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心肌祖细胞衍生的外泌体通过 Akt/mTOR 激活促进 H9C2 细胞生长。

Cardiac progenitor cell‑derived exosomes promote H9C2 cell growth via Akt/mTOR activation.

机构信息

Department of Pediatrics, The Third Xiangya Hospital, Central South University, Changsha, Hunan 410013, P.R. China.

State Key Laboratory of Advanced Welding and Joining, Harbin Institute of Technology (Shenzhen), Shenzhen, Guangdong 518055, P.R. China.

出版信息

Int J Mol Med. 2018 Sep;42(3):1517-1525. doi: 10.3892/ijmm.2018.3699. Epub 2018 May 21.

Abstract

Exosomes are cell‑derived vesicles released from a variety of mammalian cells that are involved in cell‑to‑cell signalling. It has been reported that cardiac progenitor cells (CPCs) derived from an adult heart are one of the most promising stem cell types for cardioprotection and repair. The mammalian target of rapamycin (mTOR) signalling pathway is a pivotal regulator in CPCs, therefore, CPC‑derived exosomes were used in the present study to investigate whether it can promote H9C2 cell growth through the protein kinase B (PKB, or Akt)/mTOR signalling pathway. The CPCs were isolated from Sprague‑Dawley hearts. Following treatment with a specific medium, the exosomes were purified and identified by electron micrograph and western blot assays, using CD63 and CD81 as markers. The methyl‑thiazolyl‑tetrazolium and 5‑ethynyl‑2'‑deoxyuridine methods were used to detect H9C2 cell growth. The expression of Akt and mTOR were detected by western blot analysis following treatment with 200 or 400 µg/ml of exosomes for 24 or 48 h, respectively. It was found that, compared with higher concentrations of exosomes, prolonging the duration of exposure promoted cell growth. Accordingly, CPC‑derived exosomes stimulated the expression of Akt to a marked degree; groups treated with exosomes for 48 h showed higher expression of Akt than those treated for 24 h at the same concentration. mTOR was also stimulated by CPC‑derived exosomes. The activation of mTOR increased in accordance with the treatment time at an exosome concentration of 200 µg/ml and decreased with treatment time at an exosome concentration of 400 µg/ml. In conclusion, the present study demonstrated that CPC‑derived exosomes promoted H9C2 cell growth via the activation of Akt/mTOR in a time‑dependent manner at a relatively low exosome concentration, which may provide a novel therapy for cardiovascular disease.

摘要

外泌体是一种由多种哺乳动物细胞释放的细胞衍生小泡,参与细胞间信号传递。据报道,来源于成人心脏的心肌祖细胞 (CPCs) 是最有前途的心脏保护和修复的干细胞类型之一。哺乳动物雷帕霉素靶蛋白 (mTOR) 信号通路是 CPCs 中的关键调节因子,因此,本研究使用 CPC 衍生的外泌体来研究其是否可以通过蛋白激酶 B (PKB,即 Akt)/mTOR 信号通路促进 H9C2 细胞生长。CPCs 从 Sprague-Dawley 心脏中分离出来。在经过特定培养基处理后,通过电子显微镜和 Western blot 分析,使用 CD63 和 CD81 作为标记物对细胞外泌体进行纯化和鉴定。采用甲基噻唑基四唑和 5-乙炔基-2'-脱氧尿苷法检测 H9C2 细胞生长情况。用 Western blot 分析检测 Akt 和 mTOR 的表达,方法是分别用 200 或 400μg/ml 的外泌体处理 24 或 48 h。结果发现,与较高浓度的外泌体相比,延长暴露时间可促进细胞生长。因此,CPC 衍生的外泌体显著刺激 Akt 的表达;与同一浓度的 24 h 处理组相比,48 h 处理组的 Akt 表达更高。CPC 衍生的外泌体也可刺激 mTOR。mTOR 的激活随外泌体浓度为 200μg/ml 时的处理时间增加而增加,随外泌体浓度为 400μg/ml 时的处理时间增加而减少。综上所述,本研究表明,CPC 衍生的外泌体在相对较低的外泌体浓度下通过激活 Akt/mTOR 以时间依赖性方式促进 H9C2 细胞生长,这可能为心血管疾病提供一种新的治疗方法。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/f96f/6089767/9341e0669a6b/IJMM-42-03-1517-g00.jpg

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