Analysis of Glomerular Anionic Charge Status in Children with IgA Nephropathy Using Confocal Laser Scanning Microscopy

Abstract

Background: The proteinuria resulting from IgA nephropathy is secondary to altered charge-selective and/or size-selective properties of the glomerular capillary walls. However, the functional changes occurring within the glomerular capillary network which lead to proteinuria are still poorly understood. In this study, we analyzed the participation of charged components in the glomerular capillary and their role with respect to proteinuria in childhood IgA nephropathy. Methods: We examined glomerular anionic charge in renal biopsy specimens with confocal laser scanning microscopy using FITC-conjugated poly-L-lysine as a cationic tracer. The renal specimens investigated were from 9 children with IgA nephropathy (IgAN(+)) associated with detectable proteinuria in a morning urine specimen, 8 children with IgA nephropathy (IgAN(–)) and 11 children with thin basement membrane disease (TBMD) with no detectable proteinuria. Results: The labeling intensity of glomerular fixed anionic sites from IgAN(+) was significantly lower than that of IgAN(–) and TBMD. Staining the serial sections following methylation treatment revealed that the labeling intensity for fixed anionic sites in TBMD was significantly higher than that of both IgAN(+) and IgAN(–). On the other hand, saponification treatment resulted in significantly more intensive fluorescence in TBMD and IgAN(–) than in IgAN(+). Furthermore, statistical analysis demonstrated a significant correlation between 24-hour protein excretion and the intensity of fixed anionic sites in all patients with IgA nephropathy at pH 7.0 and following staining with saponification treatment. Conclusions: These findings suggest that a reduction of glomerular anionic charge might be causally associated with the development of proteinuria in childhood IgA nephropathy. Furthermore, sulfate components such as heparan sulfate proteoglycan might be involved initially followed by carboxyl components such as sialoglycoproteins in the glomeruli of patients with IgA nephropathy contributing to the occurrence of proteinuria. We suggest that this method represents a straightforward approach to dissect the participation of charged components in the pathogenesis of childhood IgA nephropathy and their relationship to the development of glomerular proteinuria.

Reevaluation of glomerular charge selective protein-sieving function

“…Independent verification is difficult, but the CSI in Dent disease might be approximately equal to the Donnan equilibrium value of physiological GBM [10], thereby reflecting the estimated normal charge barrier function. Our findings support the results of past studies, which showed a decrease in the negative charge in GBM [12,13] and a change in morphological GBM [14,15], deduced from evidence obtained by the loss of cationic marker staining in the GBM in nephrotic syndrome.…”

Glycosylation Profile of Differently Charged IgA1 and Their Binding Characteristics to Cultured Mesangial Cells in IgA Nephropathy

“…They further showed that serum IgA from IgAN patients preferentially binds to cationic polypetides and suggested that negatively charged IgA may play a role in the recurrence of mesangial IgA deposition [11]. A recent study demonstrated that there was a reduction of GBM anionic sites in childhood IgAN [12], and the implication of this change of charge in selectivity barrier in the deposition of IgA in the mesangium remains to be investigated. The electrostatic charge of immune complexes is important in the mesangial binding as revealed in animal experiments demonstrating the preferential mesangial deposition of immune complexes of anionic charge [7].…”

Sulphur anion metabolism in Rett syndrome patients: A pilot study

“…Several studies in man have shown that heparan sulphates (HS) are the anionic polysaccharide side chains of the proteoglycans present in renal basement membranes. This sulphation is critically important for retention of protein as lower renal endothelial HS is linked with proteinuria and heparinases increase proteinuria [24,25]; there is an inverse correlation between HS stain- ing in the glomerular basement membrane and urinary protein loss [26,27]. It seems probable therefore that the protein/peptiduria in the RS patients in our study is secondary to loss of sulphate and therefore lower renal HS; general kidney dysfunction as an explanation for the results appears less feasible as, unlike the other anions, thiocyanate excretion is reduced.…”