heparitin-sulfate and Nephrotic-Syndrome

The highly sulfated domains of heparan sulfate (HS), alias HS S-domains, are made up of repeated trisulfated disaccharide units [iduronic acid (2S)-glucosamine (NS, 6S)] and are selectively remodeled by extracellular endoglucosamine 6-sulfatases (Sulfs). Although HS S-domains are critical for signal transduction of several growth factors, their roles in amyloidoses are not yet fully understood. Herein, we found HS S-domains in the kidney of a patient with transthyretin amyloidosis. In in vitro assays with cells stably expressing human Sulfs, heparin, a structural analog of HS S-domains, promoted aggregation of transthyretin in an HS S-domain-dependent manner. Interactions of cells with transthyretin fibrils and cytotoxicity of these fibrils also depended on HS S-domains at the cell surface. Furthermore, glypican-5, encoded by the susceptibility gene for nephrotic syndrome GPC5, was found to be accumulated in the transthyretin amyloidosis kidney. Our study, thus, provides a novel insight into the pathologic roles of HS S-domains in amyloidoses, and we propose that enzymatic remodeling of HS chains by Sulfs may offer an effective approach to inhibiting formation and cytotoxicity of amyloid fibrils.

Topics: Adult; Aged; Amyloid; Amyloid Neuropathies, Familial; Female; Glypicans; Heparitin Sulfate; Humans; Kidney; Male; Middle Aged; Nephrotic Syndrome; Prealbumin; Sulfotransferases

To investigate the expression of glomerular heparin sulfate (HS) in paediatric patients with minimal change nephritic syndrome (MCNS).. The kidyney tissues were collected by biopsy from 13 paediatric patients with MCNS, while 5 normal renal biopsy samples were used as control. HS in glomeruli was analysed by indirect immunofluorescence staining using four different monoclonal antibodies, Hepss1, 3G10, JM403 and 10E4, which all recognize distinct HS species and each interacts with a specific HS domain. The concentrations of urine heparan sulfate also were measured by enzyme-linked immunosorbent assay (Elisa).. Expression of HS fine domains was aberrant in paediatric patients compared with control subjects. Children with MCNS in replase showed a decreased glomerular expression of 10E4, JM403 and Hepss1 (P < 0.05). The level of urinary HS was significantly increased in peadiatric patients with MCNS when compared with that in control subjects (P < 0.01).. These results suggest that loss of heparan sulphate in renal tissue may play a role in the pathogenesis of MCNS proteinuria.

Topics: Antibodies, Monoclonal; Case-Control Studies; Child; Female; Heparitin Sulfate; Humans; Kidney Glomerulus; Male; Nephrotic Syndrome

Minimal change nephrotic syndrome (MCNS) is the most frequent form of nephrotic syndrome in childhood. In the glomerular basement membrane (GBM) of adult patients with MCNS, a reduced expression of a specific heparan sulphate (HS) domain has been reported. In children with MCNS, urinary activity of the HS-degrading enzyme heparanase was increased. It is, therefore, possible that a decreased GBM HS expression is associated with the pathogenesis of proteinuria in patients with MCNS.. In this study, HS in glomeruli of five adult and six paediatric patients with MCNS were analysed by immunofluorescence staining using four different antibodies, each defining a specific sulphated HS domain. The pediatric patients were subdivided into three groups depending on the presence or absence of podocyte foot process effacement, the level of proteinuria and prednisone administration at the time of the biopsy. In addition, kidneys of rats with adriamycin nephropathy (ADRN), a model for MCNS, were included in the study.. Expression of sulphated HS domains was not aberrant in adult or paediatric patients compared with control subjects. Children with and without proteinuria had the same HS content. In contrast, rats with ADRN showed a decreased glomerular expression of sulphated HS domains.. These results suggest that in patients with MCNS proteinuria is not associated with major changes in glomerular expression of sulphated HS domains.

Topics: Adult; Aged; Animals; Biopsy; Child; Child, Preschool; Doxorubicin; Female; Gene Expression Regulation; Heparitin Sulfate; Humans; Kidney; Kidney Glomerulus; Male; Middle Aged; Models, Biological; Nephrotic Syndrome; Podocytes; Rats; Rats, Wistar

Immune cells express heparanase, an endoglycosidase, able to degrade heparan sulfate glycosaminoglycan (HSGAG) in the glomerular capillary wall (GCW) and potentially induce proteinuria. The aim of this study was to determine whether dysregulated heparanase expression is associated with the heavy proteinuria of childhood steroid-sensitive nephrotic syndrome (SSNS).. Plasma and urinary heparanase activity and peripheral blood mononuclear cell (PBMC) mRNA heparanase levels [real-time polymerase chain reaction (PCR)] were measured in children with SSNS in relapse and remission. Plasma and urinary heparanase activity was determined in adult patients with nephrotic syndrome and in age- and gender-matched controls.. Plasma heparanase activity was reduced in SSNS with relapse (811.2 units) compared to remission (1147.96 units) (P= 0.003) and control subjects (1390.51 units) (P < 0.001). In adult nephrotic syndrome, plasma heparanase activity was significantly lower in patients compared to controls. However, there was no difference between remission and relapse states. In children, urinary heparanase activity/urinary creatinine ratio was highest in SSNS relapse (14.26 units/mg) compared with remission (7.43 units/mg) (P= 0.016) and controls (2.29) (P < 0.001). However, PBMC heparanase mRNA expression was not different between these three groups. In adult nephrotic syndrome, urinary heparanase activity/urinary creatinine levels were lower in both remission and relapse compared to controls and there was no difference between remission and relapse states.. In childhood SSNS, there is a qualitative and quantitative difference in urinary heparanase activity expression that is not paralleled in adult nephrotic syndrome. These data suggest that dysregulated heparanase expression may play a significant role in the pathogenesis of SSNS, possibly through an abnormality in post-translational control of latent heparanase activation.

Topics: Adult; Age Factors; Case-Control Studies; Child; Enzyme Activation; Female; Gene Expression Regulation, Enzymologic; Glucuronidase; Heparitin Sulfate; Humans; Male; Middle Aged; Nephrotic Syndrome; Protein Processing, Post-Translational; Recurrence; RNA, Messenger; Steroids

Lymphocytes are involved in the physiopathologic mechanism of idiopathic nephrotic syndrome (INS). We have recently demonstrated that plasma from patients with INS decreases human glomerular epithelial cell (GEC) glycosaminoglycans (GAGs), particularly heparan sulfates (HS) in vitro. In this study we investigate the effect of peripheral blood lymphocytes (PBL) from INS patients on glomerular cell GAG and HS.. Human GECs were cultured with total peripheral blood mononuclear cells (PBMCs), PBL, and monocytes from patients and controls. The amounts of GAG and HS were assessed using a cationic membrane after metabolic labeling.. In coculture with GECs, mononuclear cells from controls decreased total epithelial cell GAG (-30% with PBMC, P < 0.05; -25% with PBL, P < 0.02; -19% with monocytes, P < 0.05). Particularly HSs were decreased (-36% with PBMC, P < 0.05; -27% with PBL, P < 0.02; and -19% with monocytes, P < 0.05). When GECs were in coculture with PBL from INS patients, the decrease in GAG and HS was significantly greater in comparison to control PBL (-10%, P < 0.02; -10%, P < 0.02, respectively, for GAG and HS). Moreover, supernatants of stimulated PBMCs from patients decreased also GAG and HS in comparison with controls (-13%, P < 0.02; -15%, P < 0.02, respectively, for GAG and HS).. These data provide direct evidence that PBLs from INS patients are able to decrease GEC HS as previously shown with plasma from patients. This might be instrumental in the onset of albuminuria.

Topics: Cells, Cultured; Child; Chondroitin Sulfates; Glycosaminoglycans; Heparitin Sulfate; Humans; Kidney Glomerulus; Lymphocytes; Monocytes; Nephrotic Syndrome; Polymers; Reference Values

In idiopathic nephrotic syndrome (INS), ciclosporin A (CsA) was shown to decrease proteinuria, an effect explained by its immunologic and hemodynamic actions. In order to determine whether CsA could have a direct action on glomerular cells, we studied the effect of CsA on glomerular cells in vitro, particularly on glycosaminoglcycans (GAG) and heparan sulfates (HS) which are decreased in INS patients.. Human glomerular epithelial cells and rat mesangial cells were cultured at various concentrations of CsA. HS were quantified using a cationic membrane after metabolic labeling.. Mesangial cell GAG and HS and epithelial cell HS increased significantly when cells were cultured with CsA. For both cell types this increase was prevailing on the secreted fraction of HS in comparison with the cellular fraction. CsA induced also an increase in cellular cAMP levels, but the effect of CsA was not transduced via a cAMP pathway.. CsA is able to increase glomerular GAG and HS in vitro. As this effect of CsA was the opposite effect on glomerular cells to the effect of plasma from INS patients, we conclude that this direct action of CsA on glomerular cells could explain in part the effect of this drug in decreasing proteinuria in INS.

Topics: Animals; Cells, Cultured; Cyclic AMP; Cyclosporine; Glomerular Mesangium; Glycosaminoglycans; Heparitin Sulfate; Humans; Immunosuppressive Agents; Kidney Glomerulus; Nephrotic Syndrome; Rats; Rats, Sprague-Dawley

The physiopathological mechanisms of idiopathic nephrotic syndrome involve a circulating plasma factor and a decrease in HS in the glomerular basement membrane. Previous studies have demonstrated that plasma from patients with INS decreases glomerular cell HS in vitro. We examined the involvement of cyclic adenosine monophosphate (cAMP) in this interaction. We studied the effect of plasma from patients with INS on mesangial cell cAMP. We also determined mesangial cell HS when cAMP levels were modified using a cationic membrane after metabolic labeling. Cellular cAMP levels increased significantly when mesangial cells were incubated with plasma from patients with INS in comparison with control plasma (+77%, P = 0.01). Forskolin and IBMX, which increased cellular cAMP, decreased HS levels (-21 +/- 9% and -15 +/- 6% respectively, P < 0.05 for both), whereas dideoxyadenosine, which decreased cellular cAMP, increased HS levels (+24 +/- 7%, P < 0.05). Plasma from patients with INS decreased glomerular cell HS in comparison with control plasma (-34 +/- 8%, P < 0,05). This effect was abolished when cells were preincubated with ddAdo to prevent an increase in cAMP levels. We conclude that in mesangial cells, plasma from patients with INS increases cAMP levels, and that cAMP mediates a decrease in HS levels. Moreover, the action of plasma from patients on HS was inhibited when an increase in cAMP was prevented. cAMP may therefore be instrumental in the negative effect of the plasma factor on mesangial cell HS.

Topics: 1-Methyl-3-isobutylxanthine; Adult; Animals; Antimetabolites; Cells, Cultured; Child; Child, Preschool; Colforsin; Creatinine; Cyclic AMP; Dideoxyadenosine; Glomerular Mesangium; Heparitin Sulfate; Humans; Nephrotic Syndrome; Phosphodiesterase Inhibitors; Plasma; Rats; Rats, Sprague-Dawley; Signal Transduction; Statistics, Nonparametric; Urine

Danaparoid sodium is an antithrombin composed of 3 glycosaminoglycans: heparan sulfate, dermatan sulfate and chondroitin sulfate. Similar to heparin, danaparoid operates by activating antithrombin 3, but does not contain heparin or heparin fragments, and is therefore antigenically distinct. Danaparoid has been advocated as a safe and effective anticoagulant for heparin-associated thrombocytopenia. However, there is little experience in its use as a substitute for heparin in hemodialysis. We report 2 men, aged 82 and 73 years, respectively, who developed thrombocytopenia while undergoing hemodialysis with heparin, and who subsequently underwent successful dialysis with danaparoid. There was a rise in platelet levels in both while receiving danaparoid, and dialysis was completed without hemorrhagic or thrombotic complications. Danaparoid is a safe and effective substitute for heparin, and may be used as an anticoagulant in hemodialysis.

Topics: Aged; Aged, 80 and over; Antithrombin III; Chondroitin Sulfates; Dermatan Sulfate; Drug Combinations; Heparin; Heparitin Sulfate; Humans; Kidney Failure, Chronic; Male; Nephrotic Syndrome; Platelet Aggregation; Renal Dialysis; Thrombocytopenia

Heparan sulfate, the polysaccharide side chain of heparan sulfate proteoglycan, is important for the permselective properties of the glomerular basement membrane. In this report, we show a role for hydroxyl radicals in heparan sulfate degradation and an enhanced glomerular basement 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 antibody against heparan sulfate, whereas binding of an antibody against the core protein remained unaltered. Second, on polyacrylamide gel electrophoresis, the molecular weight of heparan sulfate exposed to radicals was reduced which indicates depolymerization. Both in enzyme-linked immunosorbent assay and gel electrophoresis, hydroxyl radicals are instrumental for heparan sulfate degradation as shown by the addition of various radical scavengers. Third, in an experimental model for human nephrotic syndrome (Adriamycin nephropathy in rats), glomerular basement membrane staining of two recently described anti-heparan sulfate antibodies (JM403 and KJ865) was reduced by 24 and 43%. Treatment of Adriamycin-exposed rats with the hydroxyl radical scavenger dimethylthiourea both reduced albuminuria by 37% (p < 0.01) and partly prevented loss of heparan sulfate staining by 53% (JM403) and 39% (KJ865) (p < 0.03). In contrast to the heparan sulfate side chains, the core protein expression and the extent of glycanation did not change in Adriamycin nephropathy. We conclude that glomerular basement membrane heparan sulfate is susceptible to depolymerization by hydroxyl radicals leading to loss of glomerular basement membrane integrity and albuminuria.

Topics: Animals; Antibiotics, Antineoplastic; Biopolymers; Cattle; Doxorubicin; Free Radical Scavengers; Heparitin Sulfate; Humans; Hydroxyl Radical; Kidney Glomerulus; Male; Nephrotic Syndrome; Rats; Rats, Wistar; Reactive Oxygen Species

We report the beneficial effects of heparan sulfate administration in nephrotic syndrome (NS) in reducing the degree of proteinuria, in ameliorating the chylomicron removal defect and the hypercoagulable state and, consequently, in slowing down the progression to uraemia.

Topics: Heparitin Sulfate; Humans; Nephrotic Syndrome

To identify the role of circulating immune factor(s) in the development of proteinuria, the supernatant of peripheral blood mononuclear cells (PBMC) were obtained from 15 patients with steroid-responsive nephrotic syndrome, and its effect on the synthesis of sulfated compound by cultured rat glomerular epithelial cell (GEC) were studied by 35S, 3H-leucine dual-isotope labelling technique. The result showed that the supernatant of PBMC from 9 patients without steroid treatment increased 35S uptake in GEC (P < 0.01), while there was no significant difference in 35S uptake between the 6 patients treated with prednisone (1.5-2.0 mg/kg.d-1) and 6 normal controls.

Topics: Adolescent; Animals; Cells, Cultured; Child; Child, Preschool; Epithelial Cells; Heparan Sulfate Proteoglycans; Heparitin Sulfate; Humans; Kidney Glomerulus; Leukocytes, Mononuclear; Nephrotic Syndrome; Prednisone; Proteoglycans; Rats; Rats, Sprague-Dawley

The congenital nephrotic syndrome of the Finnish type (CNF) is an autosomal recessive disease characterized by massive proteinuria already at birth. The gene locus defective in CNF was searched for using polymorphic markers of candidate genes coding for components of the basement membrane (BM). The linkage analyses in 17 Finnish CNF families demonstrated exclusion of obligatory recombination events between the disease and eight genes coding for BM components. The genes coding for the alpha 1(IV), alpha 2(IV), alpha 3(IV) and alpha 4(IV) chain of type IV collagen, the B1e, B2e and B2t chains of laminin, as well as the BM heparan sulfate proteoglycan core protein were all excluded in this Finnish family material. Since the defect is not in any of the genes coding for major components of BM, the identification of the gene defect will most probably reveal a new gene important for the development and function of the glomerular basement membrane.

Topics: Cell Line; Child, Preschool; Collagen; DNA Mutational Analysis; Fibroblasts; Finland; Genetic Linkage; Heparan Sulfate Proteoglycans; Heparitin Sulfate; Humans; Kidney Glomerulus; Laminin; Nephrotic Syndrome; Pedigree; Polymorphism, Genetic; Proteoglycans

To evaluate the specificity of a raised heparan sulphate (HS) excretion previously reported in four children with congenital nephrotic syndrome (CNS), we measured the urinary excretion of HS and chondroitin sulphate (CS) in seven children with Finnish-type congenital nephrotic syndrome (CNSF), seven with diffuse mesangial sclerosis (DMS), nine with focal segmental glomerulosclerosis (FSGS), 14 with steroid-sensitive nephrotic syndrome of whom eight had a biopsy confirming minimal change histology (SSNS), and 17 controls. The urine HS/CS ratio in normal children had a median of 0.36 (observed range 0.21 to 0.68) and was independent of age. HS/CS ratio was significantly greater than controls in CNSF (median 0.80, range 0.43 to 1.28), DMS (median 0.81, range 0.49 to 1.13) and FSGS children (median 0.66, range 0.38 to 1.6), but was not in SSNS (median 0.44, range 0.28 to 0.70). There was a positive correlation between the HS/CS ratio and urine albumin excretion. High HS/CS ratios are not diagnostic of a particular histological variety of CNS.

Topics: Albuminuria; Child; Child, Preschool; Chondroitin Sulfates; Creatinine; Female; Glomerulosclerosis, Focal Segmental; Glycosaminoglycans; Heparitin Sulfate; Humans; Infant; Male; Nephrosis, Lipoid; Nephrotic Syndrome; Steroids

Studies using cationic probes have suggested that a reduction in glomerular anionic sites, composed principally of the glycosaminoglycan heparan sulphate, is responsible for the abnormal glomerular permeability in the congenital nephrotic syndrome (CNS). We therefore analysed the glycosaminoglycan content of the glomerular basement membrane (GBM) from an infant who died of CNS and from an infant who died of unrelated causes. We also measured the urinary excretion of glycosaminoglycans in children with nephrotic syndrome, both congenital and acquired, and in healthy children. Heparan sulphate constituted 59% of the glycosaminoglycan content of the GBM in the normal infant, the other principal glycosaminoglycan being chondroitin sulphate. In the GBM from the infant with CNS the heparan sulphate was greatly reduced, constituting only 3% of total glycosaminoglycans. The urinary excretion of heparan sulphate was significantly increased in CNS (expressed both in relation to creatinine and to chondroitin sulphate) compared with normal children and to those with acquired nephrotic syndrome. Diminished GBM content of heparan sulphate may be responsible for the abnormal glomerular permeability in CNS and may be a consequence of defective incorporation of heparan sulphate into the GBM with subsequent loss into the urine.

Topics: Basement Membrane; Glycosaminoglycans; Heparitin Sulfate; Humans; Infant; Kidney Glomerulus; Male; Nephrotic Syndrome; Reference Values

Nephrotic patients and rats with experimentally induced nephrotic syndrome have elevated plasma triglycerides and impaired triglyceride removal. This may be due to a defective interaction of chylomicrons and very low density lipoproteins with lipoprotein lipase. Since the glycosaminoglycan, heparan sulfate, was found to stimulate the lipoprotein lipase reaction in vitro, we investigated the plasma heparan sulfate content and measured the urinary excretion of heparan sulfate in control rats and rats with experimentally induced nephrotic syndrome. In addition, we studied the effect of heparan sulfate on the rate of removal of radiolabeled chylomicrons in nephrotic rats. Glycosaminoglycan concentrations in plasma were the same in control and nephrotic rats, although 35S incorporation in high charge glycosaminoglycans was markedly reduced. In addition, in nephrotic rats there is a marked reduction in the urinary excretion of heparan sulfate and chondroitin sulfate suggesting a markedly reduced turnover of these glycosaminoglycans. This was associated with increased plasma triglycerides in nephrotic rats. Nephrotic rats showed a reduced rate of clearance of injected chylomicrons. Intravenous administration of heparan sulfate completely and immediately corrected the chylomicron removal defect. We also noted a log-dose response effect of administered heparan sulfate on chylomicron removal. This effect was not due to a release of soluble lipoprotein lipase by heparan sulfate. These findings suggest that a rapidly turning over fraction of plasma heparan sulfate may play an important role in chylomicron clearance.

Topics: Animals; Chylomicrons; Electrophoresis, Agar Gel; Glycosaminoglycans; Heparitin Sulfate; Lipoprotein Lipase; Male; Nephrotic Syndrome; Rats; Rats, Inbred Strains; Triglycerides

Topics: Child; Chondroitin Sulfates; Diabetes Mellitus; Glomerulonephritis; Glycosaminoglycans; Heparitin Sulfate; Humans; Mucopolysaccharidoses; Nephritis, Hereditary; Nephrotic Syndrome; Proteinuria

The synthesis of intact proteoglycans and their glycosaminoglycan (GAG) side chains by isolated rat glomeruli in vitro were studied both at the onset (5 days) and at the point of maximal proteinuria (7 days) of the nephrotic syndrome induced with the aminonucleoside of puromycin. Glomeruli from nephrotic animals incorporated 1.5-fold and 3.0-fold more 35SO4 label into GAG than glomeruli from control animals at 5 and 7 days, respectively, with heparan-35SO4 GAG being responsible for the majority of the increment. Both nephrotic and control incubations contained 60% of the label in the incubation medium, 40% in the glomerular fractions, and less than 1% in the glomerular basement membrane. Glomerular basement membrane from nephrotic rats had no change in their total heparan-35SO4 GAG content. The majority of intact proteoglycan(s) from the glomerular matrix and from the incubation medium of nephrotic and control animals was found in the most buoyantly dense fraction of CsCl gradients (fraction 1). 35S-labeled material isolated from glomeruli of nephrotic animals showed a consistent shift toward lower density gradient fractions, indicating a decrease in their overall carbohydrate to protein ratio. Diethylaminoethyl chromatography of fraction 1 proteoglycan showed a single biphasic peak with the nephrotic rat having an increase in the proportion contributed by the earlier component of the peak. Fraction 1 proteoglycan(s) from the nephrotic experiment was found to have a smaller average hydrodynamic size by Sepharose CL-2B chromatography without a significant change in the corresponding 35S-GAG chain sizes (molecular weight 14,000) by Sepharose CL-6B chromatography. 35S-macromolecules from glomeruli of nephrotic and control rats that appeared in the middle of the CsCl gradients (fraction 3) had similar Sepharose CL-2B elution volumes, whereas the corresponding 35S-GAG chains from incubations of glomeruli from nephrotic animals were smaller. Increased synthesis of heparan sulfate proteoglycan by glomeruli from puromycin aminonucleoside-induced nephrotic rats may be compensatory to loss of another component of the glomerular filtration barrier or may result from abnormal interaction of proteoglycan(s) from nephrotic animals with other glomerular matrix constituents.

Topics: Animals; Centrifugation, Density Gradient; Chondroitin Sulfate Proteoglycans; Chromatography, Ion Exchange; Glycosaminoglycans; Heparan Sulfate Proteoglycans; Heparitin Sulfate; Kidney Glomerulus; Male; Nephrotic Syndrome; Proteoglycans; Puromycin; Puromycin Aminonucleoside; Rats; Rats, Inbred Strains; Sepharose; Sulfur Radioisotopes

Three basement membrane components, heparan sulfate proteoglycan, laminin, and type IV collagen, were localized in renal glomeruli using light and electron microscopic immunohistochemistry in normal and aminonucleoside-treated nephrotic rats. In normal rats, type IV collagen was found in both laminae rarae and in lamina densa; laminin and heparan sulfate were found in the three layers of the basement membrane with more intense staining in both laminae rarae. In nephrotic rats laminin and type IV collagen had the same localization and distribution as in normal rats. This was in contrast to heparan sulfate proteoglycan which was lost to a considerable extent in nephrotic animals. These results support the concept that heparan sulfate proteoglycan is a major charge determinant in normal glomerular basement membrane and suggest a crucial role for this proteoglycan in the pathogenesis of the nephrotic syndrome.

Topics: Animals; Basement Membrane; Collagen; Female; Glycoproteins; Glycosaminoglycans; Heparitin Sulfate; Immune Sera; Kidney Glomerulus; Laminin; Microscopy, Electron; Nephrotic Syndrome; Rabbits; Rats; Rats, Inbred Strains

Recent work suggests that the normal barrier to penetration of the renal glomerular basement membrane by anionic plasma proteins may depend in part on the existence of negatively charged sites within the membrane. We describe an in vitro cytochemical method for the quantitative demonstration of anionic sites in the normal human glomerular basement membrane. In five normal subjects, ranging in age from 10 days to 57 years, the sites were distributed at regular intervals in the lamina rara externa, with a frequency of 23.8 +/- 6.8 sites per 1000-nm length of membrane. A similar distribution was observed in the basement membranes from three normal human fetuses. Ex vivo perfusion of one cadaver kidney revealed a similar distribution of anionic sites. The number of anionic sites in the glomerular basement membranes of five patients with the congenital nephrotic syndrome was reduced to 8.9 +/- 3.7 (P less than 0.001). Prior incubation of sections of normal kidney in purified heparinase resulted in a marked reduction in the number of anionic sites. We conclude that congenital nephrosis results from failure of heparan sulfate--rich anionic sites to develop in the lamina rara externa of the glomerular basement membrane.

Topics: Anions; Basement Membrane; Child, Preschool; Fetus; Glycosaminoglycans; Heparin Lyase; Heparitin Sulfate; Humans; Infant; Kidney Glomerulus; Nephrotic Syndrome; Perfusion; Polyethyleneimine; Polysaccharide-Lyases; Proteinuria

Previously we found that alpha 2-acid glycoprotein fraction from urine of patients with the nephrotic syndrome stimulated the lipoprotein lipase reaction in vivo and in vitro. The activator was separated from the alpha 1-acid glycoprotein and identified as a glycosaminoglycan. The studies reported here were undertaken to characterize and quantify the glycosaminoglycans contained in urine of patients with the nephrotic syndrome and to compare these to the glycosaminoglycans in urine of the control subjects. We found that free low molecular weight glycosaminoglycans, heparan sulfate and chondroitin 4-sulfate, are excreted in both patients with the nephrotic syndrome and controls however, patients with the nephrotic syndrome excreted much less of both glycosaminoglycans. The free form of heparan sulfate was found to be the activator which stimulated the lipoprotein lipase reaction in vitro in the presence of apolipoprotein CII. In addition, the urine from patients with the nephrotic syndrome contained a protein-glycosaminoglycan complex which was absent in control urine. Glycosaminoglycans in the complex could be released by papain digestion or by trichloroacetic acid. Our evidence indicates that this glycosaminoglycans fraction is a law charge form of chondroitin sulfate.

Topics: Chondroitin Sulfates; Enzyme Activation; Glycosaminoglycans; Heparitin Sulfate; Humans; Lipoprotein Lipase; Molecular Weight; Nephrotic Syndrome