Tomoya Hamaguchi

Three new mutations in the hepatocyte nuclear factor-1α gene in Japanese subjects with diabetes mellitus: clinical features and functional characterization

Aims/hypothesis. Mutations in the hepatocyte nuclear factor-1α gene are a common cause of the type 3 form of maturity-onset diabetes of the young. We examined the clinical features and molecular basis of hepatocyte nuclear factor-1α (HNF-1α) diabetes. Methods. Thirty-seven Japanese subjects with early onset Type II (non-insulin-dependent) diabetes mellitus and 45 with Type I (insulin-dependent) diabetes mellitus were screened for mutations in this gene. Functional properties of mutant HNF-1α were also investigated. Results. Three new mutations [G415R, R272C and A site of the promoter ( + 102G-to-C)] were found. Insulin secretion was impaired in the three subjects. Insulin and glucagon secretory responses to arginine in the subject with the R272C mutation were also diminished. Molecular biological studies indicated that the G415R mutation generated a protein with about 50 % of the activity of wild-type HNF-1α. The R272C mutation had no transactivating or DNA binding activity and acted in a dominant negative manner. The + 102 G-to-C mutation in the A site of the promoter activity was associated with an increase in promoter activity and it had 42–75 % more activity than the wild-type sequence. Conclusion/interpretation. Mutations in the HNF-1α gene may affect the normal islet function by different molecular mechanisms. [Diabetologia (1999) 42: 621–626]

Tissue specificity in expression and alternative RNA splicing of human phosphofructokinase-M and -L genes

Mode of the expression of phosphofructokinase (PFK) -M and -L genes was examined in various human tissues including muscle, placenta, liver, kidney, pancreas, stomach and reticulocytes. The gross level of mRNA expression of PFK-M and -L genes was estimated by Northern analysis. Polymerase chain reaction was used to detect mRNA expressed at low levels in these tissues. Tissue-specific expression of alternatively spliced PFK-M gene transcripts was also determined by polymerase chain reaction. The results indicated that alternative splicing of PFK-M gene transcripts was controlled in a tissue-specific manner.

Dominant TCR α-chain clonotypes and interferon-γ are expressed in the pancreas of patients with recent-onset insulin-dependent diabetes mellitus

In order to clarify the nature of T lymphocytes infiltrating the pancreatic islets of patients with insulin-dependent diabetes mellitus (IDDM), we analysed T cell receptor (TCR) gene transcripts expressed in pancreatic biopsy specimens of patients with recent-onset IDDM. We also investigated the expression of cytokines (interferon-gamma: IFN-gamma; tumour necrosis factor-alpha: TNF-alpha; interleukin-4: IL-4; interleukin-6: IL-6) in the same specimens. The TCR V beta repertoire was not restricted either in the pancreas or the peripheral lymphocytes of IDDM patients. In contrast, the TCR V alpha repertoire was restricted in the pancreas, but not in the peripheral blood lymphocytes, of IDDM patients. The sequence analysis of the complementarity-determining region 3 (CDR3) of the TCR alpha revealed the presence of dominant clonality in alpha chains of T cells in the patients. IFN-gamma mRNA was highly expressed in the pancreas of IDDM patients, while IL-4 mRNA was deficient. A lower level of expression of IL-6 mRNA was detected in the IDDM pancreas than in the control tissue. These results indicate that T cells bearing a distinct TCR alpha chain are selectively retained and activated within the pancreas of recent-onset IDDM.

Risk factors for the progression of microalbuminuria in Japanese type 2 diabetic patients--a 10 year follow-up study.

To clarify risk factors for the progression of microalbuminuria in Japanese type 2 diabetic patients, the longitudinal study for 10 years was conducted on 67 outpatients with type 2 diabetes, who had shown no overt proteinuria at baseline. The urinary albumin index (UAI) has been determined based on the mean of at least two random urine samples each year. Categories were defined as normoalbuminuria (UAI < 30.0 mg/g x Cr.), microalbuminuria (30.0 < or = UAI < 300.0), and macroalbuminuria (UAI > or = 300.0). Progression was defined as worsening of the category and/or more than doubling of the baseline UAI value. Multiple logistic regression analysis was performed using age, duration of diabetes, HbA1c, blood pressure, BMI, serum lipids, smoking habits, and alcohol consumption as independent variables and the progression of microalbuminuria as a dependent variable. Age and HbA1c were estimated as significant and independent variables. Furthermore, genetic polymorphisms of angiotensin I-converting enzyme (ACE) and angiotensinogen were analyzed to evaluate the genetic contribution. The D/D genotype of ACE was significantly more common in progressors than in non-progressors. These results suggest that glycemic control and age are important risk factors and the D/D genotype of ACE acts as a risk factor for the progression of microalbuminuria in Japanese type 2 diabetic patients.

Asymmetric dimethylarginine, a biomarker of cardiovascular complications in diabetes mellitus

Cardiovascular (CV) complications are an essential causal element of prospect in diabetes mellitus (DM), with carotid atherosclerosis being a common risk factor for prospective crisis of coronary artery diseases and/or cerebral infarction in DM subjects. From another point of view, asymmetric dimethylarginine (ADMA) has been established as an inhibitor of endogenous nitric oxide synthesis and the relationship between ADMA and arteriosclerosis has been reported. In our study with 87 type 2 DM (T2DM) patients, we have examined whether ADMA and other CV risk factors are the useful predictors of DMCV complications. After the measurement of the respective CV risk factors, we have followed the enrolled T2DM patients for 5 years. We have finally analyzed 77 patients. DMCV complications developed in 15 cases newly within 5 years, and 4 cases recurred. The concentrations of ADMA in plasma were markedly more elevated in 19 DM patients with CV complications than in 58 DM patients without CV complications. Urinary albumin (U-Alb), mean intimal-medial thickness (IMT) and ankle brachial index (ABI) were also higher in patients with CV complications. Multiple regression analyses showed that U-Alb had an influence on the high level of ADMA (standardized β = 6.59, P = 0.00014) independently of age, systolic BP, fibrinogen, mean IMT, plaque score, and ABI. The review indicates what is presently known regarding plasma ADMA that might be a new and meaningful biomarker of CV complications in DM subjects.

A genetic defect in muscle phosphofructokinase deficiency, a typical clinical entity presenting myogenic hyperuricemia.

Myogenic hyperuricemia is a common pathophysiologic feature of muscle glycogenoses1. Type VII glycogenosis is the one which lacks catalytic activity of phosphofructokinase, a key enzyme of glycolysis, in muscle (PFK-M)2. Energy crisis in the exercising muscles of the patient results in the abnormal purine degradation leading to a typical manifestation of myogenic hyperuricemia1,3. As a step to clarify the molecular basis of this abnormal purine metabolism, we have reported the cloning of full-length human PFK-M cDNA4. The sequence data enabled us to reconfirm the gene duplication mechanism as hypothesized in rabbit PFK-M gene5,6,7. We have also elucidated the existence of the alternative splicing and the alternative promoter system of human PFK-M gene8,9. In addition, we have recently reported the complete structure of human PFK-M gene10. In this paper, we report on the genetic defect in a patient with PFK-M deficiency as the typical clinical entity presenting myogenic hyperuricemia.

Autoantibodies to the insulin receptor impair clearance of plasma endogenous insulin

Plasma insulin clearance was studied in a patient with autoantibodies to the insulin receptor, manifesting persistent hyperinsulinemia associated with alternating hyper- and hypoglycemia. In the postabsorptive period, the plasma glucose level gradually decreased. To prevent the development of hypoglycemia, glucose was infused and the glycemic level was clamped at 50 mg/dl without insulin infusion. The plasma C-peptide level was below the detectable range during the clamp, indicating no appreciable secretion of insulin. The plasma insulin level declined exponentially with a markedly prolonged disappearance rate (half-time: 3.0 h) during the study. These results indicate that hyperinsulinemia in the postabsorptive period in this patient is attributable to the impairment of plasma insulin clearance through receptor-mediated mechanisms, and also confirm that the receptor plays the principal role in plasma insulin removal.