南方医科大学学报

南方医科大学学报 ›› 2023, Vol. 43 ›› Issue (9): 1644-1650.doi: 10.12122/j.issn.1673-4254.2023.09.24

• • 上一篇    

心脏祖细胞来源的外泌体可减轻心肌梗死小鼠的心肌损伤：基于mTOR途径诱导Treg细胞分化

何艳娟，李卓颖，申 琳，石丁华，李申堂   

  1. 长沙市第四医院儿科，湖南 长沙 410006；中南大学湘雅三医院儿科，湖南 长沙 410013
  • 出版日期:2023-09-20 发布日期:2023-09-28

Cardiac progenitor cells-derived exosomes alleviate myocardial injury by regulating Treg cell differentiation through the mTOR pathway in mice with myocardial infarction

HE Yanjuan, LI Zhuoyi, SHEN Lin, SHI Dinghua, LI Shentang   

  1. Department of Pediatrics, Fourth Hospital of Changsha, Changsha Hospital of Hunan Normal University, Changsha 410006, China; Department of Pediatrics, Third Xiangya Hospital, Central South University, Changsha 410013, China
  • Online:2023-09-20 Published:2023-09-28

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摘要： 目的 研究心脏祖细胞来源的外泌体（CPCs-Exos）对心肌梗死（MI）小鼠调节性T细胞（Treg）分化的影响及其作用机制。方法 结扎小鼠左冠状动脉前降支制备MI动物模型。所有小鼠随机分成3组，即对照组（只开胸，不结扎）、MI组（结扎左冠状动脉前降支）和CPCs-Exos组（结扎左冠状动脉前降支后尾静脉注射CPCs-Exos）。收集小鼠血清与心脏组织，流式细胞术与全自动生化仪检测Treg细胞、CD4+IL-10+T细胞百分比及LDH、CK-MB含量，TTC染色检测小鼠心肌梗死面积。收集小鼠脾脏naive CD4+T细胞，并将细胞分成对照组、CD4+T细胞活化组（CD3+CD28）、CPCs-Exos处理活化CD4+T细胞组（CD3+CD28+CPCs-Exos）、mTOR 激活剂（CD3 + CD28 + CPCs- Exos + mTOR activator）与抑制剂组（CD3 + CD28 + CPCs- Exos + mTORinhibitor）。Western blotting法检测细胞中mTOR、p-mTOR表达水平，流式细胞术检测Treg、CD4+IL-10+T细胞百分比。结果 整体动物实验结果显示，与对照组相比，MI组小鼠心肌梗死面积显著增加，血清中CK-MB（P=0.0002）、LDH（P<0.001）显著增多，Treg、CD4+IL-10+T细胞有增加趋势。与MI组相比，CPCs-Exos组小鼠梗死面积降低，CK-MB（P=0.003）、LDH（P=0.003）含量减少，Treg 细胞（P=0.001）、CD4+IL-10+T（P=0.004）细胞百分比显著增加。离体细胞水平结果显示，与Control组相比，CD3+CD28组Treg细胞（P=0.03）、CD4+IL-10+T细胞百分比（P=0.004），mTOR（P=0.04）、p-mTOR（P=0.04）表达水平有增加趋势；与 CD3+CD28 组相比，CD3+CD28+CPCs-Exos 组 Treg 细胞（P=0.01）、CD4 +IL-10+T 细胞百分比（P=0.004），mTOR（P=0.009）、p-mTOR（P=0.009）表达水平显著增加。与CD3+CD28+CPCs-Exos组相比，CD3+CD28+CPCs-Exos+mTOR activator组Treg细胞（P=0.04）、CD4+IL-10+T细胞（P=0.01）百分比显著增加，而CD3+CD28+CPCs-Exos+mTOR inhibitor组Treg细胞（P=0.04）、CD4+IL-10+T细胞百分比（P=0.04）显著降低。结论 CPCs-Exos能够诱导Treg细胞分化，其机制与mTOR途径有关。

关键词: 心脏祖细胞来源的外泌体；mTOR；调节性T细胞；心肌损伤

Abstract: Objective To investigate the effect of cardiac progenitor cells-derived exosomes (CPCs-Exo) on Treg differentiation in mice with myocardial infarction (MI). Methods Mouse models of MI established by ligation of the left anterior descending coronary artery (LAD) were treated with CPCs-Exos, and naive CD4+ T cells were isolated from the spleen of the mice and divided into control group, CD4+T cell activation group (CD3+CD28), CPCs-Exos stimulation group (CD3+CD28+CPCs-Exos), mTOR activator group (CD3+CD28+CPCs-Exos+mTOR activator) and mTOR inhibitor group (CD3+CD28+CPCs-Exos+mTOR inhibitor). Western blotting was used to detect the expression levels of mTOR and p-mTOR in the treated cells. Flow cytometry was used to analyze the percentages of Treg and CD4+ IL-10+ T cells. The infarct size of the mice were measured with 2,3,5-triphenyltetrazole chloride (TTC) staining, and serum levels of LDH and CK-MB were detected using an automatic biochemical analyzer. Results Compared with the control group, the mouse models of MI showed significantly increased release of LDH (P<0.001) and CK-MB (P=0.0002) and increased percentages of Treg and CD4+ IL-10+ T cells. Treatment with CPC-Exos effectively reduced the MI area and lowered serum levels of LDH (P=0.003) and CK-MB (P=0.003) and the percentages of Tregs (P=0.001) and CD4+ IL-10+ T cells (P=0.004) in the MI mouse models. In the isolated CD4+ T cells, CPCs-Exos treatment significantly up-regulated the percentages of Treg (P=0.01) and CD4+IL-10+ T cells (P=0.004) and increased the expression of mTOR (P=0.009) and p-mTOR (P=0.009), and these effects could be further enhanced by the mTOR activator but obviously attenuated by the mTOR inhibitor. Conclusion CPCs-Exos promotes the differentiation of Treg in mice with MI by modulating the mTOR signaling pathway.

Key words: cardiac progenitor cell-derived exosomes; mTOR; Treg; myocardial injury