Loss of heparan sulfate glycosaminoglycan assembly in podocytes does not lead to proteinuria

Abstract

Podocytes synthesize the majority of the glomerular basement membrane components with some contribution from the glomerular capillary endothelial cells. The anionic charge of heparan sulfate proteoglycans is conferred by covalently attached heparan sulfate glycosaminoglycans and these are thought to provide critical charge selectivity to the glomerular basement membrane for ultrafiltration. One key component in herparan sulfate glycosaminoglycan assembly is the Ext1 gene product encoding a subunit of heparan sulfate co-polymerase. Here we knocked out Ext1 gene expression in podocytes halting polymerization of heparin sulfate glycosaminoglycans on the proteoglycan core proteins secreted by podocytes. Glomerular development occurred normally in these knockout animals but changes in podocyte morphology, such as foot process effacement, were seen as early as 1 month after birth. Immunohistochemical analysis showed a significant decrease in heparan sulfate glycosaminoglycans confirmed by ultrastructural studies using polyethyleneimine staining. Despite podocyte abnormalities and loss of heparan sulfate glycosaminoglycans, severe albuminuria did not develop in the knockout mice. We show that the presence of podocyte-secreted heparan sulfate glycosaminoglycans is not absolutely necessary to limit albuminuria suggesting the existence of other mechanisms that limit albuminuria. Heparan sulfate glycosaminoglycans appear to have functions that control podocyte behavior rather than be primarily an ultrafiltration barrier.

Molecular conformation and filtration properties of anionic Ficoll

“…Opposing that coherent body of theory and experiment are other data suggesting that some of the apparent charge selectivity may be artifactual, due to cellular processing of charged solutes (7). More recent papers demonstrate both enhanced transport of anionic tracers compared with neutral ones and in vivo studies in which reducing the charge of the basement membrane gel had negligible effect on permselectivity to albumin (3,6,20,22,35). At least one group, Rippe and colleagues (3), has suggested that enhanced transport of carboxymethylated Ficoll might be explained by conformational changes instead of electrostatics; that is, the modification process by which an anionic charged Ficoll is produced causes the Ficoll to become less compact and more flexible.…”

Where does albuminuria come from in diabetic kidney disease?

“…This may be the result of changes in size selectivity alone without the involvement of charge selectivity. More recent studies by different laboratories have demonstrated that genetic or enzymatic removal of glomerular heparan sulfate does not involve the appearance of albuminuria [23][24][25][26][27].…”

Reduced Sulfation of Chondroitin Sulfate but Not Heparan Sulfate in Kidneys of Diabetic db/db Mice

“…Knock-out of agrin in podocytes did not result in proteinuria (Harvey et al 2007). Also, knocking out the HS polymerase system (exostosin-1, EXT1) in podocytes affected cellular morphology, but was not accompanied by albuminuria (Chen et al 2008). The complexity of this issue has been reviewed (Raats et al 2000;Kanwar et al 2007) and debated (Morita et al 2008) and concerns several factors that contribute to kidney filtration, such as nephrin slits and the glycocalyx on endothelial cells.…”