Deoxycorticosterone Acetate-salt Promotes Endothelial-mesenchymal Transition in Human Glomerular Endothelial Cells.

Endothelial cells (ECs) lose their endothelial specification and gain mesenchymal cell features during endothelial-mesenchymal transition (EndMT). Post-developmental EndMT disrupts EC homeostasis, leading to vascular dysfunction. We found that afferent arterioles from deoxycorticosterone acetate (DOCA)-salt treated mice had > 5 fold upregulation of mRNAs for preproendothelin-1, p47phox ,NOX2 and TGF-β accompanied by microvascular dysfunction. Since these may cause EndMT, we investigated the mechanism in human glomerular endothelial cells (HGECs) treated for 7-21 days with high salt and DOCA. Endothelin-1 (ET-1) in the medium was increased 2.8±0.2 fold by day 7 while the cells gained multiple mesenchymal markers with increased mRNA for alpha-smooth muscle actin (1.78±0.19 and 2.96±0.32 fold, P<0.05 and 0.01, n=3) and transgelin ( 1.96±0.14 and 2.91±0.28 fold, p<0.05 and 0.01, n=3) on day 7 and 21, respectively, and markedly downregulated mRNA for endothelial markers with decreased vascular endothelial cadherin ( 1.99± 0.27 and 2.12±0.24 fold, P<0.05 and 0.005, n=3) and platelet endothelial cell adhesion molecule 1 (1.78±0.26 and 1.94±0.23 fold, P<0.05 and 0.005, n=3). There were parallel changes in protein expression. Dihydroethidium and MitSox fluorescence probes were used to determine intracellular and mitochondria ROS. The fluorescent intensities were increased by 1.89±0.27 and 1.62±0.22 fold (P<0.01, N=6) respectively in the cells treated for 7 days with DOCA-salt accompanied by increased expression of TGF-β and phosphorylated-extracellular signal-regulated kinases (P-ERK 1/2). In conclusion, human glomerular endothelial cells treated with high salt and DOCA for 1-3 weeks have increased cellular and mitochondrial ROS, ET-1, TGF-β and P-ERK that could account for adverse changes of endothelial-mesenchymal transition and associated microvascular dysfunction.