Disruption of Glomerular Basement Membrane Charge through Podocyte-Specific Mutation of Agrin Does Not Alter Glomerular Permselectivity

Abstract

Glomerular charge selectivity has been attributed to anionic heparan sulfate proteoglycans (HSPGs) in the glomerular basement membrane (GBM). Agrin is the predominant GBM-HSPG, but evidence that it contributes to the charge barrier is lacking, because newborn agrin-deficient mice die from neuromuscular defects. To study agrin in adult kidney, a new conditional allele was used to generate podocyte-specific knockouts. Mutants were viable and displayed no renal histopathology up to 9 months of age. Perlecan, a HSPG normally confined to the mesangium in mature glomeruli, did not appear in the mutant GBM, which lacked heparan sulfate. Moreover, GBM agrin was found to be derived primarily from podocytes. Polyethyleneimine labeling of fetal kidneys revealed anionic sites along both laminae rarae of the GBM that became most prominent along the subepithelial aspect at maturity; labeling was greatly reduced along the subepithelial aspect in agrin-deficient and conditional knockout mice. Despite this severe charge disruption, the glomerular filtration barrier was not compromised, even when challenged with bovine serum albumin overload. We conclude that agrin is not required for establishment or maintenance of GBM architecture. Although agrin contributes significantly to the anionic charge to the GBM, both it and its charge are not needed for glomerular permselectivity. This calls into question whether charge selectivity is a

Heparanase induces a differential loss of heparan sulphate domains in overt diabetic nephropathy

“…Heparanase transgenic mice with complete loss of glomerular heparan sulphate only developed very mild albuminuria [11]. Furthermore, podocyte-specific agrin knockout mice [36] and podocytespecific exostosis-1 (EXT1) knockout mice [37] have been generated, both of which lack heparan sulphate in the GBM. Despite the absence of heparan sulphate, these mice do not develop proteinuria [36] either under normal conditions or when they are overloaded with albumin.…”

“…Furthermore, podocyte-specific agrin knockout mice [36] and podocytespecific exostosis-1 (EXT1) knockout mice [37] have been generated, both of which lack heparan sulphate in the GBM. Despite the absence of heparan sulphate, these mice do not develop proteinuria [36] either under normal conditions or when they are overloaded with albumin. While these recent data suggest that loss of heparan sulphate in the GBM alone does not lead to proteinuria, it is not clear whether loss of heparan sulphate under pathological conditions such as diabetic nephropathy accelerates or worsens proteinuria.…”

Heparan Sulfate Proteoglycans

“…Agrn Agrin Null allele: embryonic lethal; reduced number, size, and density of postsynaptic acetylcholine receptor aggregates in muscles; abnormal intramuscular nerve branching and presynaptic differentiation (Gautam et al 1996(Gautam et al ,1999; smaller brains (Serpinskaya et al 1999); abnormal development of interneuronal synapses (Gingras et al 2007); increased resistance to excitotoxic injury (Hilgenberg et al 2002); reduced number of cortical presynaptic and postsynaptic specializations (Ksiazek et al 2007). Floxed allele: Inactivation in podocytes does not affect glomerular charge selectivity or glomerular architecture (Harvey et al 2007).…”

Basement Membranes: Cell Scaffoldings and Signaling Platforms

“…The polyanionic heparan sulfate chains have been thought to be responsible for the charge-sieving properties of the glomerulus in which acidic macromolecules are preferentially excluded from urine (Groggel et al 1987;Groggel et al 1988). However, recent genetic evidence in which the principal GBM HSPG agrin was selectively knocked out in mouse podocytes revealed that reduction of GBM heparan sulfates did not alter chargeselectivity (Harvey et al 2007). One possibility is that the glycocalyx of either the endothelium or podocytes is responsible for charge-sieving.…”