Abstract:

Objective: This study aims to uncover the expression profile of circular RNAs (circRNAs) in peripheral blood mononuclear cells (PBMCs) from tuberculosis (TB) patients, and identify potential circRNAs as diagnostic biomarkers for TB. Methods: Microarray chip was used to get the circRNA expression profiles of PBMCs from TB patients and healthy controls (HCs). Weighted gene co-expression network analysis (WGCNA) was employed to identify critical gene modules strongly associated with TB. The functions of genes within these key modules were further explored through gene ontology (GO), Kyoto encyclopedia of genes and genomes (KEGG), and immune infiltration analysis. Subsequently, the circRNAs derived from the key modules and had significant differences between TB and HCs, were validated by real-time fluorescent quantitative PCR (qPCR) in an independent validation sample set, to verify the microarray results. The receiver operating characteristic (ROC) curve analysis was performed to assess the diagnostic performance of these candidate circRNAs. Additionally, Pearson correlation analysis was conducted for the candidate circRNAs and clinical features. Results: Through WGCNA analysis, a total of 16 gene modules were identified, with the brown module being the most strongly associated with TB. Functional enrichment analysis revealed that genes in the brown module were primarily enriched in T cell receptor signaling pathway, mitogen-activated protein kinase signaling pathway, mitochondrial autophagy in animal cells, ferroptosis, ubiquitin-mediated proteolysis and other functional pathways. Immune infiltration analysis based on the genes in the brown module indicated significant differences in the proportion of CD8⁺ T cells (P<0.001), resting memory CD4⁺ T cells (P<0.0001) and monocytes (P<0.001) between the TB and HCs group. In the brown module, nine TB-specific circRNAs were identified based on the selection criteria (P<0.05, fold change >4 or <0.25, and the mean expression value >10 in at least one group). Among them, hsa_circ_0052124’s expression level was significantly downregulated in the TB group (Z=―6.328, P<0.0001), its area under the ROC curve (AUC) was 0.976 (95%CI: 0.940-1.000, P<0.0001), with a sensitivity of 90.0% (95%CI: 73.5%-97.9%) and a specificity of 100.0% (95%CI: 88.4%-100.0%), which meant it could effectively distinguish TB patient from HC individuals. Pearson correlation analysis showed a positive correlation between hsa_circ_0052124 expression level and other clinical features (P<0.01), including apolipoprotein B (r=0.715), lipoprotein(a)(r=0.598), and glucose (r=0.575) level in the peripheral blood. Conclusion: Our study uncovered the circRNA expression profile of TB patients, identified a key gene module significantly associated with TB, and discovered a circRNA (hsa_circ_0052124) with a significant difference between TB patients and HCs.

Key words: Tuberculosis, RNA, Mycobacterium infections, Gene expression, Immunity