摘要
背景:靶向铁过载基因调节铁死亡或许是一种较快且有效延缓骨关节炎退变的方法,但目前对骨关节炎铁死亡相关分子机制及基因靶点尚不清楚。目的:通过生物信息学分析铁死亡在骨关节炎中的关键基因和途径,结合体外实验验证骨关节炎中铁死亡标记基因,探讨铁死亡在骨关节炎中的潜在作用。方法:以“Osteoarthritis”为检索词,通过GEO数据库检索2010-01-01/2021-01-01的公开数据,筛选得到基因微阵列数据集GSE55235,对GSE55235数据集进行数据矫正后分析获得差异表达基因;利用FerrDb数据库检索得到铁死亡相关基因与GSE55235数据集差异表达基因作交集,对交集基因作基因本体(Gene Ontology,GO)与京都基因和基因组数据库富集分析(Kyoto Encyclopedia of Genes and Genomes,KEGG)并绘制蛋白质-蛋白质相互作用网络,获得骨关节炎铁死亡HUB基因,筛选出铁死亡标记基因;将正常人软骨细胞设为对照组、人软骨细胞骨关节炎模型设为实验组,采用实时荧光定量PCR对两组铁死亡标记基因的mRNA表达进行验证。结果与结论:①共获得36个骨关节炎铁死亡基因,GO富集分析表明其主要参与氧化应激反应、皮质类固醇反应、类固醇激素反应、对糖皮质激素的反应和活性氧的代谢等过程;②针对产生超氧化物的NAD(P)H氧化酶及氧化还原酶活性、血红素结合、四吡咯结合中发挥作用;③KEGG富集分析表明骨关节炎铁死亡基因主要参与NOD样受体、白细胞介素17、缺氧诱导因子1、肿瘤坏死因子及叉头盒O类(FoxO)等信号通路;④构建蛋白质-蛋白质相互作用网络,进一步分析获得血管内皮生长因子A、白细胞介素6、JUN、PTGS2和DUSP1等5个骨关节炎铁死亡HUB基因,筛选出血管内皮生长因子A、白细胞介素6、PTGS2和DUSP1等4个铁死亡标记基因,体外实验证实其在对照组(正常人软骨细胞)与实验组(细胞骨关节炎模型)中有显著差异(P<0.05);⑤上述数据显示,铁死亡可能通过氧化应激、氧化还原及诱导炎症等作用于骨关节炎,机制上可能与缺氧诱导因子1等信号通路刺激NAD(P)H有关;血管内皮生长因子A、白细胞介素6、PTGS2和DUSP1可作为骨关节炎铁死亡的生物标志物。
BACKGROUND:Regulating ferro ptosis by targeting iron overload genes may be a fast and effective way to delay the degeneration of osteoarthritis.However,the molecular mechanisms and gene targets related to ferroptosis in osteoarthritis are still unclea r.OBJECTIVE:To analyze the key genes and pathways of ferroptosis in osteoarthritis by bioinfo rmatics analysis,verify the marker genes of fe rroptosis in co mbination with in vitro experiments, and explore the potential role of fe rroptosis in osteoarthritis.METHODS: "Osteoarthritis" was used as a keyword.Relevant public data from the GEO database between January 1,2010 and January 1,2021 were retrieved.A gene microarray dataset GSE55235 was yielded and differentially expressed genes were identified after data correction analysis of the GSE55235 data set.Ferro ptosis-related genes retrieved by the FerrDb database were intersected with differentially expressed genes in the GSE55235 dataset,and th intersected genes were used for Gene Ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analysis and the protein-protein intera ction network was mapped.Ferro ptosis-related HUB genes in osteoarthritis were obtained and ferroptosis marker genes were then selected.Human chondrocytes were divided into control group(normal human chondrocytes) and experimental group(cellular osteoarthritic model).The mRNA expression of ferroptosis marker genes in the two groups was detected by quantitative real-time fluorescence PCR.RESULTS AND CONCLUSION:A total of 36 ferroptosis-related genes in osteoarthritis were obtained.Gene Ontology enrichment analysis showed that these differentially expressed genes were mainly involved in oxidative stress,corticoste roids response,ste roid hormone res ponse,glucocorticoid res ponse,and reactive oxygen metabolism, and play a role in NAD(P)H oxidase and oxidoreductase activity,heme binding,and tetrapyrrole binding that produce superoxides.Kyoto Encyclopedia of Genes and Genomes enrichment analysis showed that ferroptosis-related genes in osteoarthritis were mainly involved in NOD-like recepto r,interleukin-17,hypoxia-inducible factor-1,tumor necrosis factor,forkhead box transcription factor class O. After constructing the protein-protein interaction network,ferroptosis-related HUB genes,including vascular endothelial growth factor A,interleukin-6,JUN,PTGS2,DUSP1,were identified.Four ferro ptosis marker genes,including vascular endothelial growth factor A,inte rleukin-6,PTGS2,and DUSP1,were selected.In vitro expe riments confirmed significant differences between the control and expe rimental groups(P<0.05).The overall findings indicate that ferro ptosis may act on osteoarthritis through oxidative stress,redox and induced inflammation,which may be related to NAD(P) H sti mulation by signaling pathway such as hypoxia-inducible factor 1.Vascular endothelial growth factor A,inte rleukin-6,PTGS2,and DUSP1 can be used as biomarkers of ferroptosis in osteoarthritis.
作者
李哲
袁长深
官岩兵
徐文飞
廖书宁
容伟明
梅其杰
段戡
Li Zhe;Yuan Changshen;Guan Yanbing;Xu Wenfei;Liao Shuning;Rong Weiming;Mei Qijie;Duan Kan(Guangxi University of Chinese Medicine,Nanning 530000,Guangxi Zhuang Autonomous Region,China;Orthopedic Department of the Limbs,The First Affiliated Hospital of Guangxi University of Chinese Medicine,Nanning 530023,Guangxi Zhuang Autonomous Region,China)
出处
《中国组织工程研究》
CAS
北大核心
2023年第17期2637-2643,共7页
Chinese Journal of Tissue Engineering Research
基金
国家自然科学基金(82060875),项目负责人:袁长深
国家自然科学基金(82160912),项目负责人:段戡
广西中医药大学自然科学研究项目(2019QN022),项目负责人:袁长深。
关键词
骨关节炎
铁死亡
基因
信号通路
生物信息学
软骨细胞
细胞实验
生物标志物
氧化应激
炎症
osteoarthritis
ferroptosis
gene
signaling pathway
bioinformatics
chondrocyte
cellular experiment
biomarker
oxidative stress
inflammation
