Human neutrophil-derived extracellular vesicles induce renal endothelial inflammation in critical illness: an ex vivo investigation.

Abstract

Circulating neutrophil-derived extracellular vesicles (NEVs) may contribute to the pathophysiology of acute kidney injury by causing glomerular endothelial inflammation. NEVs were first isolated from ex vivo, lipopolysaccharide stimulation of whole blood taken from healthy volunteers (median age [interquartile range {IQR}]: 32 [26-42] yr; 47% female), and also from plasma of COVID-19 patients with acute respiratory distress syndrome (median age [IQR]: 59 [52-66] yr; 45% female). NEVs were incubated for 4 h in a co-culture of peripheral blood mononuclear cells and either human umbilical vein endothelial cells or renal glomerular endothelial cells. Enzyme-linked immunoassays (tumour necrosis factor-alpha [TNF]) and flow cytometry (median fluorescence intensity [MFI]) were used to quantify cell-specific markers of inflammation/activation, in the presence/absence of pharmacological inhibitors. NEVs were internalised by monocytes, leading to their activation via the p38 mitogen-activated protein kinase pathway and increased release of TNF (median [IQR]: 676 [474-1731] pg ml^-1 after NEV internalisation, compared with controls (27 [18-29] pg ml^-1, P=0.003). This proinflammatory response increased cell adhesion molecule expression (E-selectin) on human umbilical vein endothelial cells (median MFI [IQR]; NEVs: 4120 [3671-4858] vs untreated: 1438 [1252-1708], P=0.008) and human renal glomerular endothelial cells (median MFI [IQR]; NEVs: 2960 [2471-4991] vs untreated: 931 [881-1181], P=0.003). NEVs contained substantial amounts of matrix metalloproteinase-8 and -9, inhibitors of which (doxycycline, MMP-8 inhibitor or MMP-9 inhibitor) prevented endothelial cell inflammation. Circulating NEVs may contribute to acute kidney injury through renal endothelial inflammation in a monocyte-dependent fashion in patients with acute respiratory distress syndrome.