bluuebunny/crossref_metadata_2025 huggingface dataset:
Identification of the high-affinity tolbutamide site on the SUR1 subunit of the K(ATP) channel.
Authors: Ashfield R , Gribble F M , Ashcroft S J , Ashcroft F M
Published: March 2007

ATP-sensitive potassium channels (K(ATP)) are formed from four pore-forming Kir6.2 subunits complexed with four regulatory sulfonylurea receptor subunits (SUR1 in pancreatic beta-cells, SUR2A in heart). The sensitivity of the channel to different sulfonylureas depends on the SUR isoform. In particular, Kir6.2-SUR1 but not Kir6.2-SUR2A channels are blocked by tolbutamide with high affinity. We made chimeras between SUR1 and SUR2A to identify the region of the protein involved in high-affinity tolbutamide block. Chimeric SURs were coexpressed with Kir6.2 in Xenopus oocytes, and macroscopic currents were measured in inside-out membrane patches. High-affinity tolbutamide inhibition could be conferred on SUR2A by replacing transmembrane domains (TMs) 14-16 with the corresponding region of SUR1. Conversely, high-affinity tolbutamide inhibition of SUR1 was abolished by replacing TMs 13-16 with the corresponding SUR2A sequence, or by mutating a single serine residue within this region to tyrosine (S1237Y). Binding of [3H]glibenclamide to membranes expressing SUR1 was abolished concomitantly with the loss of high-affinity tolbutamide block. These results suggest that a site in the COOH-terminal set of TMs of the SUR1 subunit of the K(ATP) channel is involved in the binding of tolbutamide and glibenclamide.
Activation of the sphingomyelinase/ceramide signal transduction pathway in insulin-secreting beta-cells: role in cytokine-induced beta-cell death.
Authors: Major C D , Gao Z Y , Wolf B A
Published: March 2007

Activation of the sphingomyelin/ceramide pathway may mediate interleukin-1-induced beta-cell death (Welsh, N: Interleuken-1beta-induced ceramide and diacylglycerol generation may lead to activation of the c-Jun NH2-terminal kinase and the transcription factor ATF-2 in the insulin-producing cell line RINm5F. J Biol Chem 271: 8307-8312, 1996). In this report, we have examined this pathway in more detail. Culture of beta-TC3 cells with 25 micromol/l ceramide analogs (N-acetyl- and N-hexanoylsphingosine) for 72 h did not significantly affect glucose- and carbachol-induced insulin secretion. Dihydroceramide (N-acetyl- or N-hexanoylsphinganine), a structurally similar analog, had no effect on agonist-induced secretion. However, ceramide analogs both time- and dose-dependently decreased cell viability, while the dihydroceramide analog had no effect. The ceramide effect on cell viability mimicked the effect of the cytokines TNF-alpha, IL-1beta, and IFN-gamma, reported stimulators of sphingomyelin hydrolysis. Cytokines, however, failed to stimulate sphingomyelin metabolism. Furthermore, using two different methods to quantitate ceramide, cytokines failed to cause an increase in beta-cell ceramide content versus unstimulated or time-matched vehicle controls. Taken together, these data suggest that although ceramide analogs mimic the cytotoxic effect of cytokines, activation of the sphingomyelin/ceramide signaling pathway is not involved in cytokine-induced beta-cell death.
Cow's milk formula feeding induces primary immunization to insulin in infants at genetic risk for type 1 diabetes.
Authors: Vaarala O , Knip M , Paronen J , Hämäläinen A M , Muona P , Väätäinen M , Ilonen J , Simell O , Akerblom H K
Published: March 2007

Insulin autoantibodies (IAAs) often appear as the first sign of islet cell autoimmunity in prediabetic children. Because cow's milk contains bovine insulin, we followed the development of insulin-binding antibodies in children fed with cow's milk formula. Bovine insulin- and human insulin-binding antibodies by enzyme immunoassay and IAA by radioimmunoassay were analyzed in 200 infants carrying HLA-DQB1*0302 but no protective alleles who participated in a Finnish population-based birth-cohort study. Based on the prospectively registered information, the first 100 infants enrolled in the study who were exposed to cow's milk formula before age 12 weeks and the first 100 infants enrolled in the study who were exclusively breast-fed for longer than their first 12 weeks of life were selected for the present study. Also, 11 children from the birth cohort who developed at least two diabetes-associated autoantibodies, 98 children with newly diagnosed type 1 diabetes, and 92 healthy children were studied. We found that the amount of IgG-antibodies binding to bovine insulin was higher at age 3 months in infants who were exposed to cow's milk formula than in infants who were exclusively breast-fed at that age (median 0.521 vs. 0.190; P < 0.0001). The antibodies binding to bovine insulin cross-reacted with human insulin. None of these infants tested positive for IAA. The levels of bovine insulin-binding antibodies declined in both groups at ages 12 and 18 months, whereas in the 11 children with at least two diabetes-associated autoantibodies the levels increased during the follow-up period (P < 0.0001). IgG antibodies correlated with IgG2 antibodies binding to bovine insulin (r = 0.43, P = 0.004) and IAA (r = 0.27, P = 0.02) in diabetic children, but not in healthy children. Cow's milk feeding is an environmental trigger of immunity to insulin in infancy that may explain the epidemiological link between the risk of type 1 diabetes and early exposure to cow's milk formulas. This immune response to insulin may later be diverted into autoaggressive immunity against beta-cells in some individuals, as indicated by our findings in children with diabetes-associated autoantibodies.
Enhancing effects of long-term elevated glucose and palmitate on stored and secreted proinsulin-to-insulin ratios in human pancreatic islets.
Authors: Björklund A , Grill V
Published: March 2007

Relative hypersecretion of proinsulin is a feature of type 2 diabetes. We investigated to what extent this feature can be induced in human pancreatic islets by elevated glucose or fatty acids, two major abnormalities of the diabetic state. A 48-h culture period with 27 mmol/l glucose increased the intraislet proinsulin-to-insulin (PI/I) ratio 5.0-fold, owing to preferential decrease of insulin. The PI/I ratio in culture medium was enhanced 1.9-fold versus islets cultured with 5.5 mmol/l glucose. This effect of elevated glucose persisted after normalization of glucose levels: during 60-min postculture incubations at a basal glucose concentration (3.3 mmol/l), the PI/I ratio of secretion increased 4.9-fold. The ratio was also increased (14-fold) after renewed postculture stimulation with 16.7 mmol/l glucose. Diazoxide was added to culture medium to block glucose-induced insulin secretion and thus investigate the importance of overstimulation. In cultures at 27 mmol/l glucose, the presence of diazoxide decreased the PI/I ratio of islet contents by 76%, the accumulated secretion to culture medium by 70%, and the release at 3.3 or 16.7 mmol/l glucose during postculture incubations by 85 and 86%, respectively. None of these PI/I-decreasing effects of diazoxide were reproduced during or after coculture with 5.5 mmol/l glucose. Culture with 0.2 mmol/l palmitate and 5.5 mmol/l glucose decreased islet contents of proinsulin and insulin and increased the secreted products in culture media without affecting PI/I ratios. During postculture conditions, however, prior palmitate culture enhanced the PI/I ratio of release at 3.3 mmol/l glucose (from 2.2 +/- 0.4 to 5.4 +/- 0.9%, P < 0.05). Culture with palmitate together with 27 mmol/l glucose decreased islet contents of proinsulin and insulin and further enhanced intraislet PI/I ratios (from 9.3 +/- 1.1 to 13.4 +/- 2.5%, P < 0.05). However, palmitate failed to affect PI/I ratios in culture medium. In contrast, in postculture incubations at 3.3 mmol/l glucose, prior palmitate culture further elevated the PI/I ratio of secretion (from 10.8 +/- 1.2 after previous 27 mmol/l glucose alone to 13.9 +/- 2.8% after palmitate and glucose, P < 0.05). We conclude that 1) long-term exposure of human islets to elevated glucose leads to preferential secretion of proinsulin, and this effect persists also after glucose normalization; 2) the glucose effect appears secondary to depletion of mature insulin granules; and 3) elevated fatty acids influence PI/I ratios of secretion by mechanisms that are, in part, incongruous with an over-stimulation effect.
Obesity-related phenotypes and the beta3-adrenoceptor gene variant in postmenopausal women.
Authors: Tchernof A , Starling R D , Walston J D , Shuldiner A R , Dvorak R V , Silver K , Matthews D E , Poehlman E T
Published: March 2007

We examined the hypothesis that postmenopausal women with the beta3-adrenoceptor gene variant (Trp64Arg) have reduced total daily energy expenditure (TEE), altered free fatty acid kinetics, and increased intra-abdominal fat. A secondary objective was to examine whether the obese state masks the effect of the variant on resting metabolic rate (RMR). There were 23 obese heterozygous women with the genetic variant (age 58 +/- 6 years; BMI 36 +/- 7 kg/m2) who were compared with 19 homozygous obese women with the normal allele (age 56 +/- 4 years; BMI 36 +/- 3 kg/m2). Daily energy expenditure was determined from doubly labeled water and indirect calorimetry, lipolysis from infusion of [1-13C]palmitate, and body fat distribution from computed tomography. No significant differences were found in TEE, RMR, energy expenditure of physical activity, the thermic effect of a meal, fat oxidation as estimated by fasting and postprandial respiratory quotients (RQs), or rate of lipolysis. Similarly, no difference was found in visceral adipose tissue and abdominal subcutaneous fat areas. When RMR was compared between obese (n = 23) and never-obese women with the Trp64Arg variant (n = 16), we found a 317 kcal/day lower RMR in never-obese women after controlling for fat mass, fat-free mass, and age (P < 0.0017). These results do not support the hypothesis that already obese women with the Trp64Arg polymorphism of the beta3-adrenergic receptor gene have lower daily energy expenditure, altered lipolysis, and increased abdominal obesity. On the other hand, the lower RMR in never-obese women suggests that the obese state may mask a moderate effect of the Trp64Arg variant on energy expenditure. Although these results need to be confirmed in other populations, the obese state may have been a confounding factor in previous studies of the beta3-adrenoceptor Trp64Arg variant and energy expenditure.
Antibodies to oxidized LDL predict coronary artery disease in type 1 diabetes: a nested case-control study from the Pittsburgh Epidemiology of Diabetes Complications Study.
Authors: Orchard T J , Virella G , Forrest K Y , Evans R W , Becker D J , Lopes-Virella M F
Published: March 2007

The pathogenesis of excess cardiovascular risk in type 1 diabetes is unclear. LDL cholesterol is only weakly predictive, and its concentration is often normal in type 1 diabetes. We therefore examined whether markers of LDL oxidation such as antibodies to oxidized LDL (Ab-OxLDL) and LDL-containing immune complexes, rather than LDL concentration, were predictive of coronary artery disease (CAD) in type 1 diabetes. This nested case-control study from an epidemiologic cohort study included 49 incident cases of myocardial infarction (MI), angina, or CAD death and 49 age-, sex-, and duration-matched control subjects. Ab-OxLDL was measured by enzyme immunoassay and the apolipoprotein B (ApoB) content of immune complexes (ApoB-IC) precipitated by polyethylene glycol by immunoelectrophoresis in baseline stored samples. Ab-OxLDL was inversely, and ApoB-IC directly, related to subsequent CAD. In multivariate analyses, Ab-OxLDL remained a significant independent predictor along with previously recognized predictors, hypertension and Beck depression score. In conclusion, oxidation of LDL and the immune response it elicits may play a role in predicting the development of CAD in type 1 diabetes and explain at least some of the enhanced CAD risk in type I diabetes.
Pro12Ala substitution in the peroxisome proliferator-activated receptor-gamma2 is not associated with type 2 diabetes.
Authors: Mancini F P , Vaccaro O , Sabatino L , Tufano A , Rivellese A A , Riccardi G , Colantuoni V
Published: March 2007

Peroxisome proliferator-activated receptor (PPAR)-gamma is a major regulator of adipogenesis and insulin sensitivity. The PPAR-gamma gene generates two isoforms through alternative splicing, PPAR-gamma1 and -gamma2, the latter having an additional stretch of 28 amino acids at its NH2-terminus in the ligand-independent activation domain. This extension renders PPAR-gamma2 more sensitive to insulin action. Since there is a Pro12Ala substitution in this domain, we tested whether it is related to type 2 diabetes or insulin resistance. Therefore, 131 type 2 diabetic patients and 312 normoglycemic control subjects were screened for the presence of the mutation and for major clinical and metabolic features. The frequency of the mutation did not differ significantly between diabetic patients and control subjects. BMI, insulin, and other metabolic and anthropometric variables were also not associated with the mutation. Although the study was carried out on a sufficiently large sample, the conclusions do not support a major role for the Pro12Ala substitution of the PPAR-gamma gene in the etiology of type 2 diabetes.
Leptin restores euglycemia and normalizes glucose turnover in insulin-deficient diabetes in the rat.
Authors: Chinookoswong N , Wang J L , Shi Z Q
Published: March 2007

Leptin has been shown to improve insulin sensitivity and glucose metabolism in normoinsulinemic healthy or obese rodents. It has not been determined whether leptin may act independently of insulin in regulating energy metabolism in vivo. The present study was designed to examine the effects of leptin treatment alone on glucose metabolism in insulin-deficient streptozotocin (STZ)-induced diabetic rats. Four groups of STZ-induced diabetic rats were studied: 1) rats treated with recombinant methionine murine leptin subcutaneous infusion with osmotic pumps for 12-14 days (LEP; 4 mg x kg(-1) x day(-1), n = 10); 2) control rats infused with vehicle (phosphate-buffered saline) for 12-14 days (VEH; n = 10); 3) pair-fed control rats given a daily food ration matching that of LEP rats for 12-14 days (PF; n = 8); and 4) rats treated with subcutaneous phloridzin for 4 days (PLZ; 0.4 g/kg twice daily, n = 10). Phloridzin treatment normalizes blood glucose without insulin and was used as a control for the effect of leptin in correcting hyperglycemia. All animals were then studied with a hyperinsulinemic-euglycemic clamp (6 mU x kg(-1) x min(-1). Our study demonstrates that leptin treatment in the insulin-deficient diabetic rats restored euglycemia, minimized body weight loss due to food restriction, substantially improved glucose metabolic rates during the postabsorptive state, and restored insulin sensitivities at the levels of the liver and the peripheral tissues during the glucose clamp. The effects on glucose turnover are largely independent of food restriction and changes in blood glucose concentration, as evidenced by the minimal improvement of insulin action and glucose turnover parameters in the PF and PLZ groups. Our results suggest that the antidiabetic effects of leptin are achieved through both an insulin-independent and an insulin-sensitizing mechanism.
Deficiency of phosphofructo-1-kinase/muscle subtype in humans is associated with impairment of insulin secretory oscillations.
Authors: Ristow M , Carlqvist H , Hebinck J , Vorgerd M , Krone W , Pfeiffer A , Müller-Wieland D , Ostenson C G
Published: March 2007

In healthy humans, insulin is secreted in an oscillatory manner. While the underlying mechanisms generating these oscillations are not fully established, increasing evidence suggests a central role for phosphofructo-1-kinase/muscle subtype (PFK1-M), which also serves as the predominantly active PFK1 subtype in the pancreatic beta-cell. The fact that normal oscillatory secretion is impaired in subjects with impaired glucose tolerance and healthy relatives of patients with type 2 diabetes suggests that this defect may be involved in the secretory dysfunction. To evaluate a possible link between inherited PFK1-M deficiency in humans (Tarui's disease or glycogenosis type VII) and altered insulin oscillations, in vivo studies were performed. We determined basal insulin oscillations during 2 h of frequent plasma sampling in two related teen-aged individuals with homozygous and heterozygous PFK1-M deficiency compared with nondeficient, unrelated control subjects. As predicted by the underlying hypothesis, normal oscillations in insulin secretion were completely abolished in the individual with homozygous deficiency of PFK1-M and significantly impaired in the heterozygous individual, as shown by spectral density and autocorrelation analyses. Thus, deficiency of PFK1-M subtype in humans appears to be associated with an impaired oscillatory insulin secretion pattern and may contribute to the commonly observed secretion defects occurring in type 2 diabetes.
Dehydroepiandrosterone suppresses the elevated hepatic glucose-6-phosphatase and fructose-1,6-bisphosphatase activities in C57BL/Ksj-db/db mice: comparison with troglitazone.
Authors: Aoki K , Saito T , Satoh S , Mukasa K , Kaneshiro M , Kawasaki S , Okamura A , Sekihara H
Published: March 2007

The effect of dehydroepiandrosterone (DHEA) on the hepatic and muscle glucose metabolizing enzymes and on blood glucose were investigated in insulin-resistant diabetic C57BL/KsJ-db/db mice and their heterozygote littermates (db/+m). The results were compared with those after troglitazone administration under the same conditions. Despite hyperinsulinemia, hepatic glucose-6-phosphatase (G6Pase) and fructose-1,6-bisphosphatase (FBPase) activities are higher in db/db than in db/+m mice. Dietary administration of DHEA and that of troglitazone for 15 days to respective groups of five mice each significantly decreased blood glucose in db/db mice and hepatic G6Pase and FBPase activities in both db/db and db/+m mice. Hepatic G6Pase and FBPase activities showed a linear relationship with blood glucose in all groups of mice, suggesting that the activities of G6Pase and FBPase are closely related to blood glucose levels. Because androstenedione, a DHEA metabolite, barely affected either of these enzyme activities or blood glucose in db/db mice, the actions of DHEA, which are similar to those of troglitazone, are presumed to be caused by DHEA itself. DHEA is considered to be a modulating agent for the activities of hepatic gluconeogenic enzymes in db/db mice.
Metabolic factors contributing to increased resting metabolic rate and decreased insulin-induced thermogenesis during the development of type 2 diabetes.
Authors: Weyer C , Bogardus C , Pratley R E
Published: March 2007

Previous studies have indicated that individuals with type 2 diabetes have an increased resting metabolic rate (RMR) but decreased insulin-induced thermogenesis (IIT) compared with those with normal glucose tolerance (NGT). When and by which mechanisms these abnormalities occur during the development of diabetes remain unknown. In 560 Pima Indians, sleeping metabolic rate (respiratory chamber) was higher not only in subjects with diabetes (+4.9%, P < 0.001) but also in those with impaired glucose tolerance (IGT) (+2.7%, P < 0.01) compared with subjects with NGT. Longitudinally, RMR (ventilated hood) increased progressively in 17 subjects in whom glucose tolerance deteriorated from NGT to IGT (+4.2%) to diabetes (+2.6%) over 5.1 +/- 1.4 years (P < 0.05 for trend). In parallel, IIT (% increase in metabolic rate during an insulin/glucose infusion) decreased during the transition from NGT (11.7%) to IGT (7.3%) to diabetes (6.5%) (P < 0.05 for trend). In 151 subjects, basal endogenous glucose output (3-3H-glucose), fasting insulin and free fatty acid concentrations, and glucose disposal (hyperinsulinemic clamp) were significant determinants of RMR, independent of body composition, age, and sex. Nonoxidative and oxidative glucose disposal, RMR, and fasting insulin and glucose concentrations were determinants of IIT. Differences in RMR and IIT between glucose tolerance groups decreased after adjusting for these factors. These findings indicate that increases in RMR and decreases in IIT occur early in the development of type 2 diabetes, and that both changes are related to the progressive metabolic abnormalities that occur during the development of the disease.
Glucose metabolism and insulin sensitivity in transgenic mice overexpressing leptin with lethal yellow agouti mutation: usefulness of leptin for the treatment of obesity-associated diabetes.
Authors: Masuzaki H , Ogawa Y , Aizawa-Abe M , Hosoda K , Suga J , Ebihara K , Satoh N , Iwai H , Inoue G , Nishimura H , Yoshimasa Y , Nakao K
Published: March 2007

Leptin acts as an adipocyte-derived blood-borne satiety factor that can increase glucose metabolism. To elucidate the therapeutic implications of leptin for obesity-associated diabetes, we crossed transgenic skinny mice overexpressing leptin (Tg/+), which we have developed recently, and lethal yellow KKAy mice (Ay/+), a genetic model for obesity-diabetes syndrome, and examined the metabolic phenotypes of F1 animals. At 6 weeks of age, plasma leptin concentrations in Tg/+ mice with the Ay allele (Tg/+:Ay/+) were significantly higher than those in Ay/+ mice. Although no significant differences in body weight were noted among Tg/+:Ay/+ mice, Ay/+ mice, and their wild-type lean littermates (+/+), glucose and insulin tolerance tests revealed increased glucose tolerance and insulin sensitivity in Tg/+:Ay/+ compared with Ay/+ mice. However, at 12 weeks of age, when plasma leptin concentrations in Ay/+ mice were comparable to those in Tg/+:Ay/+ mice, Tg/+:Ay/+ mice developed obesity-diabetes syndrome similar to that of Ay/+ mice. Body weights of 12-week-old Tg/+:Ay/+ and Ay/+ mice were reduced to those of +/+ mice by a 3-week food restriction; when plasma leptin concentrations remained high in Tg/+:Ay/+ mice but were markedly reduced in Ay/+ and +/+ mice, glucose tolerance and insulin sensitivity in Tg/+:Ay/+ mice were markedly improved as compared with Ay/+ and +/+ mice. The present study demonstrates that hyperleptinemia can delay the onset of impaired glucose metabolism and accelerate the recovery from diabetes during caloric restriction in Tg/+:Ay/+ mice, thereby suggesting the potential usefulness of leptin in combination with a long-term caloric restriction for the treatment of obesity-associated diabetes.
Relationship between insulin resistance and nonmodulating hypertension: linkage of metabolic abnormalities and cardiovascular risk.
Authors: Ferri C , Bellini C , Desideri G , Valenti M , De Mattia G , Santucci A , Hollenberg N K , Williams G H
Published: March 2007

Insulin resistance is a feature common to patients with diabetes and to some with hypertension. It is assumed that this feature confers the increased metabolic risk in hypertension. However, the state of the renin-angiotensin system might contribute to cardiovascular risk, although there is no clear mechanistic explanation. Our recent observation that insulin levels are increased in a specific subset of patients with normal/high-renin hypertension, the nonmodulators, provided the background for the current hypothesis: to ascertain whether abnormalities in lipid and carbohydrate metabolism are observed in the same patients in whom alterations in sodium transport, sodium homeostasis, and the renin-aniotensin system response have been identified. Exploration of a family history of cardiovascular risk was a secondary goal. Insulin sensitivity (assessed by a 75-g oral glucose load), lipid levels, and two defects in the renin-angiotensin system were assessed in 62 hypertensive and 14 normotensive subjects placed on a high (210 mmol/l) and a low (10 mmol/l) sodium intake for 2 weeks, to classify them as low-renin, nonmodulator, or modulating hypertensive subjects. Only in nonmodulators were the following cardiovascular risk factors significantly increased: fasting insulin (P < 0.01); increment in post-glucose load insulin (P < 0.01); total, LDL, and VLDL cholesterol and triglyceride levels (P < 0.05); and erythrocyte Na+/Li+ countertransport activity (P < 0.001). Both nonmodulators and low-renin hypertensive subjects had a significantly (P < 0.01) increased frequency of a family history of hypertension by questionnaire compared with subjects with intact modulation. However, only nonmodulators had a significantly (P < 0.02) higher frequency of a family history of myocardial infarction. Thus, there is a clustering of metabolic abnormalities in a discrete subset of the essential hypertensive population with a specific dysregulation of the renin-angiotensin system--nonmodulation. The absence of this cluster in low-renin hypertensive subjects may explain their relatively diminished cardiovascular risk. Its presence in nonmodulators likely contributes to the increased cardiovascular risk observed in normal/high-renin hypertension.
Plasma phospholipid transfer protein activity is lowered by 24-h insulin and acipimox administration: blunted response to insulin in type 2 diabetic patients.
Authors: Riemens S C , van Tol A , Sluiter W J , Dullaart R P
Published: March 2007

Cholesteryl ester transfer protein (CETP) transfers cholesteryl esters from HDL to VLDL and LDL. Phospholipid transfer protein (PLTP) transfers phospholipids between lipoproteins, converts HDL3 into larger and smaller particles, and is involved in pre-beta-HDL generation. We examined the effects of 24-h hyperinsulinemia (30 mU x kg(-1) x h(-1)) and 24-h Acipimox (250 mg/4 h) on plasma lipids as well as CETP and PLTP activities (measured with exogenous substrate assays) in eight healthy and eight type 2 diabetic subjects. After 24 h of insulin, plasma free fatty acids (FFAs), HDL cholesterol, and plasma apolipoprotein AI decreased in healthy subjects and type 2 diabetic patients (P < 0.05). Plasma triglycerides did not significantly change in either group. After 24 h of Acipimox, all parameters, including plasma triglycerides, decreased in both groups (P < 0.05). Insulin decreased plasma PLTP activity by 17.6% after 24 h in healthy subjects (P < 0.05) and 10.2% in diabetic patients (P < 0.05 vs. baseline; P < 0.05 vs. healthy subjects). Acipimox lowered PLTP activity by 10.3% in healthy subjects (P < 0.05) and 11.3% in diabetic patients (P < 0.05). When insulin was infused for 3 h after Acipimox, a further decrease was found only in healthy subjects. Plasma CETP activity decreased by 9.5% after 24 h of insulin in healthy subjects (P < 0.05), but not in diabetic patients. Acipimox did not decrease plasma CETP activity in either group. In healthy subjects, the PLTP responses with insulin and Acipimox were larger than the changes in CETP activity (P < 0.05). These findings suggest that there is a metabolic link between the regulation of plasma FFA and PLTP, but not CETP. The PLTP response to insulin is blunted in type 2 diabetes.
Characterization of glucokinase mutations associated with maturity-onset diabetes of the young type 2 (MODY-2): different glucokinase defects lead to a common phenotype.
Authors: Miller S P , Anand G R , Karschnia E J , Bell G I , LaPorte D C , Lange A J
Published: March 2007

Glucokinase (GK) is expressed in the pancreatic beta-cells and liver, and plays a key role in the regulation of glucose homeostasis. The enzymatic activity and thermal stability of wild-type (WT) GK and several mutant forms associated with maturity-onset diabetes of the young type 2 (MODY-2) were determined by a steady-state kinetic analysis of the purified expressed proteins. The eight MODY-2 mutations studied were Ala53Ser, Val367Met, Gly80Ala, Thr168Pro, Arg36Trp, Thr209Met, Cys213Arg, and Val226Met. These missense mutations were shown to have variable effects on GK kinetic activity. The Gly80Ala and Thr168Pro mutations resulted in a large decrease in Vmax and a complete loss of the cooperative behavior associated with glucose binding. In addition, the Gly80Ala mutation resulted in a sixfold increase in the half-saturating substrate concentration (S0.5) for ATP, and Thr168Pro resulted in eight- and sixfold increases in the S0.5 values for ATP and glucose, respectively. The Thr209Met and Val226Met mutations exhibited three- and fivefold increases, respectively, in the S0.5 for ATP, whereas the Cys213Arg mutation resulted in a fivefold increase in the S0.5 for glucose. These mutations also led to a small yet significant reduction in Vmax. Of all the mutations studied, only the Cys213Arg mutation had reduced enzymatic activity and decreased thermal stability. Two mutants, Ala53Ser and Val367Met, showed kinetic and thermal stability properties similar to those of WT. These mutants had increased sensitivities to the known negative effectors of GK activity, palmitoyl-CoA, and GK regulatory protein. Taken together, these results illustrate that the MODY-2 phenotype may be linked not only to kinetic alterations but also to the regulation of GK activity.
Association between type 1 diabetes age of onset and HLA among sibling pairs.
Authors: Valdes A M , Thomson G , Erlich H A , Noble J A
Published: March 2007

In this study, we report type 1 diabetes age-of-onset association with HLA class II (DRB1, DQB1, and DPB1) and class I (A) genes in 222 multiplex families from the Human Biological Data Interchange. Linear regression showed a small (R2 = 0.26) but significant correlation in the ages of onset among sib pairs. A strong association in age of onset between members of sib pairs was observed when the analysis was performed using contingency tables that split sibs into three age-at-onset ranges (0-10, 11-20, and 21-36 years). The association is strongest for sib pairs that share both haplotypes and is nonsignificant for sib pairs that do not share any DR-DQ haplotypes. A goodness-of-fit test revealed that DR-DQ haplotype sharing cannot account for all the association in age of onset among sib pairs. The age-of-onset distribution of DR-DQ haplotypes is affected by the DPB1 and A alleles present. The strongest associations were found with the A locus: DR3/DR4 genotype frequency decreases with age of onset in this data set only among A*0101- individuals, and A*2402 is strongly associated with younger age of onset in many DR-DQ haplotypes.
T-1095, an inhibitor of renal Na+-glucose cotransporters, may provide a novel approach to treating diabetes.
Authors: Oku A , Ueta K , Arakawa K , Ishihara T , Nawano M , Kuronuma Y , Matsumoto M , Saito A , Tsujihara K , Anai M , Asano T , Kanai Y , Endou H
Published: March 2007

T-1095A and T-1095 are synthetic agents derived from phlorizin, a specific inhibitor of Na+-glucose cotransporters (SGLTs). Unlike phlorizin, T-1095 is absorbed into the circulation via oral administration, is metabolized to the active form, T-1095A, and suppresses the activity of SGLTs in the kidney. Orally administered T-1095 increases urinary glucose excretion in diabetic animals, thereby decreasing blood glucose levels. Indeed, the postprandial hyperglycemia after a meal load was shown to be suppressed by this compound in streptozotocin (STZ)-induced diabetic rats. With long-term T-1095 treatment, both blood glucose and HbA1c levels were reduced in STZ-induced diabetic rats and yellow KK mice. In addition, there was amelioration of abnormal carbohydrate metabolism, i.e., hyperinsulinemia and hypertriglyceridemia, and of the development of microalbuminuria, in yellow KK mice. Thus, T-1095 may be a useful antidiabetic drug, providing a novel therapeutic approach for diabetes.
Altered hypothalamic function in response to glucose ingestion in obese humans.
Authors: Matsuda M , Liu Y , Mahankali S , Pu Y , Mahankali A , Wang J , DeFronzo R A , Fox P T , Gao J H
Published: March 2007

The hypothalamus plays a central role in the regulation of energy intake and feeding behavior. However, the presence of a functional abnormality in the hypothalamus in humans that may be related to excess energy intake and obesity has yet to be demonstrated in vivo. We, therefore, used functional magnetic resonance imaging (fMRI) to monitor hypothalamic function after oral glucose intake. The 10 obese (34 +/- 2 years of age, BMI 34.2 +/- 1.3 kg/m2) and 10 lean (32 +/- 4 years of age, BMI 22.0 +/- 0.9 kg/m2) subjects with normal glucose tolerance ingested 75 g of glucose while a midsagittal slice through the hypothalamus was continuously imaged for 50 min using a conventional T2*-weighted gradient-echo pulse sequence. After glucose ingestion, lean subjects demonstrated an inhibition of the fMRI signal in the areas corresponding to the paraventricular and ventromedial nuclei. In obese subjects, this inhibitory response was markedly attenuated (4.8 +/- 1.3 vs. 7.0 +/- 0.6% inhibition, P < 0.05) and delayed (9.4 +/- 0.5 vs. 6.4 +/- 0.5 min, P < 0.05) compared with that observed in lean subjects. The time taken to reach the maximum inhibitory response correlated with the fasting plasma glucose (r = 0.75, P < 0.001) and insulin (r = 0.47, P < 0.05) concentrations in both lean and obese subjects. These results demonstrate in vivo, for the first time, the existence of differential hypothalamic function in lean and obese humans that may be secondary to obesity.
Improvement in insulin resistance and the restoration of reduced phosphodiesterase 3B gene expression by pioglitazone in adipose tissue of obese diabetic KKAy mice.
Authors: Tang Y , Osawa H , Onuma H , Nishimiya T , Ochi M , Makino H
Published: March 2007

Phosphodiesterase (PDE) 3B is a key enzyme in the mediation of the antilipolytic action of insulin in adipocytes, and activation of this molecule results in a reduced output of free fatty acids (FFAs). An elevation of serum FFAs is known to cause insulin resistance in skeletal muscle and liver, which could be the primary cause of type 2 diabetes. To elucidate whether PDE3B is involved in this disease, we examined the PDE3B gene expression in epididymal fat tissues of obese insulin-resistant diabetic KKAy mice. We also examined the effect of an insulin-sensitizing drug, pioglitazone, on this gene expression. In adipose tissue of KKAy mice, PDE3B mRNA and its corresponding protein were reduced to 48 and 43% of those in C57BL/6J control mice. Basal and insulin-stimulated membrane-bound PDE activities were also decreased to 50 and 36% of those in the controls, respectively. Pioglitazone increased both PDE3B mRNA and protein levels by 1.8-fold of those in untreated KKAy mice. Basal and insulin-induced membrane-bound PDE activities were also increased by 1.6- and 2.0-fold, respectively. Pioglitazone reduced the elevated levels of serum insulin, glucose, FFAs, and triglyceride in KKAy mice. Thus, the reduced PDE3B gene expression in adipose tissues could be the primary event in the development of insulin resistance in KKAy mice, which was improved by pioglitazone possibly because of the restoration of the reduced PDE3B gene expression.
Autoantibody against N(epsilon)-(carboxymethyl)lysine: an advanced glycation end product of the Maillard reaction.
Authors: Shibayama R , Araki N , Nagai R , Horiuchi S
Published: March 2007

Prolonged incubation of proteins with reducing sugar produces advanced glycation end products (AGEs), which are implicated as factors for aging and diabetic complications. We previously demonstrated the presence of N(epsilon)-(carboxymethyl)lysine (CML), one of the main AGE structures, in human and animal tissues using a monoclonal anti-CML antibody (6D12). These findings suggest that CML structures present in vivo could serve as immunogens to generate autoantibodies. This suggestion was tested in the present study. First, plasma samples from diabetic rats reacted positively with AGE bovine serum albumin (BSA). These reactivities increased with the duration of diabetic states and were inhibited specifically by CML-BSA. Second, a fraction purified from plasma of diabetic patients, which bound to AGE-BSA, showed a positive reaction to CML-BSA and furthermore also to human lens proteins, which are known to undergo CML modification in vivo. Finally, patients with renal failure caused by diabetes or nondiabetic pathologies had a higher autoantibody activity against CML structure than that in normal subjects or diabetic patients without renal failure. These results indicate that CML accumulated in vivo serves as an immunological epitope to generate an autoantibody specific for CML that might be used as a potential marker for diabetic nephropathy or chronic renal failure.
Hyperketonemia can increase lipid peroxidation and lower glutathione levels in human erythrocytes in vitro and in type 1 diabetic patients.
Authors: Jain S K , McVie R
Published: March 2007

Recent studies have suggested that elevated cellular lipid peroxidation may play a role in the development of cellular dysfunction and other complications of diabetes. People with type 1 diabetes frequently encounter elevated levels of the ketone bodies acetoacetate (AA), beta-hydroxybutyrate (BHB), and acetone (ACE). This study was undertaken to test the hypothesis that ketosis might increase lipid peroxidation and lower glutathione (GSH) levels of red blood cells (RBCs) in diabetic patients. This study demonstrates that incubation of AA with normal RBCs in phosphate-buffered saline (37 degrees C for 24 h) resulted in marked GSH depletion, oxidized glutathione accumulation, hydroxyl radical generation, and increased membrane lipid peroxidation. Increases in oxygen radicals and lipid peroxidation and depletion of GSH in RBCs were not observed with BHB or ACE treatments. Similarly, there was a significant generation of superoxide ion radicals even in a cell-free buffer solution of AA, but not in that of BHB. The presence of BHB together with AA did not influence the capacity of AA to generate oxygen radicals in a cell-free solution or the increase in lipid peroxidation of RBCs incubated with AA. The antioxidants vitamin E and N-acetylcysteine (NAC) blocked increase in lipid peroxidation in AA-treated RBCs. To examine the effects of ketone bodies in vivo, studies were performed that showed a significant decrease in GSH and an increase in lipid peroxidation levels in RBCs of hyperketonemic diabetic patients, but not in normoketonemic type 1 diabetic patients, when compared with age-matched normal subjects. This study demonstrates that elevated levels of the ketone body AA can increase lipid peroxidation and lower GSH levels of RBCs in people with type 1 diabetes.
A quantitative trait locus influencing BMI maps to the region of the beta-3 adrenergic receptor.
Authors: Mitchell B D , Cole S A , Comuzzie A G , Almasy L , Blangero J , MacCluer J W , Hixson J E
Published: March 2007

The beta-3 adrenergic receptor (ADRB3) has been implicated as a regulator of energy expenditure, and a polymorphism in codon 64 of this gene (Trp64Arg) has been associated in some studies with obesity and insulin resistance. However, many studies have failed to detect an effect of this variant, and the importance of the Trp64Arg variant in human obesity remains controversial. We performed a quantitative linkage analysis of the ADRB3 and obesity, using 12 markers (including the intragenic Trp64Arg polymorphism) spanning a 57-cM region of chromosome 8. The study population consisted of 470 individuals from 10 large multigenerational families of Mexican-American ancestry residing in San Antonio, TX. In two-point analysis, logarithm of odds (LOD) scores >1.0 were observed for six markers surrounding ADRB3 in a 33-cM region spanned by markers D8S1477 and D8S1136. The multipoint LOD score was 3.21, occurring between markers D8S1121 and ADRB3, approximately 2-3 cM from ADRB3. Adjusting for the presence of the Arg64 allele or excluding from the analysis the 11 individuals homozygous for the Arg64 allele did not reduce the evidence for linkage. A genome scan was conducted at 10 cM map density to detect other loci influencing variation in BMI. Multipoint LOD scores >1.0 were observed in four other regions, including two on chromosome 17, one on chromosome 6q, and one on chromosome 2p. These data suggest that the ADRB3 should continue to be regarded as a strong candidate gene for obesity even though evidence for an effect of the Trp64Arg polymorphism could not be established. It is also possible that a gene closely linked to ADRB3 may influence susceptibility to obesity.
Conservation of an insulin response unit between mouse and human glucose-6-phosphatase catalytic subunit gene promoters: transcription factor FKHR binds the insulin response sequence.
Authors: Ayala J E , Streeper R S , Desgrosellier J S , Durham S K , Suwanichkul A , Svitek C A , Goldman J K , Barr F G , Powell D R , O'Brien R M
Published: March 2007

Because overexpression of the glucose-6-phosphatase catalytic subunit (G-6-Pase) in both type 1 and type 2 diabetes may contribute to the characteristic increased rate of hepatic glucose production, we have investigated whether the insulin response unit (IRU) identified in the mouse G-6-Pase promoter is conserved in the human promoter. A series of human G-6-Pase-chloramphenicol acetyltransferase (CAT) fusion genes was transiently transfected into human HepG2 hepatoma cells, and the effect of insulin on basal CAT expression was analyzed. The results suggest that the IRU identified in the mouse promoter is conserved in the human promoter, but that an upstream multimerized insulin response sequence (IRS) motif that is only found in the human promoter appears to be functionally inactive. The G-6-Pase IRU comprises two distinct promoter regions, designated A and B. Region B contains an IRS, whereas region A acts as an accessory element to enhance the effect of insulin, mediated through region B, on basal G-6-Pase gene transcription. We have previously shown that the accessory factor binding region A is hepatocyte nuclear factor-1, and we show here that the forkhead protein FKHR is a candidate for the insulin-responsive transcription factor binding region B.
Reevaluation of the major histocompatibility complex genes of the NOD-progenitor CTS/Shi strain.
Authors: Mathews C E , Graser R T , Serreze D V , Leiter E H
Published: March 2007

The common Kd and/or Db alleles of NOD mice contribute to the development of autoimmune diabetes, but their respective contributions are unresolved. The major histocompatibility complex (MHC) of the CTS/Shi mouse, originally designated as H2ct, shares MHC class II region identity with the H2g7 haplotype of NOD mice. However, CTS mice were reported to express distinct but undefined MHC class I gene products. Because diabetes frequency was reduced 56% in females of a NOD stock congenic for H2ct, this partial resistance may have derived from the MHC class I allelic differences. In the present report, we use a combination of serologic analysis and sequencing of MHC class I cDNAs to establish that NOD/Lt and CTS/Shi share a common H2-Kd allele but differ at the H2-D end of the MHC complex. The H2-D allele of CTS/Shi was identified as the rare H2-Ddx recently described in ALR/Lt, another NOD-related strain. These results in mouse model systems show that multiple MHC genes confer diabetes resistance and suggest that at least one of the protective MHC or MHC-linked genes in CTS mice may be at the H2-D end of the complex.
Role of adenosine in insulin-stimulated release of leptin from isolated white adipocytes of Wistar rats.
Authors: Cheng J T , Liu I M , Chi T C , Shinozuka K , Lu F H , Wu T J , Chang C J
Published: March 2007

Leptin, the ob gene product that can decrease caloric intake and increase energy expenditure, is functionally released by insulin from adipose tissue. Adenosine is thought to be an important regulator of the action of insulin in adipose tissue. The present study investigated the role of adenosine in the release of leptin by insulin in isolated rat white adipocytes. Release of leptin, measured by radioimmunoassay, from insulin-stimulated samples was seen after 30 min. Adenosine deaminase, at concentrations sufficient to metabolize endogenous adenosine, decreased insulin-stimulated leptin release. Also, the insulin-stimulated leptin release was completely blocked by the adenosine A1 receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine (DPCPX). Mediation of endogenous adenosine in this action of insulin was further supported by the assay of adenosine released into the medium from adipocytes stimulated with insulin. In addition, activation of adenosine A1 receptors by N6-cyclopentyladenosine (CPA) induced an increase in leptin release in a concentration-dependent manner that could be blocked by antagonists, either DPCPX or 8-(p-sulfophenyl)theophylline (8-SPT). In the presence of U73312, a specific inhibitor of phospholipase C (PLC), CPA-stimulated leptin secretion from adipocytes was reduced in a concentration-dependent manner, but it was not affected by U73343, the negative control for U73312. Moreover, chelerythrine and GF 109203X diminished the CPA-stimulated leptin secretion at concentrations sufficient to inhibit protein kinase C (PKC). These results suggest that, in isolated white adipocytes, the released adenosine acts as a helper and/or a positive regulator for insulin in the release of leptin via an activation of adenosine A1 receptors that involves the PLC-PKC pathway.
Evidence of islet cell autoimmunity in elderly patients with type 2 diabetes.
Authors: Pietropaolo M , Barinas-Mitchell E , Pietropaolo S L , Kuller L H , Trucco M
Published: March 2007

In light of an occurring growth of elderly people affected by type 2 diabetes and recent observations indicating that type 2 diabetes may be a disease of the innate immune system, we evaluated whether signs of islet cell autoimmunity are associated with an abnormal glucose control, the presence of insulin requirement, or an activation of the acute-phase response in older individuals with type 2 diabetes. GAD65 and IA-2 autoantibodies along with the acute-phase response markers fibrinogen and C-reactive protein were tested in 196 serum samples from patients with type 2 diabetes and in 94 nondiabetic control subjects over the age of 65 years from the Pittsburgh cohort of the Cardiovascular Health Study. Of the diabetic patients, 12% (24 of 196) had autoantibodies against GAD65 and/or IA-2, a prevalence significantly higher than that found in nondiabetic individuals (1 of 94, 1.1%; P = 0.001). Type 2 diabetic patients who were positive for GAD65 and/or IA-2 autoantibodies (Ab+), as compared with those negative for these autoantibodies (Ab-), had an abnormal oral glucose tolerance test (OGTT) (P = 0.03) before and a higher frequency of oral hypoglycemic treatment (P = 0.003) at the time of autoantibody testing. No differences were seen in the percentage of insulin requirement in the two groups. Moreover, a statistically significant increase in fibrinogen (P = 0.005) and C-reactive protein levels (P = 0.025) was found in type 2 diabetic patients with high levels of GAD65 and/or IA-2 autoantibodies as compared with Ab-patients and control subjects. In conclusion, in type 2 diabetic subjects > or =65 years old, the presence of islet cell autoimmunity is associated with an impairment of the acute-phase insulin secretion, as revealed by an OGTT. A pronounced activation of the acute-phase response, found to be associated with islet cell autoimmunity, may in part explain this defect in insulin secretion. These findings not only have direct implications for adequate classification and treatment of diabetes in the elderly, but also for understanding the autoimmune/inflammatory mechanisms involved in the pathogenesis of hyperglycemia.