Circulating Extracellular Vesicles Exert Age-Dependent Immunomodulation in Acute Hypoxic Kidney Tubular Injury

Background: Aging and paracrine secreted factors would affect systemic immune responses, and circulating extracellular vesicles (EVs) potentially play a role in these processes. This study aims to investigate the interaction of plasma EVs and bone marrow-derived macrophages (BMDMs) isolated from young and aged mice in the context of acute hypoxic renal tubular injury. Methods: Plasma EVs from young (Y_EV) and aged (O_EV) C57BL/6 mice were first isolated by precipitation, then characterized by nanoparticle tracking analysis. EV miRNA were extracted and profiled, then miRNA targets were predicted using ingenuity pathway analysis. EVs stimulated BMDMs from young (Y_BMDM) and aged (O_BMDM) mice, both in naïve conditions and following hypoxic renal tubular injury. Macrophage M1/M2-related secretomes were assessed by quantitative PCR for comparison. Results: Our results revealed distinct differences in the immunomodulatory effects of Y_EVs and O_EVs, and the corresponding responses of Y_BMDMs and O_BMDMs. Y_EVs tended to induce less pro-inflammatory cytokines, whereas O_EVs had a more varied impact, inciting both pro- and anti-inflammatory responses. However, neither EV population induced a distinct ‘M1’ or ‘M2’ phenotype. Notably, plasma EVs and whole plasma induced very different responses in BMDMs; plasma from aged mice was profoundly pro-inflammatory, whereas O_EVs were not. We further characterized and compared Y_EV and O_EV miRNA cargo, surprisingly finding only a very limited difference. In the context of hypoxic renal injury, Y_EVs suppressed inflammatory cytokine expression, but O_EVs were able to induce higher anti-inflammatory cytokine expression. However, both Y_EVs and O_EVs did not directly affect the degree of protection offered by macrophage secretomes following hypoxic renal tubular injury (Figure). Conclusion: This study showed some age-related differences in the effects of EVs on macrophage function. In hypoxic renal injury, young EVs were able to reduce macrophage pro-inflammatory gene expression and increase anti-inflammatory gene expression. Further in vivo studies are warranted to decipher the exact role of circulating EVs in kidney diseases. Funding: Government Support – Non-U.S.