Because of difficulties of propagating and cultivating glomerular endothelial cells (GEN), the role of glomerular mesangial cells (GMC) in the regulation of GEN growth is still unknown. We cloned individually GEN and GMC from the same adult bovine kidneys, and a direct coculture system was established to examine the interactions between GEN and GMC. When cocultured with GMC which was growth arrested by mitomycin C, the GEN growth was inhibited by 63% after 8 days of the beginning of coculture. No inhibitory activity was detected when media conditioned by individual culture of GMC. In contrast, media condi tioned by GEN cocultured with GMC inhibited bromodeoxyuridine (BrdU) uptake of GEN detected by flow cytometry. Addition of anti-transforming growth factor (TGF)-β antibody to media conditioned by coculture blocked the inhibitory actions of conditioned media onGEN growth. Western blotting analysis of conditioned media by coculture revealed an existence of TGF-β in the media. These results indicate that latent TGF-β is produced by these cells and activated in coculture of GEN and GMC.

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To gain an insight into the effect of erythropoietin (Epo) upon cation transporters and cytosolic free Ca²⁺ concentration ([Ca²⁺]i) of vascular smooth muscle cells (VSMC), we studied whether 1) Epo, per se, alters Ca²⁺, Na⁺, K⁺ fluxes and [Ca²⁺]i of VSMC, and 2) Epo may modify the effect of endothelin (ET-1). Using serially passaged quiescent cultured VSMC, the following results were obtained. 1) Epo had no direct effect on steady state Na^- - K⁺ transporters (Na⁺ -K⁺ pump, Na⁺ -K⁺ cotransport and Na⁺ -H⁺ antiport). 2) ET-1 alone substantially stimulated Na⁺ -K⁺ pump, Na⁺ -H⁺ antiport and ⁴⁵Ca uptake, although these effects were not potentiated in the presence of Epo. 3) Epo alone substantially stimulated ⁴⁵Cauptake, leading to an increase in [Ca²⁺]i, which effect was not seen in Ca2+ deficient medium, and was partially inhibited with diltiazem but not with TMB-8. 4)Even in the presence ofEpo, ET-1 and angiotensin II (A II)had substantial stimulatory effect on [Ca²⁺]i of cultured VSMC. The present data indicate that Epo, per se, elicits an increase in [Ca²⁺]i of VSMC through the stimulation of inward Ca⁰ flux without affecting Na⁺ -K⁺ transporters. In contrast, Epo did not potentiate ET-l's stimulatory effect on the transporters. Although the effect of Epo was subtle compared to ET-1 and A II, it may alter an overall characteristic of vascular smooth muscle cell contractility, possibly leading to blood pressure elevation in patients on maintenance dialysis.

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In order to study the effect of thiazide on urinary citrate excretion, we measured sodium dependent citrate uptake by rat renal cortical brush border membrane vesicles (BBMVs) and analyzed acid-base balance with or without thiazide treatment. 5-weeks male Wistar rats (Jcl) were housed in metabolic cages, given 0.1 mg / 100 g BW / day hydrochlorothiazide (HCTZ) by a gastric tube once a day for 3 weeks (thiazide treated rats). Arterial blood wastaken from aorta and BBMVs were prepared with divalent cation precipitation method. Citrate uptake was measured by a Millipore rapid membrane filtration technique. Oral treatment of HCTZ for 3 weeks showed a significant increase of urinary citrate excretion in both concentration and daily excretion. The sodium dependent citrate transport was observed in rat renal brush border membrane (BBM). But no significant effect of oral HCTZ was observed on acid base balance and sodium dependent citrate uptake in rat renal BBM. These data suggested that the stimulative effect of thiazide on citrate excretion was not via sodium dependent citrate transport in rat renal BBM.

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In order to clarify the abnormalities of intra-glomerular coagulation and fibrinolysis in patients with various renal diseases, urinary fibrin/fibrinogen degradation products (FDP) have been examined by several methods. We established a highly sensitive new method of enzyme-linked immunosorbent assay for urinary FDP. The results were as follow: 1) The mean±SD of urinary FDP in normal subjects was 10.30±9.08ng/ml. 2) The urinary FDP levels in chronic glomerulonephritis, nephrotic syndrome and chronic renal failure patients were significantly higher than normal subjects, and the levels in SLE, Alport's syndrome patients were higher than normal subjects. 3) The urinary FDP levels were a little bit higher in the patients with proliferative glomerulonephritis than in chronic glomerulonephritis patients with minor lesion or membranous nephropathy. 4) There was significant correlationbetween urinary FDP and urinary protein in chronic glomerulonephritis, while there was no correlation in nephrotic syndrome. 5) There was no correlation between urinary FDP and intra-glomerular fibrin deposits examined by immumofluorescent study in chronic glomerulonephritis, while in nephrotic syndrome, there were high levels of urinary FDP in the positive fibrin deposits cases. These results suggested that the most of the part of excretionof the urinary FDP in chronic glomerulonephritis is associated with the filtration of blood FDP to urine through the glomerular basement membrane, while in the nephrotic syndrome cases the origin of urinary FDP is related to the filtration and /or the intra-glomerular coagulation abnormalities.

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To evaluate cardiorespiratory endurance of patients with chronic glomerulonephritis (CGN), ventilatory threshold oxygen uptake (VT) was measured in 69 patients with chronic glomerulonephritis and 337 healthy subjects (male: 140, female: 197) varying in age from 15 to 86yrs. They underwent ramp exercise test using bicycle ergometer with breath by breath measurement of ventilation and gas exchange variables. VT was determined by visual inspection. In both healthy subjects and patients, VT declined with age and significant negative correlation was found between age and VT in each male and female. VT of the each patient was presented as percentage of the respected value of sex and age-matched healthy subjects (%VT). Mean %VT of all the patients was 88.7+21.3% and significantly lower than the healthy subjects. Mean Ccr of the patients was 84.1+31. Oml/min and mean hemoglobinconcentrantion was 13.9+1.7g/dl. Although there was no significant difference in renal function or duration of disease between the patients of sedentary life style and active life style, %VT of the patients of sedentary life style (82.7+16.0%) was significantly lower than healthy subjects and the patients of active life style (120.8+27.1%). Sedentary life style seems to be the most important factor responsible for the decline of cardiorespiratory endurance in the patients with mild impairment of renal function. Secondary, elongation of duration of disease seems to reduce VT of the patients. %VT of the patients over 1 year (86.6+21.9%) is significantly lower than healthy subjects. The decrease of hemoglobin concentration and glomerular filtration rate, and the increase of protein excretion also have tendency to reduce VT of the patients with CGN. In conclusion cardiorespiratory endurance in the patients with CGN is reduced before azotemia and the most important factor for the decline seems to be the reduction of physical activities in daily life.

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A long term corticosteroids (CS) treatment is often required for children with nephrotic syndrome (NS), which may affect their bone metabolism. In this study we evaluate the effects of CS, and also the implication of the character of NS itself on bones. We divided sixteen cases of idiopathic NS in children into two groups, such as CS responder group (R; male/female-8/4, mean age 10.9 years old), and CS non-responder group (NR; male/female =4/0, mean age 10.2 years old). We measured the bone mineral density of lumber vertebrae by quantitative computed tomography (QCT), some biochemical and endocrinological markers for about half a year. The bone mineral density was significantly decreased soon after the beginning of CS therapy. A significant decrease in the serum osteocalcine level and a slight tendency for an increase in urinary excretion of the free hydroxyproline in the both R and NR groups. In addition, NR group showed hypocalcemia, a decrease in the serum 25-hydroxyvitamin D3 and 1, 25-dihydroxyvitamin D3, and a slight tendency for an increase in the C-parathyroid hormone and highly sensitive parathyroid hormone levels. In conclusion, the results suggested that the effects of CS on bone metabolism were mainly the suppression of bone formation. In addition, the effects of CS and the character of NS itself should be evaluated separately when we consider about bone metabolism in children with NS and in greater detail.

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We studied quantitatively a morphological change in diabetic nephropathy by light microscopy. The renal biopsy specimenes were examined using color image processor in 53 patients with non-insulin-depedent diabetes mellitus (NIDDM). These patients, aged 22 to 69 years, had been NIDDM for 2 month to 27 years. Glomerular area (GA), mesangial rate (MR) and length of capillary filtration (CFL) were used as the morphological parameters. Duration of diabetes, urinary albumin excretion and creatinine clearance (Ccr) were chosen as clinical manifestations. Duration of diabetes was correlated with MR (r=0.55, p<0.01), but not GA. Glomerular hypertrophy could not be found. A significant relationship between Ccr and CFL were noticed (r=0.64, p<0.01).

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Four cases of idiopathic acute tubulointerstitial nephritis (TIN) associated with uveitis (so-called TINU syndrome) were experienced between 1986 and 1990. Patients' ages ranged from 14 to 42 years old and three were female and one was male . All cases showed general symptoms, such as general malaise, anorexia and weight loss. All patients had initially TIN and became ill uveitis four to eight months after the onset of TIN. All cases had mild proteinuria, mild anemia, the lower serum levels of potassium, hyper r-globulinemia and the reduced glomerular filtration rate with the increased β2-microglobulin in urine and serum. All renal biopsies specimens showed mild edema and diffuse infiltration of inflammatory mononuclear cells in the interstitium without any glomerular or vascular abnormalities. Furthermore, numerous CD4 positive cells, CD8 positive cells and CDllc positive cells were seen in the interstitium. Of four patients, three cases were treated with both oral administra tion and eye drop of prednisolone (PSL), another one case was therapied with eye drop PSL only. In all cases TIN had good prognosis, but two patients had recurrences of uveitis. All patients underwent tissue typing for HLA-A, B, C and DR antigens. Three patients had identical HLA-Cw3 and all four cases revealed identical HLA-A24(9). These results suggest that immunological mechanism, especially cell-mediated, and HLA system may play animportant role in the occurrence of TINU syndrome.

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In order to clarify the abnormalities of cellular immune responses in uremic patients, we studied several immunological parameters such as T cell subsets, the number of B cells, in vitro immunoglobulin production by peripheral blood mononuclear cells (PBMC) with or without addition of pokeweed mitogen or IL-2 in 19 patients on chronic hemodialysis (HD), 42 patients on continuous ambulatory peritoneal dialysis (CAPD) and 27 healthy volunteers. The CD4/CD8 ratio was significantly higher in HD and CAPD patients than in the control group. Higher activated T cell counts (T cells with DR antigen and IL-2R) were observed in the patient groups than in normal controls. The number of DR+T cells showed a positive correlation with the level of serum β₂MG in patient groups. Spontaneous synthesis of immunoglobulin in vitro in mononuclear cell culture was observed in all groups in the following order; control, CAPD and HD. The number of B cells (CD19+) was less in thepatient groups than in the control group. There was a positive correlation between the serum levels of β₂MG and the degree of activated T cell markers. It is concluded that altered function in T and B cells might be responsible for various immunological abnormalities observed in patients with chronic renal failure on HD or CAPD.

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The effect of recombinant human erythropoietin (rHuEPO) on the lymphocyte subpopulations of maintenance hemodialysis patients has been studied. Peripheral lymphocyte counts, lymphocyte subpopulations (Two color analysis) and PHA-induced proliferative response were measured before and after rHuEPO administration during 8 months in 22 stable hemodialysis patients (mean age 52 years, mean duration of dialysis 48 months). We could find a significant increase in the proportions of CD4⁺ Leu8^- cells, CD3⁺ HLA-DR⁺ cells, CD8⁺ HLA-DR⁺ cells, CD14⁺ HLA-DR⁺ cells and CD4⁺ Leu8^-/CD8⁺ CDllb⁺ cells ratios after rHuEPO therapy. In addition, a significant decrease in the proportions of CD16⁺ CD57 cells and CD20⁺ cells could be found after rHuEPO therapy. These results suggest that rHuEPO administration to stable hemodialysis patients induces increases in lymphocytes categorized into helper subset groups and in activated T lymphocytes.

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We examined the records of 101 Japanese patients with chronic renal failure (CRF) to determine the influence of established risk factors on the incidence of coronary artery disease (CAD). CAD was found in 25 patients (myocardial infarction in 18, significant coronary artery stenosis by coronary angiogram in 16). In patients with CAD, there were significantly more male (80% vs 58%, p<0.05), more smokers (80% vs 41 %, p<0. 0005), more diabetics (68% vs 32%, p<0.005) and more hypertensives (92% vs 76%, p<0.05). However, lipoprotein (a), high-density lipoprotein cholesterol, low-density lipoprotein cholesterol, or triglycerides were not significantly defferent between CAD group and control group. When significant risk factors were entered into a stepwise logistic regression analysis, we found a history of smoking (p=0.0025) and the presence of diabetes (p=0.0141) to be independent risk factors for the presence of CAD in Japanese patients with CRF.

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Cerebrovascular and cardiovascular diseases are important predictors for survival in patients undergoing continuous ambulatory peritoneal dialysis (CAPD), and account for about half the deaths in these patients. Lipoprotein(a) [Lp(a)] is known to show high values in diabetics with proteinuria, and albuminuric renal disease. The purpose of this study was to determine Lp(a) levels and to investigate the association of Lp(a) and atherosclerotic risk factors in patients treated by CAPD. Lp(a) concentration were measured in 20 CAPD patients in the age range 31 to 83 years. Mean(±SD) levels of serum Lp(a) were elevated in the CAPD patients compared to age, sex matched 17 controls (49.5±27. 7 vs. 15.5±12.4mg/dl, p<0.001). The levels of Lp(a) were significantly higher in the diabetic CAPD patients than in non-diabetics. There were significant positive correlations between serum Lp(a) concentrations and fasting blood sugar. However, when the above two groups were matched for age, sex, body mass index and FBS, Lp(a) concentrations were also significantly higher in CAPD patients than those in normal controls. We found no statistically significant correlations of Lp (a) with either age, body mass index, blood pressure, serum lipoprotein, apoprotein, glycated hemoglobin, BUN, creatinine or serum protein levels. There were no correlations between serum Lp(a) levels and albumin and LP(a) concentrations in the dialysate in all CAPD patients. Along with assessment of other known established cardiovascular risk factors such as elevated blood pressure, atherogenic abnormalities of plasma lipids and lipoproteins, and impaired glucose tolerance, we suggest that elevated levels of Lp(a) may lead to the accelerated atherosclerosis in these patients.

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In order to examine the initiation time of drug treatment for chronic renal failure by the removal of certain substances which are accumulated in the digestive tract, experiments were carried out on 60 male Sprague-Dawley rats weighing 285-325g. The rats were first subjected to 2/3, 3/4 and 4/5 nephrectomy (n=20). The experiments were begun at 2 weeks after the surgery, and were performed over an 8-week period. Half of each group of nephrectomized rats (n=10) was administered the oral adsorbent, 1 g/day of Kremezin^(R) (AST-120, Kureha Chemical Industry Co, Tokyo), and pair feeding was done in each group of nephrectomized rats. The administration of Kremezin^(R) delayed the occurrence of glomerular hypertrophy, glomerulosclerosis, hypertrophy of the glomerular epithelial cells, flatteing of the tubular cells, dilation of the tubular cavity and infiltration of monocytes into the interstitium in the 2/3 nephrectomized rats. In addition, the administration of Kremezin^(R) delayed the appear ance of proteinaceous cast formation in the tubules, ballooning of the tubular cells, an increase in systemic blood pressure and an increase in urinary protein excretion in the 3/4and 4/5 nephrectomized rats. These findings indicated that the correction of an abnormal milieu within the digestive tract in chronic renal failure can delay its progression. Since the level of the creatinine clearance in the 2/3 nephrectomized rats was equal to approximately 60% of the creatinine clearance in normal rats, it is suggested that drug treatment for chronic renal failure with Kremezin^(R) should be initiated before the level of the creatinine clearance decreases to 60% of creatinine clearance in normal human.

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We made clinicopathological studies of twelve patients with rapidly progressive glome rulonephritis (RPGN) treated at our department of Okayama University Medical School Hospital for the last eight years. By immunofluorescence microscopy, seven cases were immune complex (IC) type and five were pauci immune type. By light microscopy, the mean percentage of glomeruli with crescents was seventy five, and cellular and fibro-cellular crescents were dominant in eleven cases. Combined therapy including methylprednisolone pulse or oral steroid was effective in eleven. Hemodialysis was performed in four patients with severe uremia and three of them recovered from hemodialysis. Two cases of RPGN associated with systemic lupus erythematosus died of cerebral hemorrhage. We conclude that IC type and pauci immune type RPGN have good prognosis when treated at their active stage with intensive combined therapy and with hemodialysis if necessary.

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Therapeutic effect of hemodialysis on gouty arthritis and tophi in 2 patients with chronic renal failure (CRF) is described. One patient was 74-year-old man with CRF due to benign nephrosclerosis, and had severe secondary gout. There were many tophi in metatarsophalangeal joints of the fingers and toes. Another patient was 44-year-old man with CRF probably due to familial gout. He had many tophi in elbows, knees, wrists, fingers and toes. In both cases initition of hemodialysis treatment, at first temporaly exercerbated gouty arthritis, but thereafter gradually eliminated the tophi. From these results, dialysis treatment is considered to be quite efficient method of treatment for gouty arthritis and tophi in patient with CRF. Gouty arthritis in CRF patient, which does not respond to the conservative therapy, should be considered to be one of the indices for commencement of hemodialysis.

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