肺心汤通过调控AMPK/mTOR信号通路抑制自噬对低氧性肺动脉高压大鼠的保护作用  

Protective effects of Feixin Decoction on rats with hypoxic pulmonary hypertension by inhibiting autophagy through regulating AMPK/mTOR signaling pathway

作　　者：张超 易健 丁蓉珍[1,2] 万佳婧 谭骏岚 王飞英 周灵灵 宋岚 戴爱国 ZHANG Chao;YI Jian;DING Rongzhen;WAN Jiajing;TAN Junlan;WANG Feiying;ZHOU Lingling;SONG Lan;DAI Aiguo(Respiratory Disease Laboratory,School of Medicine,Hunan University of Chinese Medicine,Changsha,Hunan 410208,China;Key Laboratory of Vascular Biology and Translational Medicine,Education Department of Hunan Province,Changsha,Hunan 410208,China;Medical Innovation Experiment Center,The First Affiliated Hospital of Hunan University of Chinese Medicine,Changsha,Hunan 410021,China;Department of Respiratory Medicine,The First Affiliated Hospital of Hunan University of Chinese Medicine,Changsha,Hunan 410021,China;School of Medicine,Hunan University of Chinese Medicine,Changsha,Hunan 410208,China)

机构地区：[1]湖南中医药大学医学院呼吸疾病研究室,湖南长沙410208 [2]血管生物学与转化医学湖南省重点实验室/湖南省高校重点实验室,湖南长沙410208 [3]湖南中医药大学第一附属医院医学创新实验中心,湖南长沙410007 [4]湖南中医药大学第一附属医院呼吸内科,湖南长沙410007 [5]湖南中医药大学医学院,湖南长沙410208

出　　处：《湖南中医药大学学报》2024年第5期744-753,共10页Journal of Hunan University of Chinese Medicine

基　　金：国家自然科学基金面上项目(82370069,82200066);国家自然科学基金青年项目(82305214);湖南省自然科学基金项目(2021JJ30017,2022JJ40308);中国博士后科学基金面上项目(2021M690982);湖南省中医药科研计划重点项目(C2022001);湖南省研究生科研创新项目(2022CX196)。

摘　　要：目的探究肺心汤通过调控腺苷酸活化蛋白激酶(adenosine monophosphate-activated protein kinase,AMPK)/哺乳动物雷帕霉素靶蛋白(mammalian target of rapamycin,mTOR)信号通路对低氧性肺动脉高压(hypoxic pulmonary hypertension,HPH)大鼠的治疗作用及机制。方法36只SD雄性大鼠随机分为正常组、模型组、西地那非组以及肺心汤低、中、高剂量组。除正常组外,将其余5组置于氧气浓度为10.0%±0.5%的低氧舱内造模28 d,每天8 h,造模同时灌胃给药。4周后检测大鼠心肺血流动力学指标[右心室收缩压(right ventricular systolic pressure,RVSP),右心室肥厚指数(right ventricular hypertrophy index,RVHI),肺动脉加速时间/肺动脉射血时间(pulmonary acceleration time/pulmonary ejection time,PAT/PET)、三尖瓣环收缩期位移(tricuspid annulus plane systolic excursion,TAPSE)、右心室前壁厚度(right ventricular anterior wall thickness,RVAWT)];HE染色法检测肺动脉重塑;免疫荧光法检测α平滑肌肌动蛋白(α-smooth muscle actin,α-SMA)和增殖细胞核抗原(proliferating cell nuclear antigen,PCNA)蛋白共定位表达;透射电镜观察自噬溶酶体的数量;免疫组织化学法检测肺组织中p-AMPK、p-mTOR、微管相关蛋白1轻链3B(microtubule-associated protein light chain 3B,LC3B)、Beclin1和p62蛋白阳性表达。结果与正常组相比,模型组大鼠RVSP、RVHI及RVAWT显著升高(P<0.01),PAT/PET和TAPSE显著降低(P<0.01);肺小动脉壁明显增厚,管腔狭窄,肺小动脉管壁厚度占血管直径的百分比(the percentage of wall thickness of pulmonary arterioles to vascular diameter,WT%)显著升高(P<0.01);α-SMA和PCNA共定位表达显著增加;自噬溶酶体数量增加;肺组织中p-AMPK、LC3B和Beclin1蛋白阳性表达上调(P<0.01),p-mTOR及p62蛋白阳性表达下调(P<0.01)。与模型组相比,各给药组大鼠RVSP、RVHI及RVAWT显著下降(P<0.01),PAT/PET、TAPSE显著升高(P<0.05,P<0.01);肺小动脉壁增厚程度减轻,WT%显著降低(P<0.0Objective To investigate the therapeutic efficacy and mechanism of Feixin Decoction(FXD)on hypoxic pulmonary hypertension(HPH)rats through regulating adenosine monophosphate-activated protein kinase(AMPK)/mammalian target of rapamycin(mTOR)signaling pathway.Methods Thirty-six SD rats were randomized into control group,model group,sildenafil group and low-,medium-,high-dose FXD groups.Except control group,the remaining five groups were placed in a hypoxic chamber with an oxygen concentration of 10%±0.5%for modeling,8 hours per day for 28 d,with simultaneous intragastric administration.After 4 weeks,the rat cardiopulmonary hemodynamic indexes[right ventricular systolic pressure(RVSP),right ventricular hypertrophy index(RVHI),the ratio of pulmonary acceleration time to pulmonary ejection time(PAT/PET),tricuspid annulus plane systolic excursion(TAPSE),right ventricular anterior wall thickness(RVAWT)]were determined.Pulmonary artery remodeling was checked by HE staining;the co-localization ofα-smooth muscle actin(α-SMA)and proliferating cell nuclear antigen(PCNA)was examined by immunofluorescence assay;the number of autolysosomes was observed by transmission electron microscope(TEM);the positive expressions of p-AMPK,p-mTOR,microtubule-associated protein 1 light chain 3B(LC3B),Beclin1,and p62 in the lung tissue were measured by immunohistochemical assay.Results Compared with the control group,in the model group,RVSP,RVHI,and RVAWT were significantly increased(P<0.01),while PAT/PET and TAPSE were significantly decreased(P<0.01);the wall of the pulmonary arterioles was markedly thickened and the lumina narrowed,with a significant increase in the percentage of wall thickness of pulmonary arterioles to vascular diameter,WT%(P<0.01);the co-localization ofα-SMA and PCNA was significantly enhanced(P<0.01);the number of autolysosomes was increased;the protein positive expressions of p-AMPK,LC3B,and Beclin1 in the lung tissue were up-regulated(P<0.01),while those of p-mTOR and p62 were down-regulated(P<0.01).Compared with

关 键 词：肺心汤 低氧性肺动脉高压 腺苷酸活化蛋白激酶 哺乳动物雷帕霉素靶蛋白 自噬 

分 类 号：R285.5[医药卫生—中药学]