绝经后骨质疏松小鼠颌骨骨髓间充质干细胞差异miRNA检测及靶基因分析

doi:10.19439/j.sjos.2024.06.001

摘要/Abstract

摘要： 目的: 通过生物信息学方法探讨绝经后骨质疏松（postmenopausal osteoporosis,POP）小鼠模型颌骨骨髓间充质干细胞（mandibular bone marrow mesenchymstem cells, MBMSCs）的差异miRNA表达谱和miRNA预测靶基因,为 POP 的诊治和预防提供新的靶点。方法: 通过卵巢切除术建立绝经后骨质疏松小鼠模型,采用全骨髓贴壁法获取MBMSCs,利用微阵列测序技术对MBMSCs进行芯片检测。随后,对检测结果进行miRNA鉴定和预测分析,并预测miRNA的靶基因。对预测靶基因结果进行基因本体论（GO）分析、基因数据库（KEGG）富集分析以及蛋白质-蛋白质相互作用（PPI）网络分析,通过 Degree、Betweenness 和 Closeness 等3种算法筛选出关键Hub基因。采用GraphPad Prism 8.0及R语言对数据进行统计学分析。结果: 以P<0.05为阈值,获得84个差异表达miRNA,其中,上调33个,下调51个。对84个差异 miRNA 的预测靶基因 mRNA 进行GO、KEGG 富集分析,发现涉及多种生物学过程及相关通路,其中,130个靶基因 mRNA 富集于“调节干细胞多能性”的信号通路,对富集于该通路的130个预测靶基因mRNA绘制 PPI 网络图并进行 Hub 基因筛选,最终得到可靠度较高的7个Hub基因,分别是Ctnnβ1、Hras、Kras、Akt1、Mapk3、Smad3和Smad2。其中,与OP有显著相关性的基因分别是Ctnnβ1、Akt1、Mapk3、Smad3和Smad2。结论: 在POP小鼠MBMSCs 细胞中发现的差异 miRNA,有可能作为 POP 潜在的生物标志物,为POP的诊治提供新的思路及理论依据。

关键词: 绝经后骨质疏松症, 颌骨骨髓间充质干细胞, 差异miRNA表达谱, 生物信息学

Abstract: PURPOSE: To explore the differential miRNA expression profiles and predicted target genes of mandibular bone marrow mesenchymal stem cells (MBMSCs) in a postmenopausal osteoporosis (POP) mouse model using bioinformatics methods, providing new targets for diagnosis, treatment, and prevention of POP. METHODS: POP mouse model was established by performing ovariectomy surgery, and MBMSCs were obtained using whole bone marrow adherent culture method. Microarray sequencing was performed to detect the miRNA expression profile of MBMSCs. Subsequently, miRNA identification and prediction analysis were conducted, along with the prediction of target genes. Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses, and protein-protein interaction (PPI) network analysis were performed on the predicted target genes. Key hub genes were identified using algorithms such as Degree, Betweenness, and Closeness. The data were analyzed by GraphPad Prism 8.0 and R language. RESULTS: A total of 84 differentially expressed miRNAs were obtained using a threshold of P<0.05, with 33 upregulated and 51 downregulated miRNAs. GO and KEGG enrichment analyses of the 84 differentially expressed miRNAs revealed their involvement in various biological processes and pathways. Among them, 130 target gene mRNAs were enriched in the "regulation of stem cell pluripotency" signaling pathway. PPI network analysis and hub gene selection were performed for the 130 predicted target gene mRNAs, resulting in the identification of 7 reliable hub genes: Ctnnβ1, Hras, Kras, Akt1, Mapk3, Smad3, and Smad2. Among these hub genes, Ctnnβ1, Akt1, Mapk3, Smad3, and Smad2 were found to be significantly associated with POP. CONCLUSIONS: The differentially expressed miRNAs identified in MBMSCs of POP mice may serve as potential biomarkers and play important roles in the pathogenesis of POP. This study provides new research direction and theoretical basis for the diagnosis and treatment of POP.

Key words: Postmenopausal osteoporosis, Mandibular bone marrow mesenchymal stem cells, Differential miRNA expression profiles, Bioinformatics

中图分类号:

R394

杨杉杉, 张为, 胡小华, 杨晓红. 绝经后骨质疏松小鼠颌骨骨髓间充质干细胞差异miRNA检测及靶基因分析[J]. 上海口腔医学, 2024, 33(6): 561-571.

YANG Shan-shan, ZHANG Wei, HU Xiao-hua, YANG Xiao-hong. Differential miRNA profiling and target gene analysis of marrow mesenchymal stem cells in postmenopausal osteoporotic mouse mandibles[J]. Shanghai Journal of Stomatology, 2024, 33(6): 561-571.

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