解剖学报

解剖学报 ›› 2022, Vol. 53 ›› Issue (1): 28-34.doi: 10.16098/j.issn.0529-1356.2022.01.004

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CD73对体外大鼠心肌细胞增殖和功能的影响

王琰1 赵育洁1 李志营1 王前1 郭志坤1,2* 
  

  1. 1.郑州第七人民医院心内科，郑州 450000; 2.新乡医学院河南省医用组织重点实验室，河南 新乡 453003
  • 收稿日期:2020-08-20 修回日期:2020-10-11 出版日期:2022-02-06 发布日期:2022-02-06
  • 通讯作者: 郭志坤 E-mail:gzk@xxmu.edu.cn
  • 基金资助:
    国家自然科学基金;河南省卫生厅攻关项目;郑州市心脏结构研究重点实验室资助项目;河南省医学科技攻关计划联合共建项目

Effects of CD73 on proliferation and function of rat cardiomyocytes in vitro

WANG Yan1  ZHAO Yu-jie1 LI Zhi-ying1  WANG Qian1  GUO Zhi-kun1,2*   

  1. 1.Department of Cardiology, the Zhengzhou Seventh People’s Hospital, Zhengzhou 450000, China; 2.He’nan Key Laboratary of Medical Tissue Regeneration, He’nan Xinxiang 453003, China
  • Received:2020-08-20 Revised:2020-10-11 Online:2022-02-06 Published:2022-02-06
  • Contact: GUO Zhi-kun E-mail:gzk@xxmu.edu.cn

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摘要：

目的  探讨 CD73 对大鼠心肌细胞（CMs）增殖和功能的影响。   方法  体外培养原代大鼠 CMs 和大鼠心肌细胞系 H9C2，构建慢病毒 CD73 过表达载体和空载病毒组，分别转染 CMs 和 H9C2。实验分组：CD73 过表达组（OE组）和阴性对照组（NC组），即CMs-OE组和CMs-NC组；H9C2-OE组和H9C2-NC组，每组各5只。Real-time PCR 法检测转染病毒后各组细胞 CD73 基因水平表达情况；MTT 法检测细胞增殖能力变化；无损伤心肌细胞功能分析仪检测 CMs 细胞增殖曲线、倍增时间和搏动变化。   结果  慢病毒转染 CMs 和 H9C2 72 h 后，过表达组 CD73 mRNA 水平明显高于对照组（P<0.05）；CMs和 H9C2 转染后3～4 d的增殖能力为OE组大于NC组（P<0.05）；CMs 和 H9C2 细胞增殖曲线均显示 OE 组高于 NC 组，CMs 和 H9C2 倍增时间为 OE 组低于 NC 组（P<0.05）；CMs转染后72 h显示OE组细胞搏动率高于NC组， OE组转录因子T-同源盒基因5（TBX5）表达和血管内皮生长因子（VEGF）的分泌高于NC组，而缺氧诱导因子1α（HIF-1α）及肝病坏死因子α（TNF-α）的分泌低于NC组。   结论  CD73可促进 CMs 和 H9C2 的增殖，促进 CMs 搏动及TBX5表达和VEGF分泌，抑制HIF-α和TNF-α的分泌。 

关键词: CD73, 旁分泌作用, 心肌细胞, 搏动, 四甲基偶氮唑盐法, 大鼠 

Abstract:

Objective  To investigate the effect of CD73 on the proliferation and function of rat cardiac myocytes (CMs) in vitro.    Methods  The primary rat CMs and H9C2 cells were cultured in vitro, and lentivirus CD73 overexpression vector and empty vector group were constructed and transfected into CMs and H9C2 respectively. Experimental groups: CD73 overexpression group (OE group) including CMs-OE group and CMs-NC group, and negative control group (NC group) including H9C2-OE grop and H9C2-NC group, 5 in each group. After transfection, the expression of CD73 gene was detected by Real-time PCR method , and the proliferation ability was detected by MTT method , and the proliferation curve, doubling time and pulsatility of CMs were detected by non-destructive cardiomyocyte function analyzer.    Results  After 72 hours of lentivirus transfection, CD73 mRNA level in the over expression group was significantly higher than that in the control group (P<0.05); the proliferation ability of CMs and H9C2 at 3-4 days after transfection in OE group was larger than that in NC group (P<0.05); the proliferation curve of CMs and H9C2 cells in OE group was higher than that in NC group, and the doubling time of CMs and H9C2 in OE group was lower than that in NC group 72 hours after CMs transfection, the beating rate of OE group was higher than that of NC group, the expression of T-box 5(TBX5) and the secretion of vasuular endothelial growth factor VEGF) in OE group were higher than those in NC group, while the secretion of hypoxia iducible factor-1α(HIF-1α) and tumor necrosis factor-α(TNF-α) was lower than that of NC group.    Conclusion  CD73 can promote the proliferation of CMs and H9C2, promote CMs pulsation, TBX5 expression and VEGF secretion, and inhibit the secretion of HIF-α and TNF-α. 

Key words: CD73, Paracrine effect, Cardiomyocyte, Pulsation, Methyl thiazolyl tetrazolium, Rat

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