Vascular Endothelial Growth Factor-A165b Is Protective and Restores Endothelial Glycocalyx in Diabetic Nephropathy.

Diabetic nephropathy is the leading cause of ESRD in high-income countries and a growing problem across the world. Vascular endothelial growth factor-A (VEGF-A) is thought to be a critical mediator of vascular dysfunction in diabetic nephropathy, yet VEGF-A knockout and overexpression of angiogenic VEGF-A isoforms each worsen diabetic nephropathy. We examined the vasculoprotective effects of the VEGF-A isoform VEGF-A165b in diabetic nephropathy. Renal expression of VEGF-A165b mRNA was upregulated in diabetic individuals with well preserved kidney function, but not in those with progressive disease. Reproducing this VEGF-A165b upregulation in mouse podocytes in vivo prevented functional and histologic abnormalities in diabetic nephropathy. Biweekly systemic injections of recombinant human VEGF-A165b reduced features of diabetic nephropathy when initiated during early or advanced nephropathy in a model of type 1 diabetes and when initiated during early nephropathy in a model of type 2 diabetes. VEGF-A165b normalized glomerular permeability through phosphorylation of VEGF receptor 2 in glomerular endothelial cells, and reversed diabetes-induced damage to the glomerular endothelial glycocalyx. VEGF-A165b also improved the permeability function of isolated diabetic human glomeruli. These results show that VEGF-A165b acts via the endothelium to protect blood vessels and ameliorate diabetic nephropathy.

Matrix metalloproteinase 9-mediated shedding of syndecan 4 in response to tumor necrosis factor α: a contributor to endothelial cell glycocalyx dysfunction.

The endothelial surface glycocalyx is a hydrated mesh in which proteoglycans are prominent. It is damaged in diseases associated with elevated levels of tumor necrosis factor α (TNF-α). We investigated the mechanism of TNF-α-induced disruption of the glomerular endothelial glycocalyx. We used conditionally immortalized human glomerular endothelial cells (GEnCs), quantitative PCR arrays, Western blotting, immunoprecipitation, immunofluorescence, and dot blots to examine the effects of TNF-α. TNF-α induced syndecan 4 (SDC4) mRNA up-regulation by 2.5-fold, whereas cell surface SDC4 and heparan sulfate (HS) were reduced by 36 and 30%, respectively, and SDC4 and sulfated glycosaminoglycan in the culture medium were increased by 52 and 65%, respectively, indicating TNF-α-induced shedding. Small interfering (siRNA) knockdown of SDC4 (by 52%) caused a corresponding loss of cell surface HS of similar magnitude (38%), and immunoprecipitation demonstrated that SDC4 and HS are shed as intact proteoglycan ectodomains. All of the effects of TNF-α on SDC4 and HS were abrogated by the metalloproteinase (MMP) inhibitor batimastat. Also abrogated was the associated 37% increase in albumin passage across GEnC monolayers. Specific MMP9 knockdown by siRNA similarly blocked TNF-α effects. SDC4 is the predominant HS proteoglycan in the GEnC glycocalyx. TNF-α-induced MMP9-mediated shedding of SDC4 is likely to contribute to the endothelial glycocalyx disruption observed in diabetes and inflammatory states.