title: Glomerular endothelial glycocalyx constitutes a barrier to protein permeability.
abstract: Glycocalyx, composed of glycoproteins including proteoglycans, coats the luminal surface of the glomerular capillaries. Human heparanase degrades heparan sulphate glycosaminoglycans and is up-regulated in proteinuric states. In this study, we analyze the structure of the human glomerular endothelial cell glycocalyx in vitro and examine its functional relevance, especially after treatment with human heparanase. Electron microscopy of conditionally immortalized glomerular endothelial cells revealed a 200-nm thick glycocalyx over the plasma membrane, which was also demonstrated by confocal microscopy. Neuraminidase treatment removed the majority of glycocalyx, reduced trans-endothelial electrical resistance by 59%, and increased albumin flux by 207%. Heparinase III and human heparanase specifically cleaved heparan sulphate: this caused no change in trans-endothelial electrical resistance, but increased the albumin passage across the monolayers by 40% and 39%, respectively. Therefore, we have characterized the glomerular endothelial cell glycocalyx and have shown that it contributes to the barrier to flux of albumin across the cell layer. These results suggest an important role for this glycocalyx in the restriction of glomerular protein passage in vivo and suggest ways in which human heparanase levels may be linked to proteinuria in clinical disease.
authors.affiliation: Academic Renal Unit, Southmead Hospital, Paul O'Gorman Lifeline Centre, Clinical Sciences at North Bristol, University of Bristol, Bristol, UK. Anurag.Singh@bristol.ac.uk
authors.forename: Anurag
authors.initials: A
authors.lastname: Singh
authors.forename: Simon C
authors.initials: SC
authors.lastname: Satchell
authors.forename: Chris R
authors.initials: CR
authors.lastname: Neal
authors.forename: Edward A
authors.initials: EA
authors.lastname: McKenzie
authors.forename: John E
authors.initials: JE
authors.lastname: Tooke
authors.forename: Peter W
authors.initials: PW
authors.lastname: Mathieson
mesh_terms: D018929:Cell Culture Techniques; D002460:Cell Line; D017097:Electric Impedance; D042783:Endothelial Cells / Q000187:drug effects / Q000502:physiology*; D005966:Glucuronidase / Q000494:pharmacology; D019276:Glycocalyx / Q000187:drug effects / Q000502:physiology* / Q000648:ultrastructure*; D019812:Heparan Sulfate Proteoglycans / Q000502:physiology; D006801:Humans; D007678:Kidney Glomerulus / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D046529:Microscopy, Electron, Transmission; D009439:Neuraminidase / Q000494:pharmacology; D010539:Permeability; D011133:Polysaccharide-Lyases / Q000494:pharmacology; D011507:Proteinuria / Q000209:etiology; D012710:Serum Albumin, Bovine / Q000493:pharmacokinetics*; D000099175:Heparanase
chemical_list: D005966:Glucuronidase; D019812:Heparan Sulfate Proteoglycans; D009439:Neuraminidase; D011133:Polysaccharide-Lyases; D012710:Serum Albumin, Bovine; D000099175:Heparanase; C021349:heparitinsulfate lyase