Mycobacterium tuberculosis Mce2D protein blocks M1 polarization in macrophages by inhibiting the ERK signaling pathway.

Macrophages play a pivotal role in controlling Mycobacterium infection, and the pathogen thrives in the event of immune evasion and immunosuppression of macrophages. Mammalian cell entry proteins (Mce) are required for Mycobacterium tuberculosis (M. tb) growth and the host cell's initial phagocytosis and cytokine response. Mce2D protein is one of a family of proteins that infect M. tb; however, the function and mechanism of action remain unclear. In this study, we constructed the Mce2D knockout strain using Mycobacterium smegmatis to study the function of Mce2D in the infection of macrophages. The results indicated that compared to the knockout strain, the release of proinflammatory cytokines (TNF-α and IL-1ß) reduced when WT strain infected the macrophages. Moreover, Mce2D boosted the metabolism of oxidized fatty acids, increased the energy supply of TCA, and lowered the glycolysis of glucose in macrophages after bacterial infection, all of which prevented the polarization of macrophages to M1, which was driven by the fact that Mce2D blocked ERK2 phosphorylation by interacting with ERK2 through its DEF motif. This, in turn, promoted nuclear translocation of HIF-1α, allowing signal accumulation, which increased the HIF-1α transcription levels. Finally, the mouse infection experiment showed that Mce2D caused blockage of M1 polarization of alveolar macrophages, resulting in reduced bactericidal activity and antigen presentation, weakening Th1 cell-mediated immune response and helping bacteria escape the immune system. Our results reveal that Mce2D causes immune escape by blocking M1 polarization in macrophages, providing potential targets for the rational design of therapies against M. tb infection.

Hsa_circ_0007460 affects the survival of intracellular Mycobacterium tuberculosis by regulating autophagy and apoptosis of macrophages.

Circular RNA (circRNA) is a category of non-coding RNAs characterized by the absence of a 5'-cap and 3'-poly(A) tail, and participates in the physiological processes of various human diseases. Nonetheless, the diagnostic and functional significance of circRNAs in active pulmonary tuberculosis (ATB) remains uncertain. Consequently, the purpose of this study is to investigate whether hsa_circ_0007460 can be employed as a potential diagnostic biomarker in ATB patients and explore its function. The result of real-time quantitative fluorescent PCR (RT-qPCR) validated a notable increase in the expression of hsa_circ_0007460 in the peripheral blood of 32 ATB patients, as well as in THP-1 human macrophages infected with Bacillus Calmette Guerin (BCG) which is an attenuated strain of Mycobacterium bovis. Additionally, the receiver operating curve (ROC) illustrated that the area under the ROC curve (AUC), sensitivity and specificity were 0.7474, 76.67%, and 78.13% respectively. RNase R, Actinomycin D and other experiments confirmed that hsa_circ_0007460 was stabler than its linear mRNA, indicating that hsa_circ_0007460 has potential as a diagnostic biomarker of ATB. Furthermore, Western blot (WB), Cell Counting Kit-8 (CCK-8), plate counting, and immunofluorescence experiments revealed that hsa_circ_0007460 could regulate apoptosis and autophagy of macrophages. The downstream miRNAs and mRNAs were subsequently predicted using bioinformatics, and the hsa circ 0007460/hsa-miR-3127-5p/PATZ1 axis was built. These above results suggest that hsa_circ_0007460 is substantially up-regulated in the peripheral blood of patients with ATB and can be utilized as a potential diagnostic biomarker. In addition, hsa_circ_0007460 can promote apoptosis of macrophages and inhibit autophagy of macrophages, thereby promoting the survival of BCG.

Enhanced Glycosylation Caused by Overexpression of Rv1002c in a Recombinant BCG Promotes Immune Response and Protects against Mycobacterium tuberculosis Infection.

Tuberculosis (TB) is a major global health threat despite its virtual elimination in developed countries. Issues such as drug accessibility, emergence of multidrug-resistant strains, and limitations of the current BCG vaccine highlight the urgent need for more effective TB control measures. This study constructed BCG strains overexpressing Rv1002c and found that the rBCG-Rv1002c strain secreted more glycosylated proteins, significantly enhancing macrophage activation and immune protection against Mycobacterium tuberculosis (M. tb). These results indicate that Rv1002c overexpression promotes elevated levels of O-glycosylation in BCG bacteriophages, enhancing their phagocytic and antigenic presentation functions. Moreover, rBCG-Rv1002c significantly upregulated immune regulatory molecules on the macrophage surface, activated the NF-κB pathway, and facilitated the release of large amounts of NO and H2O2, thereby enhancing bacterial control. In mice, rBCG-Rv1002c immunization induced greater innate and adaptive immune responses, including increased production of multifunctional and long-term memory T cells. Furthermore, rBCG-Rv1002c-immunized mice exhibited reduced lung bacterial load and histological damage upon M. tb infection. This result shows that it has the potential to be an excellent candidate for a preventive vaccine against TB.

The hsa_circ_0002371/hsa-miR-502-5p/ATG16L1 axis modulates the survival of intracellular Mycobacterium tuberculosis and autophagy in macrophages.

Circular RNAs (circRNAs) play a critical role in pathological mechanisms of Mycobacterium tuberculosis (Mtb) and can be used as a new biomarker for active tuberculosis (ATB) diagnosis. Therefore, we identified significantly dysregulated circRNAs in ATB patients and healthy controls (HC) and explored their molecular mechanism. We found that hsa_circ_0002371 was significantly up-regulated in PBMCs of ATB patients and Mycobacterium tuberculosis H37Rv- or Mycobacterium bovis bacillus Calmette Guerin (BCG)-infected THP-1 cells. Functional experiments demonstrated that hsa_circ_0002371 inhibited autophagy in BCG-infected THP-1 cells and promoted intracellular BCG survival rate. In terms of mechanism, hsa_circ_0002371 facilitated the expression of hsa-miR-502-5p, as shown by bioinformatics and dual-luciferase reporter gene analysis, respectively. Notably, hsa-miR-502-5p inhibited autophagy via suppressing autophagy related 16 like 1 (ATG16L1) in BCG-infected macrophages and thus promoting intracellular BCG growth. In summation, hsa_circ_0002371 increased the suppression of hsa-miR-502-5p on ATG16L1 and inhibited autophagy to promote Mtb growth in macrophages. In Conclusion, our data suggested that hsa_circ_0002371 was significantly up-regulated in the PBMCs of ATB patients compared with HC. The hsa_circ_0002371/hsa-miR-502-5p/ATG16L1 axis promoted the survival of intracellular Mtb and inhibited autophagy in macrophages. Our findings suggested hsa_circ_0002371 could act as a potential diagnostic biomarker and therapeutic target.

Mycobacterium tuberculosis hijacks host macrophages-derived interleukin 16 to block phagolysosome maturation for enhancing intracellular growth.

The discovery of promising cytokines and clarification of their immunological mechanisms in controlling the intracellular fate of Mycobacterium tuberculosis (Mtb) are necessary to identify effective diagnostic biomarkers and therapeutic targets. To escape immune clearance, Mtb can manipulate and inhibit the normal host process of phagosome maturation. Phagosome maturation arrest by Mtb involves multiple effectors and much remains unknown about this important aspect of Mtb pathogenesis. In this study, we found that interleukin 16 (IL-16) is elevated in the serum samples of Tuberculosis (TB) patients and can serve as a specific target for treatment TB. There was a significant difference in IL-16 levels among active TB, latent TB infection (LTBI), and non-TB patients. This study first revealed that macrophages are the major source of IL-16 production in response to Mtb infection, and elucidated that IL-16 can promote Mtb intracellular survival by inhibiting phagosome maturation and suppressing the expression of Rev-erbα which can inhibit IL-10 secretion. The experiments using zebrafish larvae infected with M. marinum and mice challenged with H37Rv demonstrated that reducing IL-16 levels resulted in less severe pathology and improved survival, respectively. In conclusion, this study provided direct evidence that Mtb hijacks the host macrophages-derived interleukin 16 to enhance intracellular growth. It is suggesting the immunosuppressive role of IL-16 during Mtb infection, supporting IL-16 as a promising therapeutic target.