Vascular Endothelial Growth Factor-C, a Potential Paracrine Regulator of Glomerular Permeability, Increases Glomerular Endothelial Cell Monolayer Integrity and Intracellular Calcium

Abstract

We have previously reported expression of vascular endothelial growth factor (VEGF)-A and -C in glomerular podocytes and actions of VEGF-A on glomerular endothelial cells (GEnC) that express VEGF receptor-2 (VEGFR-2). Here we define VEGFR-3 expression in GEnC and investigate the effects of the ligand VEGF-C. Renal cortex and cultured GEnC were examined by microscopy, and both cell and glomerular lysates were assessed by Western blotting. VEGF-C effects on trans-endothelial electrical resistance and albumin flux across GEnC monolayers were measured. The effects of VEGF-C156S, a VEGFR-3-specific agonist, and VEGF-A were also studied. VEGF-C effects on intracellular calcium ([Ca2+]i) were measured using a fluorescence technique, receptor phosphorylation was examined by immunoprecipitation assays, and phosphorylation of myosin light chain-2 and VE-cadherin was assessed by blotting with phospho-specific antibodies. GEnC expressed VEGFR-3 in tissue sections and culture, and VEGF-C increased trans-endothelial electrical resistance in a dose-dependent manner with a maximal effect at 120 minutes of 6.8 Omega whereas VEGF-C156S had no effect. VEGF-C reduced labeled albumin flux by 32.8%. VEGF-C and VEGF-A increased [Ca2+]i by 15% and 39%, respectively. VEGF-C phosphorylated VEGFR-2 but not VEGFR-3, myosin light chain-2, or VE-cadherin. VEGF-C increased GEnC monolayer integrity and increased [Ca2+]i, which may be related to VEGF-C-S particular receptor binding and phosphorylation induction characteristics. These observations suggest that podocytes direct GEnC behavior through both VEGF-C and VEGF-A.

The Importance of Cellular VEGF Bioactivity in the Development of Glomerular Disease

“…Schumacher et al[ 13] describe the downregulation of VEGF 165 b mRNA expression in laser-microdissected glomeruli from patients with Denys-Drash disease, whilst high levels of conventional VEGF 165 remain and, as a result, the glomeruli are not capable of developing passed the s-shaped body stage. Another potential player in glomerular disease from the VEGF family of proteins is VEGF-C. We have demonstrated the expression of VEGF-C by podocytes, which has functional effects on both cultured GEnC and podocytes [14, 15]. Interestingly, VEGF-C activates VEGFR2 not VEGFR3 in cultured GEnC resulting in a reduction in macromolecular passage, which is in sharp contrast to the increased macromolecular passage by VEGF-A [14].…”

“…Another potential player in glomerular disease from the VEGF family of proteins is VEGF-C. We have demonstrated the expression of VEGF-C by podocytes, which has functional effects on both cultured GEnC and podocytes [14, 15]. Interestingly, VEGF-C activates VEGFR2 not VEGFR3 in cultured GEnC resulting in a reduction in macromolecular passage, which is in sharp contrast to the increased macromolecular passage by VEGF-A [14]. Therefore, VEGF-C has the potential to affect the local dosage of glomerular VEGF-A by competing with signalling effects through the same receptor.…”

Quantitating Glomerular Endothelial Fenestration: An Unbiased Stereological Approach

“…Podocytes are the main source of VEGF in glomeruli [14]. Glomerular endothelial cells express VEGF receptors (VEGFR) 2 and VEGFR3 [12,15]. Reduced endothelial fenestration is a known finding in conditions with severe endothelial injury, such as preeclampsia and hemolytic uremic syndrome.…”

Nephrotic syndrome unfavorable course correlates with downregulation of podocyte vascular endothelial growth factor receptor (VEGFR)-2

“…The authors suggested there was another receptor binding VEGF-C in podocytes. The same scientific team, but in a different publication, proved VEGF-C to increase glomerular cell monolayer integrity via VEGFR-2, and observed glomerular ECs to express VEGFR-3 in tissue sections and culture [16].…”