Previous studies have demonstrated that the heparan sulfate glycosaminoglycans (HS-GAG) have an integral role in the process of glomerular ultrafiltration, showing that removal of HS-GAG compromised the permeaselectivity of the ultrafiltration barrier. Subsequent studies have shown that in diseases, such as diabetes mellitus, the loss of HS-GAG from the glomerular capillary basement membrane (GBM) is associated with the onset of proteinuria and the beginning of prosclerotic changes in the renal glomerulus. Although permeaselectivity of the GBM may be one function of the HS-GAG, data from our laboratory suggests perhaps an even more critical role for GBM HS-GAGs is the control of podocyte behavior. Using Cre-lox technology to selectively delete the EXT1 gene, which encodes for the first enzyme in HS-GAG chain elongation on proteoglycan core proteins, we eliminated the assembly of all HS-GAGs made by podocytes. Conventional wisdom would predict that animals genetically manipulated in this manner should suffer perinatal mortality due to severe, acute proteinuria. However, PEXTKO (Podocyte EXT1 knockout) mice do not develop significant proteinuria. Routine light microscopy studies of young adult PEXTKO animals show no distinct differences in morphology. However at the ultrastructural level, there are significant changes in the glomerulus such as podocyte foot process effacement and enhanced GBM synthesis and turnover. Immunohistochemistry studies indicate that podocytes detach and are lost from the glomeruli of these animals. As the animals age, mild proteinuria does develop. Because of the above data, our hypothesis is that HS-GAGs directly influence podocyte behavior in normal tissues by mediating podocyte-GBM interactions and the loss of these interactions are manifested in diseases that cause the loss of HS-GAG. To test the hypothesis we propose the following specific aims: 1. To investigate the role of HS-GAGs in modulating podocyte-GBM interactions and ultimately glomerular function in developing and adult wild-type mice and PEXTKO mice;2. Investigate the mechanism by which HS-GAG influences podocyte cell behavior using immortalized EXT1 fl/fl and EXT1 fl/+ podocyte cell lines.
Diseases,such as diabetes mellitus, have profound effects on the filtration apparatus of the kidney. The primary manifestation of these diseases is gradual scarring of the filtration apparatus of the kidney, which results in dysfunction and eventual demise of the filtration apparatus. Using a novel animal model which manifests several changes seen in renal diseases we will investigate the mechanism by which glomerular cells interact with the filtration surface.
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|McCarthy, Kevin J; Wassenhove-McCarthy, Deborah J (2012) The glomerular basement membrane as a model system to study the bioactivity of heparan sulfate glycosaminoglycans. Microsc Microanal 18:3-21|
|Chen, Shoujun; Wassenhove-McCarthy, Deborah; Yamaguchi, Yu et al. (2010) Podocytes require the engagement of cell surface heparan sulfate proteoglycans for adhesion to extracellular matrices. Kidney Int 78:1088-99|