Modifications in glomerular polyanion distribution in adriamycin nephrosis

Previous reports have suggested that, in proteinuria induced by adriamycin (ADR), the functional size barrier of the glomerular basement membrane (GBM) is altered as the result of a sieving defect, whereas the functional charge barrier of the glomeruli remains intact. The aim of this study was to reevaluate the effect of ADR on anionic constituents in the glomerular capillary wall (GGW). Kidneys of nephrotic rats, induced by the injection of 7.5 mg/kg ADR, and controls were resected, and cortices were isolated 24 h and 10 days postinjection, fixed with formaldehyde, and embedded in paraffin. For the histochemical evaluation of sialyl residues, deparaffinized sections were treated with biotin-labeled peanut agglutinin (PNA), before or after neuraminidase treatment. PNA binding was visualized by the avidin-biotin-peroxidase complex and interacted with hydrogen peroxide and diaminobenzidine. For electron microscopy, kidney cortices were fixed with glutaraldehyde and embedded in araldite or LR-white. The postembedding localization of anionic sites was carried out by cationic colloidal gold (CCG), directly applied on thin LR-white sections. Although in the 24-h ADR group, kidney functions and glomerular morphology were generally unaltered, the 10-day ADR group exhibited severe proteinuria, hypoalbuminemia, and massive fusion of intercalated foot processes of the podocytes. Intense PNA binding was observed after neuraminidase treatment in the GCW of the controls. This was gradually decreased in the 24-h ADR kidneys and further decreased in the 10-day ADR, indicating a gradual decrease in glomerular sialic acid content. CCG binding in the controls was mainly restricted to the GBM, exhibiting a threefold higher concentration at the lamina rara externa as compared with the lamina rara interna. In the 24-h ADR, the CCG binding was practically similar to that of the controls, whereas in the 10-day ADR, there was a drastic reduction in CCG binding. The results indicate that ADR-induced nephrosis is associated with a reduction in the content and/or expression of the polyanionic constituents of the GCW, including sialoglycoconjugates ot the podocytes and sulfated proteoglycans of the GBM.