缺氧誘導(dǎo)因子對(duì)急性心肌梗死后心肌細(xì)胞及細(xì)胞外基質(zhì)的影響

張宇紅　鄧麗玉　林　彬　涂　勝　顧新元　唐利龍

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缺氧誘導(dǎo)因子對(duì)急性心肌梗死后心肌細(xì)胞及細(xì)胞外基質(zhì)的影響

張宇紅鄧麗玉林彬涂勝顧新元唐利龍

【摘要】急性心肌梗死是指冠狀動(dòng)脈閉塞、血流中斷，使部分心肌因嚴(yán)重的缺血缺氧而發(fā)生局部壞死。而缺氧誘導(dǎo)因子作為機(jī)體應(yīng)對(duì)缺氧應(yīng)激的適應(yīng)性反應(yīng)的調(diào)控因子，參與了急性心肌梗死時(shí)心臟的適應(yīng)性保護(hù)。該文主要介紹缺氧誘導(dǎo)因子的結(jié)構(gòu)、功能及其對(duì)急性心肌梗死后心肌細(xì)胞及細(xì)胞外基質(zhì)的影響。

【關(guān)鍵詞】急性心肌梗死；缺氧誘導(dǎo)因子；心肌細(xì)胞；細(xì)胞外基質(zhì)；心臟保護(hù)

作者單位：350108福建醫(yī)科大學(xué)附屬協(xié)和醫(yī)院心內(nèi)科(張宇紅，鄧麗玉，林彬，涂勝)；512000廣東省韶關(guān)市粵北人民醫(yī)院心內(nèi)科(顧新元)；411199湖南省湘潭市中心醫(yī)院心內(nèi)科(唐利龍)

1概述

缺氧誘導(dǎo)因子(hypoxia-inducible factor, HIF)是由α亞基和β亞基組成的異二聚體。哺乳動(dòng)物有3種HIFα亞基的亞型,包括HIF1α、2α和3α,其蛋白穩(wěn)定性受氧濃度影響；而HIF的β亞基則可穩(wěn)定地在核內(nèi)表達(dá)且不受氧濃度調(diào)節(jié)。α亞基和β亞基都是螺旋-環(huán)-螺旋肽蛋白家族的成員。

缺氧時(shí),脯氨酸羥化酶的活性受抑制,HIF1α通過HLH和PAS區(qū)域與β亞基異源二聚體化,并與轉(zhuǎn)錄的共激活劑CBP/P300相結(jié)合,形成活性轉(zhuǎn)錄復(fù)合體,轉(zhuǎn)位至細(xì)胞核,介導(dǎo)靶基因缺氧應(yīng)答部分的反向激活。HIF異二聚體可以識(shí)別并結(jié)合靶基因上的缺氧應(yīng)答序列,這些序列擁有共同的堿基片段5-(A/G)CGTG-3。

HIF1α在多細(xì)胞生物的各種類型細(xì)胞中均表達(dá)；而HIF2α僅在脊椎動(dòng)物的特定細(xì)胞如角質(zhì)形成細(xì)胞表達(dá),參與缺氧條件下新生血管形成；HIF3α也僅在某些特定細(xì)胞通過與HIF1α、HIF2α、HIF1β結(jié)合，抑制負(fù)性調(diào)節(jié)HIF1α、HIF2α的轉(zhuǎn)錄活性。迄今，大量實(shí)驗(yàn)證明HIF可以調(diào)節(jié)超過200多種基因的轉(zhuǎn)錄活性，在應(yīng)對(duì)缺氧應(yīng)激損傷的適應(yīng)性變化中發(fā)揮著重要作用。

2HIF對(duì)急性心肌梗死后心肌的影響

HIF是急性心肌梗死(AMI)后心臟適應(yīng)性變化過程中的主要調(diào)節(jié)因子之一。在人和鼠都觀察到，提高HIF活性后心肌梗死面積縮小,左室收縮功能提高，存活率提高[8-9]。HIF的心肌保護(hù)作用是由多因素介導(dǎo)的，包括一系列HIF靶基因的轉(zhuǎn)錄及其相應(yīng)信號(hào)轉(zhuǎn)導(dǎo)通路的活化。

2.1調(diào)節(jié)心肌收縮

心血管活性肽是心血管系統(tǒng)穩(wěn)態(tài)的重要調(diào)節(jié)者,也是HIF1的靶基因之一，在AMI早期有維持心肌收縮功能的作用[10]。心肌梗死后24 h內(nèi)，機(jī)體通過激活HIF通路使心血管活性肽水平急劇升高[11],心血管活性肽再通過磷酸肌醇信號(hào)途徑PKCε和細(xì)胞外信號(hào)調(diào)節(jié)激酶途徑ERK1/2,激活下游肌球蛋白輕鏈激酶。肌球蛋白輕鏈被磷酸化后，Ca2+敏感性增強(qiáng),促進(jìn)心肌肌絲交聯(lián),使心肌收縮力增強(qiáng)[12],從而實(shí)現(xiàn)心臟保護(hù)作用。

2.2調(diào)節(jié)心肌細(xì)胞能量代謝

HIF1α通過多種靶基因共同作用,減少AMI后基礎(chǔ)氧耗,提高葡萄糖利用效率。HIF介導(dǎo)AMI后心肌有氧代謝受抑,無氧酵解增強(qiáng)的過程。實(shí)驗(yàn)發(fā)現(xiàn)，HL-1心肌細(xì)胞在缺氧后24 h，葡萄糖轉(zhuǎn)運(yùn)體-1和乳酸水平分別增高5和15倍；線粒體電子傳遞鏈上的各種酶類包括復(fù)合體Ⅰ、Ⅳ和順烏頭酸酶的活性也不同程度地降低[13]。HIF1α促進(jìn)正葡萄糖轉(zhuǎn)運(yùn)蛋白-1(GLUT-1)、正葡萄糖轉(zhuǎn)運(yùn)蛋白-3(GLUT-3)和己糖激酶Ⅰ基因(HK-1)、己糖激酶Ⅱ基因(HK-2)基因表達(dá),使葡萄糖形成脫氧葡萄糖而不能進(jìn)一步代謝[14]; HIF1α促進(jìn)蛋白激酶(PDK)基因表達(dá),磷酸化丙酮酸脫氫酶(PDH)的催化區(qū)域使之失活,使丙酮酸進(jìn)入糖酵解途徑[15-16]；HIF1α促進(jìn)乳酸脫氫酶(LDH)和MOT4基因的表達(dá),使丙酮酸轉(zhuǎn)化為乳酸排出體外,同時(shí)減少線粒體源性自由基形成以減少心肌細(xì)胞死亡[17];另外正丙酮酸激酶PKM2既參與糖酵解過程，也與HIF1α結(jié)合促進(jìn)其反向激活功能，以正反饋機(jī)制實(shí)現(xiàn)缺氧狀態(tài)下心肌細(xì)胞代謝轉(zhuǎn)化[18]。

2.3調(diào)節(jié)線粒體功能

HIF通過綜合作用使線粒體產(chǎn)生的自由基減少,起到心肌保護(hù)作用。線粒體源性自由基增加會(huì)對(duì)細(xì)胞造成氧化應(yīng)激損傷[19]，HIF1有助于維持AMI后自由基的穩(wěn)態(tài)。HIF上調(diào)線粒體中線粒體蛋白BNIP3的水平,與自噬基因Beclin1競(jìng)爭(zhēng)結(jié)合癌基因Bcl2后，游離的Beclin1協(xié)同磷脂酰肌醇激酶3激活缺氧誘導(dǎo)性線粒體自溶[20]; HIF抑制線粒體電子傳遞鏈中復(fù)合體1和4的活性[21]; HIF激活微小核糖核酸miRNA-210，抑制鐵硫蛋白ISCU1/2，抑制三羧酸循環(huán)順烏頭酶和電子傳遞鏈復(fù)合體1的活性[22]。

2.4調(diào)節(jié)炎癥反應(yīng)

炎癥趨化因子(如基質(zhì)細(xì)胞衍生因子-12和單核細(xì)胞趨化蛋白-5)以及血管黏附分子(如細(xì)胞間黏附素和血管源性細(xì)胞黏附素)的mRNA表達(dá)在心肌梗死區(qū)域升高[23]。而HIF1可控制炎癥反應(yīng)，起心臟保護(hù)作用。有實(shí)驗(yàn)表明，在缺血再灌注前激活HIF活性,可減少心肌細(xì)胞表達(dá)趨化因子如角質(zhì)細(xì)胞源性趨化因子、巨噬細(xì)胞炎癥蛋白-2、單核細(xì)胞趨化因子-1、中性粒細(xì)胞趨化因子和細(xì)胞間黏附分子-1的表達(dá),從而抑制中性粒細(xì)胞的聚集，明顯降低了梗死面積[24]。

3心臟細(xì)胞外基質(zhì)組成和功能

正常情況下,細(xì)胞外基質(zhì)(ECM)由結(jié)構(gòu)蛋白、非結(jié)構(gòu)蛋白和由基質(zhì)金屬蛋白酶(MMP)、金屬蛋白酶組織抑制物(TIMP)組成的蛋白水解系統(tǒng)構(gòu)成。AMI后，左室細(xì)胞外基質(zhì)發(fā)生了一系列形態(tài)和功能的變化,稱為心室重構(gòu)。異常的心室重構(gòu)會(huì)導(dǎo)致心臟纖維化或心室過度擴(kuò)張[25-26]。

心臟細(xì)胞外基質(zhì)主要由纖維母細(xì)胞活化、增殖分化形成的心肌成纖維細(xì)胞合成[27]?；|(zhì)結(jié)構(gòu)蛋白包括膠原纖維、層黏連蛋白和纖連蛋白,基質(zhì)非結(jié)構(gòu)蛋白包括結(jié)締組織生長(zhǎng)因子、血栓調(diào)節(jié)蛋白、骨橋蛋白、骨黏連蛋白等[28]。其中,結(jié)構(gòu)蛋白參與維持心室正常結(jié)構(gòu)和功能;非結(jié)構(gòu)蛋白通過細(xì)胞表面受體、生長(zhǎng)因子、蛋白酶等參與調(diào)節(jié)結(jié)構(gòu)蛋白。

心臟細(xì)胞外基質(zhì)的MMP，主要包括MMP-1、MMP-2、MMP-3、MMP-9、MMP-14。MMP可降解心臟所有基質(zhì)成分,受促炎因子如白細(xì)胞介素-1、腫瘤壞死因子-α和促纖維化因子如轉(zhuǎn)化生長(zhǎng)因子-β、血管緊張素Ⅱ的調(diào)節(jié)[29-30]。MMP也受心肌成纖維細(xì)胞來源的TIMP調(diào)節(jié)，主要包括TIMP-1和TIMP-2[31]。TIMP是內(nèi)源性MMP活性抑制劑,在維持成纖維細(xì)胞和MMP對(duì)細(xì)胞外基質(zhì)的合成降解平衡中起重要作用[32]。

4HIF對(duì)AMI后細(xì)胞外基質(zhì)的影響

AMI時(shí)心肌細(xì)胞不可逆性死亡,梗死區(qū)域炎癥細(xì)胞浸潤(rùn),使心肌成纖維細(xì)胞迅速被激活、增殖,釋放炎癥介質(zhì)以及MMP，降解細(xì)胞外基質(zhì)并吞噬組織碎片[33-34]。HIF在該過程中通過促基質(zhì)細(xì)胞遷移、低氧條件下細(xì)胞存活、細(xì)胞分化、生長(zhǎng)因子釋放和基質(zhì)合成等發(fā)揮重要作用[35]。組織缺氧時(shí),HIF1α上調(diào)P4HA2、P4HA2和PLOD2的基因表達(dá),從而促進(jìn)膠原沉積[36]；HIF1α直接誘導(dǎo)TIMP-1、纖溶酶原激活物抑制劑-1、結(jié)締組織生長(zhǎng)因子的表達(dá)[37],促進(jìn)損傷部位膠原沉積；HIF1α激活心臟成纖維細(xì)胞內(nèi)的DNA甲基轉(zhuǎn)移酶-1、DNA甲基轉(zhuǎn)移酶-3β,引起細(xì)胞內(nèi)廣泛的DNA甲基化,激活轉(zhuǎn)化生長(zhǎng)因子-β信號(hào)通路,使細(xì)胞表達(dá)促纖維化基因產(chǎn)物如Ⅰ型、Ⅲ型膠原和α平滑肌肌動(dòng)蛋白[38]。HIF1α過度表達(dá)則可使成纖維細(xì)胞過度增殖、分化，導(dǎo)致過多的基質(zhì)形成及心臟舒張功能障礙。

5HIF促進(jìn)AMI后新生血管合成

AMI后HIF1通過刺激各種來源的促血管生成基因的轉(zhuǎn)錄和表達(dá),如刺激血管內(nèi)皮生長(zhǎng)因子、胎盤生長(zhǎng)因子、血管生成素Ⅰ和Ⅱ、血小板源性生長(zhǎng)因子和基質(zhì)細(xì)胞因子1以促進(jìn)缺血部位新生血管與側(cè)支循環(huán)形成,從而減少梗死面積、保存心功能及降低患者死亡率[39]。在冠狀動(dòng)脈疾病患者中，HIF1α水平上調(diào)與側(cè)支循環(huán)形成密切相關(guān)[40]。在大鼠心肌實(shí)驗(yàn)觀察到HIF還可通過激活絲氨酸/蘇氨酸激酶-AMP依賴的蛋白激酶信號(hào)途徑，參與微血管內(nèi)皮細(xì)胞的血管形成[41]。HIF1α可通過各種信號(hào)通路啟動(dòng)組織缺氧損傷后的組織修復(fù)進(jìn)程,但其特異性調(diào)節(jié)機(jī)制尚未了解。

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(收稿：2015-06-08修回：2015-08-03)

(本文編輯：丁媛媛)

通信作者：顧新元，Email：guxinyuanbob@sina.com

基金項(xiàng)目：國(guó)家自然科學(xué)基金項(xiàng)目資助(81170241)

doi:10.3969/j.issn.1673-6583.2016.01.013