compared to PBS-treated group.In vitro,cell proliferation examined by MTT assay were shown to reduce significantly after the injury of H/R.Pretreatment of cells with hucMSC-exosome resulted in a marked decrease in hucMSC-exosome induced mitochondrial membrane potential（ΔΨm） and apoptosis.Conclusion： Exosome secreted by hucMSC can reduce the injury of myocardial I/R effectively.