HIV Tat-Stimulated Microglial Extracellular Vesicles Are Enriched for Ferroptosis Mediators: Role of Dysregulated Autophagy.

Extracellular vesicles (EVs) serve as conduits for intercellular communication, under both physiological and pathological contexts. During disease pathogenesis, microglia-derived EVs (MEVs) play a key role in transferring pathological cargo to the recipient cells, thereby modulating their phenotype and function. The current study was aimed to explore how HIV Transactivator of transcription (Tat) protein-mediated dysregulated autophagy/ferritinophagy contributes to the expression and release of ferroptotic mediators in Tat-activated MEVs. Exposure of microglial BV2 cells to HIV Tat for 48 h resulted in significantly increased expression of the ferroptosis mediators-FTH1 and ACSL4 in Tat-MEVs. Reciprocally, pretreatment of microglia with the ferroptosis inhibitors abrogated the expression and release of these mediators in both microglial cells as well as in Tat-MEVs, without affecting the overall MEV release. Furthermore, inhibition of autophagic flux with bafilomycin A1 resulted in potentiation of HIV Tat-mediated expression and release of ferroptosis mediators in Tat-MEVs. In contrast, treatment of cells with the autophagy inducer- rapamycin, attenuated these effects. Exposure of BV2 cells to HIV Tat also resulted in increased expression of the ferritinophagy adaptor protein NCOA4, an effect mitigated by both ferroptosis inhibitors and rapamycin. These findings were also validated in brain lysates as well as brain-derived MEVs isolated from HIV Transgenic rats. Collectively, this study highlights the role of dysregulated autophagy in modulating both HIV Tat-induced ferroptosis and the release of ferroptosis cargo in MEVs. These findings advance our understanding of the molecular mechanism(s) driving HIV-associated neuropathology.