Removal of Heparan Sulfate from the Glomerular Basement Membrane Blocks Protein Passage

Abstract

Heparan sulfate (HS) within the glomerular basement membrane (GBM) is thought to play a major role in the charge-selective properties of the glomerular capillary wall. Recent data, however, raise questions regarding the direct role of HS in glomerular filtration. For example, in situ studies suggest that HS may prevent plasma macromolecules from clogging the GBM, keeping it in an "open" state. We evaluated this potential role of HS in vivo by studying the passage of protein through the glomerular capillary wall in the presence and absence of HS. Intravenous administration of neuraminidase removed neuraminic acid-but not HS-from the GBM, and this led to albuminuria. Concomitant removal of HS with heparinase III, confirmed by ultrastructural imaging, prevented the development of albuminuria in response to neuraminidase treatment. Taken together, these results suggest that HS keeps the GBM in an open state, facilitating passage of proteins through the glomerular capillary wall. The glomerular capillary wall, which consists of fenestrated endothelial cells, the glomerular basement membrane (GBM), and podocytes with foot processes interconnected by slit diaphragms, forms the major filtration barrier of the glomerulus. It excludes plasma proteins with the size of albumin (69 kD, 3.6 nm) and larger from its filtrate. 1,2 Next to size, permeability of the glomerular capillary wall is controlled by charge interactions. Neutral and cationic molecules can more easily penetrate the GBM than anionic molecules, which encounter electrostatic repulsion by HS. In addition, the negative charges of the cell coat (glycocalyx) of the endothelial cells and podocytes may play a role in the charge-dependent glomerular filtration. 3,4 HS contains multiple carboxylic groups and negatively charged N-, 2-O-, 6-O-, and 3-O-sulfate groups. 5,6 Because the majority of HS resides in the GBM, it is assumed that GBM HS is responsible for the charge-selective properties of the glomerular capillary wall. 3,7,8 This is supported by a number of studies. First, a reduction in glomerular HS has been described in a number of human kidney diseases 8 -11 and experimental animal models 8,12-14 characterized by proteinuria. Second, intravenous injection of rats with anti-HS antibody JM403 resulted in acute albuminuria. 15 Third, perfusion of isolated rat kidneys or glomeruli with the HS-degrading enzyme heparinase III resulted in penetration of ferritin and bovine serum albumin into the GBM and passage to the urinary space. 16,17 However, recent observations question the direct role of

Diabetes-Induced Reactive Oxygen Species: Mechanism of Their Generation and Role in Renal Injury

“…Moreover, heparan sulfate level of GBM was reported to be reduced in patients with both type 1 and type 2 diabetes showing remarkable proteinuria [37]. However, its restrictive protein filtration role has been debated as many studies found insignificant leakage of protein even after reduction of the heparan sulfate components through downregulation of their contributing proteoglycans agrin/perlecan [38–40]. …”

“…Interestingly, this model also showed increased secession of HS from its core proteoglycan proteins, which is a possible effect of ROS [127]. Growing body of evidences showed that the loss of HS components from GBM is the prominent reason for increased permeability of GBM resulting in proteinuria [127–129] except some contradictions [38–40]. Furthermore, HS is thought to interact with other extracellular matrix proteins of GBM including collagen IV and laminin, thereby maintaining the integrity and stability of the basement membrane.…”

Glomerular Proteinuria: A Complex Interplay Between Unique Players

“…al demonstrated that on removal of glycosaminoglycans in the GBM by perfusion of bacterial glycosaminoglycan-degrading enzymes led to increased passage of labeled bovine serum albumin through the GBM (17). In contrast, removal of the anionic sites in the GBM using heparanase did not result in albuminuria (18). Also, mice that over-expressed the enzyme heparanase, displaying a fivefold reduction of anionic sites in the GBM did not develop albuminuria (19), questioning the importance of heparan sulfates in creating charge selectivity of filtration.…”

Role of the podocyte in proteinuria

“…As the glomerular filter bears fixed negative charge, this effect can be explained by electrostatic repulsion of the anionic macromolecules within the filter meshwork and should be independent of flow. However, discussions about this concept have reemerged in studies by several groups, following charge removal from the GBM using enzymes or homologous recombination in mice [24–26]. These groups demonstrated the surprising fact that removing charge from the GBM does not significantly influence albumin permeability.…”