PstS1-loaded exosomes promote Mycobacterium tuberculosis infection via miR-122-mediated PI3K/AKT/mTOR activation and autophagy suppression.

Tuberculosis (TB), caused by Mycobacterium tuberculosis (MTB), is a chronic respiratory infectious disease. With the continuous increase in new cases, TB has become the second leading infectious disease worldwide, posing a significant challenge to public health. Previous studies have shown that exosomes play a crucial role in MTB infection, and microRNA-122 (miR-122) is highly expressed in MTB-infected cells. Exosomes secreted under the induction of phosphate-specific transport system substrate-binding protein 1 (PstS1) were characterized using transmission electron microscopy (TEM) and Western blot. Given that miR-122 expression increases upon MTB infection, quantitative real-time polymerase chain reaction (RT-qPCR) was performed to measure miR-122 levels in exosome-treated cells. Western blot, immunofluorescence (IF), and colony-forming unit assays were conducted to assess MTB survival and the expression of phosphatidylinositol 3-kinase/protein kinase B/mechanistic target of rapamycin (PI3K/AKT/mTOR) signaling pathway components and autophagy-related proteins after transfection with miR-122 inhibitor or mimic. A mouse model was established to further validate the role of exosomes in MTB infection. Enzyme-linked immunosorbent assay (ELISA) measured serum cytokine levels, and flow cytometry was used to sort immune cells. Hematoxylin and eosin (H&E) staining evaluated lung tissue pathology, while Western blot and IF analyzed signaling pathway and autophagy-related protein expression in lung tissues. A PI3K-specific inhibitor (LY294002) was used to confirm miR-122-mediated regulation of the pathway. TEM revealed the characteristic bilayer vesicle morphology of exosomes, and Western blot confirmed exosomal markers (cluster of differentiation 63, cluster of differentiation 9, tumor susceptibility gene 101) and PstS1 in PstS1-induced exosomes. RT-qPCR showed significantly elevated miR-122 levels in exosome-treated cells. Both in vitro and in vivo experiments demonstrated that miR-122 overexpression upregulated phosphorylated PI3K, AKT, and mTOR while downregulating autophagy-related proteins microtubule-associated protein 1A/1B-light chain 3-phosphatidylethanolamine conjugate and Beclin1. ELISA and flow cytometry analyses indicated that miR-122 promoted cytokine secretion and increased immune cell numbers. H&E staining confirmed that miR-122 upregulation exacerbated MTB-induced lung damage. Furthermore, LY294002 treatment confirmed that exosome-derived miR-122 activates the PI3K/AKT/mTOR pathway, suppresses autophagy, and facilitates MTB infection. PstS1-loaded exosomes upregulate miR-122, activating the PI3K/AKT/mTOR pathway to inhibit autophagy and promote MTB infection.