基于生物信息学筛选纳米二氧化硅致肺纤维化关键因子  

Identification the key factor of pulmonary fibrosis following silica nanoparticles exposure based on bioinformatics analysis

作　　者：师凡 杨晓静 熊敏 杨玉姗 张艳淑 金玉兰 Shi Fan;Yang Xiaojing;Xiong Min;Yang Yushan;Zhang Yanshu;Jin Yulan(School of Public Health,North China University of Science and Technology,Tangshan 063210,China;College of Life Sciences,North China University of Science and Technology,Tangshan 063210,China)

机构地区：[1]华北理工大学公共卫生学院,唐山063210 [2]华北理工大学生命科学学院,唐山063210

出　　处：《中华劳动卫生职业病杂志》2023年第7期497-503,共7页Chinese Journal of Industrial Hygiene and Occupational Diseases

基　　金：国家自然科学基金(81202161);华北理工大学研究生创新项目(CXZZBS2023120)。

摘　　要：目的构建巨噬细胞-成纤维细胞体外模型模拟肺纤维化过程,探讨纳米二氧化硅(SiNPs)暴露致肺纤维化的关键机制。方法于2021年1月,取对数生长期的人单核细胞白血病(THP-1)细胞,分别采用0、25、50、100μg/ml SiNPs处理24 h,收集THP-1细胞上清液分别作用于对数生长期的对照组和低、中、高剂量组人胚肺成纤维(MRC-5)细胞,继续培养24 h。检测不同组别MRC-5细胞增殖情况。检测MRC-5细胞上清液中细胞羟脯胺酸(Hyp)、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)表达水平。检测高剂量组MRC-5细胞中的差异蛋白表达情况,应用GeneCard数据库中肺纤维化相关蛋白对高剂量组MRC-5细胞进行差异肺纤维化蛋白的筛选。应用GO分析对高剂量组的差异肺纤维化蛋白进行富集分析,应用String数据库构建高剂量组的差异肺纤维化蛋白的相互作用(PPI)网络,使用CytoHubba软件筛选高剂量组的关键差异肺纤维化蛋白。使用Pearson相关分析计算关键差异肺纤维化蛋白之间的相关系数,Western blotting检测不同组别中关键差异肺纤维化蛋白表达。结果与对照组比较,低、中、高剂量组MRC-5细胞增殖率增加(P<0.05)。与对照组比较,低、中、高剂量组细胞上清液中Hyp、IL-1β的表达水平均升高(P<0.05),高剂量组细胞上清液中IL-6、TNF-α蛋白表达水平均升高(P<0.05)。用GeneCard数据库结合高剂量组差异表达蛋白共筛选出26个差异肺纤维化蛋白,GO分析显示,差异肺纤维化蛋白主要调控细胞外基质水解、细胞炎症反应、组织修复和细胞增殖等生物过程。CytoHubba软件计算出基质金属蛋白酶9(MMP9)和基质金属蛋白酶组织抑制因子1(TIMP1)是PPI网络中的关键节点蛋白。相关分析结果显示,MMP9蛋白与基质金属蛋白酶1(MMP1)、基质金属蛋白酶3(MMP3)、TIMP1和表皮生长因子受体(EGFR)蛋白表达相关(r=0.97、0.98、0.94、0.93,P<0.05)�Objective To investigate the main mechanisms of pulmonary fibrosis following silica nanoparticles(SiNPs)exposure through constructing the macrophage-fibroblast model in vitro,which simulated the process of pulmonary fibrosis.Methods In January 2021,human mononuclear leukemia cells(THP-1)were treated with 0,25,50,100μg/ml SiNPs for 24 h.The supernatant of THP-1 cells was collected and applied to human embryonic lung fibroblast cells(MRC-5)which divided into control and low,medium and high dose groups at the logarithmic growth stage for 24 h.MRC-5 cell viability was detected by CCK8.The hydroxyproline(Hyp),interleukin 6(IL-6),interleukin 1 beta(IL-1β)and tumor necrosis factor-alpha(TNF-α)expression were detected in the supernatants of MRC-5.The changed proteins were detected by liquid-phase mass spectrometry in high dose group.GeneCard database were applied to identity the differential pulmonary fibrosis proteins in high dose group.Gene Ontology(GO)was performed to identity the key biological process in differential pulmonary fibrosis proteins of high dose group.The String database was used to construct the protein-protein interactions(PPI)network of differential pulmonary fibrosis proteins.The APP of CytoHubba was applied to calculate the key protein of differential pulmonary fibrosis proteins in PPI network.Correlation coefficients between key differential pulmonary fibrosis proteins were calculated using Pearson correlation analysis.Western blotting was applied to detect the expression of key proteins of differential pulmonary fibrosis proteins in different groups.Results CCK8 results showed that MRC-5 cell viability was increasing in low,medium and high dose groups compared with control group(P<0.05).The expression levels of Hyp and IL-1βin different group were increased compared with control group,the expression levels of IL-6 and TNF-αwere increased in high dose group compared with control group(P<0.05).GeneCard database identified 26 differential pulmonary fibrosis proteins,which were mainly involved in

关 键 词：肺纤维化 纳米二氧化硅 生物信息学 基质金属蛋白酶9 基质蛋白酶组织抑制因子1 细胞外基质水解 

分 类 号：R563.9[医药卫生—呼吸系统] Q811.4[医药卫生—内科学]