博来霉素诱导的肺纤维化小鼠肺组织铁死亡相关基因表达谱分析  

Expression profile of genes related to ferroptosis in a murine model of pulmonary fibrosis induced by bleomycin

作　　者：沈忱悠 聂晓伟 卫栋[3] 杨旭生 江成 王伟[1] 盛雅婷 李桂荣[1] 陈静瑜[3] Shen Chen-You;Nie Xiao-Wei;Wei Dong;Yang Xu-Sheng;Jiang Cheng;Wang Wei;Sheng Ya-Ting;Li Gui-Rong;Chen Jing-Yu(Laboratory of Human Organ Transplantation,Wuxi People's Hospital of Nanjing Medical University,Wuxi,Jiangsu 214023,China;Shenzhen Institute of Respiratory Diseases,Shenzhen People's Hospital,Shenzhen,Guangdong 518020,China;Transplant Center,Wuxi People's Hospital of Nanjing Medical University,Wuxi,Jiangsu 214023,China)

机构地区：[1]南京医科大学附属无锡市人民医院移植实验室,江苏无锡214023 [2]深圳市人民医院呼吸疾病研究所,广东深圳518020 [3]南京医科大学附属无锡市人民医院移植中心,江苏无锡214023

出　　处：《解放军医学杂志》2023年第10期1135-1143,共9页Medical Journal of Chinese People's Liberation Army

基　　金：国家自然科学基金(82070059);江苏省自然科学基金(BK20190150)。

摘　　要：目的分析博来霉素诱导的肺纤维化模型小鼠肺组织铁死亡相关基因的表达谱并探讨其可能的作用机制。方法取12只6~7周龄雄性C57BL/6小鼠,随机分为模型组与对照组,每组6只,分别构建肺纤维化模型及进行对照处理。3周后处死小鼠,采用HE与Masson染色评估其肺部病理改变和胶原纤维堆积状况,普鲁士蓝染色观察肺组织铁聚集水平。应用小鼠铁死亡通路PCR Array筛选差异表达基因,采用生物信息学分析方法对差异基因进行GO功能富集与KEGG信号通路预测。运用实时定量PCR验证模型组与对照组肺组织铁死亡相关差异基因的表达水平。结果与对照组比较,模型组小鼠肺组织出现纤维化改变及明显的铁沉积。PCR Array检测结果显示,与对照组比较,模型组碳酸酐酶9(CA9)、半胱氨酰-tRNA合成酶1(CARS1)、热休克转录因子(HSF1)、NADPH氧化酶3(NOX3)、线粒体铁蛋白(FTMT)等5个铁死亡相关基因的表达量明显降低(P<0.05)。GO功能富集分析与KEGG信号通路预测结果显示,肺纤维化模型小鼠肺组织的差异表达基因主要与体温内稳态、NADPH氧化酶复合体、碳酸脱氢酶活性等功能有关,并在氮素代谢、氨酰合成、军团菌病等通路中富集。实时定量PCR验证结果显示,模型组小鼠肺组织中CA9、CARS1、HSF1及FTMT的表达量均明显低于对照组(P<0.05)。结论博来霉素诱导的肺纤维化模型小鼠肺组织中多个铁死亡相关基因的表达水平发生明显变化,提示铁死亡在特发性肺纤维化中发挥重要作用,其中的差异基因或可为临床治疗提供潜在靶点。Objective To study the expression profile and possible roles of ferroptosis-related genes in a bleomycin-induced murine model of pulmonary fibrosis.Methods Twelve 6-7-week-old male C57BL/6 mice were randomly divided into model group and control group,6 in each group.The model group received nasal inhalation of bleomycin,while the control group was given an equal volume of normal saline.Lung tissues were collected 3 weeks after modeling.Pathological changes and collagen deposition in lungs were observed by HE and Masson staining.Prussian blue staining was used to observe the level of iron accumulation in lung tissue.Total RNA was extracted for PCR Array to identify ferroptosis-related differentially expressed genes(DEGs).Functional analysis for DEGs was performed by Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)pathway analysis.Expression levels of DEGs were verified by RT-qPCR analysis.Results Compared with control group,fibrosis and obvious iron deposition occured in mouse lung tissue of model group.The PCR array identified five ferroptosis-related genes that expression significantly decreased in model group compared with control group including carbonic anhydrase 9(CA9),cysteinyl-tRNA synthetase 1(CARS1),heat shock transcription factor 1(HSF1),NADPH oxidase 3(NOX3)and mitochondrial ferritin(FTMT)(P<0.05).GO functional enrichment analysis and KEGG pathway analysis showed that the DEGs were mainly related to temperature homeostasis,NADPH oxidase complex,and carbonate dehydratase activity,and were involved in nitrogen metabolism,aminoacyl-tRNA biosynthesis and legionellosis signaling pathways.RT-qPCR verification confirmed that the expression levels of CA9,CARS1,HSF1 and FTMT were significantly decreased in model group than those in control group(P<0.05).Conclusions Change of the expression of genes related to ferroptosis has been confirmed in a murine model of pulmonary fibrosis induced by bleomycin.The result indicates that ferroptosis is involved in the process of idiopathic pulmonary fibros

关 键 词：肺纤维化 铁死亡 PCR列阵 生物信息学 

分 类 号：R563[医药卫生—呼吸系统]