Masato Isome

The arginine metabolite agmatine protects mitochondrial function and confers resistance to cellular apoptosis

Agmatine, an endogenous metabolite of arginine, selectively suppresses growth in cells with high proliferative kinetics, such as transformed cells, through depletion of intracellular polyamine levels. In the present study, we depleted intracellular polyamine content with agmatine to determine if attrition by cell death contributes to the growth-suppressive effects. We did not observe an increase in necrosis, DNA fragmentation, or chromatin condensation in Ha-Ras-transformed NIH-3T3 cells administered agmatine. In response to Ca(2+)-induced oxidative stress in kidney mitochondrial preparations, agmatine demonstrated attributes of a free radical scavenger by protecting against the oxidation of sulfhydryl groups and decreasing hydrogen peroxide content. The functional outcome was a protective effect against Ca(2+)-induced mitochondrial swelling and mitochondrial membrane potential collapse. We also observed decreased expression of proapoptotic Bcl-2 family members and of execution caspase-3, implying antiapoptotic potential. Indeed, we found that apoptosis induced by camptothecin or 5-fluorourocil was attenuated in cells administered agmatine. Agmatine may offer an alternative to the ornithine decarboxylase inhibitor difluoromethyl ornithine for depletion of intracellular polyamine content while avoiding the complications of increasing polyamine import and reducing the intracellular free radical scavenger capacity of polyamines. Depletion of intracellular polyamine content with agmatine suppressed cell growth, yet its antioxidant capacity afforded protection from mitochondrial insult and resistance to cellular apoptosis. These results could explain the beneficial outcomes observed with agmatine in models of injury and disease.

Clinicopathological features and the prognosis of IgA nephropathy in Japanese children on long-term observation.

AIMS Clinicopathological features were investigated to clarify the ultimate prognosis and prognostic indicators for patients with IgA nephropathy in Japanese children. METHODS We evaluated the outcomes of 181 patients in whom IgA nephropathy was diagnosed before the age of 15 years since September 1979 and followed-up at least for three years with regard to clinical data at the onset of symptoms and renal histologic data. RESULTS After mean follow-up of 7.3 years from onset, 91 patients of 181 (50.3%) were in clinical remission at the last examination, 24 (13.2%) had isolated hematuria, 59 (32.6%) had hematuria and proteinuria. Eighteen of 59 (9.9%) had proteinuria more than 1 g per 24 hours. Hypertension was observed in 12 cases and 7 (3.9%) developed end-stage renal disease. Except 7, no patient had reduced renal function and elevated serum creatinine at the final follow-up. Predicted renal survival rate from onset was 92.3% at 10 years and 89.1% at 20 years. In multivariable analysis, age at onset and chronic changes of tubulointerstitium were associated with poor outcome. CONCLUSIONS Of 181 children with IgA nephropathy, 50% regressed, remaining 46% had hematuria and/or proteinuria and 4% of patients lapsed into end-stage renal disease. Our results indicate that childhood IgA nephropathy has a benign course and the risk for end-stage renal disease is lower than that of adults. Age at onset and tubulointerstitial lesions were the strong predictors of a progressive course of childhood IgA nephropathy.

Efficacy of Multidrug Therapy Combined with Mizoribine in Children with Diffuse IgA Nephropathy in Comparison with Multidrug Therapy without Mizoribine and with Methylprednisolone Pulse Therapy

Aim: To evaluate the efficacy of prednisolone, warfarin, and dipyridamole therapy combined with mizoribine (PWDM) in the treatment of diffuse immunoglobulin A (IgA) nephropathy in comparison with prednisolone, warfarin, and dipyridamole therapy without mizoribine (PWD) and with methylprednisolone pulse therapy (PWD pulse). Methods: We collected data on 61 patients diagnosed with diffuse IgA nephropathy, and these patients were retrospectively divided into three groups without randomization. Group A included 21 patients before 1987 who were treated with PWD for 24 months, group B included 20 patients from 1987 to 1989 who were treated with PWD pulse therapy for 24 months, and group C included 20 patients after 1990 who were treated with PWDM for 24 months. Clinical features and pathological findings in each group were analyzed retrospectively. Results: The time from initiation of therapy in group A, group B, and group C was 8.9 ± 5.2, 8.1 ± 3.9, and 7.7 ± 3.8 years, respectively. At the latest follow-up examination, the mean urinary protein excretion (mg/m2/h) was 17 ± 10 in group A, 22 ± 20 in group B, and 6 ± 6 in group C and had decreased significantly in group C as compared with the other groups. The activity index in all three groups was lower at the second biopsy than that at the first biopsy (5.1 ± 0.8 vs. 6.5 ± 2.1 in group A, p < 0.05; 5.6 ± 0.9 vs. 6.6 ± 1.7 in group B, p < 0.01, and 4.5 ± 1.0 vs. 6.8 ± 1.9 in group C, p < 0.01). The chronicity index in groups A and B at second biopsy was higher than at first biopsy (7.3 ± 1.4 vs. 4.8 ± 1.0 in group A, p < 0.01, and 8.1 ± 2.0 vs. 5.3 ± 0.9 in group B, p < 0.01), but was unchanged in group C. At the latest follow-up examination, 1 patient (4.8%) in group A, 3 patients (15%) in group B, and none (0%) in group C had renal insufficiency. Conclusion: These results suggest that PWDM appears to be more effective than PWD or PWD pulse in ameliorating proteinuria and histological severity of patients with IgA nephropathy.

Coxsackievirus and Adenovirus Receptor, a Tight Junction Membrane Protein, Is Expressed in Glomerular Podocytes in the Kidney

In nephrosis, filtration slits of podocytes are greatly narrowed, and slit diaphragms are displaced by junctions with close contact. Freeze-fracture studies have shown that the newly formed junctions consist of tight junctions and gap junctions. Several tight-junction proteins are known as integral membrane components, including occludin and claudins; but none of them have been found in podocytes. Coxsackievirus and adenovirus receptor (CAR) has recently been identified as a virus receptor that is a 46-kDa integral membrane protein with two Ig-like domains in the extracellular region. In polarized epithelial cells, CAR is expressed at the tight junction, where it associates with ZO-1 and plays a role in the barrier to the movement of macromolecules and ions. In the present study, we investigated the expression and localization of CAR in rat kidneys treated with puromycin aminonucleoside (PAN) and in rat kidneys perfused for 15 minutes with protamine sulfate (PS). Both the experimental models have been used to induce tight junctions in podocytes. Ribonuclease protection assay and Western blot analysis revealed a distinct increase of CAR transcript and protein in glomeruli during PAN nephrosis but no increase in glomeruli by PS perfusion. Immunohistochemistry revealed a significant increase in CAR staining intensity along the glomerular capillary wall in PAN nephrosis and after PS perfusion. Immunoelectron microscopy demonstrated in both the models that the immunogold particles for CAR along the capillary wall were found predominantly at close cell-cell contact sites of podocytes but were rarely found at slit diaphragms. In cultured podocytes, CAR was localized at cell-cell contact sites. CAR distribution was identical to that of ZO-1 and different from that of a gap junction protein, connexin43. These findings indicate that CAR is an integral membrane component of tight junction in podocytes and that CAR expression in podocytes is regulated at the transcriptional level and in the redistribution of protein.

Expression profile of extracellular matrix and its regulatory proteins during the process of interstitial fibrosis after anti‐glomerular basement membrane antibody‐induced glomerular sclerosis in Sprague‐Dawley rats

Anti‐glomerular basement membrane (GBM) nephritis in Sprague‐Dawley (SD) rats was characterized by development of marked glomerular sclerosis and tubulointerstitial fibrosis. To elucidate sequential change of the glomerular sclerosis and tubulointerstitial fibrosis, accumulation and mRNA expression of extracellular matrix (ECM) components and transforming growth factor (TGF)‐β were examined in the glomerulus and cortex during the disease course by histology, immunostaining and ribonuclease protection assay. Mild proliferative and degenerative lesions appeared in the glomeruli by day 15 after anti‐GBM antibody binding to GBM and progressed to glomerular sclerotic lesion thereafter. Conversely, interstitial change was first recognized by infiltration of mononuclear cells after day 20, followed by marked accumulation of ECM and tubular degeneration. The interstitial fibrosis was induced without apparent binding of anti‐GBM antibody to tubular basement membrane. Accumulation of fibronectin, collagen type I and type IV was noted in the interstitium by immunofluorescence microscopy in association with enhanced expression of mRNA for these ECM components and their regulatory molecules such as matrix metalloproteinase (MMP2), tissue inhibitor of metalloproteinase (TIMP)‐1 and TGF‐β1 both in glomeruli and cortex. The glomerular expression of these mRNA increased apparently by day 15 and reached a plateau or a peak at day 20. The expression of the same mRNA increased gradually from day 15 to day 29 in the cortex. These observations show that interstitial fibrosis follows glomerular sclerosis after anti‐GBM antibody injection in SD rats, suggesting that at least a part of the mechanism for ECM accumulation in the glomerulus and interstitium is essentially the same in terms of composition of ECM and expression of its regulatory molecules.

Glomerulonephritis associated with chronic infection from long-term central venous catheterization

There have been few reports on immune complex-mediated glomerulonephritis associated with chronic infection from long-term central venous catheterization in adulthood. We report here on a 13-year-old boy with nephritis who exhibited glomerulonephritis that had been induced by the long-term use of central venous catheters, and its resolution after extraction of the central venous catheter. A diagnosis of glomerulonephritis associated with chronic infection caused by long-term central venous catheterization was made, based on the absence of clinical findings after removal of the catheter, hypocomplementemia, pathology findings resembling membranoproliferative glomerulonephritis, and detection of Staphylococcus epidermidis from culture of the removed catheter culture. For clinicians using long-term central venous access for parenteral feeding, rapid catheter exchange is necessary for patients with fever of unknown origin.

Uninephrectomy induces progressive glomerulosclerosis and apoptosis in anti-Thy1 glomerulonephritis

Administration of the anti‐Thy1 antibody in rats induces reversible glomerulonephritis resembling human mesangiolytic and mesangioproliferative diseases. The purpose of the present study was to design a model of irreversible glomerulosclerosis, using the anti‐Thy1 antibody injection after uninephrectomy, and examine it, focusing on apoptosis in the process of progressive sclerotic changes. Wistar rats were divided into three groups: one‐kidney groups (group I and III) and a two‐kidney group (group II). All groups were injected with the anti‐Thy1 antibody (OX‐7) at day 0, and group I and III were uninephrectomized at day −6. Only group III rats were given a half dose of OX‐7 as compared with group I and II. Rats were killed for histological examinations at days 7, 14 and 30. In group I, progressive glomerular lesions, such as glomerular adhesion to Bowmans capsule, crescent formation, and collapse of capillary tufts were observed at days 14 and 30. No significant differences were observed in the pathological findings between group I and III. There was a significantly higher number of glomerular terminal deoxynucleotidyltransferase‐mediated dUTP nick end labeling‐positive cells in group I as compared to group II at days 7 and 14. Moreover, the glomerular expression of transforming growth factor‐β, heparan sulfate proteoglycan and chondroitin sulfate proteoglycan significantly increased in group I as compared to group II at days 7 and 14. Progressive glomerulosclerosis can be induced in the rat by a single injection of the anti‐Thy1 antibody after unilateral nephrectomy. It is suggested that apoptosis and extracellular matrix accumulation play an important role in the development of glomerulosclerosis.

Renal Effects of Coxsackie B4 Virus in Hyper-IgA Mice

For clarification of the pathogenetic role of viral infection in chronic glomerulonephritis, the renal effects of Coxsackie B4 virus (CB4) were examined in hyper-IgA (HIGA) mice. In experiment 1, HIGA mice (n = 75) were inoculated intravenously with live CB4 and inactivated CB4 once a month from 1 to 12 mo of age. In experiment 2, HIGA mice (n = 45) were inoculated intravenously with live CB4 and inactivated CB4 once at 6 wk of age. In experiment 3, 60 mice were inoculated intravenously with carbon and live or inactivated CB4 once at 6 wk of age. Mice in the control group were inoculated with vehicle. The kidneys were extirpated from five mice of each group killed with time after inoculation for histologic evaluation. The scores for mesangial IgA deposition, PCNA-positive cells, and matrix at 20 wk were higher in mice with live CB4 than in mice with inactivated CB4 or without CB4. On electron microscopic examination, swelling and detachment of endothelial cells from 24 h after inoculation and increase of serum IFN-gamma concentration were found in mice with live CB4. Many carbon particles were present in peripheral and central zones of the mesangium from 5 to 10 d in mice with carbon and live CB4. These results suggest that CB4 provokes exacerbation of renal pathologic findings in HIGA mice via endothelial injury, IFN-gamma production, and dysfunction of the mesangial pathway.

Epstein-Barr virus-associated hemophagocytic syndrome in a patient with lupus nephritis

Hemophagocytic syndrome (HPS) is an unusual but severe illness associated with a variety of infections, as well as genetic, malignant tumors, and autoimmune diseases. We report an 11-year-old girl with systemic lupus erythematosus and nephritis who developed HPS associated with Epstein-Barr virus reactivation. In our patient, the onset of reactive HPS might be related to immunosuppressive treatment during the course of lupus nephritis.

Implication of the complement system in the induction of anti-glomerular basement membrane glomerulonephritis in WKY rats

In WKY rats, administration of a small dose of anti‐glomerular basement membrane antibody induces severe crescentic glomerulonephritis, which is characterized by infiltration of CD8‐positive lymphocytes and monocytes/macrophages into the glomeruli with crescent formation. In this study, the involvement of the complement system was examined in the induction of this model. To deplete complement, cobra venom factor (CVF) was used. By a single injection with CVF at 24 h prior to the induction of this model, plasma C3 level from days 0–2 was less than 10% compared with pre‐injection of CVF. Complement was almost depleted in the early phase of this model. No significant differences were observed in proteinuria and the frequency of glomeruli associated with the extracapillary crescentic lesions between the CVF‐treated group and the control group. In addition, the number of monocytes/macrophages and CD8‐positive lymphocyte infiltration into the glomeruli showed no significant differences between both groups. These results indicate that the possibility of complement system involvement is considered low in the induction of this model.