目的:应用转录组学技术研究脊柱结核的差异表达基因(DEGs),目的是确定脊柱结核临床治疗的新治疗目标和预后指标。
方法:在第二医院骨科就诊的患者,兰州大学于2021年1月至2023年5月入学。根据纳入和排除标准,试验组5例,对照组5例。提取总RNA并在测序平台上进行配对末端测序。用干净的读数处理测序数据并注释参考基因组后,进行FPKM归一化和差异表达分析。分析DEGs和长非编码RNA(LncRNA)的京都基因和基因组百科全书(KEGG)和基因本体论(GO)富集。预测并分析了LncRNA对差异表达mRNA(DEmRNA)的顺式调节,以建立共表达网络。
结果:这项研究确定了2366个DEG,974个基因显著上调,1392个基因显著下调。上调的基因与细胞因子-细胞因子受体相互作用有关,结核病,和TNF-α信号通路,主要富含免疫和炎症等生物过程。下调的基因与肌肉发育有关,收缩,真菌防御反应,和胶原蛋白代谢过程。对来自骨结核RNA-seq数据的LncRNAs的分析检测到总共3652个LncRNAs,356个显著上调,184个显著下调。进一步的分析确定了311个显著不同的LncRNAs,它们可以顺式调节777个靶基因,富含肌肉收缩等途径,炎症反应,和免疫反应,与骨结核密切相关。有51个基因富集在由顺式作用LncRNAs调节的免疫应答途径中。调节免疫应答相关基因的LncRNAs,例如上调的RP11-451G4.2,RP11-701P16.5,AC079767.4,AC017002.1,LINC01094,CTA-384D8.35和AC092484.1,以及下调的RP11-2C24.7可能是潜在的预后和治疗靶点.
结论:脊柱结核中的DEmRNAs和LncRNAs均与免疫调节途径相关。这些途径在机理上促进或抑制结核的感染和发展,在结核向骨组织转移的过程中发挥重要作用。
OBJECTIVE: To investigate the differential expression genes (DEGs) in spinal tuberculosis using transcriptomics, with the aim of identifying novel therapeutic targets and prognostic indicators for the clinical management of spinal tuberculosis.
METHODS: Patients who visited the Department of Orthopedics at the Second Hospital, Lanzhou University from January 2021 to May 2023 were enrolled. Based on the inclusion and exclusion criteria, there were 5 patients in the test group and 5 patients in the control group. Total RNA was extracted and paired-end sequencing was conducted on the sequencing platform. After processing the sequencing data with clean reads and annotating the reference genome, FPKM normalization and differential expression analysis were performed. The DEGs and long non-coding RNAs (LncRNAs) were analyzed for Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) enrichment. The cis-regulation of differentially expressed mRNAs (DE mRNAs) by LncRNAs was predicted and analyzed to establish a co-expression network.
RESULTS: This study identified 2366 DEGs, with 974 genes significantly upregulated and 1392 genes significantly downregulated. The upregulated genes are associated with cytokine-cytokine receptor interactions, tuberculosis, and TNF-α signaling pathways, primarily enriched in biological processes such as immunity and inflammation. The downregulated genes are related to muscle development, contraction, fungal defense response, and collagen metabolism processes. Analysis of LncRNAs from bone tuberculosis RNA-seq data detected a total of 3652 LncRNAs, with 356 significantly upregulated and 184 significantly downregulated. Further analysis identified 311 significantly different LncRNAs that could cis-regulate 777 target genes, enriched in pathways such as muscle contraction, inflammatory response, and immune response, closely related to bone tuberculosis. There are 51 genes enriched in the immune response pathway regulated by cis-acting LncRNAs. LncRNAs that regulate immune response-related genes, such as upregulated RP11-451G4.2, RP11-701P16.5, AC079767.4, AC017002.1, LINC01094, CTA-384D8.35, and AC092484.1, as well as downregulated RP11-2C24.7, may serve as potential prognostic and therapeutic targets.
CONCLUSIONS: The DE mRNAs and LncRNAs in spinal tuberculosis are both associated with immune regulatory pathways. These pathways promote or inhibit the tuberculosis infection and development at the mechanistic level and play an important role in the process of tuberculosis transferring to bone tissue.