Koji Nojima

Prevention and Treatment of Obesity, Insulin Resistance, and Diabetes by Bile Acid–Binding Resin

Bile acid–binding resins, such as cholestyramine and colestimide, have been clinically used as cholesterol-lowering agents. These agents bind bile acids in the intestine and reduce enterohepatic circulation of bile acids, leading to accelerated conversion of cholesterol to bile acids. A significant improvement in glycemic control was reported in patients with type 2 diabetes whose hyperlipidemia was treated with bile acid–binding resins. To confirm the effect of such drugs on glucose metabolism and to investigate the underlying mechanisms, an animal model of type 2 diabetes was given a high-fat diet with and without colestimide. Diet-induced obesity and fatty liver were markedly ameliorated by colestimide without decreasing the food intake. Hyperglycemia, insulin resistance, and insulin response to glucose, as well as dyslipidemia, were markedly and significantly ameliorated by the treatment. Gene expression of the liver indicated reduced expression of small heterodimer partner, a pleiotropic regulator of diverse metabolic pathways, as well as genes for both fatty acid synthesis and gluconeogenesis, by treatment with colestimide. This study provides a molecular basis for a link between bile acids and glucose metabolism and suggests the bile acid metabolism pathway as a novel therapeutic target for the treatment of obesity, insulin resistance, and type 2 diabetes.

Age-related association of MHC class I chain-related gene A (MICA) with type 1 (insulin-dependent) diabetes mellitus

To assess the contribution of the HLA class I region to susceptibility to and heterogeneity of type 1 diabetes, we investigated the association of polymorphism of MHC class I chain-related gene A (MICA) with age-at-onset as well as susceptibility to type 1 diabetes. One hundred one Japanese patients and 110 healthy control subjects were studied. The frequency of A4 allele was significantly higher and that of A6 allele was significantly lower in patients than in control subjects. The frequency of A5.1 allele was highest in early-onset patients (23.0%), intermediate in intermediate-onset patients (9.2%) and lowest in late-onset patients (7.7%) (trend chi-squared test, p = 0.0098). A5. 1 allele was strongly associated with HLA-B7 and Cw7, suggesting that MICA*A5.1-B7-Cw7 haplotype contains a gene responsible for age-at-onset. A4 allele was associated with a susceptible haplotype, DR4-DQB1*0401, and A6 allele was associated with a protective haplotype, DR2-DQB1*0601, suggesting that the association of MICA with type 1 diabetes susceptibility may be due to linkage disequilibrium with class II haplotypes. These data suggest that MICA gene is associated with age-at-onset and that a gene (or genes) responsible for age-at-onset of type 1 diabetes is located in the HLA class I region, probably near the region of MICA-B-C.

Age-dependent changes in phenotypes and candidate gene analysis in a polygenic animal model of Type II diabetes mellitus; NSY mouse.

Aims/hypothesis. The Nagoya-Shibata-Yasuda (NSY) mouse closely mimics human Type II (non-insulin-dependent) diabetes mellitus in that the onset is age-dependent, the animals are not severely obese, and both insulin resistance and impaired insulin response to glucose contribute to disease development. The aim of this study was to clarify the influence of age on the pathogenesis of diabetes and to analyse a candidate gene for Type II diabetes in this strain.¶Methods. Several phenotypic characteristics related to diabetes mellitus were monitored longitudinally in male NSY and control C3H/He mice. The nucleotide sequence of Glut4, a candidate gene for Nidd1nsy (a susceptibility gene for Type II diabetes) on Chromosome 11, encoding insulin-sensitive glucose transporter, was determined in NSY and C3H mice.¶Results. Glucose intolerance worsened with age, and fasting blood glucose and fasting plasma insulin concentration increased with age in NSY mice. Pancreatic insulin content increased until 24 weeks of age but then decreased at 48 weeks of age in NSY mice. The hypoglycaemic response to insulin was statistically significantly smaller in NSY than in C3H/He mice. The nucleotide sequence of GLUT4 cDNA was identical in NSY and C3H/He mice, but both were different from the sequence reported previously.¶Conclusion/interpretation. Insulin secretion and insulin resistance, as well as ageing possibly play an important part in the disease development in NSY mice. A decline of pancreatic insulin content in older age might cause the relative insulin deficiency in this strain. Nucleotide sequencing suggests that Glut4 is unlikely to be a candidate gene for Nidd1nsy. [Diabetologia (2000) 43: 932–938]

Involvement of microsomal triglyceride transfer protein in nonalcoholic steatohepatitis in novel spontaneous mouse model

BACKGROUND & AIMS Nonalcoholic fatty liver disease (NAFLD) is currently recognized as a global health issue and encompasses a wide spectrum of entities, ranging from simple hepatic steatosis to nonalcoholic steatohepatitis (NASH). The lack of a spontaneous animal model of NASH, however, has hampered basic research in this field. METHODS We examined the hepatic lesions in the inbred Fatty Liver Shionogi (FLS) mouse, which exhibits type 2 diabetes, and investigated the molecular mechanism leading to NAFLD/NASH. Using vector-mediated hepatic expression of microsomal triglyceride transfer protein (MTP), a key molecule for very low density lipoprotein (VLDL) assembly and export, its contribution to the hepatic lesions as well as to glucose intolerance was examined. RESULTS The FLS mouse, maintained on normal chow, exhibited excessive hepatic triglyceride (TG) accumulation due to impaired VLDL secretion, and subsequently hepatic lesions comparable to NASH, with increased expression of inflammatory molecules as well as insulin resistance. Gene expression and Western blot analyses demonstrated reduced hepatic expression of MTP in the FLS mouse. Hepatic induction of MTP resulted in a reduction in hepatic TG accumulation, improvement of VLDL export, and amelioration of NASH-like lesions, as well as glucose intolerance. CONCLUSIONS These data suggest that the FLS mouse could serve as a spontaneous model of NASH with insulin resistance, and that reduced MTP is involved in the development of NASH, pointing towards MTP as a critical target for the prevention and treatment of NASH.

Association of plasma fibrinogen level and blood pressure with diabetic retinopathy, and renal complications associated with proliferative diabetic retinopathy, in Type 2 diabetes mellitus

Aim To clarify the association of several clinical parameters, including plasma fibrinogen level, with diabetic retinopathy in patients with Type 2 diabetes mellitus (DM).

Allelic variation in class I K gene as candidate for a second component of MHC-linked susceptibility to Type 1 diabetes in non-obese diabetic mice

Aims/hypothesisRecent studies have revealed that MHC-linked susceptibility to Type 1 diabetes is determined by multiple components. In the non-obese diabetic (NOD) mouse, a second component (Idd16) has been mapped to a region adjacent to, but distinct from Idd1 in the class II region. In this study, we investigated the class I K gene as a candidate gene for Idd16.MethodsWe determined the genomic sequences of the class I K gene as well as the reactivity of K molecules with monoclonal antibodies in the NOD mouse, the Cataract Shionogi (CTS) mouse, and the NOD.CTS-H-2 congenic strain, which possesses a resistance allele to Type 1 diabetes at the Idd16 on the NOD genetic background genes.ResultsWhile the K sequence of the NOD mouse was identical to that of Kd type, ten nucleotide substitutions were identified in the CTS mouse compared with the NOD mouse. Of these, three were in exon 4, giving two amino acid substitutions, which were identical to those seen in KK type. These characteristics were retained in the NOD.CTS-H-2 congenic strain, which had a lower incidence and delayed onset of Type 1 diabetes owing to a resistance allele at Idd16. Lymphocytes from NOD.CTS-H2 congenic mice reacted with anti-Kd and anti-Kk monoclonal antibodies, reflecting the unique sequence of the K gene. The nucleotide sequence of the K gene in the non-obese non-diabetic (NON) mouse was also unique, consisting of a combination of Kk- and Kb-like sequences.Conclusions/interpretationThese data suggest that H2-K is unique in CTS and NON mice, and that allelic variation of the class I K gene may be responsible for Idd16.

Asp905Tyr Polymorphism of the Gene for the Skeletal Muscle-Specific Glycogen-Targeting Subunit of Protein Phosphatase 1 in NIDDM

OBJECTIVE To clarify the contribution of the Asp905Tyr polymorphism of the musclespecific glycogen-targeting subunit of protein phosphatase 1 (PP1G) to insulin resistance and related diseases. RESEARCH DESIGN AND METHODS We investigated the Asp905Tyr polymorphism of the PPP1R3 gene, which encodes the muscle-specific glycogen-targeting subunit of PP1G, in 259 Japanese patients with NIDDM and 194 healthy control subjects. RESULTS No significant difference was found in the genotype distribution between NIDDM patients (n = 259; Asp/Asp = 0.10, Asp/Tyr = 0.44, Tyr/Tyr = 0.46) and healthy control subjects (n = 194; Asp/Asp = 0.13, Asp/Tyr = 0.37, Tyr/Tyr = 0.50) or between patient groups subdivided by the mode of treatment: NIDDM patients with insulin therapy (Asp/Asp = 0.14, Asp/Tyr = 0.46, Tyr/Tyr = 0.40) and those without insulin therapy (Asp/Asp = 0.07, Asp/Tyr = 0.43, Tyr/Tyr = 0.50). However, NIDDM patients with the Tyr allele, which was previously reported to be associated with insulin resistance, tended to have lower BMIs than those without this allele (Asp/Asp: 24.5 ±1.1 kg/m2, Asp/Tyr: 22.6 ± 0.4 kg/m2, Tyr/Tyr: 22.8 + 0.3 kg/m2, P = 0.06 by analysis of variance). CONCLUSIONS These data suggest that the Asp905Tyr polymorphism of the PPP1R3 gene is not associated with NIDDM or high BMI, both of which are known to be insulin-resistant states, in the Japanese population.

Fatty liver and obesity : phenotypically correlated but genetically distinct traits in a mouse model of type 2 diabetes

Aims/hypothesisObesity and fatty liver are commonly associated with type 2 diabetes, but the genetic and functional bases linking fatty liver with obesity and diabetes are largely unknown. Our aim was to investigate the association of fatty liver with obesity and other diabetes-related phenotypes and to define the genetic control of obesity and fatty liver.Materials and methodsWe established 306 F2 mice by crossing Nagoya–Shibata–Yasuda (NSY) mice, an animal model of type 2 diabetes, with control C3H mice, and analysed their phenotypes. Whole-genome screening of F2 mice was performed to identify the loci responsible for fatty liver and obesity.ResultsA strong association of fatty liver with obesity, hyperinsulinaemia and hyperglycaemia was observed in F2 mice. Using whole-genome screening in 306 F2 mice, we mapped a new locus for fatty liver (Fl1n) on chromosome 6 (maximum logarithm of odds score [MLS] 10.0) and one for body weight (Bw1n) on chromosome 7 (MLS 5.1). Fl1n was linked to epididymal fat weight as well as fatty liver, but its effects were opposite in the two tissues in that the NSY allele increased liver fat but decreased epididymal fat, suggesting a role of Fl1n in partitioning of fat mass. The sequence of peroxisome proliferator-activated receptor γ (Pparg), a candidate for Fl1n, showed allelic variation between NSY and C3H mice.Conclusions/interpretationThese data suggest that fatty liver and obesity are phenotypically related but genetically independent. Loci homologous to Fl1n and Bw1n are good candidate genes for susceptibility to fatty liver and obesity in humans.

Paternal-maternal effects on phenotypic characteristics in spontaneously diabetic Nagoya-Shibata-Yasuda mice

The Nagoya-Shibata-Yasuda (NSY) mouse is an inbred strain with spontaneous development of type 2 (non-insulin-dependent) diabetes mellitus. The purpose of this study was to determine the mode of inheritance of various phenotypes related to diabetes in this strain. Two reciprocal outcrosses, female C3H/He x male NSY F1 (C3NF1) and female NSY x male C3H/He F1 (NC3F1) mice, were performed. The phenotypic characteristics in both F1 mice were investigated. The cumulative incidence of diabetes was 100% (25 of 25) in male C3NF1 mice and 97% (29 of 30) in male NC3F1 mice at 48 weeks of age, indicating that diabetes in NSY mice was transmitted to male F1 hybrids in an autosomal dominant manner. Fatty liver also showed an autosomal dominant mode of inheritance. In contrast, epididymal fat accumulation and impaired insulin secretion showed an autosomal recessive mode of inheritance. The body mass index (BMI) showed a codominant mode of inheritance. Paternal-maternal effects associated with the severity of diabetes were observed. Insulin resistance was much more severe in male F1 mice than in the parental NSY strain. These data indicate different modes of inheritance among phenotypes related to type 2 diabetes. The presence of more severe insulin resistance in F1 mice versus the parental strains suggests the interaction of both parental genomes in the development of insulin resistance. The F1 mouse is expected to be useful for studies of the pathogenesis and genetic synergism of the insulin resistance syndrome.

Direct evidence for susceptibility genes for type 2 diabetes on mouse chromosomes 11 and 14

Aims/hypothesisDiabetogenic loci for type 2 diabetes have been mapped to mouse chromosome (Chr) 11 and 14 in the Nagoya–Shibata–Yasuda (NSY) mouse, an animal model of type 2 diabetes. We aimed to obtain direct evidence of these genes on each chromosome and to clarify their function and interaction in conferring susceptibility to type 2 diabetes.MethodsWe established three consomic strains homozygous for diabetogenic NSY-Chr11, NSY-Chr14 or both on the control C3H background (C3H-11NSY, C3H-14NSY and C3H-11NSY14NSY, respectively), and monitored diabetes-related phenotypes longitudinally. The glucokinase gene was sequenced as a positional candidate gene on Chr11.ResultsC3H-11NSY mice showed hyperglycaemia associated with impaired insulin secretion and age-dependent insulin resistance without obesity. C3H-14NSY mice exhibited hyperglycaemia mainly due to insulin resistance, with a slight increase in percentage body fat. C3H-11NSY14NSY double consomic mice showed marked hyperglycaemia and obesity, which was not observed in single consomic strains. Sequences of the glucokinase gene were allelically variant between NSY and C3H mice.Conclusions/interpretationThese data provide direct evidence that Chr11 and Chr14 harbour major susceptibility genes for type 2 diabetes. These two chromosomes interact to cause more severe hyperglycaemia and obesity, which was not observed with the presence of either single chromosome, indicating different modes of gene–gene interaction depending on the phenotype. Marked changes in the phenotypes retained in the consomic strains will facilitate fine mapping and the identification of the responsible genes and their interaction with each other, other genes and environmental factors.