[Sacubitril/valsartan attenuates left ventricular remodeling and improve cardiac function by upregulating apelin/APJ pathway in rats with heart failure].

Objective: To investigate the effect and mechanism of sacubitril/valsartan on left ventricular remodeling and cardiac function in rats with heart failure. Methods: A total of 46 SPF-grade male Wistar rats weighed 300-350 g were acclimatized to the laboratory for 7 days. Rats were then divided into 4 groups: the heart failure group (n=12, intraperitoneal injection of adriamycin hydrochloride 2.5 mg/kg once a week for 6 consecutive weeks, establishing a model of heart failure); heart failure+sacubitril/valsartan group (treatment group, n=12, intragastric administration with sacubitril/valsartan 1 week before the first injection of adriamycin, at a dose of 60 mg·kg-1·d-1 for 7 weeks); heart failure+sacubitril/valsartan+APJ antagonist F13A group (F13A group, n=12, adriamycin and sacubitril/valsartan, intraperitoneal injection of 100 μg·kg-1·d-1 APJ antagonist F13A for 7 weeks) and control group (n=10, intraperitoneal injection of equal volume of normal saline). One week after the last injection of adriamycin or saline, transthoracic echocardiography was performed to detect the cardiac structure and function, and then the rats were executed, blood and left ventricular specimens were obtained for further analysis. Hematoxylin-eosin staining and Masson trichrome staining were performed to analyze the left ventricular pathological change and myocardial fibrosis. TUNEL staining was performed to detect cardiomyocyte apoptosis. mRNA expression of left ventricular myocardial apelin and APJ was detected by RT-qRCR. ELISA was performed to detect plasma apelin-12 concentration. The protein expression of left ventricular myocardial apelin and APJ was detected by Western blot. Results: Seven rats survived in the heart failure group, 10 in the treatment group, and 8 in the F13A group. Echocardiography showed that the left ventricular end-diastolic diameter (LVEDD) and the left ventricular end-systolic diameter (LVESD) were higher (both P<0.05), while the left ventricular ejection fraction (LVEF) and left ventricular fractional shortening (LVFS) were lower in the heart failure group than in the control group (both P<0.05). Compared with the heart failure group, rats in the treatment group were featured with lower LVEDD and LVESD (both P<0.05), higher LVEF and LVFS (both P<0.05), these beneficial effects were reversed in rats assigned to F13A group (all P<0.05 vs. treatment group). The results of HE staining showed that the cardiomyocytes of rats in the control group were arranged neatly and densely structured, the cardiomyocytes in the heart failure group were arranged in disorder, distorted and the gap between cells was increased, the cardiomyocytes in the treatment group were slightly neat and dense, and cardiomyocytes in the F13A group were featured similarly as the heart failure group. Masson staining showed that there were small amount of collagen fibers in the left ventricular myocardial interstitium of the control group, while left ventricular myocardial fibrosis was significantly increased, and collagen volume fraction (CVF) was significantly higher in the heart failure group than that of the control group (P<0.05). Compared with the heart failure group, the left ventricular myocardial fibrosis and the CVF were reduced in the treatment group (both P<0.05), these effects were reversed in the F13A group (all P<0.05 vs. treatment group). TUNEL staining showed that the apoptosis index (AI) of cardiomyocytes in rats was higher in the heart failure group compared with the control group (P<0.05), which was reduced in the treatment group (P<0.05 vs. heart failure group), this effect again was reversed in the F13A group (P<0.05 vs. treatment group). The results of RT-qPCR and Western blot showed that the mRNA and protein levels of apelin and APJ in left ventricular myocardial tissue of rats were downregulated in heart failure group (all P<0.05) compared with the control group. Compared with the heart failure group, the mRNA and protein levels of apelin and APJ were upregulated in the treatment group (all P<0.05), these effects were reversed in the F13A group (all P<0.05 vs. treatment group). ELISA test showed that the plasma apelin concentration of rats was lower in the heart failure group compared with the control group (P<0.05); compared with the heart failure group, the plasma apelin concentration of rats was higher in the treatment group (P<0.05), this effect was reversed in the F13A group (P<0.05 vs. treatment group). Conclusion: Sacubitril/valsartan can partially reverse left ventricular remodeling and improve cardiac function in rats with heart failure through modulating Apelin/APJ pathways.目的： 探讨沙库巴曲缬沙坦对心力衰竭（心衰）大鼠左心室重构和心功能的影响及其机制。 方法： SPF级雄性Wistar大鼠46只，体重300~350 g，于动物实验室适应性饲养7 d。然后，分为4组，即心衰组（n=12，腹腔注射盐酸阿霉素2.5 mg/kg，每周1次，连续6周，建立心衰模型）、心衰+沙库巴曲缬沙坦组（治疗组，n=12，阿霉素给药方法同心衰组，在首次注射阿霉素前1周开始给予沙库巴曲缬沙坦治疗，按照60 mg·kg-1·d-1剂量灌胃，持续7周）、心衰+沙库巴曲缬沙坦+F13A组［F13A组，n=12，阿霉素及沙库巴曲缬沙坦用药方法同治疗组，在给予沙库巴曲缬沙坦同时，腹腔注射100 μg·kg-1·d-1 血管紧张素Ⅱ1型受体相关蛋白（APJ）拮抗剂F13A，持续7周］及对照组（n=10，用相同容量的生理盐水代替阿霉素，给药方法同心衰组）。末次注射阿霉素1周后，采用超声心动图检测大鼠心脏结构及功能。超声检测完成后处死大鼠，留取血液及左心室标本。HE染色观察大鼠左心室心肌组织形态，Masson染色观察大鼠左心室心肌纤维化情况，TUNEL染色法检测大鼠左心室心肌细胞凋亡情况，实时荧光定量逆转录聚合酶链反应（RT-qRCR）检测大鼠左心室心肌Apelin和APJ的mRNA表达，酶联免疫吸附（ELISA）法检测大鼠血浆Apelin-12浓度，Western blot法检测大鼠左心室心肌Apelin和APJ的蛋白表达水平。 结果： 干预完成后心衰组、治疗组、F13A组和对照组大鼠分别存活7、10、8和10只，进入下一步实验。超声心动图结果示，与对照组比较，心衰组大鼠左心室舒张末期内径（LVEDD）和左心室收缩末期内径（LVESD）均较高（P均<0.05），而左心室射血分数（LVEF）和左心室短轴缩短率（LVFS）均较低（P均<0.05）。与心衰组比较，治疗组大鼠LVEDD和LVESD均较低（P均<0.05），而LVEF和LVFS均较高（P均<0.05）。与治疗组比较，F13A组大鼠LVESD较高（P<0.05），LVEF和LVFS较低（P均<0.05）。HE染色结果示，对照组大鼠心肌细胞排列整齐、结构致密，心衰组大鼠心肌细胞排列紊乱、扭曲且细胞间隙增加，治疗组大鼠心肌细胞排列稍整齐、致密，F13A组则与心衰组类似。Masson染色结果示，对照组大鼠左心室心肌间质仅有少量胶原纤维，而心衰组胶原纤维较多，胶原容积分数（CVF）大于对照组（P<0.05）。与心衰组比较，治疗组大鼠左心室心肌纤维化程度较轻，CVF较小（P<0.05），而F13A组大鼠的CVF则大于治疗组（P<0.05）。TUNEL检测结果示，与对照组比较，心衰组大鼠心肌细胞凋亡指数较高（P<0.05）。与心衰组比较，治疗组大鼠心肌细胞凋亡指数较低（P<0.05）。与治疗组比较，F13A组大鼠心肌细胞凋亡指数较高（P<0.05）。RT-qPCR和Western blot法检测结果示，与对照组比较，心衰组大鼠左心室心肌组织中Apelin和APJ的mRNA和蛋白水平均较低（P均<0.05）；与心衰组比较，治疗组大鼠左心室心肌组织中Apelin和APJ的mRNA和蛋白水平均较高（P均<0.05）；与治疗组比较，F13A组大鼠左心室心肌组织中APJ mRNA和蛋白以及Apelin蛋白水平均较低（P均<0.05）。ELISA检测结果示，与对照组比较，心衰组大鼠血浆Apelin浓度较低（P<0.05）；与心衰组比较，治疗组大鼠血浆Apelin浓度较高（P<0.05）；与治疗组比较，F13A组大鼠血浆Apelin浓度较低（P<0.05）。 结论： 沙库巴曲缬沙坦可部分逆转心衰大鼠左心室重构、改善心功能，而这一作用与Apelin/APJ通路上调有关。.