Endothelial progenitor cell-derived extracellular vesicles protect from complement-mediated mesangial injury in experimental anti-Thy1.1 glomerulonephritis.

ABSTRACT

BACKGROUND: Endothelial progenitor cells (EPCs) are known to induce tissue repair by paracrine mechanisms including the release of growth factors and extracellular vesicles (EVs), nanoparticles able to carry proteins and genetic information to target cells. The aim of this study was to evaluate whether EVs derived from EPCs may protect from complement-mediated mesangial injury in experimental anti-Thy1.1 glomerulonephritis. METHODS: EVs were isolated by serial ultracentrifugation from supernatants of cultured human EPCs and characterized for their protein and RNA content. In vivo, EVs were injected i.v. in the experimental rat model of mesangiolytic anti-Thy1.1 glomerulonephritis evaluating renal function, proteinuria, complement activity and histological lesions. In vitro, the biological effects of EPC-derived EVs were studied in cultured rat mesangial cells incubated with anti-Thy1.1 antibody and rat or human serum as complement source. RESULTS: After i.v. injection in Thy1.1-treated rats, EVs localized within injured glomeruli and inhibited mesangial cell activation, leucocyte infiltration and apoptosis, decreased proteinuria, increased serum complement haemolytic activity (CH50) and ameliorated renal function. EV treatment decreased intraglomerular deposition of the membrane attack complex (MAC or C5b-9) and expression of smooth muscle cell actin and preserved the endothelial antigen RECA-1 and the podocyte marker synaptopodin. The protective effect of EVs was significantly reduced by pre-treatment with a high dose of RNase (1 U/mL), suggesting a key role for EV-carried RNAs in these mechanisms. Indeed, EPC-derived EVs contained different mRNAs coding for several anti-apoptotic molecules and for the complement inhibitors Factor H, CD55 and CD59 and the related proteins. The in vitro experiments aimed to investigate the mechanisms of EV protection indicated that EVs transferred to mesangial cell mRNAs coding for Factor H, CD55 and CD59 and inhibited anti-Thy1.1 antibody/complement-induced apoptosis and C5b-9/C3 mesangial cell deposition. CONCLUSIONS: EVs derived from EPCs exert a protective effect in Thy1.1 glomerulonephritis by inhibition of antibody- and complement-mediated injury of mesangial cells.