缺氧诱导因子对急性心肌梗死后心肌细胞及细胞外基质的影响

张宇红　邓丽玉　林　彬　涂　胜　顾新元　唐利龙



缺氧诱导因子对急性心肌梗死后心肌细胞及细胞外基质的影响

张宇红邓丽玉林彬涂胜顾新元唐利龙

【摘要】急性心肌梗死是指冠状动脉闭塞、血流中断，使部分心肌因严重的缺血缺氧而发生局部坏死。而缺氧诱导因子作为机体应对缺氧应激的适应性反应的调控因子，参与了急性心肌梗死时心脏的适应性保护。该文主要介绍缺氧诱导因子的结构、功能及其对急性心肌梗死后心肌细胞及细胞外基质的影响。

【关键词】急性心肌梗死；缺氧诱导因子；心肌细胞；细胞外基质；心脏保护

作者单位：350108福建医科大学附属协和医院心内科(张宇红，邓丽玉，林彬，涂胜)；512000广东省韶关市粤北人民医院心内科(顾新元)；411199湖南省湘潭市中心医院心内科(唐利龙)

1概述

缺氧诱导因子(hypoxia-inducible factor, HIF)是由α亚基和β亚基组成的异二聚体。哺乳动物有3种HIFα亚基的亚型,包括HIF1α、2α和3α,其蛋白稳定性受氧浓度影响；而HIF的β亚基则可稳定地在核内表达且不受氧浓度调节。α亚基和β亚基都是螺旋-环-螺旋肽蛋白家族的成员。

缺氧时,脯氨酸羟化酶的活性受抑制,HIF1α通过HLH和PAS区域与β亚基异源二聚体化,并与转录的共激活剂CBP/P300相结合,形成活性转录复合体,转位至细胞核,介导靶基因缺氧应答部分的反向激活。HIF异二聚体可以识别并结合靶基因上的缺氧应答序列,这些序列拥有共同的碱基片段5-(A/G)CGTG-3。

HIF1α在多细胞生物的各种类型细胞中均表达；而HIF2α仅在脊椎动物的特定细胞如角质形成细胞表达,参与缺氧条件下新生血管形成；HIF3α也仅在某些特定细胞通过与HIF1α、HIF2α、HIF1β结合，抑制负性调节HIF1α、HIF2α的转录活性。迄今，大量实验证明HIF可以调节超过200多种基因的转录活性，在应对缺氧应激损伤的适应性变化中发挥着重要作用。

2HIF对急性心肌梗死后心肌的影响

HIF是急性心肌梗死(AMI)后心脏适应性变化过程中的主要调节因子之一。在人和鼠都观察到，提高HIF活性后心肌梗死面积缩小,左室收缩功能提高，存活率提高[8-9]。HIF的心肌保护作用是由多因素介导的，包括一系列HIF靶基因的转录及其相应信号转导通路的活化。

2.1调节心肌收缩

心血管活性肽是心血管系统稳态的重要调节者,也是HIF1的靶基因之一，在AMI早期有维持心肌收缩功能的作用[10]。心肌梗死后24 h内，机体通过激活HIF通路使心血管活性肽水平急剧升高[11],心血管活性肽再通过磷酸肌醇信号途径PKCε和细胞外信号调节激酶途径ERK1/2,激活下游肌球蛋白轻链激酶。肌球蛋白轻链被磷酸化后，Ca2+敏感性增强,促进心肌肌丝交联,使心肌收缩力增强[12],从而实现心脏保护作用。

2.2调节心肌细胞能量代谢

HIF1α通过多种靶基因共同作用,减少AMI后基础氧耗,提高葡萄糖利用效率。HIF介导AMI后心肌有氧代谢受抑,无氧酵解增强的过程。实验发现，HL-1心肌细胞在缺氧后24 h，葡萄糖转运体-1和乳酸水平分别增高5和15倍；线粒体电子传递链上的各种酶类包括复合体Ⅰ、Ⅳ和顺乌头酸酶的活性也不同程度地降低[13]。HIF1α促进正葡萄糖转运蛋白-1(GLUT-1)、正葡萄糖转运蛋白-3(GLUT-3)和己糖激酶Ⅰ基因(HK-1)、己糖激酶Ⅱ基因(HK-2)基因表达,使葡萄糖形成脱氧葡萄糖而不能进一步代谢[14]; HIF1α促进蛋白激酶(PDK)基因表达,磷酸化丙酮酸脱氢酶(PDH)的催化区域使之失活,使丙酮酸进入糖酵解途径[15-16]；HIF1α促进乳酸脱氢酶(LDH)和MOT4基因的表达,使丙酮酸转化为乳酸排出体外,同时减少线粒体源性自由基形成以减少心肌细胞死亡[17];另外正丙酮酸激酶PKM2既参与糖酵解过程，也与HIF1α结合促进其反向激活功能，以正反馈机制实现缺氧状态下心肌细胞代谢转化[18]。

2.3调节线粒体功能

HIF通过综合作用使线粒体产生的自由基减少,起到心肌保护作用。线粒体源性自由基增加会对细胞造成氧化应激损伤[19]，HIF1有助于维持AMI后自由基的稳态。HIF上调线粒体中线粒体蛋白BNIP3的水平,与自噬基因Beclin1竞争结合癌基因Bcl2后，游离的Beclin1协同磷脂酰肌醇激酶3激活缺氧诱导性线粒体自溶[20]; HIF抑制线粒体电子传递链中复合体1和4的活性[21]; HIF激活微小核糖核酸miRNA-210，抑制铁硫蛋白ISCU1/2，抑制三羧酸循环顺乌头酶和电子传递链复合体1的活性[22]。

2.4调节炎症反应

炎症趋化因子(如基质细胞衍生因子-12和单核细胞趋化蛋白-5)以及血管黏附分子(如细胞间黏附素和血管源性细胞黏附素)的mRNA表达在心肌梗死区域升高[23]。而HIF1可控制炎症反应，起心脏保护作用。有实验表明，在缺血再灌注前激活HIF活性,可减少心肌细胞表达趋化因子如角质细胞源性趋化因子、巨噬细胞炎症蛋白-2、单核细胞趋化因子-1、中性粒细胞趋化因子和细胞间黏附分子-1的表达,从而抑制中性粒细胞的聚集，明显降低了梗死面积[24]。

3心脏细胞外基质组成和功能

正常情况下,细胞外基质(ECM)由结构蛋白、非结构蛋白和由基质金属蛋白酶(MMP)、金属蛋白酶组织抑制物(TIMP)组成的蛋白水解系统构成。AMI后，左室细胞外基质发生了一系列形态和功能的变化,称为心室重构。异常的心室重构会导致心脏纤维化或心室过度扩张[25-26]。

心脏细胞外基质主要由纤维母细胞活化、增殖分化形成的心肌成纤维细胞合成[27]。基质结构蛋白包括胶原纤维、层黏连蛋白和纤连蛋白,基质非结构蛋白包括结缔组织生长因子、血栓调节蛋白、骨桥蛋白、骨黏连蛋白等[28]。其中,结构蛋白参与维持心室正常结构和功能;非结构蛋白通过细胞表面受体、生长因子、蛋白酶等参与调节结构蛋白。

心脏细胞外基质的MMP，主要包括MMP-1、MMP-2、MMP-3、MMP-9、MMP-14。MMP可降解心脏所有基质成分,受促炎因子如白细胞介素-1、肿瘤坏死因子-α和促纤维化因子如转化生长因子-β、血管紧张素Ⅱ的调节[29-30]。MMP也受心肌成纤维细胞来源的TIMP调节，主要包括TIMP-1和TIMP-2[31]。TIMP是内源性MMP活性抑制剂,在维持成纤维细胞和MMP对细胞外基质的合成降解平衡中起重要作用[32]。

4HIF对AMI后细胞外基质的影响

AMI时心肌细胞不可逆性死亡,梗死区域炎症细胞浸润,使心肌成纤维细胞迅速被激活、增殖,释放炎症介质以及MMP，降解细胞外基质并吞噬组织碎片[33-34]。HIF在该过程中通过促基质细胞迁移、低氧条件下细胞存活、细胞分化、生长因子释放和基质合成等发挥重要作用[35]。组织缺氧时,HIF1α上调P4HA2、P4HA2和PLOD2的基因表达,从而促进胶原沉积[36]；HIF1α直接诱导TIMP-1、纤溶酶原激活物抑制剂-1、结缔组织生长因子的表达[37],促进损伤部位胶原沉积；HIF1α激活心脏成纤维细胞内的DNA甲基转移酶-1、DNA甲基转移酶-3β,引起细胞内广泛的DNA甲基化,激活转化生长因子-β信号通路,使细胞表达促纤维化基因产物如Ⅰ型、Ⅲ型胶原和α平滑肌肌动蛋白[38]。HIF1α过度表达则可使成纤维细胞过度增殖、分化，导致过多的基质形成及心脏舒张功能障碍。

5HIF促进AMI后新生血管合成

AMI后HIF1通过刺激各种来源的促血管生成基因的转录和表达,如刺激血管内皮生长因子、胎盘生长因子、血管生成素Ⅰ和Ⅱ、血小板源性生长因子和基质细胞因子1以促进缺血部位新生血管与侧支循环形成,从而减少梗死面积、保存心功能及降低患者死亡率[39]。在冠状动脉疾病患者中，HIF1α水平上调与侧支循环形成密切相关[40]。在大鼠心肌实验观察到HIF还可通过激活丝氨酸/苏氨酸激酶-AMP依赖的蛋白激酶信号途径，参与微血管内皮细胞的血管形成[41]。HIF1α可通过各种信号通路启动组织缺氧损伤后的组织修复进程,但其特异性调节机制尚未了解。

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(收稿：2015-06-08修回：2015-08-03)

(本文编辑：丁媛媛)

通信作者：顾新元，Email：guxinyuanbob@sina.com

基金项目：国家自然科学基金项目资助(81170241)

doi:10.3969/j.issn.1673-6583.2016.01.013