bluuebunny/crossref_metadata_2025 huggingface dataset:
Unresponsiveness to glibenclamide during chronic treatment induced by reduction of ATP-sensitive K+ channel activity.
Authors: Kawaki J , Nagashima K , Tanaka J , Miki T , Miyazaki M , Gonoi T , Mitsuhashi N , Nakajima N , Iwanaga T , Yano H , Seino S
Published: March 2007

The insulin response to the sulfonylurea glibenclamide was markedly impaired in pancreatic beta-cell line MIN6 cells with chronic glibenclamide treatment (MIN6-Glib). The intracellular calcium concentration increased only slightly in response to glibenclamide in MIN6-Glib. While the properties of the voltage-dependent calcium channels were not altered, the conductance of the K(ATP) channels, the primary target of glibenclamide, was significantly reduced in MIN6-Glib. The ATP-sensitive K+ (K(ATP)) channels in MIN6 cells comprise inwardly rectifying K+ channel member Kir6.2 subunits and sulfonylurea receptor (SUR) 1 subunits. MIN6 cells have both high- and low-affinity binding sites for glibenclamide. The binding affinities at these two sites were unchanged, but the maximum binding capacities at both sites were similarly increased by chronic glibenclamide treatment. Both SUR1 and Kir6.2 mRNA levels were not altered, but SUR1 protein was rather increased in MIN6-Glib. In addition, electron microscopic examination revealed a majority of the SUR1 to be present in a cluster near the plasma membrane in control MIN6, while it tends to be distributed in the cytoplasm in MIN6-Glib. These data suggest that chronic glibenclamide treatment causes the defect in acute glibenclamide-induced insulin secretion by reducing the number of functional K(ATP) channels on the plasma membrane of the beta-cells.
Rapid increase in circulating leptin in ventromedial hypothalamus-lesioned rats: role of hyperinsulinemia and implication for upregulation mechanism.
Authors: Suga A , Hirano T , Kageyama H , Kashiba M , Oka J , Osaka T , Namba Y , Tsuji M , Miura M , Adachi M , Inoue S
Published: March 2007

The mechanisms of marked increase in plasma leptin soon after ventromedial hypothalamus (VMH) lesions were investigated. Although rats did not gain body weight or parametrial fat-pad mass 24 h after the operation, the acute VMH-lesioned rats exhibited substantial five- and fourfold increases in plasma leptin levels compared with sham-operated control rats in fed (22.6 +/- 3.2 vs. 5.8 +/- 1.2 ng/ml) and fasted (8.8 +/- 2.0 vs. 2.3 +/- 0.3 ng/ml) states, respectively. Plasma insulin concentration was doubled in VMH-lesioned rats compared with sham-operated controls in both fed and fasting states. Northern blot analysis revealed that mRNA of ob gene was not increased in parametrial fat pad of animals 24 h after the creation of VMH lesions. However, leptin content in the fat pad was significantly increased in VMH-lesioned rats compared with sham-operated controls (32.2 +/- 4.7 vs. 17.4 +/- 2.3 ng/g wet tissue). The leptin content in parametrial fat pad was highly correlated with plasma leptin concentrations (r = 0.898, P < 0.001). To define the effect of hyperinsulinemia on their hyperleptinemia, a small dose of streptozotocin (STZ) (25 mg/kg body wt) was intravenously administered into rats 5 days before the creation of VMH lesions. Plasma insulin levels were not increased after VMH lesions in STZ-pretreated rats. Plasma leptin levels were halved in the absence of hyperinsulinemia, but still remained twofold higher than those in their sham-operated counterparts (9.9 +/- 1.3 vs. 4.8 +/- 0.7 ng/ml). These results indicate that the destruction of VMH rapidly promotes leptin production before obesity develops through an enhanced translational process in which hyperinsulinemia occurring after VMH lesioning plays an important role. The present study also suggests that there are other mechanisms that rapidly upregulate leptin production in adipocytes in VMH-lesioned rats in which the target organ of this hormone has been destroyed.
Targeting autoimmune diabetes with gene therapy.
Authors: Giannoukakis N , Rudert W A , Robbins P D , Trucco M
Published: March 2007

The autoimmune nature of insulin-dependent, or type 1, diabetes targets the beta-cells of the pancreas for destruction and results in a lifelong commitment to insulin replacement therapy. Although the number of formulations and dosing of insulin have become more sophisticated and more efficient in recent years, insulin therapy alone is unable to prevent nephropathy, retinopathy, or vascular and heart disease, which still occur in a large number of patients. Different approaches have been attempted to eliminate the requirement of exogenous insulin administration. Historically, these have included pancreatic and islet transplants, which were later combined with treatments intended to halt the destructive process directed against the islets. Despite significant advances made in all of these areas, each approach faces a hostile immunological response that frequently ends with the loss of the islets. Gene therapy-based approaches add a new dimension to the efforts aimed at specifically blocking the immunological attack against the islets in genetically at-risk individuals (autoimmunity) or the immunological response against transplanted allogeneic islets (rejection). This new technology may have an important role in the therapy and cure of type 1 diabetes.
Diabetes-prone and diabetes-resistant BB rats share a common major diabetes susceptibility locus, iddm4: additional evidence for a "universal autoimmunity locus" on rat chromosome 4.
Authors: Martin A M , Maxson M N , Leif J , Mordes J P , Greiner D L , Blankenhorn E P
Published: March 2007

Diabetes-prone (DP) BB rats develop autoimmune type 1 diabetes spontaneously. At least five loci are linked to disease expression: the major histocompatibility complex (iddm2), two susceptibility loci (iddm4, iddm5), and, possibly, a resistance locus (iddm3). Spontaneous disease also requires homozygosity for lyp/iddm1, which causes lymphopenia. It has not been determined whether lyp/iddm1 is required for predisposition to diabetes autoimmunity in addition to being required for its spontaneous expression. We analyzed backcross rats segregating for diabetes but not lymphopenia using Wistar-Furth (WF) and diabetes-resistant (DR) BB animals. The latter are nonlymphopenic (lyp+/+) and develop diabetes only in response to immunological perturbants. Treatment of (DR-BB x WF)F1 x WF animals (all lyp+/+) using a standard induction protocol caused type 1 diabetes in 58% of progeny. Expression of type 1 diabetes was strongly linked to iddm4. The results suggest that lyp/iddm1 does not determine the predisposition to autoimmunity in BB rats and that iddm4 is a major diabetogenic locus in both DP- and DR-BB animals. The iddm4 gene maps to a region containing several major autoimmunity loci, including aia2, aia3, and cia3. We propose that BB rat diabetes requires 1) class II RT1u (iddm2) for susceptibility, 2) additional loci for disease initiation and progression in response to perturbants, and 3) lyp for spontaneous disease.
Resistance of ALR/Lt islets to free radical-mediated diabetogenic stress is inherited as a dominant trait.
Authors: Mathews C E , Leiter E H
Published: March 2007

ALS/Lt and ALR/Lt are inbred mouse strains selected for susceptibility and resistance to alloxan (AL)-induced diabetes. Within 24-h after AL administration in vivo, ALS/Lt islets were distinguished from ALR/Lt islets by more extensive necrotic changes. Within 7 days post-AL, ALS/Lt mice exhibited hyperglycemia and hypoinsulinemia, whereas ALR/Lt mice maintained normal plasma insulin and glucose levels. We have recently shown that resistance in ALR/Lt correlated with constitutively elevated systemic (and pancreatic) free radical defense status. In the present report, we examined whether ability to detoxify free radical stress extended to the level of ALR/Lt pancreatic islets. Cultured ALS/Lt islets exposed for 5 min to increasing (0-3 mmol/l) AL concentrations in vitro exhibited an 80% decline in numbers of intact islets after a subsequent 6-day culture period, as well as a 75% reduction in islet insulin content and a 94% decrease in glucose-stimulated insulin secretory capacity. In contrast, ALR/Lt islets remained viable and retained glucose-stimulated insulin secretory capacity as well as normal insulin content. This ALR/Lt islet resistance extended to hydrogen peroxide, a free radical generator whose entry into beta-cells is not dependent on glucose transporters. The elevated antioxidant defenses previously found in ALR/Lt pancreas were extended to isolated islets, which exhibited significantly higher glutathione and Cu-Zn superoxide dismutase 1 levels compared with ALS/Lt islets. A dominant genetic trait from ALR/Lt controlling this unusual AL resistance was indicated by the finding that reciprocal F1 mice of both sexes were resistant to AL administration in vivo. A backcross to ALS/Lt showed 1:1 segregation for susceptibility/resistance, indicative of a single gene controlling the phenotype. In conclusion, the ALR/Lt mouse may provide important insight into genetic mechanisms capable of rendering islets strongly resistant to free radical-mediated damage.
Evidence of a novel type 2 diabetes locus 50 cM centromeric to NIDDM2 on chromosome 12q.
Authors: Bektas A , Suprenant M E , Wogan L T , Plengvidhya N , Rich S S , Warram J H , Krolewski A S , Doria A
Published: March 2007

To replicate the recent finding of a type 2 diabetes locus (NIDDM2) on 12q, families segregating early-onset autosomal-dominant type 2 diabetes were screened for linkage. Included were 26 Caucasian and 6 non-Caucasian pedigrees with an average age at diabetes diagnosis of 37 +/- 18 years. Affected (n = 233) and nonaffected (n = 152) family members were genotyped for 17 markers covering 90 cM on chromosome 12q. While no evidence for linkage was detected at the NIDDM2 locus, a linkage peak was observed 50 cM centromeric to NIDDM2 at markers D12S375 and D12S1052. In a nonparametric analysis, the Z(all) score was 2.9 (P = 0.015) at D12S375, and increased to 3.8 (P = 0.007) among Caucasian families. Further increase in significance was observed in pedigrees with poor insulin response, with a maximum Z(all) of 6.2 (P = 0.002) at D12S375. Suggestive evidence of linkage was also detected by the parametric analysis, with the heterogeneity logarithm of odds score peaking at 2.5 (alpha = 0.15) between D12S375 and D12S1052. In summary, our data indicate that the NIDDM2 locus does not play a major role in early-onset autosomal-dominant type 2 diabetes. Rather, they strongly suggest that a previously undetected type 2 diabetes locus exists 50 cM from NIDDM2 on 12q.
Hypothalamic neuronal histamine as a target of leptin in feeding behavior.
Authors: Yoshimatsu H , Itateyama E , Kondou S , Tajima D , Himeno K , Hidaka S , Kurokawa M , Sakata T
Published: March 2007

Leptin, an ob gene product, has been shown to suppress food intake by regulating hypothalamic neuromodulators. The present study was designed to examine the involvement of brain histamine in leptin-induced feeding suppression. A bolus infusion of 1.0 microg leptin into the rat third cerebroventricle (i3vt) elevated the turnover rate of hypothalamic neuronal histamine (P < 0.05) as assessed by pargyline-induced accumulation of tele-methylhistamine (t-MH), a major metabolite of histamine. No remarkable change in the mRNA expression of histidine decarboxylase (HDC), a histamine-synthesizing enzyme, was observed in the hypothalamus after i3vt infusion of leptin. These results indicate that leptin increases histamine turnover by affecting the posttranscriptional process of HDC formation or histamine release per se. As expected, concomitant suppression in 24-h cumulative food intake was also observed after infusion of leptin. Systemic depletion of brain histamine levels by pretreatment with an intraperitoneal injection of 224 micromol/kg alpha-fluoromethylhistidine (FMH), a suicide inhibitor of HDC, attenuated the leptin-induced feeding suppression by 50.7% (P < 0.05). This attenuation of feeding suppression was mimicked by the i3vt infusion of 2.24 micromol/kg FMH before leptin treatment (P < 0.05). In addition, concentrations of hypothalamic histamine and t-MH were lowered in diabetic (db/db) mice, which are known to be deficient in leptin receptors (P < 0.05 vs. lean littermates for each amine), although the amine levels were higher in diet-induced obese rats (P < 0.05 for each amine). Leptin-deficient obese mice (ob/ob) showed lower histamine turnover (P < 0.05 vs. lean littermates), which recovered after leptin infusion. Thus, a growing body of results points to an important role for the hypothalamic histamine neurons in the central regulation of feeding behavior controlled by leptin.
Effects of graft-versus-host reaction on intrahepatic islet transplants.
Authors: Li H , Ricordi C , Inverardi L
Published: March 2007

The use of donor-specific bone marrow transplantation might represent a valuable approach for the induction of hyporesponsiveness to concurrent allogeneic organ and tissue grafts. In this setting, the occurrence of subclinical forms of graft-versus-host reaction (GVHR) can seldom be formally ruled out. When systemic administration of donor-specific bone marrow is performed together with intraportal transplantation of islets of Langerhans, GVHR might damage the implanted islets through an innocent bystander mechanism. The liver is, in fact, a prominent target of GVHR, and islets are sensitive to the noxious effects of proinflammatory mediators, including selected cytokines produced during GVHR. A parent (Lewis) to F1 (Lewis x Brown Norway) splenocyte transfer model was used to induce GVHR in rats. Islets were also obtained from Lewis donors and implanted into F1 recipients to examine the effects of GVHR on islet function in the absence of rejection. Selected doses of splenocytes were infused in F1 recipients to induce GVHR. When 150 x 10(6) cells were administered, all recipients developed clinical lethal GVHR. When 100 x 10(6) or 80 x 10(6) cells were given, lethal GVHR was observed in 40 and 20% of the F1 animals, respectively. A transient clinical syndrome occurred in the remaining animals and spontaneously subsided; histological findings consistent with persistent low-grade GVHR were observed in these animals at the time of death several months after splenocyte inoculation. When simultaneous splenocyte infusions and islet grafts were performed in chemically diabetic animals, no adverse effect of either clinical or subclinical GVHR was observed on islet function. Blood glucose profiles were normal, as were intravenous glucose tolerance test profiles. In conclusion, GVHR does not seem to adversely influence function of islets of Langerhans implanted intrahepatically in this parent to F1 experimental model.
Autoantibodies to CD38 (ADP-ribosyl cyclase/cyclic ADP-ribose hydrolase) in Caucasian patients with diabetes: effects on insulin release from human islets.
Authors: Pupilli C , Giannini S , Marchetti P , Lupi R , Antonelli A , Malavasi F , Takasawa S , Okamoto H , Ferrannini E
Published: March 2007

The type II transmembrane glycoprotein CD38 (ADP-ribosyl cyclase/cyclic ADP-ribose hydrolase) has been proposed as a mediator of insulin secretion from pancreatic beta-cells and as a candidate for autoimmune reactions in type 2 diabetes. We evaluated the presence of anti-CD38 autoantibodies in Caucasian patients with diabetes and investigated the effect of these antibodies on insulin secretion from isolated human pancreatic islets. The presence of anti-CD38 autoantibodies was evaluated by using Western blot analysis in 236 patients with type 2 diabetes (mean age 63 years), in 160 patients with type 1 diabetes (mean age 38 years), and in 159 nondiabetic subjects. Anti-CD38 autoantibody titers at least 3 SD above the mean value of the control group were found in 9.7% of type 2 diabetic patients and in 13.1% of type 1 diabetic patients (chi2 = 15.9, P = 0.0003 vs. 1.3% of control subjects). No significant differences were observed in sex distribution, current age, age at diabetes onset, BMI, fasting serum glucose, or glycemic control between anti-CD38+ and anti-CD38-diabetic patients in either the type 2 or type 1 diabetic groups. The effect of 23 anti-CD38- and 13 anti-CD38+ sera on insulin secretion at low (3.3 mmol/l) or high (16.7 mmol/l) medium glucose concentrations was evaluated in isolated human pancreatic islets. Data are medians (interquartile range). The anti-CD38+ sera potentiated insulin release both at low [95 (64) vs. 23 (12) microU/ml of control incubations, respectively, P < 0.0001] and high [271 (336) vs. a control of 55 (37) microU/ml, respectively, P = 0.001] medium glucose concentrations, whereas the anti-CD38- sera did not. Furthermore, in the pooled data from all 36 tested sera, insulin levels in the islet incubation medium were directly related to the anti-CD38 antibody titer. We conclude that autoantibodies to CD38 are associated with both type 1 and type 2 diabetes in Caucasian subjects. These autoantibodies exert a stimulatory effect on insulin secretion by cultured human islets. The role of this autoimmune reaction in the pathogenesis of diabetes remains to be elucidated.
Circulating antibodies against an exocrine pancreatic enzyme in type 1 diabetes.
Authors: Panicot L , Mas E , Thivolet C , Lombardo D
Published: March 2007

In this article, we report the identification of a new autoantigen in type 1 diabetes originating from the exocrine pancreas. This antigen is a pancreatic enzyme termed bile salt-dependent lipase (BSDL). We show that antibodies present in the sera of newly diagnosed type 1 diabetic patients recognize BSDL and more specifically the COOH-terminal mucin-like region of the protein. Therefore, we engineered the COOH-terminal peptide of BSDL and demonstrated that autoreactivity was linked to specific glycosylation sites by at least two glycosyltransferases: the Core 2 beta(1-6)N-acetylglucosaminyltransferase and the alpha(1-3) fucosyltransferase FUT7. We next examined the prevalence of circulating anti-BSDL antibodies in type 1 diabetic patients and found 73.5% positivity (25 sera among 34 patients tested) at onset, whereas only 8.4% of normal individuals (7 of 83) were positive. Within a cohort of first-degree relatives of diabetic patients followed prospectively until development of diabetes, 6 of 19 (31.6%) were also positive. Interestingly, two prediabetic individuals were already positive for anti-BSDL antibodies (Abs), while islet cell cytoplasmic Abs and antibodies to GAD65, IA-2, and insulin were not detected. Anti-BSDL autoantibodies were weakly or not detected in patients suffering from pancreatitis or pancreatic adenocarcinoma or in patients with Graves' disease. Although autoreactivity to BSDL in prediabetic and newly diagnosed diabetic patients might reflect cross-reactivity, our results strongly suggest that in addition to pancreatic beta-cells, acinar cells may be also affected in type 1 diabetes.
Increases in phosphorylation of the myosin II heavy chain, but not regulatory light chains, correlate with insulin secretion in rat pancreatic islets and RINm5F cells.
Authors: Wilson J R , Biden T J , Ludowyke R I
Published: March 2007

Although cytoskeletal proteins such as myosin II are implicated in the control of insulin secretion, their precise role is poorly understood. In other secretory cells, myosin II is predominantly regulated via the phosphorylation of the regulatory light chains (RLC). The current study was aimed at investigating RLC phosphorylation in beta-cells. In both the insulin-secreting cell line RINm5F and rat pancreatic islets, the RLC was basally phosphorylated on the myosin light chain kinase sites (Ser19/Thr18). Phosphorylation at these sites was not consistently increased by either metabolic stimuli (glyceraldehyde/glucose) or the depolarizing agent KCl. The RLC sites phosphorylated by protein kinase C (PKC) (Ser1/Ser2) were unphosphorylated in the basal state, not affected by nutrients or KCl, and only slightly increased by the PKC activator phorbol 12-myristate 13-acetate (PMA). Like the other insulin secretagogues, however, PMA did promote serine phosphorylation of the myosin heavy chain (MHC) in RINm5F cells. Phosphopeptide mapping suggested that the same peptide was phosphorylated under both PMA and glyceraldehyde stimulation, which further extends our previous study of the Ca2+-dependent phosphorylation of this protein (Wilson JR, Ludowyke RI, Biden TJ: Nutrient stimulation results in a rapid Ca2+-dependent threonine phosphorylation of myosin heavy chain in rat pancreatic islets and RINm5F cells. J Biol Chem 273:22729-22737, 1998). Overall, our results demonstrate that in RINm5F cells and rat pancreatic islets, MHC phosphorylation correlates better with insulin secretion than phosphorylation of the RLC. We therefore propose that in beta-cells, in contrast to other secretory cells, phosphorylation of the MHC is more important than that of the RLC for regulation of the myosin II protein complex during insulin secretion.
Amadori albumin in type 1 diabetic patients: correlation with markers of endothelial function, association with diabetic nephropathy, and localization in retinal capillaries.
Authors: Schalkwijk C G , Ligtvoet N , Twaalfhoven H , Jager A , Blaauwgeers H G , Schlingemann R O , Tarnow L , Parving H H , Stehouwer C D , van Hinsbergh V W
Published: March 2007

Nonenzymatic glycation is increased in diabetes. Most studies so far have focused on the role of advanced glycation end products (AGEs) in vascular complications, whereas the role of early glycation Amadori-modified proteins, which is the predominant form of glycated proteins, has not been systemically investigated in humans. We developed an antiserum against glycated human serum albumin (HSA) and used this to study the role of early glycation products in vascular complications in type 1 diabetic patients. Amadori albumin was determined to be the recognition epitope of the antiserum. The antibody recognized a specific glucose adduct and a conformational component specific for human albumin in Amadori albumin, with no recognition of AGEs. Plasma Amadori albumin levels were significantly higher in type 1 diabetic patients (n = 55) than in healthy control subjects (n = 60) (39.2+/-9.9 vs. 20.9+/-4.0 U/ml, P < 0.0005). Amadori albumin correlated with levels of plasma markers of endothelial function von Willebrand factor (r = 0.29, P < 0.05) and vascular cell adhesion molecule-1 (r = 0.41, P < 0.005), but not soluble E-selectin. In addition, Amadori albumin immunoreactivity was detected in the capillaries of retinas of diabetic patients. Plasma levels of Amadori albumin were determined in a second group of type 1 diabetic patients with long-standing diabetes with (n = 199) or without (n = 192) diabetic nephropathy. Patients with nephropathy had higher Amadori albumin levels than did those without it (50.9+/-9.5 vs. 45.1+/-6.3 U/ml, P < 0.0005). Age-, sex-, and diabetes duration-adjusted analyses showed that nephropathy was significantly associated with Amadori albumin with an odds ratio (OR [95% CI]) of 1.11 [1.08-1.15] per U/ml increase. After additional adjustment for levels of creatinine, glycated hemoglobin, cholesterol, triglycerides, blood pressure, preexistent retinopathy, and cardiovascular disease, Amadori albumin continued to be significantly associated with nephropathy (OR 1.06 [1.01-1.11]) per U/ml increase. Our results are consistent with a proposed pathophysiological role of Amadori albumin in microvascular complications of type 1 diabetic patients.
Troglitazone, an antidiabetic agent, inhibits cholesterol biosynthesis through a mechanism independent of peroxisome proliferator-activated receptor-gamma.
Authors: Wang M , Wise S C , Leff T , Su T Z
Published: March 2007

Troglitazone is an antidiabetic agent of the thiazolidinedione family. It is generally believed that thiazolidinediones exert their insulin-sensitizing activity through activation of peroxisome proliferator-activated receptor-gamma (PPAR-gamma), a member of the steroid nuclear receptor superfamily. In the present study, we examined the effect of troglitazone on cholesterol biosynthesis in cultured Chinese hamster ovary (CHO) cells. Troglitazone inhibited biosynthesis of cholesterol, but not that of total sterols, in a dose-dependent manner, with a half-maximal concentration (IC50) value of 8 micromol/l. At 20 micromol/l, troglitazone inhibited cholesterol biosynthesis by more than 80%, resulting in the accumulation of lanosterol and several other sterol products. This inhibitory effect observed in CHO cells was also reproduced in HepG2, L6, and 3T3-L1 cells, suggesting that there is a common pathway for this troglitazone action. One hour after removal of troglitazone from the culture medium, disappearance of the accumulated sterols was accompanied by restored cholesterol synthesis, indicating that those accumulated sterols are precursors of cholesterol. PPAR-gamma reporter assays showed that PPAR-gamma activation by troglitazone was completely blocked by actinomycin D and cycloheximide. In contrast, the inhibition of cholesterol synthesis by troglitazone remained unchanged in the presence of the above compounds, suggesting that this inhibition is mechanistically distinct from the transcriptional regulation by PPAR-gamma. Like troglitazone, two other thiazolidinediones, ciglitazone and englitazone, exhibited similar inhibitory effect on cholesterol synthesis; however, other known PPAR-gamma ligands such as BRL49653, pioglitazone, and 15-deoxy-delta(12,14)-prostaglandin J2 showed only weak or no inhibition. The dissociation of PPAR-gamma binding ability from the potency for inhibition of cholesterol synthesis further supports the conclusion that inhibition of cholesterol biosynthesis by troglitazone is unlikely to be mediated by PPAR-gamma.
Renal glucose production during insulin-induced hypoglycemia in humans.
Authors: Cersosimo E , Garlick P , Ferretti J
Published: March 2007

We investigated the effects of hypoglycemia on renal glucose production (RGP) and renal glucose uptake (RGU) using arteriovenous balance combined with tracer technique in humans. Our 14 healthy subjects had arterialized hand veins (artery) and renal veins (under fluoroscopy) catheterized after an overnight fast. Systemic and renal glucose kinetics were measured with infusion of [6-(2)H2]glucose, and renal plasma flow was measured by para-aminohippurate clearance. After a 150-min equilibration period, artery and renal vein samples were obtained between -30 and 0 min, and subjects received a 180-min peripheral insulin infusion (0.250 mU kg(-1) x min(-1)) with a variable infusion of [6-(2)H2]dextrose adjusted to maintain plasma glucose at either approximately 60 mg/dl (hypoglycemic clamp) or approximately 90 mg/dl (euglycemic clamp). Blood samples were obtained between 150 and 180 min during the study period. Insulin increased from 49 +/- 14 to 130 +/- 25 (hypoglycemia) and to 102 +/- 10 (euglycemia) pmol/l. Glucose decreased from 5.32 +/- 0.11 to 3.58 +/- 0.07 micromol/ml during hypoglycemia, but it did not change during euglycemia (5.20 +/- 0.19 vs. 5.05 +/- 0.15 micromol/ml). Endogenous glucose production decreased (9.30 +/- 0.70 vs. 5.65 +/- 0.50) during euglycemia but not during hypoglycemia (9.80 +/- 0.50 vs. 10.25 +/- 0.60 micromol x kg(-1) x min(-1)). During hypoglycemia, net renal glucose output increased from 0.54 +/- 0.30 to 2.31 +/- 0.40, RGP increased from 1.88 +/- 0.70 to 3.65 +/- 0.50 (P < 0.05), and RGU did not change (1.34 +/- 0.50 vs. 1.34 +/- 0.60 micromol x kg(-1) x min(-1)). During euglycemia, renal glucose balance switched from a net output of 0.72 +/- 0.20 to a net uptake of 1.70 +/- 0.92, RGP decreased from 2.31 +/- 0.50 to 1.20 +/- 0.58, and RGU increased from 1.59 +/- 0.50 to 2.90 +/- 0.70 micromol x kg(-1) x min(-1) (P < 0.05). During hypoglycemia, arterial glucagon increased from 105 +/- 6 to 129 +/- 8, epinephrine increased from 116 +/- 28 to 331 +/- 33, norepinephrine increased from 171 +/- 9 to 272 +/- 9 (all P < 0.05), and renal vein norepinephrine increased from 236 +/- 13 to 426 +/- 50 (P < 0.001). These data indicate that, in addition to counterregulatory hormones, activation of the autonomic nervous system during hypoglycemia stimulates glucose production by the kidney, which may represent an important additional component of the body's defense against hypoglycemia in humans.
Functional properties of leptin receptor isoforms: internalization and degradation of leptin and ligand-induced receptor downregulation.
Authors: Uotani S , Bjørbaek C , Tornøe J , Flier J S
Published: March 2007

Long (ObRb) and short (ObRa) leptin receptor isoforms are thought to play essential roles in mediating leptin signaling and the transport and degradation of leptin, respectively. Although the capacity of these cloned receptor species to mediate signal transduction has been reported, there is no information on the ability of individual receptor species to mediate leptin internalization and degradation or to undergo ligand-induced downregulation. We therefore studied these parameters in Chinese hamster ovary (CHO) cells stably expressing either ObRa or ObRb isoforms of the leptin receptor. We determined that both ObRa and ObRb mediated internalization of 125I-labeled leptin by a temperature- and coated pit-dependent mechanism. Both ObRa and ObRb also mediated degradation of 125I-leptin by a lysosomal mechanism, and this was more efficiently mediated by ObRa in these cells. Neither leptin internalization nor degradation by ObRa was affected by mutation of the conserved Box 1 motif. By studying deletion mutants of ObRa, we found that efficient internalization was dependent on a motif located between amino acids 8 and 29 of the intracellular domain of ObRa. Exposure of cells expressing ObRa or ObRb to unlabeled leptin for 90 min at 37 degrees C produced downregulation of available surface receptors, and this effect was of greater magnitude in cells expressing ObRb. Whereas CHO cells expressing the growth hormone receptor showed marked downregulation of ligand binding after exposure to dexamethasone (DEX) or phorbol myristic acid (PMA), PMA had no effect on expression of ObRa or ObRb, and DEX reduced binding to cells expressing ObRb by 15%. Thus, the two leptin receptor isoforms, ObRa and ObRb, mediate leptin internalization by a coated pit-dependent mechanism, leptin degradation by a lysosomal pathway, and ligand-induced receptor downregulation. The differential capacity of the two receptor isoforms may relate to the different roles of the receptor isoforms in the biology of leptin.
Microinjection of leptin into the ventromedial hypothalamus increases glucose uptake in peripheral tissues in rats.
Authors: Minokoshi Y , Haque M S , Shimazu T
Published: March 2007

We studied the effects of microinjection of leptin into the ventromedial hypothalamus (VMH) and lateral hypothalamus (LH) on glucose uptake in peripheral tissues in unanesthetized rats. The rate of glucose uptake was assessed in vivo by 2-[3H]deoxyglucose incorporation. Single injection of leptin into VMH increased glucose uptake in brown adipose tissue (BAT), heart, skeletal muscles, and spleen but not in white adipose tissue or skin. On the other hand, microinjection of leptin into LH had little effect on glucose uptake in those tissues. The plasma concentrations of glucose and insulin were unaltered by intrahypothalamic injection of leptin into either VMH or LH. Among skeletal muscles, the increase in glucose uptake induced by intrahypothalamic injection of leptin was greater in the soleus than in the extensor digitorum longus. Likewise, the increased glucose uptake in the gastrocnemius in response to leptin was more prominent in the red part than in the white part of the tissue. When surgical sympathetic denervation of the interscapular BAT was performed, the enhanced glucose uptake by BAT in response to intrahypothalamic leptin was completely suppressed. These findings suggest that intrahypothalamic injection of leptin preferentially increases glucose uptake by some peripheral tissues through activation of the VMH-sympathetic (or its neighboring medial hypothalamus-sympathetic) nervous system, thereby contributing to the maintenance of energy balance.
Contributions by kidney and liver to glucose production in the postabsorptive state and after 60 h of fasting.
Authors: Ekberg K , Landau B R , Wajngot A , Chandramouli V , Efendic S , Brunengraber H , Wahren J
Published: March 2007

Contributions of renal glucose production to whole-body glucose turnover were determined in healthy individuals by using the arteriovenous balance technique across the kidneys and the splanchnic area combined with intravenous infusion of [U-13C6]glucose, [3-(3)H]glucose, or [6-(3)H]glucose. In the postabsorptive state, the rate of glucose appearance was 11.5 +/- 0.6 micromol x kg(-1) x min(-1). Hepatic glucose production, calculated as the sum of net glucose output (9.8 +/- 0.8 micromol x kg(-1) x min(-1)) and splanchnic glucose uptake (2.2 +/- 0.3 micromol x kg(-1) x min(-1)) accounted for the entire rate of glucose appearance. There was no net exchange of glucose across the kidney and no significant renal extraction of labeled glucose. The renal contribution to total glucose production calculated from the arterial, hepatic, and renal venous 13C-enrichments (glucose M+6) was 5 +/- 2%. In the 60-h fasted state, the rate of glucose appearance was 8.2 +/- 0.3 micromol x kg(-1) x min(-1). Hepatic glucose production, estimated as net splanchnic output (5.8 +/- 0.7 micromol x kg(-1) x min(-1)) plus splanchnic uptake (0.6 +/- 0.3 micromol x kg(-1) x min(-1)) accounted for 79% of the rate of glucose appearance. There was a significant net renal output of glucose (0.9 +/- 0.3 micromol x kg(-1) x min(-1)), but no significant extraction of labeled glucose across the kidney. The renal contribution to whole-body glucose turnover calculated from the 13C-enrichments was 24 +/- 3%. We concluded that 1) glucose production by the human kidney in the postabsorptive state, in contrast to recent reports, makes at most only a minor contribution (approximately 5%) to blood glucose homeostasis, but that 2) after 60-h of fasting, renal glucose production may account for 20-25% of whole-body glucose turnover.
Glucosamine infusion in rats rapidly impairs insulin stimulation of phosphoinositide 3-kinase but does not alter activation of Akt/protein kinase B in skeletal muscle.
Authors: Kim Y B , Zhu J S , Zierath J R , Shen H Q , Baron A D , Kahn B B
Published: March 2007

Glucosamine, a metabolite of glucose via the hexosamine biosynthetic pathway, potently induces insulin resistance in skeletal muscle by impairing insulin-induced GLUT4 translocation to the plasma membrane. Activation of phosphoinositide (PI) 3-kinase is necessary for insulin-stimulated GLUT4 translocation, and the serine/threonine kinase Akt/protein kinase B (PKB) is a downstream mediator of some actions of PI 3-kinase. To determine whether glucosamine-induced insulin resistance could be due to impaired signaling, we measured insulin receptor substrate (IRS)-1 and insulin receptor tyrosine phosphorylation; PI 3-kinase activity associated with IRS-1, IRS-2, and phosphotyrosine; and Akt activity and phosphorylation in skeletal muscle of rats infused for 2 h with glucosamine (6.0 mg x kg(-1) x min(-1)) or saline. Euglycemic-hyperinsulinemic clamp studies (12 mU x kg(-1) x min(-1) insulin) in awake rats showed that glucosamine infusion resulted in rapid induction of insulin resistance, with a 33% decrease in glucose infusion rate (P < 0.01). Tissues were harvested after saline alone (basal), 1 min after an insulin bolus (10 U/kg), or after 2 h of insulin clamp in saline- and glucosamine-infused rats. After 1 min of insulin stimulation, phosphorylation of IRS-1 and insulin receptor increased 6- to 8-fold in saline-infused rats and 7- to 10-fold in glucosamine-infused rats. In saline-infused rats, 1 min of insulin stimulation increased PI 3-kinase activity associated with IRS-1, IRS-2, or phosphotyrosine 7.6-, 6.4-, and 10-fold, respectively. In glucosamine-infused rats treated for 1 min with insulin, PI 3-kinase activity associated with IRS-1 was reduced 28% (P < 0.01) and that associated with phosphotyrosine was reduced 43% (P < 0.01). Insulin for 1 min stimulated Akt/PKB activity approximately 5-fold in both saline- and glucosamine-infused rats; insulin-induced hyperphosphorylation of Akt/PKB was not different between groups. Glucosamine infusion alone had no effect on tyrosine phosphorylation of the insulin receptor or IRS-1 or on stimulation of PI 3-kinase or Akt/PKB activity. However, 2 h of insulin clamp reduced PI 3-kinase activity associated with IRS-1, IRS-2, or phosphotyrosine to <30% of that seen with 1 min of insulin. No effect of glucosamine was seen on these signaling events when compared with 2 h of insulin clamp without glucosamine. Our data show that 1) glucosamine infusion in rats is associated with an impairment in the early activation of PI 3-kinase by insulin in skeletal muscle, 2) this insulin-resistant state does not involve alterations in the activation of Akt/PKB, and 3) prolonged insulin infusion under clamp conditions results in a blunting of the PI 3-kinase response to insulin.
13C nuclear magnetic resonance study of glycogen resynthesis in muscle after glycogen-depleting exercise in healthy men receiving an infusion of lipid emulsion.
Authors: Delmas-Beauvieux M C , Quesson B , Thiaudière E , Gallis J L , Canioni P , Gin H
Published: March 2007

In healthy individuals, glycogen recovery after a strong depletion is known to be rapid and insulin independent during the initial phase, and subsequently, slow and insulin dependent. Free fatty acids (FFAs) as a putative source of insulin resistance (IR) could thus impair glycogen recovery during the second period. Using in vivo 13C nuclear magnetic resonance (NMR), we studied the effect of long-chain triglyceride emulsion on gastrocnemius glycogen resynthesis during a 3-h recovery period after 90 min of moderate exercise consisting of plantar flexion on overnight-fasted healthy men (n = 8). In separate experiments, each subject was infused with 10% Ivelip (0.015 ml x kg(-1) x min(-1)) or 10% glycerol (0.13 mg x kg(-1) x min(-1)). NMR spectra were acquired before and at the end of the exercise and during the recovery period. Whole-body glucose and lipid oxidation rates (indirect calorimetry), plasma insulin, C-peptide, glucose, lactate, beta-hydroxybutyrate, triglycerides, and FFAs were determined. Glycogen consumption was 47.6 +/- 4.5% (glycerol) and 49.7 +/- 4.8% (Ivelip) of the initial glycogen. An acquired IR in the Ivelip group was significant at the onset of the recovery period by homeostasis model assessment (P = 0.002). Glycogen resynthesis in the glycerol group appeared faster during the 1st h than during the subsequent 2nd h of the postexercise period. The glycogen resynthesis level was significantly lower in the Ivelip group than in the glycerol group during the recovery period (P = 0.04 during the 1st h and P = 0.001 during the next 2 h). During the recovery, plasma lactate and whole-body oxidation rates were similar in the two groups, whereas glycemia was significantly higher in the Ivelip group. A decreased cellular uptake of glucose as a substrate for glycogenosynthesis, rather than a competition between oxidation of carbohydrate and FFA, is discussed.
Hyperglycemia-induced embryonic dysmorphogenesis correlates with genomic DNA mutation frequency in vitro and in vivo.
Authors: Lee A T , Reis D , Eriksson U J
Published: March 2007

Congenital malformations affecting multiple organ systems are at least three times more common in infants of mothers with IDDM than in infants born to nondiabetic mothers. Numerous studies have confirmed the teratogenic effect of hyperglycemia on the developing embryo, although no direct mechanism has been determined. In this study, we aimed to correlate the frequency of lacI mutations with degree of hyperglycemic exposure and severity of malformations in mouse embryos from in vitro cultures. Day 8 transgenic mouse embryos cultured in 30 or 50 mmol/l glucose for 48 h exhibited a higher incidence of morphological abnormalities, as well as an increase in lacI mutation frequency, compared with embryos cultured in 10 mmol/l glucose with no abnormalities and a lower frequency of lacI mutations. We also used a transgenic lacI rat system to evaluate the relationship between abnormal embryonic development and DNA mutation frequency in day 11 embryos of severely diabetic rats (serum glucose >20 mmol/l). Compared with control embryos, the embryos from diabetic rats displayed significantly more malformations, shorter crown-rump lengths, fewer somites, and more than six times greater genomic DNA mutation frequency. Genetic analysis of the mutated lacI gene from both in vitro cultured mouse embryos and in vivo developed rat embryos revealed that the majority of mutations were due to base substitutions (transitions and transversions), but that the rate of large DNA mutations tended to increase in embryos exposed to a diabetic environment. Our results support the interrelationship between increased rates of congenital malformations and DNA mutations in the offspring of diabetic pregnancy.
Inhibitory effect of a growth hormone receptor antagonist (G120K-PEG) on renal enlargement, glomerular hypertrophy, and urinary albumin excretion in experimental diabetes in mice.
Authors: Flyvbjerg A , Bennett W F , Rasch R , Kopchick J J , Scarlett J A
Published: March 2007

Growth hormone (GH) and IGFs have a long and distinguished history in diabetes, with possible participation in the development of renal complications. To investigate the effect of a newly developed GH receptor (GHR) antagonist (G120K-PEG) on renal/glomerular hypertrophy and urinary albumin excretion (UAE), streptozotocin-induced diabetic and nondiabetic mice were injected with G120K-PEG every 2nd day for 28 days. Placebo-treated diabetic and nondiabetic animals were used as reference groups. Placebo-treated diabetic animals were characterized by growth retardation, hyperphagia, hyperglycemia, increased serum GH levels, reduced serum IGF-I, IGF-binding protein (IGFBP)-3, and liver IGF-I levels, increased kidney IGF-I, renal/glomerular hypertrophy, and increased UAE when compared with nondiabetic animals. No differences were seen between the two diabetic groups with respect to body weight, food intake, blood glucose, serum GH, IGF-I, and IGFBP-3 levels or hepatic IGF-I levels. Kidney IGF-I, kidney weight, and glomerular volume were normalized, while the rise in UAE was partially attenuated in the G120K-PEG-treated diabetic animals. No effect of G120K-PEG treatment on any of the parameters mentioned above was seen in nondiabetic animals. In conclusion, administration of a GHR antagonist in diabetic mice has renal effects without affecting metabolic control and circulating levels of GH, IGF-I, or IGFBP-3, thus indicating that the effect of G120K-PEG may be mediated through a direct inhibitory effect on renal IGF-I through the renal GHR. The present study suggests that specific GHR blockade may present a new concept in the treatment of diabetic kidney disease.
Effect of acute ketosis on the endothelial function of type 1 diabetic patients: the role of nitric oxide.
Authors: Avogaro A , Calò L , Piarulli F , Miola M , deKreutzenberg S , Maran A , Burlina A , Mingardi R , Tiengo A , Del Prato S
Published: March 2007

In type 1 diabetic patients, acute loss of metabolic control is associated with increased blood flow, which is believed to favor the development of long-term complications. The mechanisms for inappropriate vasodilation are partially understood, but a role of endothelium-derived nitric oxide (NO) production can be postulated. We assessed, in type 1 diabetic patients, the effect of the acute loss of metabolic control and its restoration on forearm endothelial function in 13 type 1 diabetic patients who were studied under conditions of mild ketosis on two different occasions. In study 1, after basal determination, a rapid amelioration of the metabolic picture was obtained by insulin infusion. In study 2, seven type 1 diabetic patients underwent the same experimental procedure, except that fasting plasma glucose was maintained constant throughout. Basal plasma venous concentrations of nitrites/nitrates (NO2- + NO3-) were determined both before and after intravenous insulin infusion. Endothelium-dependent and -independent vasodilation of the brachial artery was assessed by an intra-arterial infusion of N(G)-monomethyl-L-arginine (L-NMMA) and sodium nitroprusside (SNP), respectively. The same parameters were determined in 13 control subjects at baseline conditions and during a hyperinsulinemic-euglycemic glucose clamp. Baseline forearm blood flow (4.89 +/- 0.86 vs. 3.65 +/- 0.59 ml x (100 ml tissue)(-1) x min(-1)) and NO2- + NO3- concentration (30 +/- 8 vs. 24 +/- 3 micromol/l) were higher in type 1 diabetic patients than in control subjects (P < 0.05). Insulin infusion was associated with lower forearm blood flow and plasma (NO2- + NO3-) concentration (P < 0.05), irrespective of the prevailing glucose levels, as compared with patients under ketotic conditions. The responses to L-NMMA were significantly lower in type 1 diabetic patients during euglycemia and hyperglycemic hyperinsulinemia (-11 +/- 5 and -10 +/- 4%, respectively, of the ratio of the infused arm to the control arm) than in control subjects at baseline (-18 +/- 6%, P < 0.05) and during hyperinsulinemia (-32 +/- 11%, P < 0.01). We conclude that the acute loss of metabolic control is associated with a functional disturbance of the endothelial function characterized by hyperemia and increased NO release during ketosis and blunted NO-mediated vasodilatory response during restoration of metabolic control by intravenous insulin. This functional alteration is unlikely to be explained by hyperglycemia itself.
Is blood glucose predictable from previous values? A solicitation for data.
Authors: Bremer T , Gough D A
Published: March 2007

An important question about blood glucose control in diabetes is, Can present and future blood glucose values be predicted from recent blood glucose history? If this is possible, new continuous blood glucose monitoring technologies under development may lead to qualitatively better therapeutic capabilities. Not only could continuous monitoring technologies alert a user when a hypoglycemic episode or other blood glucose excursion is underway, but measurements may also provide sufficient information to predict near-future blood glucose values. A predictive capability based only on recent blood glucose history would be advantageous because there would be no need to involve models of glucose and insulin distribution, with their inherent requirement for detailed accounting of vascular glucose loads and insulin availability. Published data analyzed here indicate that blood glucose dynamics are not random, and that blood glucose values can be predicted, at least for the near future, from frequently sampled previous values. Data useful in further exploring this concept are limited, however, and an appeal is made for collection of more.