盧婷，郜攀，司良毅，趙坤

·基礎(chǔ)研究·

AMPK調(diào)控Ca2+內(nèi)流對(duì)高糖誘導(dǎo)內(nèi)皮細(xì)胞凋亡的作用及其機(jī)制研究

盧婷，郜攀，司良毅，趙坤

目的觀(guān)察AMP依賴(lài)的蛋白激酶(AMPK)對(duì)高糖刺激內(nèi)皮細(xì)胞凋亡的抑制作用，并初步探討其機(jī)制。方法體外培養(yǎng)MS-1內(nèi)皮細(xì)胞株，分別用AMPK激動(dòng)劑、AMPK抑制劑、鈣庫(kù)依賴(lài)性鈣離子通道(SOCC)抑制劑2-APB 和(或)高糖處理，另設(shè)對(duì)照組(未經(jīng)任何方式干預(yù))。采用TUNEL法檢測(cè)細(xì)胞凋亡情況，激光共聚焦顯微鏡檢測(cè)細(xì)胞內(nèi)鈣離子(Ca2+)內(nèi)流，Western blotting檢測(cè)SOCC蛋白Stim1和Orai1的表達(dá)。結(jié)果與對(duì)照組比較，高糖能夠明顯誘導(dǎo)內(nèi)皮細(xì)胞凋亡，增加Stim1和Orai1蛋白表達(dá)(P<0.05)。與高糖組比較，AMPK抑制劑+高糖能夠明顯增強(qiáng)高糖誘導(dǎo)的內(nèi)皮細(xì)胞的凋亡(P<0.05)，而AMPK激動(dòng)劑+高糖能夠明顯抑制高糖誘導(dǎo)的內(nèi)皮細(xì)胞凋亡，并降低Stim1和Orai1蛋白表達(dá)(P<0.05)。與對(duì)照組比較，高糖能夠明顯誘導(dǎo)內(nèi)皮細(xì)胞Ca2+內(nèi)流；與高糖組比較，2-APB+高糖能夠明顯抑制高糖誘導(dǎo)的內(nèi)皮細(xì)胞Ca2+內(nèi)流，并阻斷高糖對(duì)內(nèi)皮細(xì)胞凋亡的誘導(dǎo)作用，而AMPK激動(dòng)劑能夠明顯抑制高糖誘導(dǎo)的內(nèi)皮細(xì)胞Ca2+內(nèi)流。結(jié)論AMPK能夠通過(guò)降低Stim1和Orai1蛋白的表達(dá)，抑制SOCC介導(dǎo)的Ca2+內(nèi)流，進(jìn)而阻斷高糖刺激的內(nèi)皮細(xì)胞凋亡，對(duì)內(nèi)皮細(xì)胞功能起重要的保護(hù)作用。

AMP活化蛋白激酶類(lèi)；鈣離子載體；高血糖癥；鈣庫(kù)依賴(lài)性鈣離子通道；內(nèi)皮細(xì)胞

隨著中國(guó)步入老年化社會(huì)，冠心病(CHD)的發(fā)病率和致死率逐年增高[1]。動(dòng)脈粥樣硬化(atherosclerosis，AS)是CHD的病理學(xué)基礎(chǔ)[2]，其中血管壁的內(nèi)皮細(xì)胞(endothelial cells，ECs)在血管穩(wěn)態(tài)平衡中具有重要作用，研究證實(shí)ECs受損和動(dòng)脈粥樣硬化的多種傳統(tǒng)危險(xiǎn)因素均可誘導(dǎo)ECs凋亡，而ECs凋亡可能是動(dòng)脈粥樣硬化病理發(fā)生的早期事件，并可促進(jìn)粥樣硬化病變形成、斑塊侵蝕和急性冠脈綜合征形成[3]。

AMP依賴(lài)的蛋白激酶(adenosine 5'-monophosphateactivated protein kinase，AMPK)是生物能量代謝調(diào)節(jié)的關(guān)鍵分子[4]，本課題組在先前的實(shí)驗(yàn)中發(fā)現(xiàn)抑制AMPK的活性能夠誘導(dǎo)高糖對(duì)內(nèi)皮細(xì)胞的損傷[5]。近年研究發(fā)現(xiàn)，一種新型的鈣離子通道——鈣庫(kù)依賴(lài)性鈣離子通道(store operated Ca2+channel，SOCC)在調(diào)控細(xì)胞內(nèi)鈣離子內(nèi)流中發(fā)揮重要作用[6]，尤其是SOCC調(diào)節(jié)的鈣離子內(nèi)流在調(diào)控細(xì)胞周期、分化、增殖和凋亡中具有非常重要的作用。SOCC的主要組成成分是兩個(gè)相互作用的分子Orai1和Stim1，研究發(fā)現(xiàn)抑制SOCC蛋白表達(dá)可以抑制神經(jīng)細(xì)胞凋亡[7]。本實(shí)驗(yàn)擬觀(guān)察在高糖刺激下，AMPK對(duì)內(nèi)皮細(xì)胞凋亡的保護(hù)作用并初步探討SOCC通道在此過(guò)程中的作用。

1 材料與方法

1.1 主要材料及試劑 內(nèi)皮細(xì)胞采用ATCC細(xì)胞庫(kù)的小鼠胰島微血管內(nèi)皮細(xì)胞株MS-1(ATCC? CRL-2460TM，美國(guó))。DMEM培養(yǎng)基、胎牛血清(FBS)購(gòu)自美國(guó)Gibco公司，HBSS緩沖液和PBS緩沖液購(gòu)自北京鼎國(guó)生物試劑公司，TUNEL凋亡檢測(cè)試劑盒購(gòu)自瑞士Roche公司，Stim1、Orai1一抗抗體購(gòu)自英國(guó)Abcam公司，內(nèi)參GAPDH一抗、辣根過(guò)氧化物酶標(biāo)記的兔抗山羊IgG購(gòu)自美國(guó)Santa Cruz公司，RIPA蛋白裂解液購(gòu)自北京鼎國(guó)生物公司，其余試劑包括AMPK抑制劑和激動(dòng)劑等均購(gòu)自日本Sigma公司。

1.2 細(xì)胞培養(yǎng) 將購(gòu)買(mǎi)的MS-1內(nèi)皮細(xì)胞迅速用20% FBS溶解，1000r/min離心后得到的沉淀再次用含有5% FBS的DMEM培養(yǎng)基溶解，分裝到培養(yǎng)瓶中，在37℃、5%CO2培養(yǎng)孵育箱中進(jìn)行培養(yǎng)。以復(fù)蘇傳代后3～5代MS-1細(xì)胞株進(jìn)行實(shí)驗(yàn)，0.25%胰蛋白酶消化制備細(xì)胞懸液，調(diào)整細(xì)胞密度為5×107/L，接種于培養(yǎng)板備用。經(jīng)37℃、5%CO2、5%FBS預(yù)培養(yǎng)24h和0.5%胎牛血清培養(yǎng)基預(yù)培養(yǎng)處理12h后進(jìn)行分組。

1.3 細(xì)胞分組及凋亡檢測(cè) 為明確AMPK對(duì)高糖誘導(dǎo)內(nèi)皮細(xì)胞凋亡的作用，將細(xì)胞分為6組:對(duì)照組(未經(jīng)任何方式干預(yù)）、高糖組(30mmol/L葡萄糖處理48h)、高糖+AMPK抑制劑組(30mmol/L葡萄糖處理48h+20μmol/L復(fù)合物C預(yù)處理1h)、高糖+AMPK激動(dòng)劑組(30mmol/L葡萄糖處理48h+1mmol/L AICAR預(yù)處理1h)、復(fù)合物C組(20μmol/L復(fù)合物C預(yù)處理1h)和AICAR組(1mmol/L AICAR預(yù)處理1h)。為進(jìn)一步明確高糖誘導(dǎo)的鈣內(nèi)流對(duì)內(nèi)皮細(xì)胞凋亡的影響，將細(xì)胞分為4組：對(duì)照組、高糖組、高糖+SOCC抑制劑(2-APB)組(30mmol/L葡萄糖處理48h+75μmol/L 2-APB預(yù)處理1h)、2-APB組(75μmol/ L 2-APB預(yù)處理1h)。細(xì)胞經(jīng)胰酶消化后，制成細(xì)胞爬片，按照Roche凋亡試劑盒說(shuō)明書(shū)方法進(jìn)行，將載玻片取出置于水平面，用濾紙小心吸去多余液體，將細(xì)胞用4%多聚甲醛(溶于pH7.4的PBS中)在室溫下固定30min，PBS漂洗5次；浸入封閉液(3%H2O2溶于甲醇)中，室溫封閉10min后再用PBS漂洗5min×3次；浸入細(xì)胞膜通透液中，室溫下反應(yīng)30s～2min。采用TUNEL凋亡試劑盒中的標(biāo)記物標(biāo)記凋亡的細(xì)胞核，以每100個(gè)細(xì)胞中的TUNEL凋亡細(xì)胞數(shù)表示結(jié)果。

1.4 細(xì)胞分組及鈣離子內(nèi)流檢測(cè) 為明確高糖誘導(dǎo)的鈣內(nèi)流與SOCC通道的關(guān)系，將細(xì)胞分為4組：對(duì)照組、高糖組、高糖+2-APB組和2-APB組。為進(jìn)一步明確AMPK對(duì)高糖誘導(dǎo)的鈣內(nèi)流的影響，將細(xì)胞分為4組：對(duì)照組、高糖組、高糖+AICAR組和AICAR組。將內(nèi)皮細(xì)胞培育在HEPES緩沖液中，采用激光共聚焦體系檢測(cè)胞質(zhì)內(nèi)Ca2+探針Fura-2/AM熒光強(qiáng)度。細(xì)胞在6孔板中以1% FBS孵育2h，用HBSS緩沖液洗1次，加入3μmol/L的Fura-2/AM熒光探針?lè)跤?0min，用HBSS緩沖液清洗1次。采用卡爾蔡司激光共聚焦顯微鏡(Carl Zeiss GmbH，德國(guó))采集熒光探針信號(hào)，雙激發(fā)波長(zhǎng)為340、380nm，發(fā)射波長(zhǎng)為520nm。其中毒胡蘿卜素(thapsigargin，TG，實(shí)驗(yàn)濃度1μmol/L)為誘導(dǎo)內(nèi)質(zhì)網(wǎng)鈣庫(kù)釋放進(jìn)而激活SOCC鈣離子通道的SOCC激動(dòng)劑，于實(shí)驗(yàn)第2分鐘時(shí)加入。最初3min內(nèi)，培養(yǎng)基中無(wú)Ca2+([Ca2+]0)，此后于第3分鐘后開(kāi)始加入1.5mmol/ L CaCl2([Ca2+]1.5)至反應(yīng)液中，每隔10s采集一次熒光信號(hào)，采用Zen2012軟件系統(tǒng)檢測(cè)各個(gè)時(shí)間點(diǎn)的熒光強(qiáng)度并繪制曲線(xiàn)。

1.5 Western blotting檢測(cè)細(xì)胞內(nèi)SOCC蛋白表達(dá)細(xì)胞分4組：對(duì)照組、高糖組、高糖+AICAR組和AICAR組。收集各組細(xì)胞后，以RIPA法裂解細(xì)胞，采用考馬斯亮藍(lán)法進(jìn)行蛋白定量。取約40μg總蛋白，置于95℃沸水中變性5min，采用5%濃縮膠和10%分離膠行SDS聚丙烯酰胺凝膠電泳，電泳條件恒壓80mV，時(shí)間為3h。然后采用三明治法將蛋白轉(zhuǎn)移至PVD膜，轉(zhuǎn)膜采用Bio-Rad電轉(zhuǎn)儀，轉(zhuǎn)膜條件為恒流15mA，時(shí)間為90min。之后將PVDF膜用5%脫脂奶粉封閉1h，加入封閉液稀釋的一抗，山羊抗小鼠Stim1多克隆抗體(1:200稀釋)和山羊抗小鼠Orai1多克隆抗體(1:200稀釋)，封閉過(guò)夜，加辣根過(guò)氧化物酶標(biāo)記的兔抗山羊IgG(稀釋濃度1:500)室溫反應(yīng)1h后染色。用相同方法檢測(cè)內(nèi)參GAPDH蛋白表達(dá)(稀釋濃度1:1000)。用FR-200凝膠成像分析系統(tǒng)進(jìn)行定量分析。

1.6 統(tǒng)計(jì)學(xué)處理 采用SPSS 19.0軟件進(jìn)行分析。計(jì)量資料以表示，比較其方差齊性，多組間比較采用單因素方差分析，進(jìn)一步兩兩比較采用LSD-t檢驗(yàn)。P<0.05為差異有統(tǒng)計(jì)學(xué)意義。

2 結(jié) 果

2.1 AMPK對(duì)高糖誘導(dǎo)內(nèi)皮細(xì)胞凋亡的抑制作用體外培養(yǎng)MS-1內(nèi)皮細(xì)胞株，預(yù)先用復(fù)合物C或AICAR預(yù)處理，再加入高糖刺激內(nèi)皮細(xì)胞，TUNEL法檢測(cè)細(xì)胞的凋亡情況。結(jié)果顯示，與對(duì)照組比較，高糖刺激后內(nèi)皮細(xì)胞凋亡顯著增加(P<0.05)；與高糖刺激組比較，AICAR能顯著抑制高糖刺激的內(nèi)皮細(xì)胞凋亡(P<0.05)，復(fù)合物C則進(jìn)一步誘導(dǎo)內(nèi)皮細(xì)胞凋亡(P<0.05，圖1)。

圖1 AMPK對(duì)高糖誘導(dǎo)內(nèi)皮細(xì)胞凋亡的影響(TUNEL ×10)Fig.1 Effects of AMPK on high glucose-stimulated endothelial cell apoptosis (TUNEL ×10)HG. High glucose; AICAR. 5-aminoimidazole-4-carboxamide-1-beta-d-ribofuranoside (an AMPK agonist). (1)P<0.05 compared with control group; (2)P<0.05 compared with HG group

2.2 高糖誘導(dǎo)細(xì)胞鈣離子內(nèi)流([Ca2+]i)對(duì)內(nèi)皮細(xì)胞凋亡的影響 將內(nèi)皮細(xì)胞予以2-APB預(yù)處理后，再予以高糖刺激。結(jié)果顯示，與對(duì)照組相比，高糖刺激可明顯誘導(dǎo)細(xì)胞內(nèi)SOCC介導(dǎo)的Ca2+內(nèi)流，而與高糖組相比，2-APB則能夠明顯抑制高糖誘導(dǎo)的Ca2+內(nèi)流(P<0.05)。另一方面，高糖誘導(dǎo)的內(nèi)皮細(xì)胞凋亡也可被2-APB明顯抑制(P<0.05，圖2)。

圖2 內(nèi)皮細(xì)胞[Ca2+]i對(duì)內(nèi)皮細(xì)胞凋亡的影響Fig.2 Effects of calcium influx on the apoptosis of endothelial cellsHG. High glucose; 2-APB. 2-aminoethyl diphenylborinate (a blocker of store-operated Ca2+channels). A. Effects of SOCC blocker (2-APB) on high glucose-induced calcium influx; B. Effects of SOCC blocker (2-APB) on high glucose-induced endothelial cell apoptosis; C. Column chart of endothelial cell apoptosis (TUNEL). (1)P<0.05 compared with control group; (2)P<0.05 compared with HG group

2.3 AMPK對(duì)高糖誘導(dǎo)的[Ca2+]i的影響 細(xì)胞予AMPK激動(dòng)劑AICAR(1mmol/L)預(yù)處理1h，再加入30mmol/L高糖刺激4h，細(xì)胞分成4組：對(duì)照組、高糖組、高糖+AICAR組、AICAR組。結(jié)果顯示，高糖誘導(dǎo)的Ca2+內(nèi)流被AMPK激動(dòng)劑AICAR顯著抑制(P<0.05，圖3)。

2.4 AMPK對(duì)高糖誘導(dǎo)的內(nèi)皮細(xì)胞SOCC通道蛋白表達(dá)的抑制作用 細(xì)胞予AICAR預(yù)處理后，再予高糖處理，檢測(cè)各組SOCC通道蛋白分子Stim1和Orai1蛋白表達(dá)情況。與對(duì)照組比較，高糖刺激可明顯增加Stim1和Orai1蛋白的表達(dá)，而AICAR則能夠明顯抑制Stim1、Orai1蛋白的表達(dá)(P<0.05，圖4)。

3 討 論

CHD是老年人的常見(jiàn)病和多發(fā)病，也是老年患者臨床死亡的重要原因，其病理基礎(chǔ)為冠狀動(dòng)脈粥樣硬化。研究發(fā)現(xiàn)，內(nèi)皮細(xì)胞能夠釋放大量活性因子，如一氧化氮、血管緊張素和內(nèi)皮素等，可調(diào)節(jié)冠脈事件中的血栓形成、炎癥和免疫反應(yīng)[8-10]。血管內(nèi)皮在外界因素作用下出現(xiàn)損傷、凋亡，可使血管平滑肌直接暴露于血液中的脂蛋白、活性氧等刺激因素，推動(dòng)血管內(nèi)皮細(xì)胞的異常修復(fù)[11-12]，最終導(dǎo)致動(dòng)脈粥樣硬化的發(fā)生[13]。

圖3 AMPK對(duì)高糖誘導(dǎo)的[Ca2+]i的抑制作用Fig.3 Inhibitive effect of AMPK on high glucose-induced Ca2+influx to endothelial cellsHG. High glucose; AICAR. 5-aminoimidazole-4-carboxamide-1-beta-d-ribofuranoside (an AMPK agonist); TG. Thapsigargin; (1)P<0.05 compared with control group; (2)P<0.05 compared with HG group

圖4 AMPK對(duì)高糖誘導(dǎo)的內(nèi)皮細(xì)胞SOCC通道Stim1、Orai1蛋白表達(dá)的影響(Western blotting)Fig.4 Effects of AMPK on the expressions of high glucose-stimulated expression of SOCC proteins (Stim1 and Orai1) in endothelial cells (Western blotting)HG. High glucose; AICAR. 5-aminoimidazole-4-carboxamide-1-beta-d-ribofuranoside (an AMPK agonist). (1)P<0.05 compared with control group; (2)P<0.05 compared with HG group

Ca2+對(duì)維持細(xì)胞正常生理功能具有重要作用，一方面可作為第二信使介導(dǎo)細(xì)胞內(nèi)多種生物學(xué)反應(yīng)，另一方面Ca2+穩(wěn)態(tài)在維持細(xì)胞的增殖、分化和凋亡中發(fā)揮樞紐作用[14-15]。研究表明，血管內(nèi)皮細(xì)胞外的Ca2+經(jīng)細(xì)胞膜上的鈣通道流入細(xì)胞內(nèi)，導(dǎo)致細(xì)胞內(nèi)Ca2+超載，造成血管內(nèi)皮細(xì)胞損傷；毒胡蘿卜素通過(guò)抑制內(nèi)質(zhì)網(wǎng)鈣-ATP酶、排空內(nèi)質(zhì)網(wǎng)Ca2+誘導(dǎo)細(xì)胞嚴(yán)重的內(nèi)質(zhì)網(wǎng)應(yīng)激反應(yīng)，導(dǎo)致內(nèi)皮細(xì)胞凋亡[16]。本研究中，高糖刺激內(nèi)皮細(xì)胞后Ca2+內(nèi)流明顯增加，而2-APB作為SOCC通道抑制劑在抑制高糖誘導(dǎo)內(nèi)皮細(xì)胞Ca2+內(nèi)流的同時(shí)，還可抑制高糖誘導(dǎo)的內(nèi)皮細(xì)胞凋亡，提示內(nèi)皮細(xì)胞Ca2+內(nèi)流能夠明顯誘導(dǎo)內(nèi)皮細(xì)胞凋亡的發(fā)生。

AMPK是一個(gè)含有絲/蘇氨酸蛋白激酶活性的異源三聚體，由催化亞基α、調(diào)節(jié)亞基β和γ通過(guò)不同的排列和空間構(gòu)象形成。既往本課題組在觀(guān)察白藜蘆醇(一種具有抗氧化、抗炎、抗凋亡等作用的多酚類(lèi)化合物)拮抗高糖致內(nèi)皮細(xì)胞損害的研究中發(fā)現(xiàn)，抑制AMPK活性后，白藜蘆醇對(duì)內(nèi)皮細(xì)胞功能的保護(hù)作用顯著降低，說(shuō)明AMPK在內(nèi)皮細(xì)胞功能的維持中發(fā)揮重要作用[5,17]。SOCC是一種可持續(xù)誘導(dǎo)Ca2+內(nèi)流的鈣通道，可調(diào)節(jié)細(xì)胞的基因轉(zhuǎn)錄、生長(zhǎng)和分化等。SOCC的作用機(jī)制為：當(dāng)外界或內(nèi)在刺激使胞內(nèi)鈣庫(kù)中Ca2+釋放，使鈣庫(kù)中Ca2+濃度下降到一定程度時(shí)，細(xì)胞膜上SOCC開(kāi)放產(chǎn)生Ca2+內(nèi)流，引起細(xì)胞內(nèi)Ca2+濃度持續(xù)緩慢上升，從而補(bǔ)充胞質(zhì)和鈣庫(kù)中的Ca2+[18]。目前研究發(fā)現(xiàn)，Stim1和Orai是內(nèi)皮細(xì)胞最主要的SOCC蛋白成分[19-20]。本實(shí)驗(yàn)首先觀(guān)察了AMPK激動(dòng)劑AICAR和高糖刺激對(duì)鈣通道蛋白表達(dá)的影響，結(jié)果顯示高糖能夠促進(jìn)Stim1和Orai1蛋白的表達(dá)，而AICAR則能夠顯著抑制高糖誘導(dǎo)的Stim1和Orai1蛋白表達(dá)，進(jìn)一步研究發(fā)現(xiàn)AICAR可顯著抑制高糖誘導(dǎo)的Ca2+內(nèi)流，結(jié)果提示激動(dòng)AMPK后能夠顯著抑制高糖誘導(dǎo)的內(nèi)皮細(xì)胞凋亡，其機(jī)制可能與抑制SOCC介導(dǎo)的Ca2+內(nèi)流密切相關(guān)。

綜上所述，內(nèi)皮細(xì)胞凋亡在動(dòng)脈粥樣硬化發(fā)病中具有重要作用，而AMPK能夠通過(guò)抑制SOCC介導(dǎo)的Ca2+內(nèi)流而抑制內(nèi)皮細(xì)胞凋亡，對(duì)內(nèi)皮細(xì)胞功能起到重要的保護(hù)作用。后續(xù)研究應(yīng)觀(guān)察在動(dòng)物模型中AMPK對(duì)內(nèi)皮細(xì)胞凋亡的影響，并進(jìn)一步明確AMPK在抑制內(nèi)皮細(xì)胞凋亡時(shí)發(fā)揮作用的具體亞單位。

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Effects of AMPK on high glucose stimulated apoptosis of endothelial cellsviaregulation of calcium influx

LU Ting, GAO Pan, SI Liang-yi*, ZHAO Kun
Department of Geriatrics, Xinan Hospital, Third Military Medical University, Chongqing 400038, China
*< class="emphasis_italic">Corresponding author, E-mail: doctorsly@126.com

, E-mail: doctorsly@126.com
This work was supported by the National Natural Science Foundation of China (81370446, 81370007)

ObjectiveTo investigate the inhibitory effect of adenosine monophosphate (AMP)-dependent protein kinase (AMPK) on high glucose-stimulated endothelial cell apoptosis and its mechanism.MethodsMS-1 endothelial cells were culturedin vitro, and they were treated with AMPK agonist, AMPK inhibitor, 2-APB (a blocker of store operated Ca2+channel (SOCC)) and (or) high glucose, and a control group without any intervention were set up. TUNEL assay was performed to determine apoptotic cells. Laser scanning confocal microscopy was used to assess the Ca2+influx into cells, and Western-blotting was performed to determine the expressions of Stim1 and Orai1 of the store operated Ca2+channel (SOCC) proteins.ResultsApoptosis of endothelial cells was induced significantly, and the expressions of Stim1 and Orai1 were upregulated in high glucose group compared with that in control group (P<0.05). The rate of apoptosis of high glucose-induced endothelial cell was found to be increased in AMPK inhibitor group and decreased in AMPK agonist group, and the expressions of Stim1 and Orai1 were found to be downregulated in AMPK agonist group as compared with that in high glucose group (P<0.05). Compared with the control group, high glucose stimulation significantly induced the Ca2+influx to endothelial cells; compared with high glucose group, 2-APB significantly inhibited high glucose-induced Ca2+influx to endothelial cells, and blocked the inducing effect of high-glucose on endothelial cell apoptosis. Compared with high glucose group, AMPK agonist significantly inhibited high glucose-induced cell Ca2+influx.ConclusionBy reducing the expressions of Stim1 and Orai1, AMPK may inhibit SOCC-mediated Ca2+influx, and block the high glucose-stimulated endothelial cell apoptosis, thus play an important protective role in sustaining endothelial cell function.

AMP-activated protein kinases; calcium ionophores; hyperglycemia; store operated Ca2+channel; endothelial cells

R345

A

0577-7402(2015)10-0773-05

10.11855/j.issn.0577-7402.2015.10.01

2015-03-23；

2015-07-12)

(責(zé)任編輯：張小利)

國(guó)家自然科學(xué)基金(81370446、81370007)

盧婷，碩士研究生。主要從事老年心血管病方面的研究

400038 重慶 第三軍醫(yī)大學(xué)西南醫(yī)院老年病科(盧婷、郜攀、司良毅、趙坤)

司良毅，E-mail：doctorsly@126.com