目的 探討原兒茶酸（PCA）對(duì)心肌缺血再灌注損傷（MIRI）大鼠及缺氧/復(fù)氧H9c2細(xì)胞的影響及作用機(jī)制。方法 將心肌細(xì)胞H9c2分為對(duì)照組、模型組和PCA（1.562、3.125、6.250 μmol·L^-1）組，預(yù)給藥24 h后，除對(duì)照組外厭氧箱缺氧4 h、復(fù)氧12 h在體外構(gòu)建H9c2細(xì)胞損傷模型； CCK8法檢測(cè)細(xì)胞存活率； JC-1染色檢測(cè)細(xì)胞內(nèi)線粒體膜電位； DCFHDA熒光探針檢測(cè)細(xì)胞中活性氧（ROS）； Calcein AM熒光探針檢測(cè)線粒體通透性轉(zhuǎn)換孔（mPTP）開放程度；試劑盒法檢測(cè)腺嘌呤核苷三磷酸（ATP）水平； FerroOrange熒光探針檢測(cè)Fe²⁺水平檢測(cè)。SD大鼠隨機(jī)分為假手術(shù)組、單給PCA（100 mg·kg^-1）組、模型組、阿司匹林（100 mg·kg^-1）組和PCA低、中、高劑量（25、50、100 mg·kg^-1）組，假手術(shù)組和模型組ig給予0.5%羥甲基纖維素鈉溶液，其余組ig給藥，每天1次，連續(xù)2周，末次給藥24 h后，除假手術(shù)組和單給PCA組外，采用冠狀動(dòng)脈左前降支結(jié)扎手術(shù)結(jié)扎30 min制備MIRI大鼠模型；通過2，3，5-氯化三苯基四氮唑（TTC）染色分析心肌梗死面積；蘇木精-伊紅（HE）染色分析各組心肌組織結(jié)構(gòu)變化；心臟超聲評(píng)估射血分?jǐn)?shù)（EF）和左室短軸縮短率（FS）；通過ELISA法檢測(cè)血清肌酸激酶同工酶（CK-MB）、乳酸脫氫酶（LDH）、天冬氨酸氨基轉(zhuǎn)移酶（AST）、丙二醛（MDA）和谷胱甘肽（GSH）水平； Western blotting檢測(cè)心臟組織中谷胱甘肽過氧化物酶4 （GPX4）和溶質(zhì)載體家族7成員11 （SLC7A11）蛋白表達(dá)情況。結(jié)果 細(xì)胞實(shí)驗(yàn)結(jié)果表明，與模型組比較，PCA顯著升高H9c2細(xì)胞存活率（P＜0.01），升高線粒體膜電位（P＜0.01），抑制異常mPTP開放和ROS的大量產(chǎn)生（P＜0.01），改善線粒體ATP合成受損（P＜0.01）；明顯降低細(xì)胞內(nèi)Fe²⁺水平；動(dòng)物實(shí)驗(yàn)結(jié)果表明，與模型組比較，PCA組心肌梗死率顯著降低（P＜0.01），心肌組織結(jié)構(gòu)得到改善，EF和FS顯著升高（P＜0.01），心肌損傷標(biāo)志物CK-MB、LDH、AST、MDA水平顯著降低（P＜0.01），GSH水平和GPX4、SLC7A11蛋白表達(dá)水平顯著升高（P＜0.01）。結(jié)論 PCA可以改善MIRI，減輕心肌組織病理損傷，調(diào)節(jié)體內(nèi)鐵死亡，可能與升高GPX4、SLC7A11表達(dá)有關(guān)。

Objective To explore the effects and mechanisms of protocatechuic acid (PCA) on myocardial ischemia-reperfusion injury (MIRI) in rats and hypoxia/reoxygenation H9c2 cells. Methods H9c2 cells were divided into the control group, model group, and PCA (1.562, 3.125, and 6.250 μmol·L^-1) groups. After 24 h of pretreatment, except for the control group, the cells were subjected to 4 h of hypoxia and 12 h of reoxygenation in an anaerobic chamber to establish the H9c2 cell injury model in vitro. Cell viability was detected by CCK-8 assay; mitochondrial membrane potential was detected by JC-1 staining; Reactive oxygen species (ROS) in cells were detected by DCFH-DA fluorescence probe; The opening degree of mitochondrial permeability transition pore (mPTP) was detected by Calcein AM fluorescence probe; Adenine nucleoside triphosphate (ATP) levels were detected by kit method; Fe²⁺ levels were detected by FerroOrange fluorescence probe. SD rats were randomly divided into sham operation group, single PCA (100 mg·kg^-1) group, model group, aspirin (100 mg·kg^-1) group, and PCA low, medium, and high dose (25, 50, 100 mg·kg^-1) groups. The sham operation group and model control group were ig administered 0.5% sodium carboxymethyl cellulose solution, and the other groups were ig administered drugs once a day for two weeks. 24 h after the last administration, except for the sham operation group, the MIRI rat model was established by ligating the left anterior descending coronary artery for 30 min. Myocardial infarction area was analyzed by 2, 3, 5-triphenyltetrazolium chloride (TTC) staining; Myocardial tissue structure changes were analyzed by hematoxylin-eosin (HE) staining; Left ventricular ejection fraction (EF) and fractional shortening (FS) were evaluated by cardiac ultrasound; Serum creatine kinase isoenzyme (CK-MB), lactate dehydrogenase (LDH), aspartate aminotransferase (AST), malondialdehyde (MDA), and glutathione (GSH) levels were detected by ELISA; The expression of glutathione peroxidase 4 (GPX4) and solute carrier family 7 member 11 (SLC7A11) proteins in cardiac tissue was detected by Western blotting. Results The cell experiment results showed that compared with the model group, PCA significantly increased the survival rate of H9c2 cells (P<0.01), increased mitochondrial membrane potential (P<0.01), inhibited abnormal mPTP opening and excessive ROS production (P<0.01), improved mitochondrial ATP synthesis impairment (P<0.01), and significantly reduced intracellular Fe²⁺ levels. The animal experiment results showed that compared with the model group, the myocardial infarction rate in the PCA group was significantly reduced (P<0.01), myocardial tissue structure was improved, EF and FS were significantly increased (P<0.01), myocardial injury markers CK-MB, LDH, AST, and MDA levels were significantly decreased (P<0.01), and GSH levels and GPX4 and SLC7A11 protein expression levels were significantly increased (P<0.01). Conclusion PCA ameliorates myocardial reperfusion injury, attenuates myocardial histopathological damage, and regulates iron death in vivo, which may be related to the inhibition of GPX4 and SLC7A11 expression.

R285.5

中國醫(yī)學(xué)科學(xué)院醫(yī)學(xué)與健康科技創(chuàng)新工程項(xiàng)目（2021-I2M-1-071）