HYDROXYL RADICALS DEPOLYMERIZE GLOMERULAR HEPARAN-SULFATE IN-VITRO AND IN EXPERIMENTAL NEPHROTIC SYNDROME

Heparan sulfate, the polysaccharide side chain of heparan sulfate prot eoglycan, is important for the permselective properties of the glomeru lar basement-membrane, In this report, we show a role for hydroxyl rad icals in heparan sulfate degradation and an enhanced glomerular baseme nt membrane permeability, First, in enzyme-linked immunosorbent assay, exposure of coated heparan sulfate (proteoglycan) to reactive oxygen species resulted in a +/-50% decrease of binding of a monoclonal antib ody against heparan sulfate, whereas binding of an antibody against th e cope protein remained unaltered, Second, on polyacrylamide gel elect rophoresis, the molecular weight of heparan sulfate exposed to radical s was reduced which indicates depolymepization. Both in enzyme-linked immunosorbent assay and gel electrophoresis, hydroxyl radicals are ins trumental for heparan sulfate degradation as shown by the addition of various radical scavengers, Third, in an experimental model for human nephpotic syndrome (Adriamycin nephropathy in rats), glomerular baseme nt membrane staining of two recently described anti-heparan sulfate an tibodies (JM403 and KJ865) was reduced by 24 and 43%, Treatment of Adr iamycin-exposed rats with the hydroxyl radical scavenger dimethylthiou rea both reduced albuminuria by 37% (p < 0.01) and partly prevented lo ss of heparan sulfate staining by 53% (JM403) and 39% (KJ865) (p < 0.0 3) In contrast to the heparan sulfate side chains, the core protein ex pression and the extent of glycanation did not change in Adriamycin ne phropathy, We conclude that glomerular basement membrane heparan sulfa te is susceptible to depolymerization hydroxyl radicals leading to los s of glomerular basement membrane integrity and albuminuria.