The secretion pattern of perivascular fat cells is different from that of subcutaneous and visceral fat cells.

AIMS/HYPOTHESIS: We have previously found that the mass of perivascular adipose tissue (PVAT) correlates negatively with insulin sensitivity and post-ischaemic increase in blood flow. To understand how PVAT communicates with vascular vessels, interactions between perivascular, subcutaneous and visceral fat cells with endothelial cells (ECs) were examined with regard to inflammatory, metabolic and angiogenic proteins. To test for possible in vivo relevance of these findings, circulating levels of the predominant secretion product, hepatocyte growth factor (HGF), was measured in individuals carefully phenotyped for fat distribution patterns. METHODS: Mono- and co-cultures of human primary fat cells with ECs were performed. mRNA expression and protein production were studied using Luminex, cytokine array, RealTime Ready and ELISA systems. Effects of HGF on vascular cells were determined by WST assays. In patients, HGF levels were measured by ELISA, and the mass of different fat compartments was determined by whole-body MRI. RESULTS: In contrast with other fat cell types, PVAT cells released higher amounts of angiogenic factors, e.g. HGF, acidic fibroblast growth factor, thrombospondin-1, serpin-E1, monocyte chemotactic protein-1 and insulin-like growth factor-binding protein -3. Cocultures showed different expression profiles from monocultures, and mature adipocytes differed from pre-adipocytes. HGF was preferentially released by PVAT cells and stimulated EC growth and smooth muscle cell cytokine release. Finally, in 95 patients, only PVAT, not visceral or subcutaneous mass, correlated independently with serum HGF levels (p = 0.03; r = 0.225). CONCLUSIONS: Perivascular (pre-)adipocytes differ substantially from other fat cells with regard to mRNA expression and protein production of angiogenic factors. This may contribute to fat tissue growth and atherosclerotic plaque complications. Higher levels of angiogenic factors, such as HGF, in patients with increased perivascular fat mass may have pathological relevance.

Visceral obesity modulates the impact of apolipoprotein C3 gene variants on liver fat content.

OBJECTIVE: It has not been solved whether subjects carrying the minor alleles of the -455T>C or -482C>T single nucleotide polymorphisms (SNPs) in the apolipoprotein-C3-gene (APOC3) have an increased risk for developing fatty liver and insulin resistance. We investigated the relationships of the SNPs with hepatic APOC3 expression and hypothesized that visceral obesity may modulate the effects of these SNPs on liver fat and insulin sensitivity (IS). METHODS: APOC3 mRNA expression and triglyceride content were determined in liver biopsies from 50 subjects. In a separate group (N=330) liver fat was measured by (1)H-magnetic resonance spectroscopy. IS was estimated during an oral glucose tolerance test (OGTT) and the euglycemic, hyperinsulinemic clamp (N=222). RESULTS: APOC3 mRNA correlated positively with triglyceride content in liver biopsies (r=0.29, P=0.036). Carriers of the minor alleles (-455C and -482T) tended to have higher hepatic APOC3 mRNA expression (1.80 (0.45-3.56) vs 0.77 (0.40-1.64), P=0.09), but not higher triglyceride content (P=0.76). In 330 subjects the genotype did not correlate with liver fat (P=0.97) or IS (OGTT: P=0.41; clamp: P=0.99). However, a significant interaction of the genotype with waist circumference in determining liver fat was detected (P=0.02) in which minor allele carriers had higher liver fat only in the lowest tertile of waist circumference (P=0.01). In agreement, during a 9-month lifestyle intervention the minor allele carriers of the SNP -482C>T in the lowest tertile also had less decrease in liver fat (P=0.04). CONCLUSIONS: APOC3 mRNA expression is increased in fatty liver and is regulated by SNPs in APOC3. The impact of the APOC3 SNPs on fatty liver is small and depends on visceral obesity.

Glucocorticoid receptor gene polymorphisms are associated with reduced first-phase glucose-stimulated insulin secretion and disposition index in women, but not in men.

AIM: Glucocorticoids are efficacious anti-inflammatory agents, but, in susceptible individuals, these drugs may induce glucose intolerance and diabetes by affecting ß-cell function and insulin sensitivity. We assessed whether polymorphisms in the glucocorticoid receptor gene NR3C1 associate with measures of ß-cell function and insulin sensitivity derived from hyperglycaemic clamps in subjects with normal or impaired glucose tolerance. METHODS: A cross-sectional cohort study was conducted in four academic medical centres in the Netherlands and Germany. Four hundred and forty-nine volunteers (188 men; 261 women) were recruited with normal glucose tolerance (n=261) and impaired glucose tolerance (n=188). From 2-h hyperglycaemic clamps, first- and second-phase glucose-stimulated insulin secretion, as well as insulin sensitivity index and disposition index, were calculated. All participants were genotyped for the functional NR3C1 polymorphisms N363S (rs6195), BclI (rs41423247), ER22/23EK (rs6189/6190), 9ß A/G (rs6198) and ThtIIII (rs10052957). Associations between these polymorphisms and ß-cell function parameters were assessed. RESULTS: In women, but not in men, the N363S polymorphism was associated with reduced disposition index (P=1.06 10(-4) ). Also only in women, the ER22/23EK polymorphism was associated with reduced first-phase glucose-stimulated insulin secretion (P=0.011) and disposition index (P=0.003). The other single-nucleotide polymorphisms were not associated with ß-cell function. Finally, none of the polymorphisms was related to insulin sensitivity. CONCLUSION: The N363S and ER22/23EK polymorphisms of the NR3C1 gene are negatively associated with parameters of ß-cell function in women, but not in men.

Insulin resistant phenotype of polycystic ovary syndrome does not seem to be caused by variation in FTO.

Genetic variation in the FTO gene is associated with increased body weight and reduced insulin sensitivity. We investigated whether genetic variation in FTO is associated with polycystic ovary syndrome (PCOS), a condition also characterized by insulin resistance. Furthermore, we tested whether insulin resistance is specifically associated with genetic variation in FTO in women with PCOS. Sixty-two nondiabetic patients with PCOS defined by the Rotterdam criteria were compared to BMI and age-matched women. Each PCOS case was matched to 2 controls. All participants underwent an oral glucose tolerance test and were genotyped for the single nucleotide polymorphism rs8050136 in the FTO gene. There was no difference in the frequency of FTO genotypes between the PCOS and the non-PCOS groups. In non-PCOS participants, genetic variation in FTO is associated with insulin sensitivity (p=0.03). This association remained significant after adjustment for age and/or BMI (p<= 0.03). In subjects with PCOS, however, FTO did not associate with insulin sensitivity (p=0.67). Genetic variation in FTO does not have an impact on insulin sensitivity in women with PCOS and is therefore not involved in the pathogenesis of the insulin resistant phenotype seen in patients with PCOS.

Effects of a lifestyle intervention in metabolically benign and malign obesity.

AIMS/HYPOTHESIS: We and others recently characterised metabolically benign or healthy obesity (MHO). In the present study we investigated whether a lifestyle intervention is sufficient to place obese insulin-resistant (OIR) individuals in a position where the possible metabolic consequences are similar to those for MHO individuals. METHODS: A total of 262 non-diabetic individuals participated in a 9 month lifestyle intervention programme. Obese individuals (BMI ≥ 30.0 kg/m(2)) were stratified, based on their insulin sensitivity (IS) estimated from an OGTT, into MHO (IS in the upper quartile, n = 26) and OIR (IS in the lower three quartiles, n = 77). Total body and visceral fat were measured by magnetic resonance (MR) tomography and liver fat by (1)H-MR spectroscopy. RESULTS: During the intervention, visceral fat decreased significantly in both groups (both p ≤ 0.009), whereas total body and liver fat decreased only in the OIR group (p < 0.0001; MHO p = 0.12 for total body fat and p = 0.47 for liver fat). IS improved in the OIR group (p < 0.0001), but remained essentially unchanged in the MHO group (p = 0.30). However, despite the significant increase in the OIR group, IS at follow-up barely exceeded 50% of the IS of the MHO group (OIR 9.30 ± 0.53 arbitrary units [AU]; MHO 16.41 ± 1.05 AU; p < 0.0001). CONCLUSIONS/INTERPRETATION: IS improves during the lifestyle intervention in OIR individuals. However, it does not reach a level where adequate protection from type 2 diabetes and cardiovascular disease is expected. Thus, stratification of obese individuals based on their metabolic phenotype is important to identify those who are likely to need early pharmacological treatment in addition to the lifestyle intervention.

A polymorphism in the gene encoding AdipoR1 affects olfactory recognition.

Polymorphisms in the gene encoding adiponectin receptor 1 (AdipoR1) are associated with insulin resistance, fatty liver, increased risk for type 2 diabetes and cardiovascular disease. AdipoR1 is expressed in the central nervous system and in the olfactory mucosa of mice and humans. We therefore hypothesized that a common polymorphism in AdipoR1 might alter olfactory function. We investigated a group of 222 healthy subjects (male: n = 147, female: n = 75) for olfactory recognition, and genotyped them for the polymorphism rs6666089 in the human AdipoR1 gene. This polymorphism has been previously shown to be associated with insulin resistance. Olfactory recognition was tested using standardized sniffing sticks, and parameters of glucose metabolism and serum adiponectin levels were assessed. We found a significant olfactory impairment in carriers of the AdipoR1 polymorphism rs6666089 (olfactory recognition: GG: 89.4 ± 1.2%, GA: 86.9 ± 1.4%, AA: 77.2 ± 4.8%, additive model, P = 0.0004, adjusted for age). Adiponectin levels had no impact on olfactory recognition. Fasting plasma glucose, fasting plasma insulin, body mass index and HbA1c did not differ between the genotype groups. In conclusion, the presence of a genetic variation in AdipoR1 is associated with decreased olfactory recognition in healthy subjects. Adiponectin signalling may have an important role in olfactory function and regulation of appetite.

Genetic variants affecting incretin sensitivity and incretin secretion.

Recent genome-wide association studies identified several novel risk genes for type 2 diabetes. The majority of these type 2 diabetes risk variants confer impaired pancreatic beta cell function. Though the molecular mechanisms by which common genetic variation within these loci affects beta cell function are not completely understood, risk variants may alter glucose-stimulated insulin secretion, proinsulin conversion, and incretin signals. In humans, the incretin effect is mediated by the secretion and insulinotropic action of two peptide hormones, glucose-dependent insulinotropic polypeptide and glucagon-like peptide-1. This review article aims to give an overview of the type 2 diabetes risk loci that were found to associate with incretin secretion or incretin action, paying special attention to the potential underlying mechanisms.

The ENPP1 K121Q polymorphism determines individual susceptibility to the insulin-sensitising effect of lifestyle intervention.

AIMS/HYPOTHESIS: The K121Q (rs1044498) single nucleotide polymorphism (SNP) in the ENPP1 gene has shown association with insulin resistance and type 2 diabetes in various ethnic populations. We hypothesised that K121Q may predict the success of lifestyle intervention in terms of improvement of insulin sensitivity. METHODS: We genotyped 1,563 participants with an increased risk of type 2 diabetes for K121Q and performed correlational analyses with anthropometric data and variables of insulin sensitivity. For metabolic characterisation, all participants underwent an OGTT. A subgroup of 506 participants additionally underwent a euglycaemic-hyperinsulinaemic clamp. In 342 participants, metabolic traits and anthropometric data were re-evaluated after a 9 month lifestyle intervention. RESULTS: In the overall cohort, K121Q was not associated with measures of obesity, indices of glucose tolerance during OGTT and insulin sensitivity estimated from the OGTT or derived from a euglycaemic-hyperinsulinaemic clamp after appropriate adjustment. However, K121Q did significantly influence the change in insulin sensitivity during lifestyle intervention after appropriate adjustment (p (additive) = 0.0067, p (dominant) = 0.0027). Carriers of the minor allele had an impaired increase in OGTT-derived insulin sensitivity. A similar trend was obtained for clamp-derived insulin sensitivity, but did not reach significance. CONCLUSIONS/INTERPRETATION: In our population of European ancestry, the ENPP1 SNP K121Q influenced the change in insulin sensitivity during lifestyle intervention. Thus, this SNP may determine susceptibility to environmental changes and could predict the success of lifestyle intervention.

A common genetic variant in WFS1 determines impaired glucagon-like peptide-1-induced insulin secretion.

AIMS/HYPOTHESIS: WFS1 type 2 diabetes risk variants appear to be associated with impaired beta cell function, although it is unclear whether insulin secretion is affected directly or secondarily via alteration of insulin sensitivity. We aimed to investigate the effect of a common WFS1 single-nucleotide polymorphism on several aspects of insulin secretion. METHODS: A total of 1,578 non-diabetic individuals (534 men and 1,044 women, aged 40 +/- 13 years, BMI 28.9 +/- 8.2 kg/m(2) [mean +/- SD]) at increased risk of type 2 diabetes were genotyped for rs10010131 within the WFS1 gene. All participants underwent an OGTT (and a subset additionally an IVGTT [n = 319]) and a hyperglycaemic clamp combined with glucagon-like peptide-1 (GLP-1) and arginine stimuli (n = 102). RESULTS: rs10010131 was associated with reduced OGTT-derived insulin secretion (p = 0.03). In contrast, insulin secretion induced by an i.v. glucose challenge in the IVGTT and hyperglycaemic clamp was not different between the genotypes. GLP-1 infusion combined with a hyperglycaemic clamp showed a significant reduction of the insulin secretion rate during the first and second phases of GLP-1-induced insulin secretion in carriers of the risk allele (reduction of 36% and 26%, respectively; p = 0.007 and p = 0.04, respectively). CONCLUSIONS/INTERPRETATION: A common genetic variant in WFS1 specifically impairs GLP-1-induced insulin secretion independently of insulin sensitivity. This defect might explain the impaired insulin secretion in carriers of the risk allele and confer the increased risk of type 2 diabetes.

The risk allele load accelerates the age-dependent decline in beta cell function.

AIMS/HYPOTHESIS: Among the novel type 2 diabetes risk loci identified by genome-wide association studies, TCF7L2, HHEX, SLC30A8 and CDKAL1 appear to affect beta cell function. In the present study we examined the effect of these genes' risk alleles on the age-dependent decline in insulin secretion. METHODS: The SNPs rs7903146 (TCF7L2), rs7754840(CDKAL1), rs7923837 (HHEX) and rs13266634 (SLC30A8) were genotyped in 1,412 non-diabetic patients, who were subsequently grouped according to their number of risk alleles. All participants underwent an OGTT. Insulin secretion was assessed by validated indices and proinsulin conversion by calculating AUC(proinsulin)/AUC(insulin). RESULTS: The number of risk alleles revealed a Gaussian distribution, with most participants carrying four risk alleles. Stratification into groups with low (LAL, up to three alleles), median (MAL, four alleles) and high (HAL, five to eight alleles) allele load resulted in MAL and HAL participants displaying significantly lower insulin secretion and proinsulin conversion than LAL participants (p

Protein kinase C iota mediates lipid-induced apoptosis of human coronary artery endothelial cells.

Apoptosis is involved in the development and progression of atherosclerotic lesions. Protein kinase C (PKC) signalling is of importance in atherosclerosis as well as apoptosis. Therefore, we tested the involvement of PKC in lipid-induced apoptosis of human coronary artery endothelial cells (HCAEC). Protein expression of PKC isoforms alpha, beta I, delta, epsilon, and iota was detected, whereas no relevant protein amounts of PKC isoforms beta II, gamma, eta, theta, and zeta were found. Inhibition of classical and novel PKC isoforms by treatment with bisindolylmaleimide or PKC down-regulation by long-term treatment with 12-O-tetradecanoyl phorbol-13-acetate (TPA) could not prevent apoptosis induced by palmitate or stearate. In contrast, a specific myristoylated, cell-permeable PKC zeta/iota pseudosubstrate prevented lipid-induced apoptosis in HCAEC. Furthermore, saturated fatty acids activated PKC iota as evidenced by PKC iota down-regulation upon long-term treatment with stearate. Our data provide evidence that PKC iota is activated by saturated fatty acids and mediates lipid-induced apoptosis of HCAEC.

Pegvisomant treatment in gigantism caused by a growth hormone-secreting giant pituitary adenoma.

BACKGROUND: Gigantism is rare with the majority of cases caused by a growth hormone (GH)-secreting pituitary adenoma. Treatment options for GH-secreting pituitary adenomas have been widened with the availability of long-acting dopamine agonists, depot preparations of somatostatin analogues, and recently the GH receptor antagonist pegvisomant. CASE REPORT: A 23-year-old male patient presented with continuous increase in height during the past 6 years due to a GH-secreting giant pituitary adenoma. Because of major intracranial extension and failure of octreotide treatment to shrink the tumour, the tumour was partially resected by a trans-frontal surgical approach. At immunohistochemistry, the tumour showed a marked expression of GH and a sparsely focal expression of prolactin. Somatostatin receptors (sst) 1-5 were not detected. Tumour tissue weakly expressed dopamine receptor type 2. The Gs alpha subunit was intact. Conversion from somatostatin analogue to pegvisomant normalized insulin-like-growth-factor-I (IGF-I) levels and markedly improved glucose tolerance. CONCLUSION: Pegvisomant is a potent treatment option in patients with pituitary gigantism. In patients who do not respond to somatostatin analogues, knowledge of the SST receptor status may shorten the time to initiation of pegvisomant treatment.

Adiponectin is functionally active in human islets but does not affect insulin secretory function or beta-cell lipoapoptosis.

CONTEXT: The adipokine adiponectin has insulin-sensitizing, antiatherogenic, and antiinflammatory properties. Mouse and human adiponectin receptor-1 and -2 have been cloned, both of which are expressed in various tissues and mediate effects of globular and full-length adiponectin. Whether adiponectin affects insulin secretion and beta-cell apoptosis and whether plasma adiponectin is associated with beta-cell function in humans is under investigation. DESIGN AND METHODS: In human islets from multiorgan donors, we investigated expression of adiponectin receptor-1 and -2. Furthermore, glucose-stimulated insulin secretion was determined by RIA. In addition, we investigated fatty acid-induced beta-cell apoptosis by terminal dUTP nick end labeling and flow-cytometric cell cycle analysis (sub-G1 formation). In humans in vivo, insulin secretory function was measured during hyperglycemic clamps in 65 normal glucose-tolerant subjects. We determined first and second phase of glucose-stimulated, glucagon-like peptide-1-stimulated, and arginine-stimulated insulin secretion. RESULTS: Adiponectin receptor-1 and -2 are expressed in human islets at the mRNA and protein level. Moreover, full-length adiponectin induces phosphorylation of acetyl coenzyme A carboxylase. However, adiponectin did not affect basal or glucose-stimulated insulin secretion or basal or fatty acid-induced beta-cell apoptosis. In vivo, fasting plasma adiponectin concentrations were not associated with glucose-stimulated first- and second-phase insulin secretion or with glucagon-like peptide-1- or arginine-stimulated insulin secretion (all P > 0.42). CONCLUSIONS: These data support a regulatory role of adiponectin in human islets; however, adiponectin does not seem to affect insulin secretion or basal/fatty acid-induced beta-cell apoptosis in humans.

Functional significance of the UCSNP-43 polymorphism in the CAPN10 gene for proinsulin processing and insulin secretion in nondiabetic Germans.

Recently, an association of the G allele in UCSNP-43 of calpain 10 with type 2 diabetes and decreased glucose disposal was reported. Calpain 10 is also expressed in pancreatic islets. It is not known, however, whether and how this polymorphism contributes to the biological variation of beta-cell function. We studied 73 nondiabetic subjects from the southwest region of Germany (G/G, n = 41; G/A, n = 29; and A/A, n = 3) using a modified hyperglycemic clamp (10 mmol/l glucose, added glucagon-like peptide 1, final arginine bolus). The genotype distribution was not different between subjects with normal glucose tolerance (n = 56) and those with impaired glucose tolerance (n = 17; P = 0.74, chi2 test). First-phase insulin secretion (adjusted for sex and insulin sensitivity from hyperglycemic clamp) was greater in G/G (2,747 +/- 297 pmol/min) than in G/A + A/A (1,612 +/- 156 pmol/min, P = 0.003). Insulin secretion in response to arginine (adjusted for insulin sensitivity) was also greater in G/G (9,648 +/- 1,186 pmol/min) than in G/A + A/A (5,686 +/- 720 pmol/min, P = 0.04). The acute poststimulus proinsulin-to-insulin ratio was lower in G/G (1.6 +/- 0.4% first phase; 1.6 +/- 0.2% arginine) than in G/A + A/A (4.0 +/- 0.5% first phase, P < 0.001; 2.5 +/- 0.4% arginine, P = 0.03). In conclusion, it appears unlikely that any association of the UCSNP-43 polymorphism alone with type 2 diabetes involves impairment of insulin secretion in our population of German Caucasians. This may be entirely different with specific haplotype combinations.

Pro12Ala polymorphism in the peroxisome proliferator-activated receptor-gamma2 gene is associated with increased antilipolytic insulin sensitivity.

The Pro12Ala polymorphism of the peroxisome proliferator-activated receptor (PPAR)-gamma2 is associated with reduced transcriptional activity in vitro and increased insulin sensitivity in humans in vivo. The mechanism by which this polymorphism influences insulin sensitivity in humans is unclear. PPAR-gamma2 is mainly expressed in adipocytes, and free fatty acids released from adipose tissue are key mediators of peripheral insulin resistance. Therefore, we examined insulin suppression of lipolysis in 51 subjects without (Pro/Pro) and 17 subjects with the polymorphism (X/Ala). Both groups were lean (BMI <27.0 kg/m2) and matched for age, BMI, waist-to-hip ratio, and sex. The isotopically (infusion of d5 glycerol) determined glycerol rate of appearance was used as an index of lipolysis. Insulin sensitivity of lipolysis was expressed as the insulin concentration resulting in half-maximal suppression (EC50). This was directly determined during a three-step hyperinsulinemic-euglycemic clamp (n = 21) or estimated indirectly during a standard hyperinsulinemic-euglycemic clamp (n = 47). The insulin sensitivity index (ISI) of glucose disposal was 0.095+/-0.006 micromol x kg(-1) x min(-1) x pmol(-1) x l(-1) in the control group and 0.129+/-0.008 micromol x kg(-1) x min(-1) x pmol(-1) x l(-1) in the X/Ala group (P = 0.003). The EC50 was 56+/-2 pmol/l in the control group and 44+/-3 pmol/l in the X/Ala group (P = 0.001). The EC50 of lipolysis and ISI was significantly correlated (r = 0.42, P = 0.002). In conclusion, in lean subjects, the Pro12Ala polymorphism is associated with increased insulin sensitivity of glucose disposal and suppression of lipolysis. This result suggests that an altered transcriptional activity of PPAR-gamma2 in X/Ala subjects either causes a more efficient suppression of lipolysis in adipose tissue, which in turn results in improved insulin-stimulated glucose disposal in muscle, or, alternatively, beneficially affects insulin signaling in both tissues independently of one another.

The Gly972Arg polymorphism in the insulin receptor substrate-1 gene contributes to the variation in insulin secretion in normal glucose-tolerant humans.

The Gly972Arg polymorphism in the insulin receptor substrate (IRS)-1 was found in some studies to have a higher prevalence in type 2 diabetic subjects than in control subjects. Previously, transfection of IRS-1 with this polymorphism into insulin-secreting cells resulted in a marked reduction of glucose-stimulated insulin secretion compared with the wild-type transfected cells. In the present study, we compared insulin secretion in well-matched normal glucose-tolerant subjects with and without this polymorphism. Several validated indexes of beta-cell function from the oral glucose tolerance test were significantly lower in X/Arg (n = 31) compared with Gly/Gly (n = 181) (P between 0.002 and 0.05), whereas insulin sensitivity (measured with a euglycemic clamp) was not different. During a modified hyperglycemic clamp, insulin secretion rates were significantly lower in Gly/Arg (n = 8) compared with Gly/Gly (n = 36) during the first phase (1,711+/-142 vs. 3,014+/-328 pmol/min, P = 0.05) and after maximal stimulation with arginine (5,340+/-639 vs. 9,075+/-722 pmol/min, P = 0.03). In summary, our results suggest that the Gly972Arg polymorphism in IRS-1 is associated with decreased insulin secretion in response to glucose but not with insulin sensitivity. It is possible that this polymorphism causes insulin resistance at the level of the beta-cell and contributes to the polygenic etiology of type 2 diabetes.

Genetic determinants of insulin action in polycystic ovary syndrome.

INTRODUCTION: Insulin resistance is associated with both type 2 diabetes (T2 D) and polycystic ovary syndrome (PCOS). There is an association of T2 D with several polymorphisms in candidate genes related to insulin resistance. However, there is limited information about the association of these polymorphisms with PCOS. METHODS: 57 non-diabetic women with PCOS and a control group of 567 healthy non-diabetic women underwent an oral glucose tolerance test (OGTT). They were genotyped for the polymorphisms Gly972Arg in IRS-1, Gly1057Asp in IRS-2, SNP 43, 44, and 45 in CAPN10, Pro12Ala in PPAR(gamma)2, C-512 T in FOXC2, and T45 G in adiponectin. RESULTS: Women with PCOS had higher 2-h blood glucose levels (6.5 +/- 0.2 vs. 6.0 +/- 0.06 mmol/l, p = 0.03) compared to control women, higher fasting insulin (79 +/- 7 vs. 53 +/- 2 pmol/l, p = 0.02), and a lower insulin sensitivity estimated from the OGTT (12.4 +/- 1.1 vs. 19.1 +/- 0.5 U, p = 0.0001). More homozygous G allele carriers of the T45 G polymorphism in the adiponectin gene were found in women with PCOS compared to controls (13.2 % vs. 2.6 %, p = 0.008). In women with PCOS, G-allele carriers had lower fasting insulin levels than TT carriers (61 +/- 9 vs. 88 +/- 10, p = 0.02) in contrast to controls (p = 0.03 for interaction genotype x PCOS). The other polymorphisms were distributed equally among women with PCOS and controls (all p > 0.5). SUMMARY: We found a higher prevalence of the T45 G polymorphism in the adiponectin gene in women with PCOS compared to controls. This was not associated with a more insulin resistant phenotype in PCOS, however. Other frequent polymorphisms in genes related to insulin resistance and type 2 diabetes showed no association with PCOS.

Inhibition of Ret oncogene activity by the protein tyrosine phosphatase SHP1.

Germline mutations in the Ret protooncogene give rise to the inherited endocrine cancer syndromes MEN types 2A and 2B and familiar medullary thyroid carcinoma. Although it is well accepted that the constitutive active tyrosine kinase of Ret oncogenes ultimately leads to malignant transformation, it is not clear whether a decrease in the autophosphorylation of oncogenic Ret forms can affect the mitogenic and transforming activities of Ret. Potential modulators of the tyrosine kinase activity of Ret could be tyrosine phosphatases that are expressed in human thyroid tissue. Therefore, we investigated the impact of the tyrosine phosphatases SHP1 and SHP2 on the intrinsic tyrosine kinase activity and oncogenic potency of Ret with a 9-bp duplication in the cysteine-rich domain (codons 634-636), which was described in a patient with MEN type 2A recently. SHP1 and SHP2 were stably overexpressed in NIH3T3 fibroblasts together with Ret-9bp. Coexpression of SHP1 with Ret-9bp reduced the autophosphorylation of Ret-9bp by 19 +/- 7% (P = 0.01, n = 4), whereas no effect was seen with SHP2. Furthermore, Ret-9bp could be coimmunoprecipitated with SHP1 but not with SHP2 antibodies. Suppression of the Ret-9bp tyrosine kinase activity by SHP1 caused a decrease in activation of Erk2 (extracellular signal-regulated kinase) and abolished PKB/Akt (protein kinase B) phosphorylation. In addition, diminished Ret-9bp autophosphorylation led to reduced phosphorylation of the transcription factor jun-D. Finally, the inhibitory effect on Ret-9bp signaling resulted in a 40-60% reduction of [(3)H]thymidine incorporation and in reduced ability of NIH3T3 cells to form colonies in soft agar. In conclusion, the data suggest that SHP1 caused a moderate reduction of Ret autophosphorylation, which led to a strong suppression of the Ret oncogene activity.

The prevalent Gly1057Asp polymorphism in the insulin receptor substrate-2 gene is not associated with impaired insulin secretion.

Disruption of the insulin receptor substrate-2 was shown to cause type 2 diabetes in mice. This could be largely attributed to abnormal beta-cell development. In humans, a prevalent polymorphism in insulin receptor substrate-2 (Gly1057Asp) was not found be associated with type 2 diabetes in linkage and association studies. We tested the hypothesis that an extreme challenge of the beta cell might reveal subtle abnormalities in carriers of this polymorphism undetected by conventional insulin secretion tests. Therefore, in addition to assessing beta-cell function by oral glucose tolerance testing (n = 318, normal glucose tolerance), we measured the secretory response to maximal stimulation by hyperglycemia (10 mM), glucagon-like peptide-1, and arginine administered in an additive fashion (n = 77, nondiabetic). The allelic frequency of the Asp allele was approximately 37%. Neither the beta-cell function indices from the oral glucose tolerance test nor the secretory response during the hyperglycemic clamp differed measurably between carriers and controls. Moreover, maximal plasma C-peptide concentrations in response to the combined glucose, glucagon-like peptide-1, and arginine stimulus was not different between Gly/Gly (10,745 +/- 1,186 pmol/liter) and X/Asp (10,800 +/- 490 pmol/liter, P = 0.99). In conclusion, our findings strongly suggest that the Gly1057Asp polymorphism in insulin receptor substrate-2 is not associated with beta-cell dysfunction. The normal maximal insulin secretory response makes it unlikely that this common polymorphism results in abnormal beta-cell development.