自噬在矽肺模型大鼠肺部炎症反应中的作用  

The role of autophagy in pulmonary inflammatory response in silicosis model rats

作　　者：张玉[1] 耿晓[1] 赛林霖[1] 杨玉婷 薄存香[1] ZHANG Yu;GENG Xiao;SAI Lin-lin;YANG Yu-ting;BO Cun-xiang(Shandong Academy of Occupational Health and Occupational Medicine,Shandong First Medical University&Shandong Academy of Medical Science,Jinan Shandong 250014 China)

机构地区：[1]山东省职业卫生与职业病防治研究院山东第一医科大学/山东省医学科学院,山东济南250014

出　　处：《中国职业医学》2022年第2期153-157,164,共6页China Occupational Medicine

基　　金：中国煤矿尘肺病防治基金会项目(201915J039);山东省医药卫生科技发展计划项目(2017ws438);山东省自然科学基金(ZR2019PH116,ZR2017YL001);山东省重点研发项目(2018GSF118212)。

摘　　要：目的研究自噬在矽肺大鼠肺部炎症反应中的作用。方法选择无特定病原体级健康雄性Wistar大鼠随机分为4组,每组10只。采用非暴露式气管滴注法,二氧化硅(SiO_(2))组、SiO_(2)+3-甲基腺嘌呤(3-MA)组、SiO_(2)+雷帕霉素(RAPA)组大鼠均一次性予质量浓度为0.05 mg/L的SiO_(2)悬浮液(1.0 mL/只)建立矽肺染尘大鼠模型,对照组大鼠予等体积的0.9%氯化钠溶液。于SiO_(2)染毒前2 d,SiO_(2)+3-MA组大鼠予腹腔注射剂量为2.5 mg/kg体质量的3-MA,SiO_(2)+RAPA组大鼠予腹腔注射剂量为1.0 mg/kg体质量的RAPA,每天1次;矽肺造模后隔1 d腹腔注射1次,共注射10次。采用苏木精-伊红和Masson染色法观察各组大鼠肺组织病理变化;采用酶联免疫吸附测定法检测各组大鼠肺组织中转化生长因子-β(TGF-β)、白细胞介素-1β(IL-1β)和肿瘤坏死因子-α(TNF-α)相对表达水平;采用蛋白质印迹法检测各组大鼠肺组织中微管相关蛋白1轻链3(LC3)和酵母自噬相关基因6(Beclin1)相对表达水平。结果肺组织病理观察结果显示,SiO_(2)组大鼠肺组织中可见散在炎性结节,间质中可见散在炎性细胞;SiO_(2)+3-MA组大鼠肺组织中炎症较严重且肺泡腔有黏性分泌物;SiO_(2)+RAPA组大鼠肺组织炎症较SiO_(2)组减轻且炎性结节变小。与对照组比较,SiO_(2)组大鼠肺组织中TNF-α、IL-1β、TGF-β、Beclin1蛋白相对表达水平和LC3Ⅱ/Ⅰ比值均升高(P值均<0.05)。与SiO_(2)组比较,SiO_(2)+3-MA组大鼠肺组织中TNF-α和TGF-β相对表达水平均升高(P值均<0.05),LC3Ⅱ/Ⅰ比值和Beclin1蛋白相对表达水平均下降(P值均<0.05);SiO_(2)+RAPA组大鼠肺组织中TNF-α、IL-1β和TGF-β相对表达水平均下降(P值均<0.05),LC3Ⅱ/Ⅰ比值和Beclin1蛋白相对表达水平均升高(P值均<0.05)。与SiO_(2)+3-MA组比较,SiO_(2)+RAPA组大鼠肺组织中TNF-α、IL-1β和TGF-β相对表达水平均下降(P值均<0.05),LC3Ⅱ/Ⅰ比值和Beclin1蛋白相对表达水平均升高(P�ObjectiveTo evaluate the role of autophagy in inflammatory response in the lungs of silicosis rats.MethodsThespecific pathogen-free healthy male Wistar rats were randomly divided into four groups with 10 rats in each group.The rats in thecontrol group were given an equal volume of 0.9%sodium chloride solution;all rats in the silicon dioxide(SiO_(2))group,the SiO_(2)+3-methyladenine(3-MA)group,and the SiO_(2)+rapamycin(RAPA)group were given a mass concentration of 0.05 mg/L of SiO_(2) suspension(1.0 mL/rat)to establish a rat model of silicosis using non-exposed tracheal instillation method.Two days beforeSiO_(2) exposure,the rats in SiO_(2)+3-MA group were intraperitoneally injected with 3-MA at a dose of 2.5 mg/kg body weight.The ratsin the SiO_(2)+RAPA group were intraperitoneally injected with a dose of RAPA 1.0 mg/kg body weight,once a day,and once everyother day after modeling,for a total of 10 injections.The pathological changes in the lung tissue of the rats in each group wereevaluated using hematoxylin-eosin and Masson staining.The relative expression levels of transforming growth factor-β(TGF-β),interleukin-1β(IL-1β)and tumor necrosis factor-α(TNF-α)in the lung tissue of the rats in each group were detected usingenzyme-linked immunosorbent assay.The relative expression levels of microtubule-associated protein 1 light chain 3(LC3)andyeast autophagy-related gene 6(Beclin1)in lung tissue of rats in each group were detected by Western blotting.ResultsThepathological observation of rat lung tissue showed that scattered inflammatory nodules and interstitial inflammatory cells were found in the lung tissue of the rats in the SiO_(2) group.In the SiO_(2)+3-MA group,the inflammation in the lung tissue was moresevere and the alveolar cavity had viscous secretions.The rats in the SiO_(2)+RAPA group had less inflammation and smallerinflammatory nodules than the SiO_(2) group.Compared with the control group,the TNF-α,IL-1β,TGF-β,Beclin1 protein relativeexpression levels and LC3Ⅱ/Ⅰratio in the lung tissue

关 键 词：炎症反应 矽肺 二氧化硅 自噬 雷帕霉素 3-甲基腺嘌呤 大鼠 

分 类 号：R135.2[医药卫生—劳动卫生]