Panel of novel urine biomarkers for incident microalbuminuria in people with type 2 diabetes mellitus.

AIM: The need to identify novel biomarkers for early diagnosis and treatment of diabetic nephropathy is widely recognized. However, only a few studies have investigated the association between biomarkers and the onset of albuminuria. In this study, we aimed to investigate a panel of biomarkers suitable for predicting microalbuminuria. METHODS: Some 346 Japanese people with type 2 diabetes exhibiting normoalbuminuria were studied. The endpoint was defined as the onset of microalbuminuria. Thirty biomarkers were selected from among urinary biomarkers described in previous studies. A panel of biomarkers was selected using least absolute shrinkage and selection operator (LASSO). The prognostic performance of the developed panel was evaluated. RESULTS: During a mean follow-up of 6.2 years, 45 people progressed to microalbuminuria. A composite panel of six biomarkers (monocyte chemoattractant protein-1, osteopontin, soluble human tumour necrosis factor receptor-1, tenascin C, vascular endothelial growth factor-A and kidney injury molecule-1) was developed using LASSO. Compared with the basal model comprising estimated GFR and urinary albumin-to-creatinine ratio, addition of these six biomarkers significantly increased the overall C index from 0.773 to 0.824 (P = 0.019). Net reclassification improvement and integrated discrimination improvement were estimated to be 0.412 (P = 0.049) and 0.040 (P = 0.040), respectively. Decision curve analysis also showed that the model with the six novel biomarkers had a better prognostic value for predicting the onset of microalbuminuria. The optimism was moderate or negligible according to measures. CONCLUSIONS: The panel consisting of six novel urinary biomarkers effectively predicted incident microalbuminuria in people with type 2 diabetes.

Differences in risk factors for the onset of albuminuria and decrease in glomerular filtration rate in people with Type 2 diabetes mellitus: implications for the pathogenesis of diabetic kidney disease.

AIMS: To determine differences in predictors of albuminuria and decreased estimated GFR in Japanese people with Type 2 diabetes mellitus without chronic kidney disease. METHODS: This single-centre observational cohort study involved 1802 Japanese people with Type 2 diabetes with normoalbuminuria and estimated GFR ≥ 60 ml/min/1.73 m(2) (740 women; mean ± sd age 58 ± 12 years). Two separate outcomes were evaluated: onset of albuminuria ( ≥ 30 mg/g creatinine, albuminuria cohort; n = 1655) and decrease in estimated GFR ( < 60 ml/min/1.73 m(2) ; estimated GFR cohort; n = 1777). A Cox proportional hazards model was used to identify significant predictors for each outcome. RESULTS: During a median follow-up period of 6.9 years for the albuminuria cohort and 8.0 years for the estimated GFR cohort, 181 and 316 individuals reached the respective outcome. The 5-year cumulative incidence of albuminuria was 8.3%, and that of decreased estimated GFR was 10.4%. In the multivariate Cox model, greater urinary albumin-to-creatinine ratio, presence of diabetic retinopathy and higher HbA1c levels were associated with both outcomes. Unique risk factors for onset of albuminuria were male gender and higher uric acid levels; those for decreased estimated GFR were older age, greater systolic blood pressure, and lower baseline estimated GFR and HDL cholesterol levels. CONCLUSIONS: Identification of both common and distinct predictive factors for onset of albuminuria and decreased estimated GFR support the hypothesis that both common and distinct pathophysiological mechanisms are involved in the development of these two manifestations of chronic kidney disease in diabetes.

Comparison of the prevalence of chronic kidney disease in Japanese patients with Type 1 and Type 2 diabetes.

AIMS: The relationship between type of diabetes and risk of chronic kidney disease has not been studied in detail. We conducted this study to determine the prevalence of chronic kidney disease in Japanese adults with diabetes, with a particular emphasis on the comparison of Type 1 and Type 2 diabetes. METHODS: We studied 3,575 Japanese patients with diabetes, 504 with Type 1 (mean +/- SD age 38 +/- 13 years; 350 women and 154 men) and 3071 with Type 2 diabetes (60 +/- 13 years; 1187 women and 1884 men). Prevalence rates of albuminuria [urinary albumin/creatinine ratio (> or = 30 mg/g], decreased estimated glomerular filtration rate (eGFR < 60 ml/min/1.73 m(2)) and chronic kidney disease (defined as albuminuria and/or decreased eGFR) were compared between the two diabetic groups. RESULTS: The prevalence of albuminuria was higher in Type 2 than Type 1 diabetic patients by both Fisher's exact test (36.1 vs. 15.9%, P < 0.001) and multivariate logistic regression analysis [adjusted odds ratio (OR) = 1.482, 95% confidence interval (CI) = 1.050-2.091, P = 0.025]. The prevalence of decreased eGFR was also higher in Type 2 diabetic patients (25.2 vs. 7.9%, P < 0.001); however, the statistical significance disappeared after adjusting for covariates, including age (OR = 0.656, 95% CI = 0.395-1.088, P = 0.102). The prevalence of chronic kidney disease was also higher in Type 2 diabetic patients (46.0 vs. 19.1%, P < 0.001); however, the statistical significance disappeared in the multivariate analysis. CONCLUSIONS Type 2 diabetic patients are more than twice as likely as Type 1 diabetic patients to have chronic kidney disease due to an age-independent higher prevalence of albuminuria and age-dependent decreased eGFR.

Relationship between chronic kidney disease and silent cerebral infarction in patients with Type 2 diabetes.

AIMS: Silent cerebral infarction (SCI) is an independent risk factor for future symptomatic stroke. Although the prevalence of SCI is closely related to kidney function in non-diabetic individuals, evidence is lacking whether albuminuria and/or reduced estimated glomerular filtration rate (eGFR) independently increase the risk of SCI in diabetic patients. We therefore examined the relationships between albuminuria, eGFR and SCI in patients with Type 2 diabetes mellitus (T2DM). METHODS: We studied 786 T2DM patients with an eGFR > or = 15 ml/min 1.73/m(2), including 337 women and 449 men [mean (+/- sd), age 65 +/- 11 years]. All patients underwent cranial magnetic resonance imaging (MRI) to detect SCI. GFR was estimated using the modified three-variable equation for Japanese subjects. Albuminuria was defined as a first morning urinary albumin-to-creatinine ratio (ACR) > or = 30 mg/g. RESULTS: SCI was detected in 415 (52.8%) of the subjects. The prevalence of SCI was significantly associated with both elevated ACR and decreased eGFR in univariate analysis. In multivariate logistic regression analysis, urinary ACR remained independently associated with SCI after adjusting for conventional cardiovascular risk factors [odds ratio (OR) of urinary ACR per logarithmical value: 1.89, 95% confidence interval (CI) = 1.41-2.51, P < 0.001]; however, eGFR was no longer significantly associated with SCI (OR per ml/min 1.73/m(2) = 0.99, 95% CI = 0.98-1.00, P = 0.095). CONCLUSION: In conclusion, albuminuria but not decreased eGFR may be an independent predictor of prevalent SCI in patients with T2DM.

Altered expression and subcellular localization of diacylglycerol-sensitive protein kinase C isoforms in diabetic rat glomerular cells.

Protein kinase C (PKC) is implicated in the pathogenesis of diabetic nephropathy. This study was designed to identify the expression of diacylglycerol (DAG)-sensitive PKC-alpha, -betaII, -delta, and -epsilon isoforms in normal and diabetic rat glomerular cells and to determine the effects of high glucose and insulin on PKC isoform cellular compartmentalization and PKC activity. Diabetic rats treated with or without insulin and normal rats were examined 2 and 4 weeks after streptozotocin/vehicle injection. Renal cortical tissue immunogold-labeled with anti-PKC-alpha, -betaII, -delta, or -epsilon antibody was visualized by electron microscopy. From isolated glomeruli, total cell lysate and cytosol and membrane fractions were immunoblotted with the same anti-PKC isoform antibodies. PKC activity in isolated glomeruli was measured by 32P-phosphorylation of the epidermal growth factor (EGF)-receptor substrate. Immunogold labeling revealed expression of the four PKC isoforms by glomerular visceral epithelial, endothelial, and mesangial cells of both normal and diabetic rats. Immunoblot analysis of the diabetic rat glomeruli at 2 weeks demonstrated a significant increase in membrane-associated PKC-alpha, -delta, and -epsilon and a significant decrease in membrane PKC-betaII content compared with normal, which were similar at 4 weeks. Insulin treatment normalized membrane PKC isoform contents and caused a significant decrease in the cytosol content of PKC-alpha, -betaII, and -delta and total cellular PKC-alpha compared with normal. Although PKC activity in the cells of diabetic rat glomeruli was increased by 20% compared with normal, the difference did not reach statistical significance. In insulin-treated diabetic rat glomeruli, PKC activity was significantly decreased compared with non-insulin-treated diabetic rat glomeruli. In conclusion, DAG-sensitive PKC-alpha, -betaII, -delta, and -epsilon isoforms are all found in the three major glomerular cell types in rats, and the expression, compartmentalization, and activity are modulated independently by high glucose and insulin.

Modulation of prostaglandin metabolism by K-MAP and prevention of toxic effect of cyclosporin on pancreatic islet cells.

The injection of 25 mg/kg i.p. cyclosporin (CsA) for 3 wk caused marked functional and morphological deteriorations of pancreatic islet cells in Wistar rats that were prevented by the combined administration of p-aminobenzoic acid-N-D-mannoside sodium salt (K-MAP). In this article, the toxic effect of CsA on pancreatic islet cells and the preventive effect of K-MAP on CsA-associated islet cell toxicity were investigated. Prolonged hyperglycemia and depressed insulin secretion after the glucose challenge observed in CsA-treated rats could be prevented by the combined administration of 300 and 900 mg/kg K-MAP. Cytoplasmic vacuolizations and a decrease in the number of mitochondria, intact endoplasmic reticula, secretory granules, and insulin-positive cells, as revealed by peroxidase-antiperoxidase staining, could also be prevented by the administration of 900 mg/kg K-MAP. This preventive effect of K-MAP on CsA-associated islet cell toxicity may suggest the combined use of K-MAP with CsA in pancreas transplantation and treatment of insulin-dependent diabetes.

The telmisartan renoprotective study from incipient nephropathy to overt nephropathy--rationale, study design, treatment plan and baseline characteristics of the incipient to overt: angiotensin II receptor blocker, telmisartan, Investigation on Type 2 Diabetic Nephropathy (INNOVATION) Study.

We planned the INNOVATION study to determine whether telmisartan, an angiotensin-2-receptor blocker, delays the progression of renal disease from incipient nephropathy to overt nephropathy in hypertensive or normotensive Japanese patients with type 2 diabetes mellitus. The INNOVATION study is a randomized, double-blind, placebo-controlled trial. Eligible patients must have incipient nephropathy (defined as a urinary albumin to creatinine ratio of 100-300 mg/g creatinine) and a serum creatinine concentration of < 1.5 mg/dl for men and < 1.3 mg/dl for women. Patients who need treatment with angiotensin II receptor blockers or angiotensin-converting enzyme inhibitors are excluded. Eligible patients are randomly assigned to three groups: telmisartan titrated to 40 mg; telmisartan titrated to 80 mg; or placebo. The primary endpoint is the time from baseline visit to first detection of overt nephropathy (defined by a urinary albumin to creatinine ratio that is > 300 mg/g creatinine and 30% higher than the baseline on at least two consecutive visits). A total of 1855 patients have been enrolled from 160 study centres. In 527 randomized patients (28.4% of the enrolled patients), mean (SD) urinary albumin to creatinine ratio and serum creatinine concentration at baseline were 173.3 (47.2) mg/g creatinine and 0.78 (0.19) mg/dl. Sixty-eight per cent of the patients had hypertension at baseline. Mean (SD) systolic and diastolic blood pressures at baseline were 137.1 (14.6) and 77.5 (10.3) mmHg. The INNOVATION study will determine whether telmisartan, an angiotensin II receptor blocker, provides clinical benefits in hypertensive or normotensive patients with diabetes mellitus and diabetic nephropathy.

High glucose stimulates hyaluronan production by renal interstitial fibroblasts through the protein kinase C and transforming growth factor-beta cascade.

Deposition of hyaluronan has been implicated in the pathogenesis of diabetic glomerulosclerosis. We hypothesized the involvement of hyaluronan in diabetic tubulointerstitial fibrosis. We investigated high-glucose effect on hyaluronan production by rat renal interstitial fibroblasts (normal rat kidney [NRK] cells) and examined the role of hyaluronan in NRK cell proliferation. The involvement of protein kinase C (PKC) and transforming growth factor-beta (TGF-beta) in this response was also examined. After 24 hours of incubation in medium containing 25.6 mmol/L glucose, production of hyaluronan by NRK cells was significantly increased compared with medium containing 5.6 mmol/L glucose (P <.01). L-glucose and mannitol had no effect on hyaluronan production. High glucose enhanced basal in situ PKC activity (P <.01), and both an activator of PKC (phorbol 12-myristate 13-acetate; [PMA]) and TGF-beta 1 were able to increase hyaluronan production by NRK cells (P <.01). The effect of high glucose on hyaluronan production was diminished by coincubating cells with PKC inhibitors (staurosporine [Stp] or calphostin C [CpC]) or with an anti-TGF-beta neutralizing antibody. Stimulation of hyaluronan production by PMA was also normalized by anti TGFbeta neutralizing antibody, but the effect of TGF-beta1 was not affected by inhibition of PKC. Finally, incubating quiescent NRK cells with 50 or 100 ng/mL hyaluronan for 24 hours significantly increased NRK cell number (P <.01). In conclusion, high glucose stimulates hyaluronan production through the PKC/TGF-beta cascade. Increased hyaluronan promotes NRK cell proliferation. These results suggest that hyaluronan may play a role in the pathogenesis of interstitial fibrosis in diabetic kidney disease.

Circulating proinsulin levels in insulin-dependent diabetic patients after whole pancreas-kidney transplantation.

Disproportional hyperproinsulinemia is a sensitive marker for beta-cell dysfunction. The objective of this study was to assess the proinsulin profile in persons with insulin-dependent diabetes mellitus (IDDM) after pancreas-kidney transplantation. We determined serum insulin, C-peptide, and proinsulin concentrations during an oral glucose challenge in five pancreas-kidney transplant recipients, nine nondiabetic kidney transplant recipients, and 17 normal subjects. Basal proinsulin concentrations were significantly increased in pancreas-kidney recipients (geometric mean [+/-1 SE range], 6.0 [5.5 to 6.4] pmol/L) and kidney recipients (6.4 [5.4 to 7.5] pmol/L) compared with the normal subjects (2.8 [2.5 to 3.2] pmol/L). Integrated proinsulin concentrations during the oral glucose load were also higher in pancreas-kidney recipients (1.4 [1.1 to 1.8] nmol/L x min) and kidney recipients (1.5 [1.2 to 2.0] nmol/L x min) versus normal subjects (0.8 [0.7 to 0.9] nmol/L x min). There was no difference in basal or integrated proinsulin concentrations between the two transplant groups. Even after adjustment for the glomerular filtration rate (GFR), basal and incremental proinsulin concentrations continued to be higher in the transplant groups than in the normal subjects. Proinsulin to C-peptide molar ratios both before and after the glucose load were similar in the three groups. From these findings, we conclude that pancreas-kidney transplantation provokes proportional hyperproinsulinemia, which is closely associated with its reduced clearance in the kidneys.

Clinical profile of Japanese dialysis patients with diabetic nephropathy, diagnosed as having diabetes before the age of thirty.

Diabetic nephropathy is the leading cause of death in young diabetic patients. There are a large number of patients with non-insulin-dependent diabetes mellitus (NIDDM) who are diagnosed before the age of 30 in Japan. We investigated 36 patients with young-onset diabetes who started dialysis between 1978 and 1987 in our hospital. Of the 36 patients, 12 (33.3%) were classified as having insulin-dependent diabetes mellitus (IDDM), 22 (61.1%) had NIDDM, and 2 (5.6%) could not be classified clinically. The percentages of the different types of diabetes in our series of dialysis patients were almost identical with those in Nagai's series of 551 diabetic patients diagnosed before the age of 30 at the Diabetes Center of Tokyo Women's Medical College from 1976 to 1981. The present study showed the young-onset NIDDM in Japan was associated with almost the same incidence of end-stage diabetic nephropathy as was IDDM. However, the number of NIDDM patients diagnosed under 30 years of age was almost double that of IDDM patients. Thus, we have to pay greater attention to the development of diabetic nephropathy in young-onset NIDDM patients than has been thought necessary in the past.

An IDDM patient who complained of chest oppression with ischemic changes on ECG in insulin-induced hypoglycemia.

A 34-year-old female IDDM patient complained of chest oppression in hypoglycemic episodes and electrocardiograms revealed reversible ischemic changes occurring concomitantly with hypoglycemia. The ECG changes improved and the chest oppression disappeared following increasing blood glucose level by glucose intake. Master's double load test and treadmill load test were positive for ischemic changes. Radioisotopic myocardial scintigraphy by thallium and BMIPP did not show any filling defects and coronary angiography revealed no remarked stenosis in the coronary arteries. She had no mitochondrial tRNA(Leu) (A-->G) gene mutation at nucleotide position 3243, but both the patient and her mother had a G-to-A transition within the replication origin of the light strand at nucleotide position 5744 of the mitochondrial gene. As the patient's maternal family had no history of ischemic heart disease, it is not clear whether mitochondrial gene mutation at nucleotide position 5744 reflects the occurrence of cardiac ischemia. Some disorders of microcirculation in capillary vessels in cardiac muscles may occur in such patients.

Cardiovascular risk factors in diabetic patients undergoing continuous ambulatory peritoneal dialysis.

To determine the impact of continuous ambulatory peritoneal dialysis (CAPD) on cardiovascular risk factors in diabetic patients, we evaluated serum lipid profiles and plasma levels of coagulation and fibrinolysis parameters in 23 diabetic patients on long-term CAPD (aged 55 +/- 14 years, mean +/- SD), and compared them with those of diabetic patients undergoing hemodialysis (n = 62, 56 +/- 10 years) or kidney transplantation (n = 14, 43 +/- 14 years), and 40 normal subjects (39 +/- 10 years). All of the parameters were compared using analysis of covariance to adjust for the difference in age among the four groups in males and females separately. In the male CAPD patients, there were no significant differences in the serum concentrations of lipids, lipoproteins, and apolipoproteins. In contrast, in the female CAPD patients, the levels of triglyceride and apolipoprotein (apo) B and the low-density lipoprotein (LDL) cholesterol/ high-density lipoprotein (HDL) cholesterol ratio were significantly higher than those in the normal females. Lipoprotein (a) did not differ significantly among the four male and female groups. The plasma levels of fibrinogen and von Willebrand factor were higher both in the male and in the female CAPD patients than in the other corresponding groups. There was no significant difference in the levels of plasminogen activator inhibitor-1 among the four groups. In conclusion, CAPD is associated with a more atherogenic lipid profile than are hemodialysis and kidney transplantation in female diabetic patients, but not in male diabetic patients. Both male and female diabetic patients on CAPD have a hypercoagulability state but not a decreased fibrinolysis state.

[The effect of sharing medical costs for the aged 70 or more on the financial balance of the health insurance society].

The rapid increase of the proportion of older persons in society has made the problem of medical costs more important. Although medical cost-sharing for those aged 70 or more is decided by their medical costs and by the current proportion of such persons in the Society, it is not constant throughout the Society. The study evaluated the contribution ratio of medical cost-sharing for those aged 70 or more to the financial balance of the Society. In addition, characteristics of the Society associated with medical cost-sharing for those aged 70 or more were analyzed. The results are as follows. 1) Medical cost-sharing for those aged 70 or more was the greatest factor in the financial balance of the Society with a contribution ratio of as much as 55.2%. 2) Multiple regression analysis disclosed that dependent ratio, the average monthly salary, the area, the rate of those aged 70 or more and the average age of the Society members are significant in medical cost-sharing for those aged 70 or more. 3) Dependent ratio and the average monthly salary are the most important characteristics among those associated with medical cost-sharing for those aged 70 or more. A higher dependent ratio and a lower average monthly salary are related to higher cost-sharing. This is also related to geographic factors as cost-sharing in western Japan is higher than in eastern Japan. The lower rate of those aged 70 or more and the higher average age of the Society members are related to the higher cost-sharing.

[Maintenance therapy on the anemia in continuous ambulatory peritoneal dialysis (CAPD) patients using subcutaneous administration of recombinant human erythropoietin fortnightly--a multicenter trial].

Eighty-seven CAPD patients whose hematocrit (Ht) level was maintained by recombinant human erythropoietin (rHuEPO) were enrolled in this trial for a new formulation of rHuEPO suitable for subcutaneous injection. 6000IU rHuEPO was administered every 2 weeks for 12 weeks. Fortnightly doses were increased to 9000IU or 12000IU at 4 or 6 weeks if the Ht level decreased by 2% or more. During the study period, Ht values were maintained at the appropriate level in 88% of patients. 6000IU or lower was selected as a maintenance dose given every 2 weeks in 57 (76.0%) patients, 9000IU was selected in 8 patients and 12000IU was chosen in one patient. In 9 patients, the Ht could not be maintained during the study and the appropriate dose, therefore, remained unclear. Hypertension was observed in 2 patients as a side effect, and headache occurred in 2 other patients during the trial. Cutaneous abnormalities were not observed in the course of the subcutaneous injections. We conclude that anemia in most CAPD patients could be controlled with fortnightly injections in the dose range of 6000 to 12000IU rHuEPO given subcutaneously.

[Combined pancreas and kidney transplantation for IDDM patients with diabetic renal failure].

We performed 7 cases of pancreas transplantation (PTX), simultaneous pancreas and kidney transplantation in 4 cases, and PTX after kidney transplantation in 3 cases. The pancreas and kidney were extirpated after in situ perfusion using UW solution and stored in UW solution. The pancreas was transplanted in the left iliac fossa with bladder drainage, and the kidney was placed in the contralateral iliac fossa. The immunosuppressive regimen consisted of cyclosporine, methylprednisolone, azathioprine and antilymphocyte globulin. Gabexate mesilate (30-40 mg/kg/day) and PGE1 (5 ng/kg/min) was administered intravenously to prevent the vascular thrombosis. The original diseases of 7 patients were insulin-dependent diabetes mellitus (IDDM) with chronic renal failure, retinopathy and neuropathy. Six out of 7 patients became insulin-free after PTX, while one patient developed the vascular thrombosis in the pancreatic graft which was removed after 12 hours after the transplantation. All patients became dialysis-free and serum creatinine was ranging from 1.5 to 2.0 mg/dl. HbAlc remained within normal range in 6 out of 7 patients, who showed normal to borderline glucose tolerance in 75g oral glucose tolerance test. Although further investigation will be required, PTX from cardiac-arrest donor will be promising as one of the therapeutic modalities for IDDM patients.

[Current status and future prospect of pancreas and islet transplantation].

As of November 1997, 9,891 pancreas transplantation were reported to the International Pancreas Transplant Registry. In the United States, for all 1994-97, SPK, PAK, and PTA transplants, one-year graft survival rates were 82%, 71%, and 62%, respectively. The 1994-97 pancreas survival rates in all categories were higher than in previous eras. The improvement in graft survival rates has been associated with the introduction of FK506 and MMF, but excellent results are seen with cyclosporine, so the improvement may in part be due to the increasing experience that centers now have with pancreas transplantation. Although the standard surgical procedure remains pancreas-exocrine bladder drainage, the number of enteric drainage cases is increasing. It has been reported that the portal venous and enteric drainages are safe, with outcomes similar to those of standard technique. It appears that these will become the standard methods in the near future. Although 15 pancreas transplantations have been carried out in Japan, they ceased after 1994. Currently, social debate to determine the rules governing such procedures is ongoing. As of December 1995, 306 adult islet allotransplantation were reported to the Islet Transplant Registry. One-year islet survival rates were 27% in cases for 1990-94. Islet transplantation has the potential to be the most physiological advantage for the treatment of diabetes mellitus. However, the endocrine function provided by these transplants has been far from optimal.