REDUCTION OF HEPARAN SULFATE-ASSOCIATED ANIONIC SITES IN THE GLOMERULAR-BASEMENT-MEMBRANE OF RATS WITH STREPTOZOTOCIN-INDUCED DIABETIC NEPHROPATHY

Heparan sulphate-associated anionic sites in the glomerular basement m embrane were studied in rats 8 months after induction of diabetes by s treptozotocin and in age- and sex-matched control rats, employing the cationic dye cuprolinic blue. Morphometric analysis at the ultrastruct ural level was performed using a computerized image processor. The hep aran sulphate specificity of the cuprolinic blue staining was demonstr ated by glycosaminoglycan-degrading enzymes, showing that pretreatment of the sections with heparitinase abolished all staining, whereas cho ndroitinase ABC had no effect. The majority of anionic sites (74% in d iabetic and 81% in control rats) were found within the lamina rara ext erna of the glomerular basement membrane. A minority of anionic sites were scattered throughout the lamina densa and lamina rara interna, an d were significantly smaller than those in the lamina rara externa of the glomerular basement membrane (p < 0.001. and p < 0.01 for diabetic and control rats, respectively). Diabetic rats progressively develope d albuminuria reaching 40.3 (32.2-62.0) mg/24 h after 8 months in cont rast to the control animals (0.8 (0.2-0.9) mg/24 h, p < 0.002). At the same time, the number of heparan sulphate anionic sites and the total anionic site surface (number of anionic sites x mean anionic site sur face) in the lamina rara externa of the glomerular basement membrane w as reduced by 19% (p < 0.021) and by 26% (p < 0.02), respectively. Num ber and total anionic site surface in the remaining part of the glomer ular basement membrane (lamina densa and lamina rara internal were not significantly changed. We conclude that in streptozotocin-diabetic ra ts with an increased urinary albumin excretion, a reduced heparan sulp hate charge barrier/density is found at the lamina rara externa of the glomerular basement membrane.