Nobuhiro Miyamura

Effect on insulin sensitivity of angiotensin converting enzyme inhibitors with or without a sulphydryl group: bradykinin may improve insulin resistance in dogs and humans.

SummaryThe present study compared the effect on insulin sensitivity of ACE inhibitors with a sulphydryl group (captopril) or those without a sulphydryl group (delapril and enalapril) during the hyperinsulinaemic euglycaemic clamp test in both animal and clinical experiments. A possible contribution of bradykinin to the improvement of insulin sensitivity by ACE-inhibition was also studied. In healthy control and depancreatized dog experiments, administration of captopril either intravenously (3.0 mmol · kg−1) or orally (5.0 mmol · kg−1) increased insulin sensitivity indices and plasma bradykinin concentrations. In comparison, intravenous administration of an active metabolite of delapril (3.0 mmol · kg−1) and oral administration of either delapril or enalapril (5.0 mmol · kg−1) showed slight, but not significant increases in insulin sensitivity indices and plasma bradykinin concentrations. Infusion of a bradykinin antagonist (N-α-adamantaneacetyl-d-Arg-[Hyp3, Thi5,8,d-Phe7]-bradykinin) (0.5 nmol · kg−1 · min−1) abolished the effect of captopril on insulin sensitivity. Furthermore, intravenous administration of bradykinin (0.1 nmol · kg−1 · min−1) increased insulin sensitivity indices. In clinical experiments, insulin sensitivity indices decreased in the following order: normotensive healthy subjects, hypertensive non-diabetic patients, normotensive NIDDM patients and hypertensive NIDDM patients. In these four groups, oral administration of captopril (2.0 mmol · kg−1) significantly increased insulin sensitivity indices, and a concomitant increase in plasma bradykinin concentrations was observed. By contrast, oral administration of enalapril or delapril showed slight, but not significant effects on insulin sensitivity indices and plasma bradykinin concentrations. From these studies, it is concluded that ACE inhibitors with a sulphydryl group have more potent action on the improvement in insulin sensitivity than those without a sulphydryl group. Bradykinin may also possibly be involved in the mechanism underlying the improvement in insulin sensitivity associated with ACE-inhibition.

Bradykinin enhances GLUT4 translocation through the increase of insulin receptor tyrosine kinase in primary adipocytes: Evidence that bradykinin stimulates the insulin signalling pathway

SummaryIt has been suggested that bradykinin stimulates glucose uptake in experiments in vivo and in cultured cells. However, its mechanism has not yet been fully elucidated. In this study, the effects of bradykinin on the insulin signalling pathway were evaluated in isolated dog adipocytes. The bradykinin receptor binding study revealed that dog adipocytes possessed significant numbers of bradykinin receptors (Kd=83 pmol/l, binding sites = 1.7×104 site/cell). Reverse transcription-polymerase chain reaction amplification showed the mRNA specific for bradykinin B2 receptor in the adipocytes. Bradykinin alone did not increase 2-deoxyglucose uptake in adipocytes; however, in the presence of insulin (10−7 mol/l) it significantly increased 2-deoxyglucose uptake in a dose-dependent manner. Bradykinin also enhanced insulin stimulated GLUT4 translocation from the intracellular fraction to the cell membrane, and insulin induced phosphorylation of the insulin receptor Β subunit and insulin receptor substrate-1 (IRS-1) without affecting the binding affinities or numbers of cell surface insulin receptors in dog adipocytes. The time-course of insulin stimulated phosphorylation of the insulin receptor Β subunit revealed that phosphorylation reached significantly higher levels at 10 min, and stayed at the higher levels until 120 min in the presence of bradykinin, suggesting that bradykinin delayed the dephosphorylation of the insulin receptor. It is concluded that bradykinin could potentiate insulin induced glucose uptake through GLUT4 translocation. This effect could be explained by the potency of bradykinin to upregulate the insulin receptor tyrosine kinase activity which stimulates phosphorylation of IRS-1, followed by GLUT4 translocation.

Inherited adrenocorticotropin-independent macronodular adrenal hyperplasia with abnormal cortisol secretion by vasopressin and catecholamines : Detection of the aberrant hormone receptors on adrenal gland ()

ACTH-independent macronodular adrenal hyperplasia (AIMAH) is a rare disorder and an unusual cause of Cushing’s syndrome, of which familial transmission has rarely been reported. In this study, a mother and her son, the former affected with definite AIMAH and the latter with possible AIMAH, are described. Although the mother manifested overt Cushing’s syndrome, her son remained with no stigmata of Cushing’s syndrome except for bilateral adrenal tumor and mild hypertension, and a full suppression of plasma cortisol by low-dose dexamethasone was observed in him. Recently, aberrant expression of adrenal receptors for various ligands has been noted in AIMAH patients. In our cases, provocation tests in vivo suggested that AVP and catecholamines promoted cortisol production through V1a and/or V1b receptors and via β-adrenergic receptor, respectively. Reverse transcriptional-PCR analysis of the operated adrenal tissues of mother revealed the abnormal expression of mRNA of receptors for V1b, V2, and LH/hCG, none of which was observed in a normal control. Inherited AIMAH is very rare, and the son might be at the earliest developmental stage of AIMAH among the cases reported so far. An intervention could be tried to prevent the development of overt Cushing’s syndrome by suppression of the possible endogenous ligands or by blockade of the receptors that may be aberrantly expressed in his adrenal glands.

Expression of insulin receptor on clonal pancreatic alpha cells and its possible role for insulin-stimulated negative regulation of glucagon secretion

SummaryIn pancreatic alpha cells, the existence and function of the insulin receptor has not yet been fully established. In this study, to confirm the expression of functional insulin receptors in pancreatic alpha cells, we performed: 1) insulin receptor binding assay, 2) Northern blot analysis and RT-PCR (reverse transcription-polymerase chain reaction) amplification of insulin receptor mRNA, 3) immunocytochemical staining, 4) biosynthetic labelling of insulin receptor protein using [35S]methionine, 5) analysis of insulin-stimulated autophosphorylation of the insulin receptor in glucagon secreting cell lines, In-R1-G9 and αTC clone 6 cells. Glucagon secretion decreased with the addition of insulin in both cells. The receptor binding studies using [125I-Tyr-A14] insulin revealed that both cells possessed a significant number of insulin receptors (In-R1-G9: K1=2.1×109mol/l−1, K2=6.2×107 mol/l−1, R1=0.2×104, R2=1.86×104 sites/cell; αTC clone 6: K1=2.1×109 mol/l−1, K2=7.3×107 mol/l−1, R1=0.27×104, R2=1.95×104 sites/cell). Northern blot analysis as well as RT-PCR amplification showed the mRNA specific for insulin receptor in both cells. By immunocytochemical staining using anti-insulin receptor α-subunit antibody, positive immunostaining for insulin receptor was observed in both cells. [35S]Methionine labelling of both cells followed by immunoprecipitation using anti-insulin receptor antibody showed the correct size of the insulin receptor protein. The insulin receptor expressed in these cells underwent autophosphorylation by insulin stimulation. It is concluded that functional insulin receptors are properly expressed in In-R1-G9 and αTC clone 6 cells.

A carboxy-terminal truncation of human alpha-galactosidase A in a heterozygous female with Fabry disease and modification of the enzymatic activity by the carboxy-terminal domain. Increased, reduced, or absent enzyme activity depending on number of amino acid residues deleted.

Fabry disease is an X-linked disorder of glycosphingolipid metabolism caused by a deficiency of alpha-galactosidase A (alpha-Gal A). We identified a novel mutation of alpha-Gal A gene in a family with Fabry disease, which converted a tyrosine at codon 365 to a stop and resulted in a truncation of the carboxy (C) terminus by 65 amino acid (AA) residues. In a heterozygote of this family, although the mutant and normal alleles were equally transcribed in cultured fibroblasts, lymphocyte alpha-Gal A activity was approximately 30% of the normal control and severe clinical symptoms were apparent. COS-1 cells transfected with this mutant cDNA showed a complete loss of its enzymatic activity. Furthermore, those cotransfected with mutant and wildtype cDNAs showed a lower alpha-Gal A activity than those with wild type alone (approximately 30% of wild type alone), which suggested the dominant negative effect of this mutation and implied the importance of the C terminus for its activity. Thus, we generated mutant cDNAs with various deletion of the C terminus, and analyzed. Unexpectedly, alpha-Gal A activity was enhanced by up to sixfold compared with wild-type when from 2 to 10 AA residues were deleted. In contrast, deletion of 12 or more AA acid residues resulted in a complete loss of enzyme activity. Our data suggest that the C-terminal region of alpha-Gal A plays an important role in the regulation of its enzyme activity.

Nifedipine Induces Peroxisome Proliferator-Activated Receptor-γ Activation in Macrophages and Suppresses the Progression of Atherosclerosis in Apolipoprotein E-Deficient Mice

Objective—Nifedipine, an L-type calcium channel blocker, protects against the progression of atherosclerosis. We investigated the molecular basis of the antiatherosclerotic effect of nifedipine in macrophages and apolipoprotein E-deficient mice. Methods and Results—In macrophages, nifedipine increased peroxisome proliferator-activated receptor-&ggr; (PPAR&ggr;) activity without increasing PPAR&ggr;-binding activity. Amlodipine, another L-type calcium channel blocker, and 1,2-bis-(o-aminophenoxy)-ethane-N,N,-N′,N′-tetraacetic acid tetraacetoxy-methyl ester (BAPTA-AM), a calcium chelator, decreased PPAR&ggr; activity, suggesting that nifedipine does not activate PPAR&ggr; via calcium channel blocker activity. Inactivation of extracellular signal-regulated kinase 1/2 suppressed PPAR&ggr;2-Ser112 phosphorylation and induced PPAR&ggr; activation. Nifedipine suppressed extracellular signal-regulated kinase 1/2 activation and PPAR&ggr;2-Ser112 phosphorylation, and mutating PPAR&ggr;2-Ser112 to Ala abrogated nifedipine-mediated PPAR&ggr; activation. These results suggested that nifedipine inhibited extracellular signal-regulated kinase 1/2 activity and PPAR&ggr;2-Ser112 phosphorylation, leading to PPAR&ggr; activation. Nifedipine inhibited lipopolysaccharide-induced monocyte chemoattractant protein-1 expression and induced ATP-binding cassette transporter A1 mRNA expression, and these effects were abrogated by small interfering RNA for PPAR&ggr;. Furthermore, in apolipoprotein E-deficient mice, nifedipine treatment decreased atherosclerotic lesion size, phosphorylation of PPAR&ggr;2-Ser112 and extracellular signal-regulated kinase 1/2, and monocyte chemoattractant protein-1 mRNA expression and increased ATP-binding cassette transporter A1 expression in the aorta. Conclusion—Nifedipine unlike amlodipine inhibits PPAR&ggr;-Ser phosphorylation and activates PPAR&ggr; to suppress monocyte chemoattractant protein-1 expression and induce ATP-binding cassette transporter A1 expression in macrophages. These effects may induce antiatherogenic effects in hypertensive patients.

An acylic polyisoprenoid derivative, geranylgeranylacetone protects against visceral adiposity and insulin resistance in high-fat-fed mice

Induction of heat shock protein (HSP)72 improves insulin resistance and obesity in diabetic animal models. Geranylgeranylacetone (GGA), known as an antiulcer drug, induces HSP72 and protects organs against several cellular stresses. This study investigated whether GGA administration would induce HSP72 in liver and render physiological protection against high-fat feeding in mice. A single and 4-wk oral administration of 200 mg/kg GGA was performed in high-fat diet (HFD)-fed mice. Metabolic parameters, cytokines, and gene expressions related to insulin signaling were evaluated. A single administration of GGA induced HSP72 in liver of normal chow-fed and HFD-fed mice. Insulin resistance after HFD was slightly ameliorated. Four weeks of GGA administration also increased HSP72 in liver and significantly improved insulin resistance and glucose homeostasis upon glucose challenge. Activation of c-jun NH₂-terminal kinase (JNK) was attenuated, and insulin signaling was improved in the liver of HFD mice. Visceral adiposity was decreased in GGA-treated mice, accompanied by reduced leptin and increased adiponectin levels. GGA can be a novel therapeutic approach to treat metabolic syndrome as well as type 2 diabetes by improving insulin signaling and reducing adiposity. These beneficial effects of GGA could be mediated through HSP72 induction and JNK inactivation in the liver.

Sudden onset of diabetes with ketoacidosis in a patient treated with FK506/tacrolimus

We report a 43-year-old man who presented diabetic ketoacidosis 1 year after receiving kidney transplantation. He was a recipient of renal transplantation treated with metyl-prednisolone and tacrolimus regimen. The serum level of tacrolimus was 12.4 ng/ml, and he showed hyperphagia before a month of admission. A week before admission, he was aware of polydipsia, polyuria, and general fatigue. He visited our hospital and was found to have severe hyperglycemia (925 mg/dl), significant ketosis and mild metabolic acidosis (pH 7.341), although he had not been diagnosed as diabetes mellitus. He administrated in our hospital, and was treated with insulin for 5 weeks. He was not obese (BMI = 18.2 kg/m(2)) and had no family history of type 2 diabetes. He was finally treated with diet therapy alone. The 24 h urine C-peptide secretion on the third hospital day was low (8.4 microg per day). However, no autoantibodies against pancreatic islets were positive, and his insulin secretion was recovered at discharge suggesting that he was not type 1 diabetes. Although, tacrolimus has been reported to cause or worsen diabetes mellitus, the present case suggests that it could cause severe decrease in insulin secretion which leading to diabetic ketoacidosis in lean subject without previous history of diabetes mellitus.

Preclinical Cushing's syndrome resulting from black adrenal adenoma. A case report.

A 59-year-old Japanese woman, admitted for the treatment of diabetes mellitus and hypertension, was incidentally discovered to have a solid mass of 1.4 cm in diameter by CT scan with the attenuation value of 38 Hounsfield units, relatively higher for ordinary adrenal adenomas. Magnetic resonance imaging revealed no reduction of signal intensity on opposite-phase image on T1-weighted sequence. Adrenal scintigraphy imaging with 131I-adosterol did not show any uptake of the isotope in the area corresponding to both adrenals. Although she had no characteristic feature of overt Cushing’s syndrome, her serum cortisol level was not suppressed after an overnight dexamethasone administration. She was diagnosed as having preclinical Cushing’s syndrome. Left adrenalectomy was performed, revealing the well-circumscribed black tumor, mainly consisted of compact cell, in which cytoplasm was filled with numerous granules pigmented with dark to golden brown colors on hematoxylin-eosin staining. These findings suggested that her incidentaloma was a black adrenal adenoma. Production of steroid hormones was confirmed by immunohistochemical analysis of steroidogenic enzymes and by measurement of the tissue contents of hormones, whose levels were comparable with those in adenomas of overt Cushing’s syndrome. This is the first case report of preclinical Cushing’s syndrome resulting from black adrenal adenoma.

Heat shock treatment with mild electrical stimulation safely reduced inflammatory markers in healthy male subjects

SUMMARY OBJECTIVE Obesity induces chronic inflammation, which contributes to the development and progression of insulin resistance, diabetes and atherosclerosis. We have recently shown that induction of heat shock protein 72 by mild electric current and thermo (MET) treatment in mouse model of type 2 diabetes ameliorated glucose homeostasis and insulin resistance accompanied by reduced adiposity. For clinical application of MET, we confirmed its safety in healthy subjects. METHODS MET was applied for 10 healthy Japanese male (12 V, 55 pulses/s, 30 min at 42 °C) twice a week for 8 weeks. Fat volume was measured by CT scan and several parameters were investigated. RESULTS MET did not induce any adverse effects nor muscle contraction/pain. There were no significant alterations in glucose homeostasis or insulin resistance. Visceral and subcutaneous fat volume showed a trend of decrease without significant difference (-3.9% and -4.3%, respectively), which were restored 8 weeks after withdrawal of MET. Interestingly, serum tumor necrosis factor-α (TNF-α: 0.91 ± 0.05 pg/mL vs. 0.67 ± 0.06 pg/mL; p = 0.006) and high sensitivity C-reactive protein (hs-CRP: 521.9 ± 73.9 ng/mL vs. 270.8 ± 43.7 ng/mL; p = 0.023) levels, both of which are associated with chronic inflammation, were significantly decreased. CONCLUSION MET may be beneficial for the reduction of an inflammatory response observed in diabetes and metabolic syndrome.