Urine Of Nephrotic Patients Research Articles

Rituximab as Initial Therapy in Adult Patients With Minimal Change Disease

Rituximab as Initial Therapy in Adult Patients With Minimal Change Disease

Proteolytic activation of the epithelial sodium channel (ENaC) by factor VII activating protease (FSAP) and its relevance for sodium retention in nephrotic mice

Proteolytic activation of the epithelial sodium channel (ENaC) by aberrantly filtered serine proteases is thought to contribute to renal sodium retention in nephrotic syndrome. However, the identity of the responsible proteases remains elusive. This study evaluated factor VII activating protease (FSAP) as a candidate in this context. We analyzed FSAP in the urine of patients with nephrotic syndrome and nephrotic mice and investigated its ability to activate human ENaC expressed in Xenopus laevis oocytes. Moreover, we studied sodium retention in FSAP-deficient mice (Habp2−/−) with experimental nephrotic syndrome induced by doxorubicin. In urine samples from nephrotic humans, high concentrations of FSAP were detected both as zymogen and in its active state. Recombinant serine protease domain of FSAP stimulated ENaC-mediated whole-cell currents in a time- and concentration-dependent manner. Mutating the putative prostasin cleavage site in γ-ENaC (γRKRK178AAAA) prevented channel stimulation by the serine protease domain of FSAP. In a mouse model for nephrotic syndrome, active FSAP was present in nephrotic urine of Habp2+/+ but not of Habp2−/− mice. However, Habp2−/− mice were not protected from sodium retention compared to nephrotic Habp2+/+ mice. Western blot analysis revealed that in nephrotic Habp2−/− mice, proteolytic cleavage of α- and γ-ENaC was similar to that in nephrotic Habp2+/+ animals. In conclusion, active FSAP is excreted in the urine of nephrotic patients and mice and activates ENaC in vitro involving the putative prostasin cleavage site of γ-ENaC. However, endogenous FSAP is not essential for sodium retention in nephrotic mice.

Proteomic analysis of urine from proteinuric patients shows a proteolitic activity directed against albumin

Nephrotic syndrome is a condition that is clinically associated with poor outcome. In this study, we compared different techniques of urine sample preparation in order to develop a robust analytical protocol to define the differential urinary proteome of urinary abnormalities compared to nephrotic proteinuria. We recruited 5 normal control subjects, 16 patients with urinary abnormalities and 16 patients with nephrotic syndrome. Proteins from normal urine were processed using three different protocols [acetone, ultrafiltration and trichloroacetic acid (TCA) precipitation], depletion of albumin and IgGs and then analysed by two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) gels and mass spectrometry. Comparing the three extraction methods by visual inspection of gels after 2D gel electrophoresis, the acetone precipitation and TCA methods yielded the best quality of protein extraction, while the acetone precipitation method was the most efficient. Furthermore, we tested three commercial kits for albumin and IgG depletion. We applied the optimized acetone extraction protocol to compare urinary samples from nephrotic patients (NP) to urinary samples obtained from patients presenting with urinary abnormalities (UAP). We observed a proteolytic activity directed against albumin. This observation was more prevalent in urinary samples from NP than from UAP. Within both groups, there was some inter-individual variability in the observed proteolytic activity. An increased concentration of alpha1 antitrypsin was also observed in urine of NP. We analysed albumin fragmentation by 1D and 2D western blots in the same samples skipping the albumin and IgG depletion steps to avoid the possible confound of albumin fragment removal. The analysis confirmed a stronger proteolytic activity in the nephrotic group. The proteolytic activity against albumin and the anti-proteolytic activity of alpha1 antitrypsin are likely linked and could play an important role in the nephrotic process. If replicated in larger samples, this methodology may lead to a better understanding of the underlying pathophysiological process of nephrotic syndrome.

Elevated Interleukin-18 Levels in the Urine of Nephrotic Patients

Background/Aim: The etiology of minimal-change nephritic syndrome (MCNS) is obscure. It has been speculated that T cells play a role in the pathogenesis of MCNS. Interleukin (IL)-18, a novel immunoregulatory cytokine with potent inferon-γ-inducing activities, may play an important role in T-helper type 1-mediated immune responses. To examine further the possible role of IL-18 in nephrotic syndrome (NS), in the present study we measured IL-18 levels in the urine in different clinical stages of MCNS. The aim of this study was to investigate the involvement of IL-18 in MCNS. Methods: Urine samples were obtained from 20 MCNS patients. The disease controls included 20 patients with IgA nephropathy. The samples were assayed for IL-18 protein by a sandwich enzyme-linked immunosorbent assay. Results: Compared with normal controls, significantly increased urinary levels of IL-18 were detected in MCNS patients with the NS. The urinary IL-18 (uIL-18) levels correlated with the degree of proteinuria in MCNS patients. Moreover, when individual MCNS patients were followed through their clinical illness, uIL-18 levels were increased during the active phase and decreased as the patients went into remission. Conclusions: These results indicate that uIL- 18 is detectable in a subgroup of patients with active NS and correlates to their disease activity in patients with MCNS. Our findings support the notion that IL-18 may play a role in the pathophysiology of NS.

Soluble CD14 from urine copurifies with a potent inducer of cytokines.

Here we report that soluble CD14 isolated from the urine of nephrotic patients (uCD14) contains a potent cytokine inducing activity. CD14 derived from urine appeared to consist of two major polypeptides of about 54 and 48 kDa. In uCD14 isolated from three different nephrotic patients the cytokine-inducing activity appeared to co-migrate with the 48-kDa polypeptide which upon sequencing had the same N-terminal sequence as native CD14. Treatment of human monocytes and the human astrocytoma cell line U373 with uCD14 resulted in a strong secretion of tumor necrosis factor (TNF) and interleukin-6, respectively. The cytokine-inducing activity of the uCD14 preparations was unaffected by the absence of serum. This is in contrast to the activation of human monocytes and U373 cells by lipopolysaccharide (LPS) which is highly dependent on the presence of serum. The cytokine-inducing activity was not affected by LPS-binding protein (LBP) or polyclonal rabbit antibodies against LBP. The TNF-inducing activity of uCD14 was also heat labile in contrast to the cytokine-inducing activity of LPS, which was relatively heat resistant. The results suggest that CD14 may exist in at least two forms of which one is involved in cytokine induction.

Blood Coagulation and the Nephrotic Syndrome: Deficiency or Excess?

Normal hemostasis can be disrupted in patients with nephrotic syndrome. More commonly a thrombotic diathesis is seen but there are also sporadic reports of abnormal bleeding. These abnormalities ap...

Insulinlike growth factors in patients with active nephrotic syndrome.

The serum concentrations of insulinlike growth factors 1 and 2 (IGF-1 and IGF-2) were measured by specific radioimmunoassays in 25 nephrotic patients. The serum concentration of IGF-1 in nephrotic patients (mean +/- SEM, 169 +/- 17 ng/mL) was significantly lower than that observed in 20 control subjects matched for sex and age (338 +/- 36 ng/mL). The IGF-2 serum concentration was significantly lower in nephrotic patients (343 +/- 22 ng/mL) than in control subjects (898 +/- 80 ng/mL). The IGF-1 and IGF-2 150-kd and 45-kd carrier protein complexes were found in the urine of nephrotic patients, whereas there was no binding of radiolabeled IGF-1 or IGF-2 to IGF carrier proteins in the urine of control subjects. The low serum IGF-1 and IGF-2 levels observed in nephrotic patients could be partially due to the increased urinary losses of the IGF carrier proteins.

Pneumococcal polysaccharide immunization in patients with active nephrotic syndrome.

We have studied the IgG and IgM antibody responses to pneumococcal polysaccharides types 3 and 19 in eleven patients with active nephrotic syndrome and fifteen normal adult controls. Immunization in both patients with nephrotic syndrome and normal subjects resulted in significant increases in serum IgM antibody to types 3 and 19 pneumococcal polysaccharides. While controls also had a significant rise in IgG antibody to both types 3 and 19, patients with nephrotic syndrome had an increase in IgG antibody only to type 3. Also, after immunization, geometric mean titers of IgG antibodies to both types 3 and 19 were lower in patients with active nephrotic syndrome than in normal subjects (p less than 0.001). In patients with nephrotic syndrome, a significant correlation was found between serum albumin level and type 3 (p less than 0.01) and type 19 (p less than 0.05) antipneumococcal IgG antibody concentration. Furthermore, antipneumococcal IgG antibody was found in the urine of nephrotic patients, while no IgM antibody was excreted in the urine of nephrotic and adult controls. Our data demonstrate that patients with active nephrotic syndrome are able to mount a normal IgM immune response to pneumococcal polysaccharide antigens. The low IgG antibody levels are likely due to increased urinary losses and/or to a partial inability of these patients to produce IgG antibodies. Moreover, since protection may depend on antipneumococcal IgG antibody, these data raise questions as to the benefits of pneumococcal vaccination in patients with active nephrotic syndrome.

A simultaneous study of the polymorphism of five proteins in the serum and the urine of nephrotic patients.

The phenotypes of the haptoglobin (Hp), ceruloplasmin (Cp), group-specific component (Gc), transferrin (Tf), and third component of complement (C3) were determined simultaneously in the serum and urine of patients with proteinuria secondary to nephrotic syndrome of various types. In a large number of cases the patterns of Hp, Cp, and C3 phenotypes in the urine showed marked deviations from those in the corresponding serum either in the mobility or the number of their electrophoretic bands. The monomeric Hp and the Cp were found to have a very augmented urine/serum ration in some cases. Such differences were not detected in the electrophoretic appearance of the Gc and Tf phenotypes. Our results imply that in the proteinuria of the nephrotic syndrome, factors other than molecular weight interfere in the passage of proteins through the glomerular wall.

Urinary excretion of 25-hydroxyvitamin D in health and the nephrotic syndrome.

Patients with the nephrotic syndrome may exhibit low serum 25-OH-D concentrations. We developed a method for isolation of 25-OH-D from urine, with measurement by competitive binding assay. Daily urinary 25-OH-D excretion in healthy subjects averaged 0.17 +/- 0.15 nmol/day. Among nephrotic patients, urinary 25-OH-D excretion ranged from 0.27 to 10 nmol/day, in direct relation to the severity of proteinuria (r=0.76) and averaged 3.7 +/- 3.5 nmol/day. The 25-OH-D in the urine of nephrotic patients was unconjugated, implying that it was excreted with the serum VDBG, which has been shown to have a molecular weight and isoelectric point similar to that of albumin. We conclude that low serum 25-OH-D concentrations among nephrotic patients are principally the result or urinary losses of steroid. (J Lab Clin Med 99:325, 1982.)

A clinical method for the determination of human albumin by means of a precipitin reaction.

Abstract The results of routine estimation of human albumin by different methods were compared. It was found that the immunologic method appeared to be superior to the chemical methods and gave reliable results with sera containing different amounts of albumin. Because of the minute quantity of albumin required for each analysis and the specificity of the test, the immunologic method also has been found useful to estimate the excretion of albumin in the urine of nephrotic patients and to determine the amount of albumin in the spinal fluid. 9