AUTHOR=Wei Jing , Guo Fangzheng , Song Yamin , Xu Kun , Lin Feiyang , Li Kangsheng , Li Baiqing , Qian Zhongqing , Wang Xiaojing , Wang Hongtao , Xu Tao 

TITLE=Transcriptional analysis of human peripheral blood mononuclear cells stimulated by Mycobacterium tuberculosis antigen

JOURNAL=Frontiers in Cellular and Infection Microbiology

VOLUME=Volume 13 - 2023

YEAR=2023

URL=https://www.frontiersin.org/journals/cellular-and-infection-microbiology/articles/10.3389/fcimb.2023.1255905

DOI=10.3389/fcimb.2023.1255905

ISSN=2235-2988

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