基于PERK/eIF2α/ATF4/CHOP诱导的肺泡上皮细胞凋亡探讨人参皂苷Rg1减轻脓毒症急性肺损伤的作用和机制  

Role and mechanism of ginsenoside Rg1 in ameliorating sepsis-induced acute lung injury based on PERK/eIF2α/ATF4/CHOP-induced alveolar epithelial cell apoptosis

作　　者：仲凯强 黄荫桂 陈秀萍 陈瑞[1,3] 邬楚雯 邹佳桢 奚小土 李俊 闫春江 ZHONG Kai-qiang;HUANG Yin-gui;CHEN Xiu-ping;CHEN Rui;WU Chu-wen;ZOU Jia-zhen;XI Xiao-tu;LI Jun;YAN Chun-jiang(Second Clinical College of Guangzhou University of Chinese Medicine;the Second Affiliated Hospital of Guangzhou University of Chinese Medicine;Guangdong Provincial Key Laboratory of Research on Emergency in Traditional Chinese Medicine;School of Life Sciences and Biopharmaceutics,Guangdong Pharmaceutical University;the First Affiliated Hospital of Guangzhou University of Chinese Medicine)

机构地区：[1]广州中医药大学第二临床医学院,广东广州510405 [2]广州中医药大学第二附属医院,广东广州510120 [3]广东药科大学生命科学与生物制药学院,广东广州510006 [4]广东省中医急诊研究重点实验室,广东广州510006 [5]广州中医药大学第一附属医院,广东广州510405

出　　处：《中国中药杂志》2024年第14期3837-3847,共11页China Journal of Chinese Materia Medica

基　　金：国家自然科学基金青年科学基金项目(82204936);广东省中医药管理局专项(20213007);广东省基础与应用基础研究项目(2023A04J0476);广东省中医急症研究重点实验室课题(YN2023JZ18);广东省中医院1010专项(YN10101908);广东省中医院院内专项(YN2023MS42)。

摘　　要：探讨人参皂苷Rg1(GRg1)对脓毒症急性肺损伤(SALI)的干预疗效与作用机制。采用盲肠结扎穿孔术(CLP)建立SALI小鼠模型,随机分组给予GRg1干预,记录存活与体质量变化情况,小鼠无创肺功能检测系统检测肺功能,苏木精-伊红(HE)染色观察肺组织损伤情况,酶联免疫吸附实验(ELISA)、实时荧光定量PCR(qRT-PCR)检测炎症因子含量与表达,流式细胞术、TUNEL染色等检测凋亡情况,蛋白质印迹(Western blot)、qRT-PCR检测凋亡相关分子胱天蛋白酶3(caspase-3)、B淋巴细胞瘤-2(Bcl-2)、Bcl-2相关X蛋白(Bax)与内质网应激相关分子蛋白激酶R样内质网激酶(PERK)、真核翻译起始因子2α激酶(eIF2α)、活化转录因子4(ATF4)、C/EBP同源蛋白(CHOP)的活化表达情况。此外,利用脂多糖(LPS)体外诱导小鼠肺泡上皮细胞损伤模型,给予内质网应激激动剂衣霉素(TUN)验证GRg1的作用机制。结果显示,与模型组相比,GRg1干预可提高CLP小鼠生存时间,减缓其体质量下降,改善其受损的肺功能指标。同时,与模型组相比,GRg1给药组小鼠肺组织病理损伤评分降低,肺组织湿干比和肺泡灌洗液蛋白含量降低,血清白细胞介素-6(IL-6)、白细胞介素-1β(IL-1β)、肿瘤坏死因子-α(TNF-α)水平及肺组织中上述细胞因子的mRNA表达下降,肺泡上皮细胞凋亡比例明显下降,caspase-3、Bax表达下调,Bcl-2表达上调,PERK、eIF2α、ATF4、CHOP活化及表达下调。体外结果显示联合TUN后,GRg1降低凋亡细胞比例、下调凋亡相关蛋白表达的作用被抑制。综上,GRg1可减少肺泡上皮细胞凋亡,抑制肺部炎症渗出,减轻肺损伤,改善肺功能,可能与PERK/eIF2α/ATF4/CHOP通路相关。The study investigates the therapeutic effects and mechanisms of ginsenoside Rg_1(GRg_1)on sepsis-induced acute lung injury(SALI).A murine model of SALI was created using cecal ligation and puncture(CLP)surgery,and mice were randomly assigned to groups for GRg_1 intervention.Survival and body weight changes were recorded,lung function was assessed with a non-invasive lung function test system,and lung tissue damage was evaluated through HE staining.The content and expression of inflammatory factors were measured by ELISA and qRT-PCR.Apoptosis was examined using flow cytometry and TUNEL staining.The activation and expression of apoptosis-related molecules cysteinyl aspartate specific proteinase 3(caspase-3),B-cell lymphoma-2(Bcl-2),Bcl-2 associated X protein(Bax),and endoplasmic reticulum stress-related molecules protein kinase R-like endoplasmic reticulum kinase(PERK),eukaryotic initiation factor 2α(eIF2α),activating transcription factor 4(ATF4),and C/EBP homologous protein(CHOP)were studied using Western blot and qRT-PCR.In addition,an in vitro model of lipopolysaccharide(LPS)-induced lung alveolar epithelial cell injury was used,with the application of the endoplasmic reticulum stress inducer tunicamycin to validate the action mechanism of GRg_1.Results indicated that,when compared to the model group,GRg_1 intervention significantly enhanced the survival time of CLP mice,mitigated body weight loss,and improved impaired lung function indices.The GRg_1-treated mice also displayed reduced lung tissue pathological scores,a reduced lung tissue wet-to-dry weight ratio,and lower protein content in the bronchoalveolar lavage fluid.Serum levels of interleukin-6(IL-6),interleukin-1β(IL-1β),and tumor necrosis factor-α(TNF-α),as well as the mRNA expressions of these cytokines in lung tissues,were decreased.There was a notable decrease in the proportion of apopto-tic alveolar epithelial cells,and down-regulated expressions of caspase-3,Bax,PERK,eIF2α,ATF4,and CHOP and up-regulated expression of Bcl-2 were observed.In

关 键 词：脓毒症 人参皂苷RG1 急性肺损伤 内质网应激 细胞凋亡 

分 类 号：R285.5[医药卫生—中药学]