Kinsuke Tsuda

Inhibition of gastric inhibitory polypeptide signaling prevents obesity

Secretion of gastric inhibitory polypeptide (GIP), a duodenal hormone, is primarily induced by absorption of ingested fat. Here we describe a novel pathway of obesity promotion via GIP. Wild-type mice fed a high-fat diet exhibited both hypersecretion of GIP and extreme visceral and subcutaneous fat deposition with insulin resistance. In contrast, mice lacking the GIP receptor (Gipr−/−) fed a high-fat diet were clearly protected from both the obesity and the insulin resistance. Moreover, double-homozygous mice (Gipr−/−, Lepob/Lepob) generated by crossbreeding Gipr−/− and obese ob/ob (Lepob/Lepob) mice gained less weight and had lower adiposity than Lepob/Lepob mice. The Gipr−/− mice had a lower respiratory quotient and used fat as the preferred energy substrate, and were thus resistant to obesity. Therefore, GIP directly links overnutrition to obesity and it is a potential target for anti-obesity drugs.

Effects of Troglitazone (CS-045) on insulin secretion in isolated rat pancreatic islets and HIT cells: an insulinotropic mechanism distinct from glibenclamide

SummaryIn order to elucidate the direct effects of (±)-5-[4-(6-hydroxy-2, 5, 7, 8-tetramethylchroman-2-yl-methoxy) benzyl]-2,4-thiazolidinedione (Troglitazone), a newly-developed oral hypoglycaemic agent, on pancreatic beta-cell function, in vitro investigation of isolated rat pancreatic islets and a hamster beta-cell line (HIT cell) were performed. Troglitazone stimulates both glucose, and glibenclamide-induced insulin release at a concentration of 10−6 mol/l in these cells but, conversely, inhibits insulin secretion at 10−4 mol/l. Glucose uptake in HIT cells is similarly enhanced by 10−6 mol/l Troglitazone, but is reduced in the presence of 10−4 mol/l Troglitazone. However, a quantitative immunoblot analysis with a specific antibody for GLUT 2 glucose transporter revealed no significant change in GLUT 2 protein in HIT cells with 10−6 mol/l Troglitazone. Specific binding of [3H]-glibenclamide to beta-cell membranes is replaced by Troglitazone in a non-competitive manner, but 10−6 mol/l Troglitazone failed to eliminate ATP-sensitive K++ channel activity. These results suggest that Troglitazone has a putative non-competitive binding site at, or in the vicinity of, the sulphonylurea receptor in rat pancreatic islets and HIT cells and that the dual effect of Troglitazone on insulin secretory capacity is mediated through the modulation of glucose transport activity, possibly due to the modification of intrinsic activity in glucose transporter in pancreatic beta cells by this novel agent. [Diabetologia (1995) 38: 24–30]

Abnormality in fibre type distribution of soleus and plantaris muscles in non-obese diabetic Goto-Kakizaki rats

1. Fibre type distributions of the slow soleus and fast plantaris muscles were investigated in 5‐, 9‐ and 20‐week‐old male Goto‐Kakizaki (GK) rats, as an animal model of non‐obese diabetes, and were compared with those of age‐matched non‐diabetic Wistar rats.

Identification of Two Missense Mutations in the GIP Receptor Gene: A Functional Study and Association Analysis with NIDDM: No Evidence of Association with Japanese NIDDM Subjects

Gastric inhibitory polypeptide (GIP) potently stimulates insulin secretion from pancreatic islets in the presence of glucose as an incretin. Because the insulinotropic effect of GIP is reduced in NIDDM, it should be clarified whether defects in the GIP receptor gene contribute to the impaired insulin secretion in NIDDM. Using genomic DNA samples from Japanese NIDDM and non-NIDDM subjects, we have investigated the entire coding region of the GIP receptor gene by polymerase chain reaction-single strand conformational polymorphism (PCR-SSCP). We have identified two missense mutations, Gly198→Cys (Gly198Cys) in exon 7 and Glu354→Gln (Glu354Gln) in exon 12. Investigation of the function of GIP receptor with either of these mutations reveals a half-maximal stimulation value of GIP-induced cAMP response in Chinese hamster ovary cells expressing the GIP receptor with Gly198Cys of 6.3 ± 1.2 × 10−10 mol/l (n = 3), which was considerably higher than that of the normal GIP receptor, 9.4 ± 3.8 × 10−12 mol/l GIP (n = 3), whereas that of the GIP receptor with Glu354Gln was not significantly different from that of the normal GIP receptor. To assess the possible role of the GIP receptor gene in genetic susceptibility to NIDDM, we have examined the allelic frequencies of Gly198Cys and Glu354Gln in NIDDM and control subjects. Association studies show no relationship between NIDDM and either of the two mutations.

Inhibitor for advanced glycation end products formation attenuates hypertension and oxidative damage in genetic hypertensive rats.

Objective A recent study demonstrated that free radicals were involved in the maintenance of hypertension in stroke-prone spontaneously hypertensive rats (SHRSP). Advanced glycation end-products (AGEs) accumulate progressively in the vasculature with ageing, and have been identified to be relevant mediators for various vascular complications. To elucidate the role of AGEs in genetic hypertension, we investigated the effect of OPB-9195, a novel inhibitor of AGEs, on hypertension and oxidative damage in SHRSP. Methods Five-week-old male SHRSP were divided into a control group, fed a control diet and two, OPB-9195, (±)-2-isopropylidenehydrazono-4-oxo-thiazolidin-5-ylacetanilide, treatment groups, fed a diet supplemented with OPB-9195 at the concentration of 0.5 (OPB-L) or 2 mg/g (OPB-H) mixed chow for 10 weeks. Results The plasma of OPB-9195-treated SHRSP had lower levels of glycated albumin as compared with that of control SHRSP. OPB-9195 lowered the systolic blood pressure (SBP) by the fourth week of administration, and this effect was maintained throughout the study. We also confirmed SBP and diastolic blood pressure (DBP) rhythms, monitored by telemetry, were significantly lower in the OPB-H group than in the control group. Urinary nitric oxide (NO) excretion as well as the expression of endothelial NO synthase (eNOS) mRNA, and eNOS activity in the aorta were significantly increased in OPB-9195-treated groups compared with the control group. The levels of 8-hydroxydeoxyguanosine (8-OHdG), produced from deoxyguanosine under conditions of oxidative stress, in the urine of OPB-9195-treated SHRSP was significantly lower than in the control SHRSP. We also confirmed that the expression of glutathione peroxidase in the aorta was significantly increased in OPB-9195 treated SHRSP. Conclusions Because long-term administration of a AGEs inhibitor reduces blood pressure and oxidative damage in SHRSP, this study suggests a role for AGEs in the progression or maintenance of hypertension and related diseases in genetic hypertension.

Starvation-induced Changes of Somatostatin, Glucagon, and Insulin Secretion from the Isolated Perfused Rat Pancreas

The effect of 16- and 48-h fasting on pancreatic somatostatin, insulin, and glucagon secretion was studied, using the isolated perfused rat pancreas. In the presence of 4.4 mM glucose, basal somatostatin and insulin concentrations in the perfusate were significantly lower in 48-h fasted rats than in fed animals, whereas basal glucagon secretion was significantly elevated in fasted rats. The infusion of 19 mM arginine significantly augmented secretion of somatostatin and glucagon and attenuated insulin secretion in 48-h fasted rats. It is concluded that fasting causes a decrease in basal pancreatic somatostatin secretion in vitro, although the response to arginine is rather exaggerated. Insulin and glucagon secretion also changed during the fasting. These results suggest that not only insulin and glucagon, but also somatostatin contribute to nutrient homeostasis.

Synergistic effect of polymorphisms of uncoupling protein 1 and β3-adrenergic receptor genes on autonomic nervous system activity

OBJECTIVE: To investigate the association of the promoter region −3826 A to G polymorphism of the uncoupling protein 1 (UCP1) gene with autonomic nervous system (ANS) activity and the interaction of the polymorphism with the Trp64Arg polymorphism of the β3 adrenergic receptor (β3AR).SUBJECTS: Three-hundred and forty-nine young (mean age 20.4±2.1 y old), healthy Japanese males.MEASUREMENTS: DNA was extracted from whole blood and genotyped by polymerase chain reaction restriction fragment length polymorphism. Plasma glucose, plasma insulin and body mass index (BMI) were measured. Frequency of family history of diabetes or obesity was determined by interview. Subjects randomly chosen from each genotype were examined for ANS activity during supine rest and standing by electrocardiogram power spectral analysis of heart rate variability.RESULTS: UCP1 or β3AR polymorphism was not associated with BMI, plasma glucose, plasma insulin and frequency of family history of diabetes or obesity. The inhibitory effect of UCP1 polymorphism on ANS activity was observed only with occurrence of the variant of β3AR. The very low frequency component associated with thermoregulation in the sympathetic nervous system of homozygotes of UCP1 (GG) at supine rest was significantly lower than normal (AA, 203.2±50.3 vs 462.2±83.6 ms2; mean±s.e., P=0.021). A higher response to postural change to standing was also observed in both sympathetic and parasympathetic nervous activities of AA than of GG.CONCLUSION: While UCP1 polymorphism alone does not affect ANS activity, it has a synergistic effect with β3AR polymorphism in decreasing sympathetic nervous system activity.

The cyclic AMP response element modulator family regulates the insulin gene transcription by interacting with transcription factor IID.

We analyzed a mechanism of transcriptional regulation of the human insulin gene by cyclic AMP response element modulator (CREM) through four cyclic AMP response elements (CREs). We isolated two novel CREM isoforms (CREMΔQ1 and CREMΔQ2), which lack one of the glutamine-rich domains, Q1 and Q2 respectively, and six known isoforms (CREMτα, CREMα, inducible cyclic AMP early repressor (ICER) I, ICER Iγ, CREM-17X, and CREM-17) from rat pancreatic islets and the RINm5F pancreatic β-cell line. CREM isoforms functioned as efficient transcriptional activators or repressors to modulate insulin promoter activity by binding to all of the insulin CREs. The binding activity of repressors is higher than that of activators and suppressed not only basal activity but also activator-induced activities. Furthermore, CREM activator interacted directly with the transcription factor IID components hTAFII130 and TATA box-binding protein (TBP). These results suggest that the activation of the insulin gene transcription by CREM activator is mediated by not only direct binding to the CREs but also by recruiting transcription factor IID to the insulin promoter via its interaction with hTAFII130 and TBP. On the other hand, the CREM repressor ICER competitively interrupts the binding of the activators to CREs and does not interact with either TBP or hTAFII130; therefore, it might fail to stabilize the basal transcriptional machinery and repress transactivation.

Suppressive effect of GABA on insulin secretion from the pancreatic beta-cells in the rat.

In order to investigate a possible role of GABA in the regulation of insulin secretion, we have studied the effect of GABA on insulin secretion from the isolated perfused rat pancreas in vitro and on the changes in the cytoplasmic Ca2+ of Beta-cells from the isolated rat islets. When glucose is present, GABA caused a dose dependent inhibition of the first phase of arginine-induced insulin secretion during the range of 10-1000 microM, but GABA did not affect arginine-induced insulin secretion in the absence of glucose. GABA inhibited not only the first phase but also the second phase of glucose-induced insulin secretion. A GABAB-receptor agonist, baclofen, also inhibited both phases of insulin secretion induced by 16.7 mM glucose. Furthermore, GABA inhibited the rise in cytoplasmic Ca2+ of Beta-cells in response to 16.7 mM glucose. These studies indicate that GABA decreases Beta cell secretory activity mainly in response to glucose. These inhibitory effects of GABA on insulin secretion may be mediated through GABAB-receptor and the inhibition of the rise in cytoplasmic Ca2+.

Somatostatin-like Immunoreactivity in Human Peripheral Plasma Measured by Radioimmunoassay Following Affinity Chromatography

Somatostatin-like immunoreactivity (SLI) concentrations were determined in human peripheral plasma using affinity chromatography followed by radioimmunoassay. In normal subjects, fasting SLI ranged from 2.9 to 22.0 pg/ml with a mean ± SE value of 10.2 ± 2.1 pg/ml. In totally pancreatectomized or gastrectomized patients, fasting SLI levels were not different from the values in normal subjects. In patients with medullary thyroid carcinoma, fasting SLI ranged from 11.8 to 71.0 pg/ml with a mean of 29.3 ± 12.3 pg/ml, which was significantly higher than normal values (P < 0.01). Following meal ingestion, plasma SLI increased significantly in normal subjects from a basal level of 9.1 ± 2.1 pg/ml to a peak value of 15.4 ± 2.9 pg/ml (P < 0.02). These results indicate that radioimmunoassay combined with affinity chromatography provides an accurate method of measuring SLI in human plasma.