Mitsumasa Nagase

A randomized open-label comparative study of conventional therapy versus mizoribine onlay therapy in patients with steroid-resistant nephrotic syndrome (postmarketing survey)

BackgroundA previous double-blind 24-week clinical trial of mizoribine (MZ) vs placebo in steroid-resistant primary nephrotic syndrome (SRPNS) showed that MZ was more effective than placebo in reducing the rate of deterioration of renal function. The present study was conducted to evaluate the efficacy and safety of MZ in patients with SRPNS after 2 years’ treatment.MethodsA multicenter randomized open-label controlled trial in patients with SRPNS was conducted as a 2-year prospective postmarketing study.ResultsThere was a significant imbalance in the baseline serum albumin level (s-Alb) between the conventional therapy (CT) and MZ onlay therapy groups. Early dropouts were more frequent in the subset of patients in the CT group having a baseline s-Alb ≤3u2009g/dl. Therefore, the primary analysis (urinary protein level (UP)-improving effect) was performed using a mixed-effects model, with stratification according to the baseline s-Alb value. The analysis revealed that, in the subset of 34 patients with membranous nephropathy (MN) within the stratum of patients with baseline s-Alb ≤3u2009g/dl (n = 52), the rate of change (slope of change in the UP level/month), in terms of the log (UP+0.2), was −0.0577 in those allocated to the MZ group and −0.0227 in those allocated to the CT group (P = 0.058). In the stratum of patients with a baseline s-Alb >3u2009g/dl (n = 97), there were no significant differences in the UP between the two treatment groups. Hence, MZ onlay therapy was not considered to be efficacious in this group of patients. No serious adverse reactions to the drug were observed.ConclusionsThe present study yielded significant results, in that it suggested the possibility that long-term MZ therapy may afford further reduction of the UP, in addition to that obtained following CT, in particular, in MN patients in a severe nephrotic state.

Clinicopathologic Study on Prognostic Markers in IgA Nephropathy

Background/Aim: Few prognostic markers have found general acceptance in IgA nephropathy (IgAN). The aim of the present study was to search for significant predictor(s) at the time of biopsy. Methods: Fifty-five patients with IgAN undergoing evaluation and treatment at our institution were examined regarding clinicopathologic features at the time of renal biopsy and, if possible, at follow-up. Factors predictive of outcome were evaluated. Renal histopathology was quantified using a glomerulosclerosis index (GSI), a tubulointerstitial index (TII), and a crescent index (CI). Results: The serum creatinine concentration (S-Cr) showed positive correlations with proteinuria and serum total cholesterol concentration, as well as with histopathologic findings. Heavy proteinuria (≧3.0 g/24 h) was associated with higher S-Cr and greater severity of pathologic abnormalities than with milder proteinuria. At follow-up, 6 patients progressed to chronic renal insufficiency, in whom the S-Cr increased by at least 50% to reach or exceed 1.5 mg/dl (132.6 µmol/l). By univariate analysis, elevated GSI, TII, and S-Cr, presence of nephrotic syndrome, elevated CI, and elevated total cholesterol were found to be negative predictors, in descending order of odds ratio. In multivariate analysis, however, only TII independently predicted unfavorable outcome. Conclusion: Renal biopsy in IgAN may be the most powerful predictor for renal outcome; an advanced tubulointerstitial lesion is unfavorable.

Protection of α3 integrin-mediated podocyte shape by superoxide dismutase in the puromycin aminonucleoside nephrosis rat

Because reactive oxygen species (ROS) are involved in the development of puromycin aminonucleoside nephrosis (PAN), we examined whether superoxide dismutase (SOD) could ameliorate this condition. Phosphatidyl choline-bound SOD (PC-SOD) has higher affinity for the cell membrane than recombinant human SOD (rhSOD). In this study, PC-SOD had a longer half-life in the circulation and also higher affinity to renal fractions (glomerulus, brush border, and tubulus) than rhSOD. PAN was induced in rats with single injections of puromycin aminonucleoside. Rats were divided into four groups: group P, PAN rats without treatment; group PC-T and group rh-T, PAN rats treated with 30,000 U/kg PC-SOD and rhSOD, respectively; and group C, normal controls. The effect of PC-SOD versus rhSOD on PAN was evaluated by morphological podocyte changes (podocyte density along the GBM) and alpha(3) integrin expression at days 4 and 10. Proteinuria was measured over time until day 14. Distribution and quantitation of alpha(3) integrin were studied by confocal laser scan microscopy. On day 4, glomerular ROS was measured by chemiluminescence without stimulation. PC-SOD decreased proteinuria to the control level, but rhSOD only decreased proteinuria by 31%. PC-SOD significantly improved podocyte density (P < 0.05 versus group P). Total alpha(3) integrin expression decreased in the P and rh-T groups at day 4 and then had recovered by day 10, but the polarity of the site of expression did not recover. PC-T preserved both the amount and polarity of integrin expression on days 4 and 10. PC-SOD significantly suppressed ROS generation in PAN (P < 0.05). These findings suggest that alpha(3) integrin regulates glomerular permeability by maintaining podocyte shape and adhesion, which is disrupted by ROS overproduction.

Increased serum and urinary neopterin in nephrotic syndrome indicate cell-mediated immune dysfunction

T-cell-mediated immune disturbances are likely but not certain to cause the nephrotic syndrome. Because neopterin (NP) production is closely related to activation of cell-mediated immunity, we addressed the question by measuring serum NP concentrations and urinary NP/creatinine (Cr) ratios, as well as by assessing interstitial lymphocyte and monocyte infiltration in the kidney and activation of the same cell types in peripheral blood. Finally, we observed whether urinary NP/Cr ratios in nephrotic syndrome are changed by steroid therapy. Seventy-four patients with primary glomerulonephritis were divided into 4 groups based on presence or absence of nephrotic syndrome and presence or absence of mesangial proliferation and expansion. Serum and urinary NP concentrations were measured chromatographically. Infiltrating cells in the kidney were identified by immunohistochemistry, and activation of peripheral blood cells was examined by fluorescent surface marker antibodies and flow cytometry. Irrespective of the pathohistology of glomeruli, nephrotic groups showed significantly higher urinary NP/Cr ratios and serum NP concentrations. Nephrotic groups also exhibited more activation of T cells in peripheral blood than did nonnephrotic groups or a healthy control group. Serum NP did not correlate with extent of interstitial renal infiltrates. Steroid therapy decreased urinary NP/Cr ratios in steroid-responsive patients, but not in steroid-resistant patients. Increased serum NP concentrations and urinary NP/Cr ratios may reflect disordered cell-mediated immunity in the nephrotic syndrome, irrespective of glomerular histology or interstitial cell infiltration.

In vivo effect of OPC15161, a superoxide scavenger, on anti-Thy1 nephritis

The in vivo effect of 6-(1 H-indol-3-ylmethyl)-5-methoxy-3-(2-methylpropyl)-2(1 H)-pyrazinone, 4-oxide (OPC15161), a superoxide scavenger, was studied in rats with anti-Thy1 nephritis. Rats were divided into 4 groups: G-1, normal control; G-2, anti-Thy1 nephritis; G-3 anti-Thy1 nephritis and treated with OPC15161 (50 mg/kg/day) starting at day 0; and G-4, anti-Thy1 nephritis and treated with OPC15161 starting 3 days before antibody injection. At weeks 2 and 8, rats were killed for morphological study and at week 8 for renal clearance. Results were compared among the 4 groups. OPC15161 suppressed urinary albumin/day. Total glomerular cells, mesangial cells, ED-1-positive cells/glomerulus and glomerular volume all increased and the increases were suppressed by OPC15161. Tubulointerstitial index, assessed by point counting, was improved by OPC15161 (P < 0.05 G-3, 4, vs. G-2, not significant vs. G-1). Glomerular filtration rate decreased in all nephritic animals, but the decrease in renal blood flow was less in the treated groups. These findings indicate a favorable effect of OPC15161 on the glomerular and interstitial lesions of anti-Thy1 nephritis.

Transforming growth factor-β receptors in self-limited vs. chronic progressive nephritis in rats

Increases in transforming growth factor‐β (TGF‐β) expression and extracellular matrix accumulation are transient in acute self‐limited mesangial proliferative glomerulonephritis induced by a single injection of anti‐thymocyte serum (ATS), while these increases persist following repeated injections that produce chronic progressive sclerosing glomerulonephritis with tubulointerstitial lesions. However, little is known about the expression of TGF‐β receptors (TβRs) in cells involved in the proliferative and sclerosing renal lesions. A study of protein and mRNA expression for type I (TβRI), type II (TβRII), and type III (TβRIII) TβR in both forms of nephritis was therefore carried out by immunohistochemistry and in situ hybridization. Inhibition of cell proliferation and stimulation of matrix production by TGF‐β1 were assessed in isolated glomeruli using [3H]thymidine incorporation and [3H]proline metabolic labelling, respectively. In acute self‐limited nephritis, expression of TβRI, TβRII, and TβRIII increased in the glomerular and Bowmans capsular epithelial cells comprising the glomerular tuft adhesions to Bowmans capsules. However, TβRII expression was not prominent in proliferating mesangial cells. Glomeruli isolated from rats with acute self‐limited nephritis at day 7, when mesangial cell proliferation was maximal, were partially resistant to the mitoinhibitory effects of TGF‐β1. In contrast, expression of all three TβRs was elevated in glomerular and tubulointerstitial lesions in chronic progressive nephritis, and glomeruli isolated from rats with chronic progressive nephritis 7 days after the second ATS injection were sensitive to TGF‐β1. These data suggest that distinct cellular responses to TGF‐β1 resulting from differential expression of TβR underlie the difference between acute self‐limited mesangial proliferative and chronic progressive sclerosing ATS nephritis in the development of proliferative and sclerotic renal lesions. Copyright

Membranous Glomerulonephritis Associated with Renal Cell Carcinoma: Failure to Detect a Nephritogenic Tumor Antigen

A 57-year-old man with renal cell carcinoma associated with membranous glomerulonephropathy (MGN) developed a transient amelioration of the nephrotic syndrome after excision of the tumor. We tried to identify a nephritogenic tumor antigen using the immunoblotting technique in this patient with MGN, since previous studies examined the interaction between tumor antigens and IgG eluted from the kidney tissue using immunofluorescence or immunodiffusion techniques, and no studies have identified the specific tumor antigen with the immunoblotting method. In the present study, no significant immunoreactivity was noted between the IgG eluted from renal cortical tissues of the patient and renal cell carcinoma proteins. Further studies are necessary to establish the pathogenic mechanism of MGN associated with malignancy.

Source of Reactive Oxygen Species in Anti-Thy1 Nephritis

In proliferative glomerulonephritis, both macrophages and mesangial cells generate reactive oxygen species (ROS), contributing to the development of glomerular injury. We have attempted to determine which cell produces ROS during anti-Thy1 nephritis (ATN) in rats. The generation of ROS was studied using luminol amplified chemiluminescence (GCL) on isolated glomeruli. Immunohistochemical studies used avidin-biotin complex (ABC) to label macrophages and mesangial cells. Immediately after ATN induction, mesangiolysis and infiltration with ED-1 positive cells (referred to as macrophage) was noted with a peak at day 1. After day 4, mesangial proliferation appeared with a decrease of the ED-1 positive cells and a prominent increase of PCNA positive cells (regarded as mesangial cells). In the early phase of ATN, GCL, reflecting ROS generation, increased along with the appearance of ED-1 positive cells. GCL subsequently decreased as mesangial cells increased. This suggested that macrophage were the principal participants in ROS generation in the early phase of ATN although mesangial cells cannot be completely disregarded in the generation of ROS and development of glomerular injury.

Glomerular handling of immune complex in the acute phase of active in situ immune complex glomerulonephritis employing cationized ferritin in rats : Ultrastructural localization of immune complex, complements and inflammatory cells

Abstractu2002The ultrastructural localization of immune complex (IC) and inflammatory mediator systems in the glomerulus was investigated in active in situ IC glomerulonephritis employing cationized ferritin in rats. Glomerulonephritis was induced by unilateral renal perfusion of cationized ferritin as antigen (Ag) in preimmunized rats, and anti-ferritin antibody (Ab), C3 and the rat C5b–9 complex were localized by means of immunogold electron microscopy. Ag–Ab complexes were initially formed subendothelially, associated with C3, and attracted platelets, polymorphonuclear leucocytes (PMN) and monocytes. Then Ag–Ab complexes, without C3, passed across the glomerular basement membrane to re-aggregate subepithelially accompanied by C3 deposition after 1 day. Ag–Ab complexes without C3 accumulated in the inter-podocyte space within 1 day and were seen in the epithelial cells at 6 h. C5b–9 complexes were found in subepithelial immune deposits and in membrane vesicles of the epithelial cells, but only in very small amounts in subendothelial immune deposits. Accumulated platelets, PMN, and monocyte were in direct contact with endothelial cells or subendothelial IC. PMN and monocytes contained Ag, Ab and C3 in intracytoplasmic vacuoles. Ag–Ab complexes were also found in the mesangial matrix adjacent to the subendothelial region after 2 h and increased slightly in number, with expansion of the mesangial area thereafter. Most ICs formed in the subendothelial space rapidly formed lattices of a size that activated C3 and were then translocated to the subepithelial space. The potential ability of C3 to solubilize ICs in the subendothelial region may be important in this process. Endocytosis of subendothelial ICs by PMN and/or monocytes and the movement of ICs to the mesangial matrix may also contribute to the removal of IC from the subendothelial space.

Nephrotoxic serum nephritis in nude rats: the roles of host immune reactions.

A description is made of renal lesions in rats induced by heterologous (rabbit) nephrotoxic serum with or without subsequent host immune reactions against it and the effects of immune reactions on the course of classical nephrotoxic serum (Masugi) nephritis are discussed. The disease was induced by injecting congenitally athymic ACI nude rats (rnu/rnu) and their normal heterozygous littermates (rnu/+) with rabbit anti‐rat glomerular basement membrane (GBM) antiserum. In the autologous phase, rat IgG and immunoglobulins were localized in a linear pattern along capillary walls only in nephritic heterozygous rats. In the indirect plaque‐forming cell (PFC) assay against rabbit immunoglobulins in the autologous phase, significantly more PFC could be detected in nephritic heterozygous rats than in nephritic nude rats. The nude and heterozygous rats were essentially the same with respect to the amount of urinary protein, histological change and clinical course. At least in classical nephrotoxic serum nephritis in rats, host immune reactions against GBM bound heterologous nephrotoxic serum were concluded to have no effect on the course of the disease.