Introduction: This study aimed to examine the association between severity of diabetic neuropathy and weight loss during hospitalization in overweight participants with type 2 diabetes. Patients and Methods: Participants of this study comprised 193 patients who were hospitalized for type 2 diabetes treatment. The participants were divided into two groups in the study, based on whether or not reduction of bodyweight was at least 3% during hospitalization. Using Cox models, the association between severity of neuropathy and effectiveness of weight loss under a controlled diet was analyzed. Autonomic neuropathy was assessed on patient admission by R-R interval, as measured in an electrocardiogram (CVRR), and sensory neuropathy was assessed using both 128-Hz tuning-fork vibration and Achilles tendon re fl ex (ATR). Results: The adjusted hazard ratio for weight loss of at least 3% for CVRR was 1.17 (95% con fi dence interval 1.07 – 1.28, P=0.0006) and for vibration time 1.93 (1.01 – 3.68, P=0.045). After dividing CVRR and vibration time into tertiles based on participant number, the adjusted hazard ratio for the high tertile of CVRR was 2.17 (1.29 – 3.62, P=0.003), and for the long tertile of vibration time 1.84 (1.10 – 3.08, P=0.02), compared with the low and short tertiles, respectively. No association was detected between ATR category and weight loss. Conclusion: Severity of diabetic neuropathy was found to be a determinant in weight loss under a caloric restriction regimen for patients with type 2 diabetes. The results of the study suggest that the peripheral nervous system is involved in responses to medical intervention for treatment for type 2 diabetes including bodyweight management. SU, sulfonylureas; TZD, thiazolidine- dione; BG, biguanide; α -GI, alpha-glucosidase inhibitors; DPP-4I, dipeptidyl peptidase-4 inhibitors; SGLT2I, sodium-glucose linked transporter 2 inhibitors; GLP1RA, glucagon-like peptide 1 receptor agonist.

Aims. - It is well-known that chronic exposure to large amounts of ligand leads to downregulation of its receptor. It is not known, however, whether a GLP-1R agonist downregulates its receptor. For this reason, our study examined whether GLP-1R expression is reduced after long-term exposure to dulaglutide (Dula) in non-diabetic and diabetic mice. Methods. - Seven-week-old male db/db and db/m mice were given either Dula (0.6 mg/kg x 2/week) or a control vehicle (CTL) for 17 weeks. Various metabolic parameters, such as glucose-stimulated insulin secretion (GSIS), insulin and TG content in islets, were evaluated after the intervention. beta-cell-related gene expression was also analyzed by real-time RT-PCR. Results. - In db/m mice, GLP-1R expression in beta-cells did not decrease, not even after long-term administration of Dula, compared with control mice, while GLP-1R expression in 24-week-old db/db mice treated with Dula was augmented, rather than downregulated, compared with 24-week-old CTL db/db mice. This was probably due to improved glycaemic control. In db/db mice treated with Dula, food intake and blood glucose levels were significantly decreased up to 24 weeks of age compared with CTL db/db mice, and their expression levels of various beta-cell-related genes, insulin content and GSIS were also enhanced. In contrast, oxidative and endoplasmic reticulum stress, inflammation, fibrosis and apoptosis were suppressed with Dula treatment. Conclusion. - Dula exerts beneficial effects on glycaemic control and has long-lasting protective effects on pancreatic beta-cells. GLP-1R expression levels were not reduced at all in non-diabetic as well as diabetic mice despite long-term dulaglutide exposure. (C) 2017 Elsevier Masson SAS. All rights reserved.

Diabetes & Metabolism - In Press.Proof corrected by the author Available online since lundi 8 juin 2015

We studied the steroidogenetic action and concomitant subcellular redistribution of protein kinase C (PKC) in cortisol hypersecreting adrenal adenoma cells obtained from two patients with Cushing`s syndrome. Isolated adenoma cells were treated with 10-9 M and 10-6 M 12-0-tetradecanoyl phorbol-13-acetate (TPA) or 10-6 M ACTH. Treatment of the isolated adenoma cells with TPA resulted in cortisol secretion equivalent to that with ACTH-treatment. Immunoblot analysis of PKC during treatment with ACTH or TPA showed that PKC f3 translocated from cytosol to membrane. A small amount of PKCa, but not membrane-associated PKCa, was detectable in the cytosolic fraction. It appeared that TPA-induced cortisol secretion mimicked ACTH-induced cortisol secretion, and that PKC3 translocated from cytosol to membrane on stimulation by both ACTH and TPA. We suggested that ACTH-induced cortisol secretion in human cortisol hypersecreting adrenal adenoma is mediated by PKCI3 activation.

Background Aldosterone has essential roles in regulating intravascular volume and blood pressure, and is suggested to influence cardiac structure. However, the association of polymorphisms in the aldosterone synthase gene (CYP11B2) with hypertension or cardiac hypertrophy remains controversial. Objective To evaluate the distribution of polymorphisms in the CYP11B2 gene and the possible associations between genotypes and blood pressure, urinary excretion of aldosterone or electrolytes and echocardiographic measurements, in a Japanese population. Methods and results We examined the association of two common diallelic polymorphisms within CYP11B2, one in the promoter −344T/C and the other an intron 2 gene conversion, with blood pressure, 24-h urinary excretion of aldosterone and electrolytes, and echocardiographic measurements, in a Japanese population. We confirmed significant linkage disequilibrium between these polymorphic loci and ethnic differences in frequency of the alleles. The −344C and −344T haplotypes apparently diverged before the intron conversion polymorphism was generated on the latter haplotype. Allele frequencies did not differ between 535 normotensive and 360 hypertensive individuals or between hypertensive individuals with higher and lower concentrations of renin. The only significant correlation was a positive correlation of left ventricular mass with 24-h urinary excretion of sodium, which occurred only in individuals with the −344CC genotype or the intron 2 conversion (−/−) genotype. Conclusions The −344CC or intron 2 conversion (−/−) genotype in CYP11B2 may be a risk factor for developing sodium-sensitive cardiac hypertrophy. Ethnic differences in the distribution of CYP11B2 genotypes combined with differences in salt intake might account for inconsistencies between previous reports.

To clarify tissue-specificity of pancreatic beta cells, comparison of mRNA expression in various conditions of the tissue of multiple organisms is important. Although the developed methodologies for mRNA monitoring such as microarray, rely on the growth of dbEST (database of expressed sequence tag), a large number of unknown genes in the genome, especially in the rat, have not been shown to be expressed. In this study, we have established the first database of ESTs from rat pancreatic islet and RINm5F cells. Two cDNA libraries were constructed using mRNAs from rat pancreatic islet and RINm5F cells to cover a wider spectrum of expressed genes. Over 40,000 clones were randomly selected from the two libraries and partially sequenced. The sequences obtained were subjected to BLAST database analyses. This large-scale sequencing generated 40,710 3'-ESTs. Clustering analysis and homology search of nucleotide and peptide databases using both 3'- and 5'-ESTs revealed 10,406 non-redundant transcripts representing 4078 known genes or homologs and 6328 unknown genes. To confirm actual expression, the unknown sequences were further subjected to dbEST search, resulting in the identification of 5432 significant matches to those from other sources. Interestingly, of the remaining sequences showing no match, 779 were found to be encoded by exon-intron organization in the corresponding genomic sequences, suggesting that these are newly found as actually expressed in this study. Since many genes are up- or down-regulated in differing conditions, applications of the expression profile should facilitate identification of the genes involved in cell-specific functions in normal and disease states.

CONTEXT:CYP11B2, the gene encoding aldosterone synthase, has several frequent polymorphisms. In particular, the Lys173Arg (K173R) polymorphism is in complete genetic linkage disequilibrium with the -344T/C polymorphism in the promoter of CYP11B2 that involves a binding site for the steroidogenic factor-1 transcription factor. These polymorphisms have been associated with cardiovascular parameters, including hypertension, but not directly with gene expression.OBJECTIVE:The objective of this study was to correlate CYP11B2 genotype with gene expression in adrenal tissue.DESIGN:We measured mRNA levels of CYP11B2 [presented as a ratio against glyceraldehyde-3-phosphate dehydrogenase (B2/G)] and CYP11B1 in relation to the K173R polymorphism.SUBJECTS:We studied 28 subjects with aldosterone-producing adenomas (APA) and 18 subjects with normal adrenals.MAIN OUTCOME MEASURE:The main outcome measure was CYP11B2 expression levels.RESULTS:Preoperative treatment with spironolactone or beta-blocker in five APA patients was associated with higher B2/G. The B2/G and B2/B1 ratios were much higher even in the remaining 23 APA patients than in subjects with normal adrenals. The B2/G and B2/B1 ratios in normal adrenals and APA were higher in the KK genotype than in the RR genotype. In patients with APA, urinary aldosterone excretion was higher in those with the KK genotype than in those with the KR genotype. Measurement of cDNA band intensities from normal and APA samples of the KR genotype revealed that the R173 allele was transcribed at levels 46.6 +/- 12.2% (mean +/- sd; n = 7) and 49.1 +/- 20.8% (n = 6), respectively, those of the K173 allele.CONCLUSIONS:A CYP11B2 haplotype including -344T and K173 is associated with higher gene expression than the -344C/R173 haplotype, supporting reported associations of -344T with higher aldosterone production and blood pressure.

We measured serum DHEA-sulfate (DHEA-S) concentration, fasting plasma glucose (FPG), 2 hr-plasma glucose after 75g oral glucose loading (2hrPG), fasting insulin (FIRI) and HbA(1C) of 34 male adults in Kokufu-cho, most longevity district (mean life span in male: 80.4 yr-old) in Japan. and those of 559 male adults in Kasamatsu-cho (mean life span: 77.6 yr-old) as control. In addition, daily oral treatment with 25-50 mg DHEA for 2 weeks resulted in decreases of glucose levels after 75 g oral glucose tolerance test at 90 and 120 min, but no significant change of insulin levels in non-diabetic male adults. In Kokufu-cho, longevity district of Japan, serum DHEA-S concentrations of male adults was significantly higher than in Kasamatsu-cho as control, if adjusted for age, BMI, blood glucose (p < 0.0001). Negative correlation of DHEA-S with age was expectedly found (P < 0.0001) in total 593 male adults. In conclusion, DHEA may contribute to longevity in elderly aged male adults.

Two isozymes of the 11beta-hydroxysteroid dehydrogenase (11-HSD) are responsible for the interconversion of cortisol (F) and cortisone (E). The type 1 isozyme, 11-HSD1, acts mainly as a reductase in vivo, activating E to F, whereas the type 2, 11-HSD2, acts as a dehydrogenase, inactivating F to E. 11-HSD1 is the most abundant in the liver and 11-HSD2 in the kidney. In this study, we attempted to determine which isozyme and organs primarily contribute to equilibrium of plasma F and E concentrations in the peripheral circulation and to clarify differences in 11-HSD activities among adrenocortical disorders. Upon selective catheterizations for adrenocortical and renovascular disorders, plasma F and E concentrations in the femoral vein were closer to those in the renal vein than those in the hepatic vein. Values for mean plasma F/E ratios in the peripheral vein were in-between those of the adrenal and renal veins. A double reciprocal plot between peripheral plasma F and E concentrations in patients with various adrenocortical tumors was almost identical to that in normal subjects. Mean plasma F/E ratio in peripheral blood was higher in patients with Cushing's syndrome and was lower in patients with primary aldosteronism and nonfunctioning adrenocortical adenoma than that in normal subjects. These results suggest that renal 11-HSD2 is a main factor controlling the equilibrium of plasma F and E concentrations in the periphery and that cortisol and aldosterone excess do not change the equilibrium of plasma F and E concentrations in the peripheral circulation, but may alter expression of 11-HSD2. Alternation of 11-HSD2 activities as well as corticosteroid levels may be important in the pathophysiology of adrenocortical disorders.

A 69-year-old woman was referred to our department for evaluation of hypokalemia, which had been treated by oral potassium for more than ten years. She complained of headache, knee joint pain, sleeplessness and paresthesia in extremities and, most prominently, depression. Laboratory data suggested Gitelman's syndrome, which is caused by mutations in the gene encoding the thiazide-sensitive Na-Cl cotransporter. Direct sequencing of the gene in this patient revealed homozygous mutation R964Q in exon 25. Intravenous supplement of MgSO4 dramatically improved both the depression and the paresthesia, suggesting that hypomagnesemia played a role in the clinical manifestations.

Although tumor necrosis factor alpha (TNFalpha) decreases the expression of peroxisome proliferator-activated receptor gamma (PPARgamma), its mechanism is not understood. We evaluated the effect of ceramide, the second messenger of TNFalpha, on the expression of PPARgamma in primary cultured adipocytes. PPARgamma mRNA and aP2 mRNA levels were measured with real-time PCR. The PPARgamma protein level was measured with immunoblot. C6- and C2-ceramide, but not dihydroC6-ceramide, reduced the expression of PPARgamma in a time and concentration dependent manner. The application of 1 microM C6-ceramide for 36 h reduced PPARgamma mRNA level, aP2 mRNA level, and PPARgamma protein level to 56.3%, 80.4% and 62.1%, respectively. Since ceramide is known to activate atypical PKC, we also studied the role of atypical PKC on the PPARgamma reducing effect. Overexpression of wild type PKCzeta magnified and accelerated the effect of TNFalpha and C6-ceramide on PPARgamma mRNA levels, whereas overexpression of dominant negative PKCzeta abolished the effect. We also found that the overexpression of constitutive active PKCzeta reduced PPARgamma mRNA level, aP2 mRNA level, and PPARgamma protein level to 61.4%, 70.3% and 81.6%, respectively. Furthermore, TNFalpha activated nuclear factor-kappaB (NF-kappaB), known as a downstream effector of PKCzeta to 256.6%, which was enhanced with overexpression of wild-type PKCzeta. On the other hand, treatment with phorbol 12-myristate 13-acetate, another activator of NF-kappaB, also reduced the expression of PPARgamma to 57.8%. These results indicate that the reducing effect of TNFalpha is mediated through ceramide, atypical PKC and NF-kappaB pathway.