Extracellular Vesicles derived from Endothelial Progenitor Cells inhibit complement- and cytokine-mediated injury of renal glomerular endothelial cells and podocytes

Abstract

Glomerulonephritis are renal inflammatory processes characterized by increased permeability of the Glomerular Filtration Barrier (GFB), with consequent hematuria and proteinuria. Glomerular endothelial cells (GEC) and podocytes are part of the GFB and contribute to maintaining its structural and functional integrity by interacting with each other through paracrine mediators. Immune-related complement cascade activation and pro-inflammatory cytokines (CK) such as TNF-alpha and IL-6 alter GFB by causing acute glomerular injury and progression toward chronic kidney disease. Endothelial Progenitor Cells (EPC) are bone-marrow-derived hematopoietic stem cells circulating in peripheral blood that repair injured endothelium by releasing paracrine mediators, such as Extracellular Vesicles (EV). EVs are microparticles involved in intercellular communication by transferring proteins, lipids, and genetic material (mRNA, microRNA, lncRNA). We previously demonstrated EPC-derived EVs activate an angiogenic program in quiescent endothelial cells in different experimental models. This study evaluates EPC-derived EVs' protective effects on GFB through tests on GECs and podocytes in vitro in detrimental conditions with CKs (TNF-alpha/IL-6) and complement protein C5a. First, EVs internalize in GECs and podocytes through different integrins and L-selectin. In GECs, EVs trigger angiogenesis, the formation of capillary-like structures, and cell migration by modulating gene expression and inducing the release of growth factors, such as VEGF and HGF. In the presence of CKs, EVs protect GECs from apoptosis by decreasing oxidative stress and prevent leukocyte adhesion by inhibiting adhesion molecules' expression (ICAM-1, VCAM-1, E-selectin). On podocytes, EVs inhibit cell death and prevent nephrin shedding induced by CKs. Last, in a co-culture model mimicking GFB, EV-mediated biological action on GECs protects podocytes indirectly from CK-mediated damage. RNase pre-treatment of EVs abrogated their protective effects, suggesting the crucial role of RNA transfer from EVs to target glomerular cells. In conclusion, EPC-derived EVs protect GFB from complement- and cytokine-induced damage, indicating their potential role as therapeutic agents for glomerulonephritis.