lncRNA GAS5调控miR-452-5p/Mcl-1通路对急性肺损伤大鼠肺组织细胞焦亡的影响  

Effect of lncRNA GAS5 on Lung Tissue Cell Pyroptosis in Rats with Acute Lung Injury via Regulation of the miR-452-5p/Mcl-1 Pathway

作　　者：柯有力 罗东 刘旭[2] 别磊 Ke Youli;Luo Dong;Liu Xu(Department of Thoracic Surgery,Affiliated Integrated Traditional Chinese and Western Medical Hospital of Tongji Medical College,Huazhong University of Science and Technology,Wuhan 430033,China;Department of Respiratory and Critical Care Medicine,Affiliated Integrated Traditional Chinese and Western Medical Hospital of Tongji Medical College,Huazhong University of Science and Technology,Wuhan 430033,China)

机构地区：[1]华中科技大学同济医学院附属武汉中西医结合医院,武汉市第一医院胸外科,武汉430033 [2]华中科技大学同济医学院附属武汉中西医结合医院,武汉市第一医院呼吸及危重症医学科,武汉430033

出　　处：《华中科技大学学报（医学版）》2024年第5期621-628,共8页Acta Medicinae Universitatis Scientiae et Technologiae Huazhong

基　　金：武汉市医学科研西医药类面上项目(No.WX21D24)。

摘　　要：目的探讨长链非编码RNA生长抑制特异因子5(lncRNA GAS5)调控miR-452-5p/髓样细胞白血病序列-1(Mcl-1)通路对急性肺损伤(ALI)大鼠肺组织细胞焦亡的影响。方法取SD大鼠随机分成6组(每组10只):对照组、模型组、pUC57-lncRNA GAS5组、miR-452-5p抑制剂组、pUC57-NC+miR-452-5p-NC组、pUC57-lncRNA GAS5+miR-452-5p激活剂组。模型组和各干预组大鼠以腹腔注射脂多糖的方法构建ALI模型,对照组大鼠腹腔注射等量生理盐水,造模同时进行分组干预。然后检测大鼠肺功能,比较各组用力肺活量(FVC)、0.3秒用力呼气容积(FEV0.3)/FVC、潮气量(VT);HE染色观察大鼠肺组织病理损伤,并以Holfbauer评分评估其损伤程度;酶联免疫吸附法(ELISA)测定大鼠肺泡灌洗液(BALF)和血清炎症因子白细胞介素(IL)-18、IL-1β水平;免疫印迹法检测大鼠肺组织焦亡相关指标[消皮素D(GSDMD)、核苷酸结合寡聚化结构域样受体蛋白3(NLRP3)、天冬氨酸特异性半胱氨酸蛋白酶(Caspase)-1、Caspase-11]及Mcl-1蛋白表达;实时荧光定量PCR检测大鼠肺组织lncRNA GAS5、miR-452-5p及Mcl-1表达;双荧光素酶报告基因实验验证大鼠肺泡上皮细胞L2中lncRNA GAS5对miR-452-5p的靶向调节。结果与对照组比较,模型组大鼠FVC、FEV0.3/FVC、lncRNA GAS5表达、Mcl-1蛋白及mRNA表达降低(均P<0.05),VT、Holfbauer评分、IL-18及IL-1β水平、GSDMD、NLRP3、Caspase-1及Caspase-11蛋白表达、miR-452-5p表达升高(均P<0.05)。与模型组比较,pUC57-lncRNA GAS5组、miR-452-5p抑制剂组大鼠FVC、FEV0.3/FVC、Mcl-1蛋白及mRNA表达均升高(均P<0.05),VT、Holfbauer评分、IL-18及IL-1β水平、GSDMD、NLRP3、Caspase-1及Caspase-11蛋白表达、miR-452-5p表达均降低(均P<0.05);pUC57-NC+miR-452-5p-NC组大鼠各指标无显著差异(均P>0.05)。与pUC57-lncRNA GAS5组比较,pUC57-lncRNA GAS5+miR-452-5p激活剂组大鼠FVC、FEV0.3/FVC、Mcl-1蛋白及mRNA表达降低(均P<0.05),VT、Holfbauer评分、IL-18及IL-1β水平�Objective To investigate the effect of long non-coding RNA growth arrest-specific transcript 5(lncRNA GAS5)on lung tissue cell apoptosis in acute lung injury(ALI)rats via regulation of the miR-452-5p/myeloid cell leukemia sequence 1(Mcl-1)pathway.Methods SD rats were randomly separated into 6 groups(10 in each group):control group,model group,pUC57 lncRNA GAS5 group,miR-452-5p antagonist group,pUC57 NC+miR-452-5p NC group,and pUC57 lncRNA GAS5+miR-452-5p agonist group.The ALI model was constructed via the intraperitoneal injection of lipopolysaccharide in the model group and drug intervention group,whereas the control group rats were intraperitoneally injected with an equal dose of physiological saline.At the same time,the intervention was performed,and the lung function of the rats was subsequently measured.The forced vital capacity(FVC),forced expiratory volume in 0.3 second(FEV0.3)/FVC,and tidal volume(VT)were compared among the groups.HE staining was used to observe pathological damage in the lung tissue,and the degree of damage was evaluated via the Holfbauer score.An enzyme-linked immunosorbent assay(ELISA)was used to measure the levels of bronchoalveolar lavage fluid(BALF)and the serum inflammatory factors interleukin-18 and IL-1βin the rats.Immunoblotting was used to detect the expression of pyroptosis-related indicator proteins[(GSDMD),nucleotide binding oligomeric domain-like receptor protein 3(NLRP3),cysteine-containing aspartate-specific protease(Caspase-1),and Caspase-11]and Mcl-1 in rat lung tissue.Real-time fluorescence quantitative PCR was used to detect the expression of lncRNA GAS5 and miR-452-5p in rat lung tissue.A dual-luciferase reporter gene experiment was performed to validate the targeted regulation of miR-452-5p by lncRNA GAS5 in rat alveolar epithelial cell L2.Results Compared with those in the control group,the FVC,FEV0.3/FVC,expression of lncRNA GAS5,expression of Mcl-1 protein and mRNA in the model group decreased(P<0.05),and the VT,Holfbauer score,IL-18 and IL-1βlevels,GSDMD,NLRP3

关 键 词：lncRNA GAS5 miR-452-5p/Mcl-1通路 急性肺损伤 肺组织细胞焦亡 

分 类 号：R563[医药卫生—呼吸系统]