Sanki Kodera

Detection of Serum IL-6 in Patients with Diabetic Nephropathy

Yasuhiko Tomino. MD, Division of Nephrology, Department of Medicine, Juntentto University School of Medicine, 2-1-1 Hongo, Bunkyo-ku, Tokyo 113 (Japan) Dear Sir, A study on the detection of serum IL-6 in patients with non-insulin-dependent diabetes mellitus (NIDDM) with or without nephropathy is described. IL-6 is generally regarded as a multifunctional cytokine which has a variety of biological activities, including the ability to stimulate bone marrow stem cell proliferation, B cell differentiation, immuno-globulin secretion, T cell activation, and acute phase protein synthesis [1, 2], IL-6 is also produced by the renal glomerular mesan-gial cells. Cytokines are known to play an important role in autoimmunity and appear to be involved in the pathogenesis of insulin-dependent diabetes mellitus (IDDM). However, Cavallo et al. [3] reported that detectable levels of serum IL-6 were observed in only 10% of IDDM patients. Serum samples were obtained from 9 patients with NIDDM with nephropathy (diabetic nephropathy), 9 patients with NIDDM without nephropathy and 29 patients with chronic glomerulonephritis (CGN). NIDDM was diagnosed with a 75-gram glucose tolerance test. Patients with diabetic nephropathy continuously showed more than 200 mg/24 h. Serum IL-6 levels were measured with ELISA as described previously [4]. Mouse monoclonal anti-IL-6 antibody (HH61-10) and monoclonal horse radish peroxidase-conjugated anti-IL-6 antibody (HH61-2 Fab’) were used in a double-antibody sandwich ELISA [5]. Levels of serum IL-6 of healthy controls were less than 4.0 pg/ml [5]. The mean levels of serum IL-6 in all patients with NIDDM were significantly higher than those in patients with CGN (p < 0.05). The levels of serum IL-6 in patients with diabetic nephropathy were significantly higher than those in cases of CGN or NIDDM without nephropathy (p < O.Ol and p < 0.05, respectively; table 1). It appears that the presence of IL-6 in the patients’ sera may reflect increased localized production of this cytokine at the pancreatic and/or glomerular me-sangial levels. The measurement in serum IL-6 may add

Genetic Dissection of the Effects of Stimulatory and Inhibitory IgG Fc Receptors on Murine Lupus

Immune complex (IC)-mediated tissue inflammation is controlled by stimulatory and inhibitory IgG Fc receptors (FcγRs). Systemic lupus erythematosus is a prototype of IC-mediated autoimmune disease; thus, imbalance of these two types of FcγRs is probably involved in pathogenesis. However, how and to what extent each FcγR contributes to the disease remains unclear. In lupus-prone BXSB mice, while stimulatory FcγRs are intact, inhibitory FcγRIIB expression is impaired because of promoter region polymorphism. To dissect roles of stimulatory and inhibitory FcγRs, we established two gene-manipulated BXSB strains: one deficient in stimulatory FcγRs (BXSB.γ−/−) and the other carrying wild-type Fcgr2b (BXSB.IIBB6/B6). The disease features were markedly suppressed in both mutant strains. Despite intact renal function, however, BXSB.γ−/− had IC deposition in glomeruli associated with high-serum IgG anti-DNA Ab levels, in contrast to BXSB.IIBB6/B6, which showed intact renal pathology and anti-DNA levels. Lymphocytes in BXSB.γ−/− were activated, as in wild-type BXSB, but not in BXSB.IIBB6/B6. Our results strongly suggest that both types of FcγRs in BXSB mice are differently involved in the process of disease progression, in which, while stimulatory FcγRs play roles in effecter phase of IC-mediated tissue inflammation, the BXSB-type impaired FcγRIIB promotes spontaneous activation of self-reactive lymphocytes and associated production of large amounts of autoantibodies and ICs.

Detection of Apoptotic Cells in Glomeruli of Patients with IgA Nephropathy

The purpose of the present study was to determine the relationship between the existence of apoptotic cells in glomeruli, clinical or histopathological findings and response to treatment in patients with IgA nephropathy. Renal biopsy specimens were obtained from 23 patients with IgA nephropathy. These patients were divided into two groups: mild glomerular damage (12 patients), and severe glomerular damage (11 patients). The nick end-labelling method (TUNEL) and fluorescent staining (Hoechst 33258) were used for the detection of apoptotic cells. Five of 6 patients with apoptotic cells in the glomeruli detected by TUNEL were in the severe glomerular damage group, and only 1 patient was in the mild glomerular damage group. Apoptotic cells in glomeruli were also detected by fluorescent staining in 3 of 5 patients in the severe glomerular damage group who showed apoptotic cells in TUNEL. However, no apoptotic cells were detected in patients in the mild glomerular damage group in fluorescent staining. Mean levels of urinary protein excretion at the time of renal biopsy in the patients with apoptotic cells were significantly higher than those in patients without apoptotic cells (p < 0.01). The mean levels of creatinine clearance (Ccr) in the patients with apoptotic cells were slightly lower than those in patients without such cells. There were no significant differences in the levels of serum creatinine (s-Cr) and BUN in patients with or without apoptotic cells. In the severe glomerular damage group, urinary protein excretion after treatment in the patients with apoptotic cells was significantly improved compared with that in the patients without such cells (p < 0.01). It appears that the levels of proteinuria and renal function tests might be influenced by apoptosis in patients with IgA nephropathy. It is postulated that apoptosis may induce reduction of excess proliferative mesangial cells and/or infiltrated cells, and tissue repair. Thereafter, these histological alterations may improve proteinuria and renal function.

Ocular Fundus Lesions in Systemic Lupus Erythematosus Model Mice

To evaluate spontaneous development of the ocular fundus abnormalities associated with collagen disease, we investigated the ocular fundus lesions in systemic lupus erythematosus (SLE) models. (NZW x BXSB) F1 mice were employed as SLE models with antiphospholipid syndrome. The abnormal findings in the ocular fundus were recorded with a fundus camera for small animals (KOWA Co., Ltd.), and the chorioretinal lesions were studied histopathologically. As in the systemic symptoms of SLE, the incidence of ocular fundus abnormalities in these (NZW x BXSB) F1 mice was significantly higher in males than in females, suggesting the influence of the Yaa (Y chromosome-linked autoimmune acceleration) gene. Lesions in the fundus appeared in the form of white spots, which increased in number along with the course of the disease. The lesion developed into retinal detachment in some animals. Dilatation of veins and narrowing of arteries were marked. These lesions were very similar to multifocal posterior pigment epitheliopathy (MPPE) in humans in that white spots appear first and then develop into exudative retinal detachment caused by retinal pigment epithelial disorder. Histopathological findings included 1. structural destruction of the photoreceptor cell layer, 2. degeneration and loss of the retinal pigment epithelium, and 3. narrowing and occlusion of the choriocapillaris associated with thrombus formation, cellular infiltration into the surrounding tissues, and wall thickening of the choroidal arterioles. The study of these SLE mouse may contribute to the elucidation of abnormalities in the fundus associated with collagen diseases, including the relationship between thrombus formation and antiphospholipid syndrome.

A Case of Lupus Nephritis Showing Good Clinical Course and Apoptosis in Glomerular Cells Detected by the Nick End Labeling Method

We report here an adult patient with lupus nephritis who had a good clinical course under long-term observation. Apoptotic bodies in the glomeruli were determined in serial renal biopsy specimens by the nick end labeling method. Apoptotic bodies in the proliferated glomerular cells were detected in the 3rd renal biopsy but not in the 2nd biopsy. The clinical activities of lupus nephritis fluctuated until the time of the 3rd renal biopsy. The 3rd renal biopsy was performed because of increased proteinuria and an increased amount of hyaline, granular and red blood cell casts, with impairment of renal function. The levels of proteinuria, creatinine clearance and serum complements were improved after the 3rd renal biopsy. It appears that apoptosis might control glomerular cell proliferation and also influence the clinical course of lupus nephritis.

Effect of low-protein diet on glomerular changes in ddY mice: a spontaneous animal model of IgA nephropathy.

Yasuhiko Tomino, MD, Division of Nephrology, Department of Medicine, Juntendo University School of Medicine, 2-1-1 Hongo, Bunkyo-Ku, Tokyo 113 (Japan) Dear Sir, Long-term dietary protein restriction is generally considered to reduce the levels of urinary protein and ameliorate the glomerular injuries in patients with various glomeru-lonephritides. We usually start dietary protein restriction after the definite diagnosis, although restriction from an early stage might show a good prognosis in such diseases. The authors reported that the ddY mouse strain can serve as a spontaneous animal model for IgA nephropathy [1, 2]. Marked depositions of IgA and C3 in glo-meruli and glomerular mesangial expansion were observed in the ddY mice after 40 weeks of age [1]. We attempted to compare the glomerular changes between the low-protein and the high-protein diets in ddY mice. Twenty ddY mice were fed a standard diet containing 22% protein until 40 weeks of age. These ddY mice were divided into two diet groups, i.e. low protein (6%) and high protein (50%). ddY mice of both groups were sacrificed at 70 weeks of age. Urinary protein was measured every 2 weeks according to the method of Knight et al. [3]. Renal sections stained with hematoxylin and eosin, and periodic acid-Schiff were examined by light microscopy. Renal cryostat sections were stained with FITClabelled goat anti-mouse IgA, IgG, IgM and C3 antisera at room temperature for 45 min. Type IV and I collagens, ñbronectin and laminin were also stained. The sections were examined using a Fig. 1. a Glomerular enlargement and mesangial expansion were observed in the high-protein diet ddY mouse. PAS. × 400. b These light-microscopic findings were improved in the lowprotein diet mouse. PAS. × 400. c Glomerular deposition of IgA was marked in the high-protein diet ddY mouse. × 400. d The intensity of glomerular IgA deposition was decreased in the lowprotein diet mouse. × 400. KAIIGER E-Mail karger@karger.ch Fax+ 41 61 306 12 34

Apoptotic cells in six patients with IgA nephropathy in repeat biopsies

SUMMARY: Programmed cell death is a selective process of physiological cell deletion and is known as apoptosis. The purpose of the present study was to determine the relationship between the existence of apoptotic cells in glomeruli and the clinical or histopathological findings obtained in repeat renal biopsies of patients with IgA nephropathy. Repeat renal biopsy specimens were obtained from six patients with IgA nephropathy. The nick end labelling method (TUNEL) was used for the detection of apoptotic cells. Clinical laboratory data, i.e. urinary protein excretion, creatinine clearance (Ccr), blood urea nitrogen (BUN) and serum creatinine, (s‐Cr) were obtained from these patients. At the first renal biopsy, apoptotic cells in the glomeruli were observed in three out of six patients using TUNEL. These patients were classified as the severe glomerular damage group. The other three patients without apoptotic cells were in the mild glomerular damage group. Mean levels of urinary protein excretion at the first renal biopsy in the patients with apoptotic cells were slightly higher than those in patients without apoptotic cells. Levels of Ccr in patients with apoptotic cells were lower than those in patients without apoptotic cells. There were no significant differences in the levels of BUN and s‐Cr in patients with or without apoptotic cells. Two patients with apoptotic cells in glomeruli at the first renal biopsy did not show apoptotic cells at the second renal biopsy. These two patients showed improvement not only in clinical laboratory findings but also in histological findings at the second biopsy. Only one patient with apoptotic cells at the first and second biopsies exhibited deterioration at the second biopsy. All three patients without apoptotic cells at the first renal biopsy also showed deterioration of the clinical laboratory and histopathological findings. It is postulated that various factors other than apoptosis might induce progression of renal injuries in such patients. It appears that the clinical laboratory data, i.e. proteinuria, renal function and histopathological findings, might be influenced by apoptosis in patients with IgA nephropathy. It is postulated that apoptosis may induce reduction of excess proliferative glomerular mesangial cells and/or infiltrating cells and tissue repair.