解剖学报

›› 2012, Vol. 43 ›› Issue (6): 778-783.doi: 10.3969/j.issn.0529-1356.2012.06.011

• 细胞和分子生物学 • 上一篇    下一篇

PI3K/Akt在线粒体ATP敏感性钾通道开放抑制缺氧复氧大鼠心肌微血管内皮细胞FKN表达中的作用 

陈秋萍*; 黄赛赛; 周伟伟; 沈施仁; 曹苏   

  1. 南通大学附属医院麻醉科，江苏 南通 226001
  • 收稿日期:2012-01-09 修回日期:2012-03-19 出版日期:2012-12-06
  • 通讯作者: 陈秋萍

Role of PI3K/Akt signal pathway in opening mito-KATP to inhibit the expression of FKNin rat myocardium microvascular endothelia cells exposed to hypoxia-reoxygenation

  1. Department of Anesthesiology,Affilited Hospital of Nantong University, Jiangsu Nantong 226001, China
  • Received:2012-01-09 Revised:2012-03-19 Online:2012-12-06
  • Contact: CHEN Qiu-ping

摘要： 目的 观察PI3K/Akt信号在线粒体ATP敏感性钾通道（mito-KATP）开放抑制缺氧复氧大鼠心肌微血管内皮细胞（MMECs）fractalkine(FKN)表达中的作用。方法 培养SD大鼠离体MMECs，建立缺氧复氧损伤模型24皿，随机分为4组( EM>n /EM>=6)：正常对照组（N组）、缺氧/复氧组（H/R组）、二氮嗪预处理+缺氧/复氧组（DZ组）、LY294002+二氮嗪预处理+缺氧/复氧组（LY294002+DZ组）。DZ组加入100μmol/L二氮嗪预处理2h，LY294002+DZ组在加入 100μmol/L LY294002预处理2h后再加入100μmol/L二氮嗪预处理2h，然后和缺氧复氧组同样进行缺氧2h、复氧2h。Hoechst染色观察凋亡细胞显微结构，四甲基偶氮唑盐（MTT）法测定细胞活力，RT-PCR检测Akt和FKNmRNA水平， Western blotting检测Akt蛋白水平。结果 与N 组比较，H/R组细胞增殖率显著降低（EM>P /EM><0.01）、凋亡率显著升高（EM>P/EM> <0.01）， FKN mRNA和FKN蛋白表达显著增加（P <0.01）、Akt mRNA和Akt蛋白升高（ EM>P/EM> <0.05）。与H/R组比较，DZ组细胞增殖率显著升高（EM>P/EM> <0.01）、凋亡率显著降

关键词: 线粒体ATP敏感性钾通道, 心肌微血管内皮细胞, PI3K, Akt, Fractalkine, 缺氧, 复氧, 反转录-聚合酶链反应, 免疫印迹法, 大鼠

Abstract: Objective To investigate the role of PI3K/Akt signal pathway in openned mito-KATP inhibit the expression of fractalkine (FKN) in rat myocardium microvascular endothelial cells exposed to hypoxia-reoxygenation. Methods Cardiac microvascular endothelial cells of the SD rat were cultured to establish a hypoxia-reoxygenation model. Twenty-four dishes were randomly divided into 4 groups ( EM>n/EM>=6 each).The normal control group (group N ), the cells of H/R group, diazoxide pretreatment +H/R group (group DZ) and diazoxide pretreatment+LY294002+ H/R group (group DZ+LY294002). BR>The cell vitality, the cell apoptotic rate and expression of Akt, FKN mRNA were detected with MTT. Hoechst dyeing RT-PCR and Western blotting, respectively. Results Compared with group N, the cell vitality significantly decreased ( EM>P /EM><0.01) and the apoptosis rate increased( EM>P/EM> <0.01). The expression of FKN mRNA and protein ( EM>P/EM> <0.01) was up-regulated in H/R group, the expression of Akt mRNA and protein significantly rised ( EM>P /EM><0.05).Compared with group H/R, the cell vitality significantly increased ( EM>P/EM> <0.05), the apoptosis rate significantly decreased (EM>P/EM><0.01).The expression of Akt mRNA and protein( EM>P/EM> <0.01and P <0.05) was up-regulated,fractalkine mRNA and significantly decreased ( EM>P/EM> <0.01) in group DZ. Compared with group DZ, Akt protein and mRNA significantly decreased (EM> P/EM> <0.01), BR>FKN mRNA and protein significantly increased ( EM>P /EM><0.01). Conclusion Openned mito-KATP through the PI3K/Akt signal pathway may regulate downstream of the transcription and expression of fractalkine mRNA inhibit cell apoptosis and improve the proliferation rate to protect the rat cardiac microvascular endothelial cells exposed on hypoxia-reoxygenation. BR

Key words: mito-KATP, Cardiac microvascular endothelial cell, PI3K, Akt, FKN, Hypoxia-reoxygenation, RT-PCR, Western blotting, Rat

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