肺纤维化中1,25-(OH)_2D_3对内质网应激及ATG12表达的影响  

Effects of 1,25-(OH)_2D_3 on endoplasmic reticulum stress and ATG12 expression in pulmonary fibrosis

作　　者：刘营[1] 刘乃国[1] 王楠[1] LIU Ying;LIU Nai-guo;WANG Nan(Clinical Medicine Laboratory,Affiliated Hospital of Binzhou Medical University,Binzhou 256603,Shandong,China)

机构地区：[1]山东滨州医学院附属医院临床医学实验室,滨州医学硕士研究生256603

出　　处：《医学研究生学报》2018年第4期391-397,共7页Journal of Medical Postgraduates

基　　金：山东省医药卫生科技发展计划项目(2015WS0490)

摘　　要：目的 1,25-(OH)2D3是否影响肺纤维化中内质网应激及自噬仍不清楚。文章探讨内质网应激及自噬在大鼠肺纤维化发生发展中的作用,研究了1,25-(OH)2D3对内质网应激相关分子及ATG12表达的影响。方法雄性SD大鼠区组随机分为对照组、模型组和治疗组,每组30只。模型组、治疗组经气管灌注博来霉素(5.0 mg/kg),对照组经气管灌注等渗盐水200μL/只。自气管灌注后第2天起,治疗组腹腔注射1,25-(OH)2D32μg/kg,模型组腹腔注射1,25-(OH)2D3溶剂(0.1%乙醇、99.9%丙二醇)200μL/只,对照组腹腔注射等渗盐水200μL/只,各种注射均为隔天1次,直至处死。用实时定量PCR方法检测PERK、ATF4及ATG12的mRNA表达状况,用免疫组化方法检测PERK、ATF4的蛋白质表达水平。结果与对照组14、21、28 d的PERK水平(1.01±0.23、1.05±0.09、1.04±0.08)比较,模型组(2.30±0.19、3.59±0.27、4.63±0.19)和治疗组(1.44±0.34、1.92±0.17、2.52±0.15)表达均增加(P<0.05);与对照组14、21、28 d的ATF4水平(1.04±0.07、1.05±0.08、1.03±0.10)比较,模型组(2.10±0.12、3.91±0.14、6.20±0.28)和治疗组(1.49±0.27、2.52±0.42、4.02±0.31)表达均增加(P<0.05)。模型组和治疗组14、21、28 d组内不同时间两两比较差异均有统计学意义(P<0.05);与对照组相比,14 d模型组、治疗组中ATG12的mRNA表达明显升高(P<0.05),而21、28 d ATG12的mRNA表达均显著降低(P<0.05)。模型组肺组织中PERK、ATF4蛋白质在肺泡上皮细胞、肺间质细胞,尤其是肺巨噬细胞中表达,可见大量棕褐色颗粒,较正常肺组织中两种蛋白质阳性信号明显增强。14、21、28 d模型组和治疗组中PERK、ATF4的蛋白质表达均明显高于对照组(P<0.05),且随建模时间的延长蛋白质的表达递增,14、21、28 d组内两两比较差异具有统计学意义(P<0.05)。结论 1,25-(OH)2D3可能通过抑制PERK-e IF2α-ATF4信号通路来抑制肺纤维化的发生发展。Objective It is not yet clear whether 1,25-(OH)2 D 3 acts on endoplasmic reticulum stress(ERS)and autophagy in pulmonary fibrosis(PF).This study aimed to investigate the roles of ERS and autophagy in the development and progression of pulmonary fibrosis in rats and the effects of 1,25-(OH)2 D 3 on the expressions of ERS-related molecules and autophagy-related gene 12(ATG12).Methods Ninety male SD rats were randomly divided into a control,a PF model and a treatment group of equal number.Bleomycin was injected into the tracheas of the latter two groups of rats to induce PF.On the second day after modeling,the rats of the treatment group were injected intraperitoneally with 1,25-(OH)2 D 3 at 2μg/kg,those of the PF model group with 1,25-(OH)2 D 3 solvent at 200μL,and those of the control group with isotonic saline at 200μL,all once 2 days.Then the mRNA expressions of PERK,ATF4 and ATG12 were measured by real-time PCR and the protein expressions of PERK and ATF4 detected by immunohistochemistry.Results At 14,21 and 28 days after treatment,the expression levels of PERK were significantly higher in the PF model group(2.30±0.19,3.59±0.27,and 4.63±0.19)and treatment group(1.44±0.34,1.92±0.17,and 2.52±0.15)than in the control(1.01±0.23,1.05±0.09,and 1.04±0.08)(P<0.05),and so were the expression levels of ATF4 in the PF models(2.10±0.12,3.91±0.14,and 6.20±0.28)and treated rats(1.49±0.27,2.52±0.42,and 4.02±0.31)than in the controls(1.04±0.07,1.05±0.08,and 1.03±0.10)(P<0.05).Compared with the control group,the PF model and treatment groups showed markedly increased expression levels of ATG12 mRNA at 14 days(P<0.05),but decreased at 21 and 28 days(P<0.05).Both the expressions of PERK and ATF4 proteins were remarkably higher in the model and treatment groups than in the control at 14,21 and 28 days(P<0.05),increasing in a time-dependent manner.Conclusion By suppressing the PERK-eIF2α-ATF4 signaling pathway,1,25-(OH)2 D 3 inhibits the development and progression of pulmonary fibrosis.

关 键 词：内质网应激 肺纤维化 PERK-eIF2α-ATF4通路 ATG12 1 25-(OH)2D3 

分 类 号：R563[医药卫生—呼吸系统]