激素性股骨头坏死的生物标志与免疫浸润分析△

Biomarkers and immune infiltration analysis of steroid-induced necrosis of the femoral head

摘要:

[目的] 探讨参与激素性股骨头坏死 （steroid-induced osteonecrosis of femoral head, SONFH） 的潜在长链非编码 RNA （lncRNA） 和信号通路，并研究其分子机制。[方法] 从 NCBI-GEO 数据库 （http://www.ncbi.nlm.nih.gov/geo） 下载微阵列数据 （GSE123568），并使用生物信息学工具对其进行分析。通过分析差异表达基因 （differential expressed genes, DEG）、京都基因和基因组百科全书扩增通路、基因本体论，鉴定出蛋白质-蛋白质相互作用网络，确定了 3 个关键非编码基因和 6 个关键 mrna。 并进一步研究激素性股骨头坏死 mRNAs、miRNAs 和 lncRNAs 的共表达谱，建立 SONFH 特异性竞争内源性 RNA（ceRNA）网络，分析免疫浸润，探索 DEG 与免疫细胞的相关性。最后用 GSE26316 进行验证。[结果] 在基因芯片中总共获得了 374 个 mRNA，258 个下调，116 个上调。获得 7 个 lncRNA，其中 C20orf197、MIR22HG、XIST 是差异显著的 lncRNA（P＜0.05）。生物过程分析结果表明，DEG 在炎症反应中富集明显；分子功能分析表明，DEG 在泛素-蛋白质转移酶活性、泛素蛋白连接酶结合、 受体活性富集显著；对于细胞组分分析表明，DEG 主要富集于细胞质。PPI 网络模块中的基因主要在 GO 的血影蛋白相关细胞骨架、质膜和细胞骨架的结构成分中富集显著，而在 KEGG 中，主要在 Toll 样受体信号通路中富集。本研究共得到了 6 个关键基因。通过分析其免疫浸润，发现与正常组织相比，SONFH 组织含有更多比例的 CD4 原始 T 细胞（P＜0.05），从 GSE26316 数据集验证了关键基因 FOXO3 的表达水平。[结论] C20orf197、MIR22HG、XIST 是 SONFH 发病过程中的潜在标志物，基因轴 XIST/ Has-miR-217/FOXO3 在 SONFH 的发生发展过程中起到重要作用。

Abstract:

[Objective] To identify the potential long non-coding RNAs (lncRNAs) and signaling pathways involved in steroid-induced osteonecrosis of femoral head (SONFH), and investigate their molecular mechanisms. [Methods] Microarray data (GSE123568) were down- loaded from NCBI-GEO and analyzed using bioinformatics tools. By analyzing Differentially Expressed Genes (DEG), Kyoto Encyclopedia of Genes and Genomes (KEGG) amplification pathways, Gene Ontology (GO), and finally identified a protein-protein interaction (PPI) net- work and identified 3 key noncoding genes and 1 key mRNA. We further studied the co-expression profiles of mRNAs, miRNAs and ln- cRNAs in SONFH, established a specific competitive endogenous RNA (ceRNA) network for SONFH, analyzed immune infiltration, and ex- plored the relationship between DEG and immune cells, finally verified with GSE26316. [Results] A total of 374 mRNAs were obtained in the microarray, including 258 down-regulated and 116 up-regulated. Seven lncRNAs were obtained, among which C20orf197, MIR22HG and XIST were significantly different (P＜0.05). The results of biological process analysis showed that DEG was obviously enriched in in- flammatory response. Molecular function analysis showed that DEG was significantly enriched in ubiquitin- protein transferase activity, ubiquitin- protein ligase binding and receptor activity. Analysis of cell components showed that DEG was mainly enriched in cytoplasm. Genes in PPI network modules were mainly enriched in the structural components of GO^,s blood shadow protein-related cytoskeleton, plas- ma membrane and cytoskeleton, while in KEGG, they were mainly enriched in toll-like receptor signaling pathways. Six key genes were identified in this study. By analyzing the immune infiltration, it was found that SONFH tissues contained a higher proportion of CD4 primi- tive T cells (P＜0.05), and verified the expression level of FOXO3 in GSE26316 data set. [Conclusion] C20orf197, MIR22HG and XIST are potential markers in the pathogenesis of SONFH, and the gene axis XIST/Has-miR-217/FOXO3 plays an important role in the occurrence and development of SONFH.