Host Cell-Derived Extracellular Vesicles Regulate Iron Uptake in Recipient Macrophages during <i>Mycobacterium abscessus</i> Infection

The role of host cell-derived extracellular vesicles (EVs) in host-pathogen interactions remains to be defined during mycobacterial infection. In this study, we characterized EVs from mouse RAW 264.7 cells uninfected or infected with <i>Mycobacterium abscessus</i> (<i>M</i>.<i>ab</i>), one of the most common nontuberculous mycobacterial (NTM) pathogens in humans. Our results show that <i>M</i>.<i>ab</i> infection increased the release of EVs but had no effect on the size distribution and total protein abundance of EVs from RAW 264.7 cells. Interestingly, EVs released by <i>M</i>.<i>ab</i>-infected RAW 264.7 cells significantly enhanced <i>M</i>.<i>ab</i> growth within recipient macrophages in cell culture. The proteomic analysis found that transferrin receptor was enriched in EVs from <i>M</i>.<i>ab</i>-infected RAW 264.7 cells compared to EVs from uninfected cells. Iron assay further indicates that EVs from <i>M</i>.<i>ab</i>-infected RAW 264.7 cells increased total iron level in recipient macrophages but not EVs from uninfected RAW 264.7 cells. However, iron abundance within EVs remained similar across both conditions. Finally, EVs from <i>M</i>.<i>ab</i>-infected RAW 264.7 cells failed to improve <i>M</i>.<i>ab</i> growth within recipient macrophages in the presence of an iron-chelating agent, deferoxamine, in cell culture. Therefore, our data suggest that <i>M</i>.<i>ab</i>-infected mouse macrophages likely regulate <i>M</i>.<i>ab</i> growth by upregulating iron uptake in recipient cells via released EVs.