F. Shimizu

Suppressive Effect of Superoxide Dismutase on Adriamycin Nephropathy

A single intravenous injection of adriamycin (ADR) results in marked proteinuria and glomerular morphological changes that are similar to minimal-change disease in humans. We examined the effect of superoxide dismutase (SOD) on ADR-induced proteinuria. ADR in a dose of 7.5 mg/kg body weight significantly increased urinary protein by day 14; proteinuria rapidly increased thereafter. Concurrent administration of SOD (50 mg/kg) over 30 min prior to and 30 min following ADR injection markedly reduced proteinuria. Twenty-one days after the treatment with SOD, the amount of urinary protein was 108.6 +/- 43.1 mg/24 h in the experimental animals, while it was 221.6 +/- 102.9 mg/24 h in the ADR control group (p less than 0.05). There were also less severe glomerular morphologic changes in the SOD group versus ADR controls. The protective effects of SOD provide indirect evidence that oxygen free radicals are important mediators of ADR-induced proteinuria.

Mesangial sclerotic change with persistent proteinuria in rats after two consecutive injections of monoclonal antibody 1-22-3.

Irreversible mesangial changes with persistenl proteinuria were induced in rats given two consecutive injections 2 weeks apart of a MoAb 1–22–3 to rat mesangial cell. The characteristics of the resulting lesions were investigated and compared with those of the reversible change induced by a single injection. At 24 h after the second injection, mesangiolytic changes similar to those after a single injection were evident. The accumulation of macrophagc‐like cells in glomeruli observed at I week after the first injection was not evident during the experimental period after the second injection. Hypercellularity with the characteristics of intrinsic mesangial cell and increased mesangial matrix were already present I week after the second injection. And mesangial sclerotic change progressed up to 6 months. Deposition of collagen type I and type III and accumulation of collagen fibril at the ullraslructural level were evident in rats 6 months after the second injection. Proteinuria started immediately and continued for more than 6 months after the second injection. The mesangial sclerotic change with persistent proteinuria described here is considered to be a better model for investigating the mechanism of chronic progression of human mesangial proliferative glomerulo‐nephritis.

Quantitative studies of monoclonal antibody 5-1-6-induced proteinuric state in rats

Murine monoclonal antibody (MoAb) 5‐1‐6 was already reported to bind to epithelial cell fool processes and to cause proteinuria in rats. In vivo kinetics of the injected MoAb 5‐1‐6, relationship between the quantity of kidney‐binding antibody and proteinuria. and changes in the amount of antigenic molecule recognized by this MoAb in the proteinuric state were studied. The amount of total kidney‐binding antibody (TK Ab) as determined I h after a 2 mg administration was 50.8 ± 10.4 μg/2 kidneys, and TKAb declined to 1.9 ± 0.4 at day 15. The minimum dose of MoAb required to induce proteinuria was 125 μg as the injected dose. This dose corresponded to 12‐8 μg of TKAb at I h and 0.34 μg of TKAb at day 5. The amount of MoAb 5‐1 ‐6 binding to isolated normal glomeruli was also shown to exceed 147‐7 μg/76000 glomeruli, indicating proteinuria to be induced provided more than 8.7% (= 12.8/147.7) of the critical epitopes is specifically occupied by MoAb. The total amount of MoAb 5‐1‐6 bound to glomeruli in vivo and in vitro was assayed with [125I]‐labelled anti‐mouse IgG. The amount of [125I] anti‐mouse IgG bound to glomeruli was 6.93 ± 0.45 μg/10 000 glomeruli from rat 5 days after this MoAb injection and 26.58 ± 0.66 μg/10000 control glomeruli, indicating the decrease in the number of MoAb 5‐1‐6‐rccognizcd antigen molecules in glomeruli isolated from the rat in proteinuric state induced by this MoAb. Thus, the MoAb 5‐1‐6‐recognized molecule itself may principally function to regulate the permeability of the glomerular capillary wall and the decrease of the molecule may lead to proteinuria.

Quantitative study of mesangial injury with proteinuria induced by monoclonal antibody 1-22-3

Murine MoAb 1‐22‐3 has already been reported to bind to the mesangial cell surface and to cause transient proteinuria and mesangial morphological changes characterized by mesangiolysis. subsequent mesangial cell proliferation and mesangial matrix increase by a single i.v. injection. In this study, MoAb‐induced glomerulopathy was quantitatively analysed. No correlation between the severity of mesangial morphological changes and the degree of proteinuria was detected (r= 0.190). The minimum dose injected to induce abnormal proteinuria was 25 μg. This dose corresponded to 1.79 μg/2 kidneys 30 min after MoAb injection. The highest average level of proteinuria was observed in rats injected with 500 μg of MoAb, and less proteinuria was observed in rats injected with 10.0, 5.0 and 2.0mg. Although the amounts of kidney‐fixing MoAb and the subsequent deposition of rat C3 in the high‐dose‐injected group were larger than in the 500 ng injected group, the numbers of infiltrating inflammatory cells were the same in both groups. No correlations between the degrees of such mediators and proteinuria were observed.

Genetic Regulation of the Development of Glomerular Sclerotic Lesions in the BUF/Mna Rat

BUF/Mna strain rats spontaneously develop renal glomerular sclerotic lesions (RSL) at a nearly 100% incidence, diagnosed by hyperalbuminuria (greater than 500 mg/dl) and glomerular lesions morphologically resembling one type of human focal glomerular sclerosis (FGS). Genetic segregation of RSL development was studied by crossing the BUF/Mna strain with two other rat strains, WKY/NCrj and ACI/NMs, which were free of RSL. Two autosomal recessive genes in the BUF/Mna rats were found to determine the susceptibility to RSL in both combinations of crosses.

Effect of chlorpromazine on kinetics of injected monoclonal antibody in MoAb-induced glomerular injury.

The effect of chlorpromazine, one of several calmodulin antagonists that inhibit cytoskeletal movement, on the local kinetics of injected proteinuria‐inducing MoAb 5–1–6 was examined to test the hypothesis that proteinuria is inhibited if the antigen recognized by MoAb 5–1–6 or injected MoAb remains on the surface of epithelial foot processes. MoAb 5–1–6 was injected into both chlorpromazine‐treated (5 mg/100 g body weight) and untreated rats. As a positive control for the chlorpromazine treatment, anti‐Fx 1A serum was also injected into other chlorpromazine‐treated and untreated rats. Chlorpromazine inhibited neither the change in localization of injected MoAb 5–1–6 nor proteinuria, although it showed an inhibitory effect on redistribution of immune complex and the fixation of complement in passive Heymann glomerulonephritis induced by injection of anti‐Fx 1A serum. We conclude that the kinetics of bound MoAb 5–1–6 are regulated by a system different from that operating in passive Heymann glomerulonephritis.

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.

Altered localization of antigen recognized by proteinuria-inducing monoclonal antibody in experimental nephrosis

SummaryChange in the localization of the antigen recognized by the proteinuria-inducing monoclonal antibody (MA) 5-1-6 in experimental nephrosis was studied by indirect and biotin-avidin immunofluorescence, and immunoperoxidase at light and electron microscopical levels. The proteinuric state was induced by the administration of the aminonucleoside of puromycin (PAN) or adriamycin. The antigen decreased in quantity and/or its distribution changed with an increase in the amount of protein excreted in both experimental models. Recovery from the alterations observed during the development and proteinuria appeared to occur when PAN-induced proteinuria subsided. This antigenic molecule may thus be essential for maintaining the normal permselectivity of glomerular capillary walls.

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

By immunization with rabbit immunoglobulins and the injection of a subnephritogenic dose of rabbit nephrotoxic serum (NTS), accelerated‐type nephrotoxic serum nephritis (NTN) was induced in heterozygous (rnu/+) rats but not in athymic nude (rnu/rnu) rats. By transferring rat antibody against rabbit immunoglobulins, marked proteinuria was induced also in nude rats (202.0 ± 98.4 mg/ day on day 3) as in rnu/+ rats (122.6 ± 35.3 mg/day on day 3). No marked differences in histological findings could be found between both groups. The most marked increase in the number of intraglomerular infiltrating cells was observed in heterozygous rats indicating that the presence of thymus‐derived cells leads to the accumulation of more cells in glomeruli. We conclude that humoral immunity alone is enough to accelerate the pathogenic mechanism which induces glomerular injury with heavy proteinuria in this model.

Cell and Matrix in Monoclonal Antibody-Induced Mesangiolysis

In order to investigate the mutual relationship between mesangial cells and matrix, we used a monoclonal antibody (mAb) to develop an experimental model in rats, in which the rats have transient but massive proteinuria and mesangiolysis, followed by an increase in cell number and an increase of matrix size in the mesangial area.