Transcriptional analysis of human peripheral blood mononuclear cells stimulated by Mycobacterium tuberculosis antigen (Mtb-Ag)

Background: Mycobacterium tuberculosis antigen (Mtb-Ag) is a polypeptidecomponent with a molecular weight of 10-14 kDa that is obtained from thesupernatant of the H37Ra strain after heat treatment. It stimulates the activationand proliferation of gdT cells in the blood to produce an immune responseagainst tuberculosis. Mtb-Ag is therefore crucial for classifying and detecting the central genes and key pathways involved in TB initiation and progression.Methods: In this study, we performed high-throughput RNA sequencing ofperipheral blood mononuclear cells (PBMC) from Mtb-Ag-stimulated andcontrol samples to identify differentially expressed genes and used them forgene ontology (GO) and a Kyoto Encyclopedia of Genomes (KEGG) enrichmentanalysis. Meanwhile, we used PPI protein interaction network and Cytoscapeanalysis to identify key genes and qRT-PCR to verify differential gene expression. Single-gene enrichment analysis (GSEA) was used further to elucidate the potential biological functions of key genes. Analysis of immune cell infiltration and correlation of key genes with immune cells after Mtb-Ag-stimulated using R language.Results: We identified 597 differentially expressed genes in Mtb-Ag stimulatedPBMCs. KEGG and GSEA enrichment analyzed the cellular pathways related toimmune function, and DEGs were found to be primarily involved in the TNFsignaling pathway, the IL-17 signaling pathway, the JAK-STAT signaling pathway, cytokine-cytokine receptor interactions, and the NF-kB signaling pathway. Wayne analysis using GSEA, KEGG, and the protein-protein interaction (PPI) network showed that 34 genes, including PTGS2, IL-1b, IL-6, TNF and IFN-g et al., were co-expressed in the five pathways and all were up-regulated by Mtb-Ag stimulation. Twenty-four DEGs were identified using qRT-PCR, including fourteen up-regulated genes (SERPINB7, IL20, IFNG, CSF2, PTGS2, TNF-a, IL36G, IL6, IL10, IL1A, CXCL1, CXCL8, IL4, and CXCL3) and ten down-regulatedgenes (RTN1, CSF1R CD14, C5AR1, CXCL16, PLXNB2, OLIG1, EEPD1, ENG , and CCR1). These findings were consistent with the RNA-Seq results.Conclusion: The transcriptomic features associated with Mtb-Ag provide thescientific basis for exploring the intracellular immune mechanisms against Mtb.However, more studies on these DEGs in pathways associated with Mtb-Agstimulation are needed to elucidate the underlying pathologic mechanisms ofMtb-Ag during Mtb infection.