Polycation binding to glomerular basement membrane. Effect of biochemical modification.
The polycation hexadimethrine (HDM) binds to anionic sites in the glomerular basement membrane (GBM) and causes heavy proteinuria when infused in vivo. An in vitro assay of 3H-HDM binding to isolated dog GBM was developed, to permit further analysis of the GBM components binding HDM. 3H-HDM binding to isolated GBM was saturable, reversible in dose-dependent fashion by competing polycations, and inhibited by increasing salt concentration and low pH. The pH dependence of binding suggested that most of the HDM binds to carboxyl groups rather than to the sulfate groups of proteoglycans. Removal of heparan sulfate by heparinase or purified heparatinase had no detectable effect on HDM binding. Treatment of GBM with neuraminidase, hyaluronidase, or chondroitinase reduced binding of HDM by a maximum of 20 to 38%. However, substitution of carboxyl anions with nonionizable glycine methyl ester residues resulted in complete elimination of HDM binding. Parallel results were obtained in studies of glomerular localization of cationized ferritin (CatF), pI 8.5. After carboxyl substitution, GBM did not bind CatF; heparinase-treated GBM bound CatF in a distribution not demonstrably different from normal. Cellulose acetate electrophoresis of glycosaminoglycan fractions prepared from treated GBM confirmed that carboxyl modification did not alter the content or charge of the heparan sulfate of GBM, but heparinase treatment removed at least 90% of heparan sulfate. The results indicate that carboxyl groups are quantitatively more important than heparan sulfate for binding of HDM in vitro. Since HDM causes proteinuria in vivo, carboxyl groups may be important for maintenance of normal permselectivity.
D000818:Animals; D001485:Basement Membrane / Q000378:metabolism* / Q000648:ultrastructure; D004285:Dogs; D006025:Glycosaminoglycans / Q000378:metabolism*; D006583:Hexadimethrine Bromide / Q000378:metabolism* / Q000494:pharmacology; D007672:Kidney Cortex / Q000378:metabolism*; D007678:Kidney Glomerulus / Q000378:metabolism* / Q000648:ultrastructure; D007700:Kinetics; D008854:Microscopy, Electron; D009994:Osmolar Concentration; D011073:Polyamines / Q000378:metabolism*; D014316:Tritium
D006025:Glycosaminoglycans; D011073:Polyamines; D014316:Tritium; D006583:Hexadimethrine Bromide