Investigating the roles of macrophage age and circulating extracellular vesicles in cytoprotection of hypoxic renal tubular cell stress.

BACKGROUND: Acute kidney injury (AKI) involves complex crosstalk between tubular epithelial cells (TECs) and immune cells. Extracellular vesicles (EVs) and ageing have emerged as modulators of this process. METHODS: We investigated the effects of circulating plasma EVs from young (Y_EVs) and old (O_EVs) mice on hypoxic TEC stress, using bone marrow–derived macrophages (BMDMs) from young or old mice, which were pre-conditioned with EVs. First, direct effects of EVs on TEC viability were assessed following hypoxia/reoxygenation injury. We then examined macrophage responses to hypoxic TEC secretomes with or without EV priming, and tested whether EV-primed BMDM secretomes modulated TEC damage, in terms of viability, cytokine expression, and epithelial–mesenchymal transition (EMT). RESULTS: EVs exerted no direct protective effect on hypoxic TECs, and no significant differences were observed between Y_EVs and O_EVs in any condition. Instead, macrophage donor age was the most significant determinant of outcomes. Conditioned medium from old BMDMs consistently improved TEC viability compared to young BMDMs. In agreement with this, the old mouse BMDM secretome also suppressed hypoxia-induced cytokine expression (Il6, Ccl2) and EMT marker induction (Acta2, Fn1) in hypoxic TECs. EV pre-treatment of BMDMs did not significantly alter these effects in either macrophage group. Thus, BMDM donor age, rather than EV donor age, was the critical driver of cytoprotection and inflammatory modulation in this model. CONCLUSION: Circulating plasma EV age had no significant direct or indirect impacts on TEC viability, inflammatory responses or EMT processes under the tested conditions. This contrasts with prior studies which have suggested broadly rejuvenating effects of young EVs and detrimental effects of old EVs. CLINICAL TRIAL NUMBER: Not applicable.