ABSTRACT: Objective: The purpose of this study is to investigate the differential expression of genes 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 & Clinical Medical School, Lanzhou Univeristy from January 2021 to May 2023 were enrolled. Based on the inclusion and exclusion criteria, there were 5 patients in the experimental group (test group) and 5 in the control group. Total RNA was extracted and paired-end sequencing was conducted using the KAPA Stranded mRNA-Seq Kit on the Illumina Novaseq 6000 sequencing platform. After processing the sequencing data with clean reads and annotating the reference genome, FPKM normalization and differential expression analysis were performed. The differentially expressed genes (DEGs) and long non-coding RNAs (LncRNAs) were analyzed for KEGG and GO enrichment. The cis-regulation of differentially expressed mRNAs by LncRNAs was predicted and analyzed to establish a co-expression network between the two. Finally, WGCNA analysis was conducted. Results: High-throughput sequencing analysis identified 2366 differentially expressed genes, with 974 genes significantly upregulated and 1392 genes significantly downregulated. KEGG enrichment revealed that 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. Downregulated genes are related to muscle development, contraction, fungal defense response, and collagen metabolism processes. Analysis of lncRNAs from bone tuberculosis RNA-seq data using an RPKM threshold of >0.5 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. Multiple molecules related to immune regulation identified through WGCNA analysis can serve as markers for early diagnosis and molecular targets for the treatment of bone tuberculosis. Conclusion: The differentially expressed mRNAs and LncRNAs in spinal tuberculosis are both associated with immune regulatory pathways. These pathways not only promote or inhibit the binding infection and development at the mechanistic level but may also serve as important markers for the earliest detection of tuberculosis metastasis to bone tissue at the clinical level.
