bluuebunny/crossref_metadata_2025 huggingface dataset:
In Vivo Evidence for Increased Oxidation of Circulating LDL in Impaired Glucose Tolerance
Authors: Kopprasch Steffi , Pietzsch Jens , Kuhlisch Eberhard , Fuecker Katja , Temelkova-Kurktschiev Theodora , Hanefeld Markolf , Kühne Helmut , Julius Ulrich , Graessler Jürgen
Published: March 2007

Oxidized LDL (oxLDL) is a key mediator in atherogenesis and a marker of coronary artery disease (CAD). Type 2 diabetes is associated with excessive cardiovascular morbidity and mortality. Because atherogenesis starts before diabetes is diagnosed, we investigated whether circulating oxLDL levels are increased in impaired glucose tolerance (IGT). OxLDL levels were measured in 376 subjects with normal glucose tolerance (NGT), 113 patients with IGT, and 54 patients with newly diagnosed type 2 diabetes. After correction for age and BMI, serum levels of oxLDL were significantly increased in IGT versus NGT subjects (P = 0.002). OxLDL levels were not associated with the following parameters of the oxidative/antioxidative balance in the blood: total antioxidant capacity, urate-to-allantoin ratio, and circulating phagocyte oxygenation activity. In stepwise multivariate analysis, LDL cholesterol (P < 0.0005) and triglycerides (P < 0.0005) were the strongest predictors of circulating oxLDL levels, followed by HDL cholesterol (P = 0.003), 2-h postchallenge C-peptide (P = 0.011), fasting free fatty acids (P = 0.013), and serum paraoxonase activity (P = 0.035). The strong correlation of oxLDL with LDL cholesterol and triglycerides indicates that LDL oxidation in IGT is preferentially associated with dyslipidemia. OxLDL increase may explain the high atherogenic potency of dyslipidemia in the prediabetic state.
Components of the “Metabolic Syndrome” and Incidence of Type 2 Diabetes
Authors: Hanson Robert L. , Imperatore Giuseppina , Bennett Peter H. , Knowler William C.
Published: March 2007

The combination of insulin resistance, dyslipidemia, hypertension, and obesity has been described as a “metabolic syndrome” that is a strong determinant of type 2 diabetes. Factor analysis was used to identify components of this syndrome in 1,918 Pima Indians. Prospective analyses were conducted to evaluate associations of identified factors with incidence of diabetes. Factor analysis identified 4 factors that accounted for 79% of the variance in the original 10 variables. Each of these factors reflected a proposed component of the metabolic syndrome: insulinemia, body size, blood pressure, and lipid metabolism. Among 890 originally nondiabetic participants with follow-up data, 144 developed diabetes in a median follow-up of 4.1 years. The insulinemia factor was strongly associated with diabetes incidence (incidence rate ratio [IRR] for a 1-SD difference in factor scores = 1.81, P < 0.01). The body size and lipids factors also significantly predicted diabetes (IRR 1.52 and 1.37, respectively, P < 0.01 for both), whereas the blood pressure factor did not (IRR 1.11, P = 0.20). Identification of four unique factors with different associations with incidence of diabetes suggests that the correlations among these variables reflect distinct metabolic processes, about which substantial information may be lost in the attempt to combine them into a single entity.
The Role of Autoimmunity in Islet Allograft Destruction
Authors: Makhlouf Leila , Kishimoto Koji , Smith Rex N. , Abdi Reza , Koulmanda Maria , Winn Henry J. , Auchincloss Hugh , Sayegh Mohamed H.
Published: March 2007

Although it has often been assumed that transplanted allogeneic islets can be destroyed by recurrent autoimmunity in recipients with type 1 diabetes, definitive evidence is lacking and the settings in which this may occur have not been defined. To address these issues, we compared the survival of islet transplants (subject to tissue-specific autoimmunity) with cardiac transplants (not subject to tissue-specific autoimmunity) from various major histocompatibility complex (MHC)-matched and -mismatched donors transplanted into autoimmune NOD recipients. We found that when recipients were treated with combined B7 and CD154 T-cell costimulatory blockade, hearts survived best with better MHC matching, whereas islets survived worst when the donor and recipient shared MHC class II antigens. In the absence of full or MHC class II matching, there was no difference in the survival of islet and cardiac allografts. We also found that the tendency of NOD mice to resist tolerance induction by costimulation blockade is mediated by both CD4+ and CD8+ T-cells, not directly linked to the presence of autoimmunity, and conferred by non-MHC background genes. These findings have clinical importance because they suggest that under some circumstances, avoiding MHC class II sharing may provide better islet allograft survival in recipients with autoimmune diabetes, since mismatched allogeneic islets may be resistant to recurrent autoimmunity. Our results may have implications for the design of future clinical trials in islet transplantation.
Uncoupling Protein 2 Knockout Mice Have Enhanced Insulin Secretory Capacity After a High-Fat Diet
Authors: Joseph Jamie W. , Koshkin Vasilij , Zhang Chen-Yu , Wang Jing , Lowell Bradford B. , Chan Catherine B. , Wheeler Michael B.
Published: March 2007

Uncoupling protein 2 (UCP2) may act as an important regulator of insulin secretion. In this study, β-cell function in UCP2-deficient mice was examined after a 45% high-fat diet (HFD) to assess its role during the development of diet-induced type 2 diabetes. HFD-fed UCP2 (−/−) mice have lower fasting blood glucose and elevated insulin levels when compared with wild-type (WT) mice. UCP2 (−/−) mice also have enhanced β-cell glucose sensitivity compared with WT mice after HFD, a result that is due in part to the deterioration of glucose responsiveness in WT mice. HFD-fed UCP2 (−/−) mice have increased insulin secretory capacity as a result of increased pancreatic β-cell mass and insulin content per islet. Islets from WT mice exposed to 0.5 mmol/l palmitate for 48 h have significantly reduced mitochondrial membrane potential, ATP concentrations, and glucose responsiveness compared with UCP2 (−/−) islets, suggesting that elevated UCP2 in WT mice increases proton leak and decreases mitochondrial ATP production. Highly increased carnitine palmitoyl transferase-1 gene expression in UCP2 (−/−) mice is suggestive of enhanced fatty acid oxidizing capacity, particularly after HFD stress. These results further establish UCP2 as a component in glucose sensing and suggest a possible new aspect of UCP2 function during the progression of type 2 diabetes.
Glucagon Is Required for Early Insulin-Positive Differentiation in the Developing Mouse Pancreas
Authors: Prasadan Krishna , Daume Erica , Preuett Barry , Spilde Troy , Bhatia Amina , Kobayashi Hiroyuki , Hembree Mark , Manna Pradip , Gittes George K.
Published: March 2007

The embryonic pancreas is thought to develop from pluripotent endodermal cells that give rise to endocrine and exocrine cells. A key guidance mechanism for pancreatic development has previously been found to be epithelial-mesenchymal interaction. Interactions within the epithelium, however, have not been well studied. Glucagon is the earliest peptide hormone present at appreciable levels in the developing pancreatic epithelium (embryonic day [E]-9.5 in mouse). Insulin accumulation begins slightly later (E11 in mouse), followed by a rapid accumulation during the “second wave” of insulin differentiation (∼E15). Here we found that blocking early expression and function of glucagon, but not GLP-1, an alternate gene product of preproglucagon mRNA, prevented insulin-positive differentiation in early embryonic (E11) pancreas. These results suggest a novel concept and a key role for glucagon in the paracrine induction of differentiation of other pancreatic components in the early embryonic pancreas.
Induction of Autoimmune Diabetes Through Insulin (but Not GAD65) DNA Vaccination in Nonobese Diabetic and in RIP-B7.1 Mice
Authors: Karges Wolfram , Pechhold Klaus , Al Dahouk Sascha , Riegger Ines , Rief Matthias , Wissmann Andrea , Schirmbeck Reinhold , Barth Christoph , Boehm Bernhard O.
Published: March 2007

Insulin has been used to modify T-cell autoimmunity in experimental models of type 1 diabetes. In a large clinical trial, the effect of insulin to prevent type 1 diabetes is currently investigated. We here show that insulin can adversely trigger autoimmune diabetes in two mouse models of type 1 diabetes, using intramuscular DNA vaccination for antigen administration. In female nonobese diabetic (NOD) mice, diabetes development was enhanced after preproinsulin (ppIns) DNA treatment, and natural diabetes resistance in male NOD mice was diminished by ppIns DNA vaccination. In contrast, GAD65 DNA conferred partial diabetes protection, and empty DNA plasmid was without effect. In RIP-B7.1 C57BL/6 mice (expressing the T-cell costimulatory molecule B7.1 in pancreatic β-cells), autoimmune diabetes occurred in 70% of animals after ppIns vaccination, whereas diabetes did not develop spontaneously in RIP-B7.1 mice or after GAD65 or control DNA treatment. Diabetes was characterized by diffuse CD4+CD8+ T-cell infiltration of pancreatic islets and severe insulin deficiency, and ppIns, proinsulin, and insulin DNA were equally effective for disease induction. Our work provides a new model of experimental autoimmune diabetes suitable to study mechanisms and outcomes of insulin-specific T-cell reactivity. In antigen-based prevention of type 1 diabetes, diabetes acceleration should be considered as a potential adverse result.
Relationship of Phospholipid Transfer Protein Activity to HDL and Apolipoprotein B-Containing Lipoproteins in Subjects With and Without Type 1 Diabetes
Authors: Colhoun Helen M. , Taskinen Marja-Riitta , Otvos James D. , van den Berg Paul , O’Connor John , Van Tol Arie
Published: March 2007

Patients with type 1 diabetes have greatly increased phospholipid transfer protein (PLTP) activity and have an altered HDL subclass distribution. In 195 patients with type 1 diabetes and in 194 men and women aged 30–55 years, we examined the relationship of PLTP activity to HDL and examined whether PLTP activity contributes to differences in HDL found in type 1 diabetes. PLTP activity was measured using an exogenous substrate assay. Average HDL particle size and HDL subclasses were measured using nuclear magnetic resonance spectroscopy. Apolipoprotein AI (apoAI) and apoAII were measured by immunoturbidimetry. The amount of apoAI present in LpAI was measured using a differential electroimmunoassay, and the amount of apoAI in LpAIAII was inferred from the apoAI and LpAI data. Higher PLTP activity was associated with more large HDL (P < 0.001) and less small HDL (P < 0.01), more apoAI and apoAII (both at P < 0.001), and more apoAI in both LpAI and LpAIAII (P = 0.02 and P < 0.001, respectively). These associations were independent of other lipids and enzyme activities. Adjusting for PLTP activity halved the difference between subjects with and without diabetes in apoA1 (from 10.1 mg/dl higher in subjects with diabetes to 4.6 mg/dl higher) and large HDL (2.4 μmol/l higher to 1.2 μmol/l higher) and reduced the difference in HDL size (from 0.31 nm higher to 0.26 nm higher). PLTP activity was also positively associated with apoB, total VLDL and LDL particle number, and IDL level in subjects with diabetes. These data support the idea that PLTP is a major factor in HDL conversion and remodeling in humans and that higher PLTP activity makes an important contribution to the higher apoAI levels and altered HDL subclass distribution in type 1 diabetes. They also support a role for PLTP in the metabolism of apoB-containing lipoproteins.
Association of IL4R Haplotypes With Type 1 Diabetes
Authors: Mirel Daniel B. , Valdes Ana Maria , Lazzeroni Laura C. , Reynolds Rebecca L. , Erlich Henry A. , Noble Janelle A.
Published: March 2007

We have investigated, in 282 multiplex Caucasian families (the Human Biological Data Interchange Repository), the association of type 1 diabetes with polymorphisms in the IL4R gene. IL4R encodes a subunit of the interleukin-4 receptor, a molecule critical to T-helper cell development. By genotyping eight different IL4R single-nucleotide polymorphisms (SNPs) and identifying haplotypes (complex alleles) in the multiplex type 1 diabetic families who were stratified for HLA genotype, we have observed significant evidence of linkage and association of the IL4R gene to type 1 diabetes. In particular, we have identified a specific haplotype that appears to be protective and observed that this protective effect is strongest among individuals not carrying the HLA DR3/DR4 genotype (which confers the strongest genetic risk for type 1 diabetes). These findings suggest an important role for the IL4R gene in immune-related disease susceptibility and illustrate the value of using multi-SNP haplotype information in association studies.
The Rise of Childhood Type 1 Diabetes in the 20th Century
Authors: Gale Edwin A.M.
Published: March 2007

The incidence of childhood type 1 diabetes increased worldwide in the closing decades of the 20th century, but the origins of this increase are poorly documented. A search through the early literature revealed a number of useful but neglected sources, particularly in Scandinavia. While these do not meet the exacting standards of more recent surveys, tentative conclusions can be drawn concerning long-term changes in the demography of the disease. Childhood type 1 diabetes was rare but well recognized before the introduction of insulin. Low incidence and prevalence rates were recorded in several countries over the period 1920–1950, and one carefully performed study showed no change in childhood incidence over the period 1925–1955. An almost simultaneous upturn was documented in several countries around the mid-century. The overall pattern since then is one of linear increase, with evidence of a plateau in some high-incidence populations and of a catch-up phenomenon in some low-incidence areas. Steep rises in the age-group under 5 years have been recorded recently. The disease process underlying type 1 diabetes has changed over time and continues to evolve. Understanding why and how this produced the pandemic of childhood diabetes would be an important step toward reversing it.
Interleukin-6 Induces Cellular Insulin Resistance in Hepatocytes
Authors: Senn Joseph J. , Klover Peter J. , Nowak Irena A. , Mooney Robert A.
Published: March 2007

Interleukin (IL)-6 is one of several proinflammatory cytokines that have been associated with insulin resistance and type 2 diabetes. A two- to threefold elevation of circulating IL-6 has been observed in these conditions. Nonetheless, little evidence supports a direct role for IL-6 in mediating insulin resistance. Here, we present data that IL-6 can inhibit insulin receptor (IR) signal transduction and insulin action in both primary mouse hepatocytes and the human hepatocarcinoma cell line, HepG2. This inhibition depends on duration of IL-6 exposure, with a maximum effect at 1–1.5 h of pretreatment with IL-6 in both HepG2 cells and primary hepatocytes. The IL-6 effect is characterized by a decreased tyrosine phosphorylation of IR substrate (IRS)-1 and decreased association of the p85 subunit of phosphatidylinositol 3-kinase with IRS-1 in response to physiologic insulin levels. In addition, insulin-dependent activation of Akt, important in mediating insulin’s downstream metabolic actions, is markedly inhibited by IL-6 treatment. Finally, a 1.5-h preincubation of primary hepatocytes with IL-6 inhibits insulin-induced glycogen synthesis by 75%. These data suggest that IL-6 plays a direct role in insulin resistance at the cellular level in both primary hepatocytes and HepG2 cell lines and may contribute to insulin resistance and type 2 diabetes.
Intrahepatic Islet Transplantation in Type 1 Diabetic Patients Does Not Restore Hypoglycemic Hormonal Counterregulation or Symptom Recognition After Insulin Independence
Authors: Paty Breay W. , Ryan Edmond A. , Shapiro A.M. James , Lakey Jonathan R.T. , Robertson R. Paul
Published: March 2007

Islet allotransplantation can provide prolonged insulin independence in selected individuals with type 1 diabetes. Whether islet transplantation also restores hypoglycemic counterregulation is unclear. To determine if hypoglycemic counterregulation is restored by islet transplantation, we studied hormone responses and hypoglycemic symptom recognition in seven insulin-independent islet transplant recipients using a 3-h stepped hypoglycemic clamp, and compared their responses to those of nontransplanted type 1 diabetic subjects and nondiabetic control subjects. Glucagon responses of islet transplant recipients to hypoglycemia were significantly less than that observed in control subjects (incremental glucagon [mean ± SE]: −12 ± 12 vs. 64 ± 22 pg/ml, respectively; P < 0.05), and not significantly different from that of nontransplanted type 1 diabetic subjects (−17 ± 10 pg/ml). Epinephrine responses and symptom recognition were also not restored by islet transplantation (incremental epinephrine [mean ± SE]: 195 ± 128 [islet transplant recipients] vs. 238 ± 73 [type 1 diabetic subjects] vs. 633 ± 139 pg/ml [nondiabetic control subjects], P < 0.05 vs. control; peak symptom scores: 3.3 ± 0.9 [islet transplant recipients] vs. 3.1 ± 1.1 [type 1 diabetic subjects] vs. 6.7 ± 0.8 [nondiabetic control subjects]). Thus the results indicate that despite providing prolonged insulin independence and near-normal glycemic control in these patients with long-standing type 1 diabetes, hypoglycemic hormonal counterregulation and symptom recognition were not restored by intrahepatic islet transplantation.
Exogenous Nitric Oxide and Endogenous Glucose-Stimulated β-Cell Nitric Oxide Augment Insulin Release
Authors: Smukler Simon R. , Tang Lan , Wheeler Michael B. , Salapatek Anne Marie F.
Published: March 2007

The role nitric oxide (NO) plays in physiological insulin secretion has been controversial. Here we present evidence that exogenous NO stimulates insulin secretion, and that endogenous NO production occurs and is involved in the regulation of insulin release. Radioimmunoassay measurement of insulin release and a dynamic assay of exocytosis using the dye FM1-43 demonstrated that three different NO donors—hydroxylamine (HA), sodium nitroprusside, and 3-morpholinosydnonimine (SIN-1)—each stimulated a marked increase in insulin secretion from INS-1 cells. Pharmacological manipulation of the guanylate cyclase/guanosine 3′,5′-cyclic monophosphate pathway indicated that this pathway was involved in mediating the effect of the intracellular NO donor, HA, which was used to simulate endogenous NO production. This effect was further characterized as involving membrane depolarization and intracellular Ca2+ ([Ca2+]i) elevation. SIN-1 application enhanced glucose-induced [Ca2+]i responses in primary β-cells and augmented insulin release from islets in a glucose-dependent manner. Real-time monitoring of NO using the NO-sensitive fluorescent dye, diaminofluorescein, was used to provide direct and dynamic imaging of NO generation within living β-cells. This showed that endogenous NO production could be stimulated by elevation of [Ca2+]i levels and by glucose in both INS-1 and primary rat β-cells. Scavenging endogenously produced NO-attenuated glucose-stimulated insulin release from INS-1 cells and rat islets. Thus, the results indicated that applied NO is able to exert an insulinotropic effect, and implicated endogenously produced NO in the physiological regulation of insulin release.
Hyperglycemia Inhibits Capacitative Calcium Entry and Hypertrophy in Neonatal Cardiomyocytes
Authors: Pang Yi , Hunton Dacia L. , Bounelis Pam , Marchase Richard B.
Published: March 2007

Hyperglycemia alters cardiac function and often leads to diabetic cardiomyopathy as cardiomyocyte apoptosis causes a hypertrophied heart to deteriorate to dilation and failure. Paradoxically, many short-term animal models of hyperglycemia protect against ischemia-induced damage, including apoptosis, by limiting Ca2+ overload. We have determined that, like nonexcitable cells, both neonatal and adult cardiomyocytes respond to depletion of sarcoplasmic/endoplasmic reticulum Ca2+ stores with an influx of extracellular Ca2+ through channels distinct from voltage-gated Ca2+ channels, a process termed capacitative Ca2+ entry (CCE). Here, we demonstrate that in neonatal rat cardiomyocytes, hyperglycemia decreased CCE induced by angiotensin II or the Ca2+ATPase inhibitor thapsigargin. Hyperglycemia also significantly blunted Ca2+-dependent hypertrophic responses by ∼60%, as well as the Ca2+-sensitive nuclear translocation of a chimeric protein bearing the nuclear localization signal of a nuclear factor of activated T-cells transcription factor. The attenuation of CCE by hyperglycemia was prevented by azaserine, an inhibitor of hexosamine biosynthesis, and partially by inhibitors of oxidative stress. This complements previous work showing that increasing hexosamine metabolites in neonatal cardiomyocytes also inhibited CCE. The inhibition of CCE by hyperglycemia thus provides a likely explanation for the transition to diabetic cardiomyopathy as well as to the protection afforded to injury after ischemia/reperfusion in diabetic models.
Early Complement Activation and Decreased Levels of Glycosylphosphatidylinositol-Anchored Complement Inhibitors in Human and Experimental Diabetic Retinopathy
Authors: Zhang Jing , Gerhardinger Chiara , Lorenzi Mara
Published: March 2007

Diabetic retinal microangiopathy is characterized by increased permeability, leukostasis, microthrombosis, and apoptosis of capillary cells, all of which could be caused or compounded by activation of complement. In this study, we observed deposition of C5b-9, the terminal product of complement activation, in the wall of retinal vessels of human eye donors with 9 ± 3 years of type 2 diabetes, but not in the vessels of age-matched nondiabetic donors. C5b-9 often colocalized with von Willebrand factor in luminal endothelium. C1q and C4, the complement components unique to the classical pathway, were not detected in the diabetic retinas, suggesting that C5b-9 was generated via the alternative pathway, the spontaneous activation of which is regulated by complement inhibitors. The diabetic donors showed a prominent reduction in the retinal levels of CD55 and CD59, the two complement inhibitors linked to the plasma membrane by glycosylphosphatidylinositol anchors, but not in the levels of transmembrane CD46. Similar complement activation in retinal vessels and selective reduction in the levels of retinal CD55 and CD59 were observed in rats with a 10-week duration of streptozotocin-induced diabetes. Thus, diabetes causes defective regulation of complement inhibitors and complement activation that precede most other manifestations of diabetic retinal microangiopathy. These are novel clues for probing how diabetes affects and damages vascular cells.
FOXC2 mRNA Expression and a 5′ Untranslated Region Polymorphism of the Gene Are Associated With Insulin Resistance
Authors: Ridderstråle Martin , Carlsson Emma , Klannemark Mia , Cederberg Anna , Kösters Christina , Tornqvist Hans , Storgaard Heidi , Vaag Allan , Enerbäck Sven , Groop Leif
Published: March 2007

The human transcription factor FOXC2 has recently been shown to protect against diet-induced insulin resistance in transgenic mice. We investigated the expression of FOXC2 in fat and muscle and performed a genetic analysis in human subjects. FOXC2 mRNA levels were increased in visceral compared with subcutaneous fat from obese subjects (12 ± 4-fold; P = 0.0001), and there was a correlation between whole-body insulin sensitivity and FOXC2 mRNA levels in visceral fat (fS-insulin R = −0.64, P = 0.01, and homeostasis model assessment of insulin resistance [HOMA-IR] R = −0.68, P = 0.007) and skeletal muscle (fS-insulin R = −0.57, P = 0.03, and HOMA-IR R = −0.55, P = 0.04). Mutation screening of the FOXC2 gene identified a common polymorphism in the 5′ untranslated region (C-512T). The T allele was associated with enhanced insulin sensitivity (HOMA-IR P = 0.007) and lower plasma triglyceride levels in females (P = 0.007). Also, the higher expression of FOXC2 in visceral than in subcutaneous fat was restricted to subjects homozygous for the T allele (P = 0.03 vs. P = 0.7). Our data suggest that increased FOXC2 expression may protect against insulin resistance in human subjects and that genetic variability in the gene may influence features associated with the metabolic syndrome.
Variants Within the Calpain-10 Gene on Chromosome 2q37 (NIDDM1) and Relationships to Type 2 Diabetes, Insulin Resistance, and Impaired Acute Insulin Secretion Among Scandinavian Caucasians
Authors: Rasmussen Søren K. , Urhammer Søren A. , Berglund Lars , Jensen Jan N. , Hansen Lars , Echwald Søren M. , Borch-Johnsen Knut , Horikawa Yukio , Mashima Hirosato , Lithell Hans , Cox Nancy J. , Hansen Torben , Bell Graeme I. , Pedersen Oluf
Published: March 2007

Variations in the calpain-10 gene (CAPN10) have been identified among Mexican-Americans, and an at-risk haplotype combination (112/121) defined by three polymorphisms, UCSNP-43, -19, and -63, confers increased risk of type 2 diabetes. Here we examine the three polymorphisms in 1,594 Scandinavian subjects, including 409 type 2 diabetic patients, 200 glucose-tolerant control subjects, 322 young healthy subjects, 206 glucose-tolerant offspring of diabetic patients, and 457 glucose-tolerant 70-year-old men. The frequency of the 112/121 combination was not significantly different in 409 type 2 diabetic subjects compared with 200 glucose-tolerant control subjects (0.06 vs. 0.05; odds ratio 1.32 [95% CI 0.58–3.30]). In glucose-tolerant subjects, neither the single-nucleotide polymorphisms individually nor the 112/121 combination were associated with alterations in plasma glucose, serum insulin, or serum C-peptide levels at fasting or during an oral glucose tolerance test, estimates of insulin sensitivity, or glucose-induced insulin secretion. In conclusion, the frequency of the 112/121 at-risk haplotype of CAPN10 is low among Scandinavians and we were unable to demonstrate significant associations between the CAPN10 variants and type 2 diabetes, insulin resistance, or impaired insulin secretion.
A Quantitative Trait Locus Influencing Type 2 Diabetes Susceptibility Maps to a Region on 5q in an Extended French Family
Authors: Martin Lisa J. , Comuzzie Anthony G. , Dupont Sophie , Vionnet Nathalie , Dina Christian , Gallina Sophie , Houari Mouna , Blangero John , Froguel Philippe
Published: March 2007

Type 2 diabetes is a heterogeneous disorder of glucose metabolism characterized by insulin resistance, β-cell dysfunction, and increased glucose production by the liver. Given the high degree of genetic heterogeneity, multiple genes with small to moderate effects may influence susceptibility to diabetes. To circumvent this limitation, we searched for quantitative trait loci (QTLs) that explain the variation in susceptibility of type 2 diabetes in a single extended family, as these individuals are likely to share polymorphisms. We collected genotypic and phenotypic data on 152 individuals ascertained through a multimedia campaign in France to find diabetes-prone families for genetic studies. The effects of genes and covariates (age and sex) on diabetes status were estimated using a threshold model and a maximum likelihood variance component approach. We obtained suggestive evidence of linkage (logarithm of odds [LOD] = 2.4) for diabetes status on chromosome 5q. Within the 1-LOD unit support interval, there are two strong candidates: PCSK1 and CAST. Furthermore, we have obtained a replication (LOD = 1.6) for a QTL for type 2 diabetes on chromosome 11 detected by Hanson and colleagues (1998).
Cellular Basis of Diabetic Nephropathy
Authors: Huang Chunmei , Kim Youngki , Caramori Maria Luiza A. , Fish Alfred J. , Rich Stephen S. , Miller Michael E. , Russell Gregory B. , Mauer Michael
Published: March 2007

Transforming growth factor-β (TGF-β) may be critical in the development of diabetic nephropathy (DN), and genetic predisposition is an important determinant of DN risk. We evaluated mRNA expression levels of TGF-β system components in cultured skin fibroblasts (SFs) from type 1 diabetic patients with fast versus slow development of DN. A total of 125 long-standing type 1 diabetic patients were ranked by renal mesangial expansion score (MES) based on renal biopsy findings and diabetes duration. Patients in the highest quintile of MES who were also microalbuminuric or proteinuric (n = 16) were classified as “fast-track” for DN, while those in the lowest quintile who were also normoalbuminuric (n = 23) were classsified as “slow-track” for DN. Twenty-five normal subjects served as control subjects. SFs were cultured in medium with 25 mmol/l glucose for 36 h. SF mRNA expression levels for TGF-β1, TGF-β type II receptor (TGF-β RII), thrombospondin-1, and latent TGF-β binding protein-1 (LTBP-1) were measured by real-time RT-PCR. LTBP-1 mRNA expression was reduced in slow-track (0.99 ± 0.38) versus fast-track patients (1.65 ± 0.52, P = 0.001) and control subjects (1.41 ± 0.7, P = 0.025). mRNA levels for TGF-β1, TGF-β RII, and thrombospondin-1 were similar in the three groups. Reduced LTBP-1 mRNA expression in SFs from slow-track patients may reflect genetically determined DN protection and suggests that LTBP-1 may be involved in the pathogenesis of DN through the regulation of TGF-β activity.
Transcriptional Regulation of Adipocyte Hormone-Sensitive Lipase by Glucose
Authors: Smih Fatima , Rouet Philippe , Lucas Stéphanie , Mairal Aline , Sengenes Coralie , Lafontan Max , Vaulont Sophie , Casado Marta , Langin Dominique
Published: March 2007

Hormone-sensitive lipase (HSL) catalyzes the rate-limiting step in the mobilization of fatty acids from adipose tissue, thus determining the supply of energy substrates in the body. HSL mRNA was positively regulated by glucose in human adipocytes. Pools of stably transfected 3T3-F442A adipocytes were generated with human adipocyte HSL promoter fragments from −2,400/+38 to −31/+38 bp linked to the luciferase gene. A glucose-responsive region was mapped within the proximal promoter (−137 bp). Electromobility shift assays showed that upstream stimulatory factor (USF)-1 and USF2 and Sp1 and Sp3 bound to a consensus E-box and two GC-boxes in the −137-bp region. Cotransfection of the −137/+38 construct with USF1 and USF2 expression vectors produced enhanced luciferase activity. Moreover, HSL mRNA levels were decreased in USF1- and USF2-deficient mice. Site-directed mutagenesis of the HSL promoter showed that the GC-boxes, although contributing to basal promoter activity, were dispensable for glucose responsiveness. Mutation of the E-box led to decreased promoter activity and suppression of the glucose response. Analogs and metabolites were used to determine the signal metabolite of the glucose response. The signal is generated downstream of glucose-6-phosphate in the glycolytic pathway before the triose phosphate step.
Effects of Free Fatty Acids and Glycerol on Splanchnic Glucose Metabolism and Insulin Extraction in Nondiabetic Humans
Authors: Shah Pankaj , Vella Adrian , Basu Ananda , Basu Rita , Adkins Aron , Schwenk W. Frederick , Johnson C. Michael , Nair K. Sreekumaran , Jensen Michael D. , Rizza Robert A.
Published: March 2007

The present study sought to determine whether elevated plasma free fatty acids (FFAs) alter the ability of insulin and glucose to regulate splanchnic as well as muscle glucose metabolism. To do so, FFAs were increased in 10 subjects to ∼1 mmol/l by an 8-h Intralipid/heparin (IL/Hep) infusion, whereas they fell to levels near the detection limit of the assay (<0.05 mmol/l) in 13 other subjects who were infused with glycerol alone at rates sufficient to either match (n = 5, low glycerol) or double (n = 8, high glycerol) the plasma glycerol concentrations observed during the IL/Hep infusion. Glucose was clamped at ∼8.3 mmol/l, and insulin was increased to ∼300 pmol/l to stimulate both muscle and hepatic glucose uptake. Insulin secretion was inhibited with somatostatin. Leg and splanchnic glucose metabolism were assessed using a combined catheter and tracer dilution approach. Leg glucose uptake (21.7 ± 3.5 vs. 48.3 ± 9.3 and 57.8 ± 11.7 μmol · kg−1 leg · min−1) was lower (P < 0.001) during IL/Hep than the low- or high-glycerol infusions, confirming that elevated FFAs caused insulin resistance in muscle. IL/Hep did not alter splanchnic glucose uptake or the contribution of the extracellular direct pathway to UDP-glucose flux. On the other hand, total UDP-glucose flux (13.2 ± 1.7 and 12.5 ± 1.0 vs. 8.1 ± 0.5 μmol · kg−1 · min−1) and flux via the indirect intracellular pathway (8.4 ± 1.2 and 8.1 ± 0.6 vs. 4.8 ± 0.05 μmol · kg−1 · min−1) were greater (P < 0.05) during both the IL/Hep and high-glycerol infusions than the low-glycerol infusion. In contrast, only IL/Hep increased (P < 0.05) splanchnic glucose production, indicating that elevated FFAs impaired the ability of the liver to autoregulate. Splanchnic insulin extraction, directly measured using the arterial and hepatic vein catheters, did not differ (67 ± 3 vs. 71 ± 5 vs. 69 ± 1%) during IL/Hep and high- and low-glycerol infusions. We conclude that elevated FFAs exert multiple effects on glucose metabolism. They inhibit insulin- and glucose-induced stimulation of muscle glucose uptake and suppression of splanchnic glucose production. They increase the contribution of the indirect pathway to glycogen synthesis and impair hepatic autoregulation. On the other hand, they do not alter either splanchnic glucose uptake or splanchnic insulin extraction in nondiabetic humans.
T-Cell Tolerance by Dendritic Cells and Macrophages as a Mechanism for the Major Histocompatibility Complex-Linked Resistance to Autoimmune Diabetes
Authors: Thiessen Shari , Serra Pau , Amrani Abdelaziz , Verdaguer Joan , Santamaria Pere
Published: March 2007

For poorly understood reasons, the development of autoimmune diabetes in humans and mice is dominantly inhibited by major histocompatibility complex (MHC) class II molecules with diverse antigen-binding sites. We have previously shown that thymocytes expressing a highly diabetogenic I-Ag7-restricted T-cell receptor (TCR) (4.1-TCR) undergo negative selection in mice carrying one copy of the antidiabetogenic H-2b haplotype in an I-Ab-dependent but superantigen-independent manner. Here, we show that 4.1-TCR-transgenic thymocytes undergo different forms of tolerance in NOD mice expressing antidiabetogenic I-Ad, I-Ag7PD, or I-Eαk transgenes. The ability of protective MHC class II molecules to induce thymocyte tolerance in 4.1-TCR-transgenic NOD mice correlates with their ability to prevent diabetes in non-TCR-transgenic mice and is associated with polymorphisms within positions 56–67 of their β1 domains. The 4.1-thymocyte tolerogenic activity of these MHC class II molecules is mediated by dendritic cells and macrophages but not by B-cells or thymic epithelial cells and is a peptide-dependent process. Antidiabetogenic MHC class II molecules may thus afford diabetes resistance by presenting, on dendritic cells and macrophages, tolerogenic peptides to a subset of highly diabetogenic and MHC-promiscuous CD4+ T-cells that play a critical role in the initiation of diabetes.
Feedback Control of the ATP-Sensitive K+ Current by Cytosolic Ca2+ Contributes to Oscillations of the Membrane Potential in Pancreatic β-Cells
Authors: Rolland Jean-François , Henquin Jean-Claude , Gilon Patrick
Published: March 2007

During glucose stimulation, pancreatic β-cells display membrane potential oscillations that correspond to intermittent Ca2+ influx, leading to oscillations of the cytosolic free calcium concentration ([Ca2+]c) and insulin secretion. The role of ATP-sensitive K+ (K+-ATP) channels in the control of these oscillations was investigated by measuring the K+-ATP current (IKATP) with the perforated mode of the patch-clamp technique. No oscillations of IKATP were observed when glucose-stimulated β-cells were kept hyperpolarized, thus with low and stable [Ca2+]c. However, increasing [Ca2+]c by Ca2+ influx (depolarizing pulses) or Ca2+ mobilization (acetylcholine) transiently augmented IKATP. This effect was abolished by tolbutamide, attenuated by increasing the glucose concentration in the medium, and prevented by abrogation of the [Ca2+]c rise, which demonstrates that the current is really IKATP and that its increase is Ca2+-dependent. Injection of a current of a similar amplitude to that of the Ca2+-induced increase in IKATP was sufficient to repolarize glucose-stimulated β-cells. These results suggest that, in the absence of [Ca2+]c oscillations, no metabolic oscillations affect IKATP in pancreatic β-cells. In contrast, [Ca2+]c oscillations evoke IKATP oscillations. This mechanism may constitute the feedback loop controlling the glucose-induced oscillating electrical activity in β-cells.
Endocrine Pancreas Development in Growth-Retarded Human Fetuses
Authors: Béringue Frédérique , Blondeau Bertrand , Castellotti Marie Claire , Bréant Bernadette , Czernichow Paul , Polak Michel
Published: March 2007

Glucose intolerance in adults born with intrauterine growth retardation (IUGR) may involve peripheral insulin resistance and/or abnormal endocrine pancreas development during fetal life. We quantified insulin-containing cells in deceased human fetuses with IUGR (<10th percentile, n = 21) or normal growth (control fetuses, n = 15). Paraffin-embedded pancreatic tissues from fetuses older than 32 weeks were obtained from two fetopathology departments. Mean gestational age was 36 weeks in both groups. Tissues with lysis and those fetuses with defects, aneuploidy, or genetic abnormalities were excluded. For each subject, six pancreatic sections spaced evenly throughout the organ were immunostained with anti-insulin antibody. Total tissue and insulin-positive areas were measured by computer-assisted quantitative morphometry. Results were expressed in percentages. To evaluate islet morphogenesis, the percentages of β-cells inside and outside islets were determined. Islet density was similar in the two groups (P = 0.86). The percentage of pancreatic area occupied by β-cells (β-cell fraction) was not correlated with gestational age (r = 0.06 and P = 0.97 in IUGR fetuses; r = 0.12 and P = 0.67 in control fetuses) or body weight (r = 0.16 and P = 0.47 in IUGR fetuses; r = 0.24 and P = 0.39 in control fetuses). Mean β-cell fraction was 2.53% in the IUGR fetuses and 2.86% in the control fetuses (P = 0.47). The percentage of β-cells located within islets was identical in the two groups (mean 35%). Our data militate against a primary developmental pancreatic abnormality in human IUGR, leaving peripheral insulin resistance as the most likely mechanism of glucose intolerance in adults born with IUGR.
Glucagon Replacement via Micro-Osmotic Pump Corrects Hypoglycemia and α-Cell Hyperplasia in Prohormone Convertase 2 Knockout Mice
Authors: Webb Gene C. , Akbar Murtaza S. , Zhao Chongjian , Swift Hewson H. , Steiner Donald F.
Published: March 2007

Prohormone convertase 2 (PC2) plays an essential role in the processing of proglucagon to mature active glucagon in pancreatic α-cells (J Biol Chem 276:27197–27202, 2001). Mice lacking PC2 demonstrate multiple defects, including chronic mild hypoglycemia and dramatic hyperplasia of the pancreatic α-cells. To define the contribution of mature glucagon deficiency to the hypoglycemia and α-cell hyperplasia, we have attempted to correct the defects by delivery of exogenous glucagon by micro-osmotic pumps. Intraperitoneal delivery of 0.5 μg glucagon/h in PC2−/− mice resulted in the normalization of blood glucose concentrations. Islet remodeling through the loss of hyperplastic α-cells was evident by day 11 after pump implantation; by 25 days postimplantation, PC2−/− islets were indistinguishable from wild-type islets. These rapid changes were brought about by induction of apoptosis in the α-cell population. Morphological normalization of islets was also accompanied by marked downregulation of endogenous preproglucagon gene expression, but with little or no change in the level of preproinsulin gene expression. Exogenous glucagon delivery also normalized hepatic expression of the gluconeogenic enzyme PEPCK. These results demonstrate that the lack of mature glucagon in PC2−/− mice is responsible for the aberrant blood glucose levels, islet morphology, and gene expression, and they confirm the role of glucagon as a tonic insulin antagonist in regulating glycemia.
Increased Expression of Antioxidant and Antiapoptotic Genes in Islets That May Contribute to β-Cell Survival During Chronic Hyperglycemia
Authors: Laybutt D. Ross , Kaneto Hideaki , Hasenkamp Wendy , Grey Shane , Jonas Jean-Christophe , Sgroi Dennis C. , Groff Adam , Ferran Christiane , Bonner-Weir Susan , Sharma Arun , Weir Gordon C.
Published: March 2007

Hypertrophy is one mechanism of pancreatic β-cell growth and is seen as an important compensatory response to insulin resistance. We hypothesized that the induction of protective genes contributes to the survival of enlarged (hypertrophied) β-cells. Here, we evaluated changes in stress gene expression that accompany β-cell hypertrophy in islets from hyperglycemic rats 4 weeks after partial pancreatectomy (Px). A variety of protective genes were upregulated, with markedly increased expression of the antioxidant genes heme oxygenase-1 and glutathione peroxidase and the antiapoptotic gene A20. Cu/Zn-superoxide dismutase (SOD) and Mn-SOD were modestly induced, and Bcl-2 was modestly reduced; however, several other stress genes (catalase, heat shock protein 70, and p53) were unaltered. The increases in mRNA levels corresponded to the degree of hyperglycemia and were reversed in Px rats by 2-week treatment with phlorizin (treatment that normalized hyperglycemia), strongly suggesting the specificity of hyperglycemia in eliciting the response. Hyperglycemia in Px rats also led to activation of nuclear factor-κB in islets. The profound change in β-cell phenotype of hyperglycemic Px rats resulted in a reduced sensitivity to the β-cell toxin streptozotocin. Sensitivity to the toxin was restored, along with the β-cell phenotype, in islets from phlorizin-treated Px rats. Furthermore, β-cells of Px rats were not vulnerable to apoptosis when further challenged in vivo with dexamethasone, which increases insulin resistance. In conclusion, β-cell adaptation to chronic hyperglycemia and, hence, increased insulin demand is accompanied by the induction of protective stress genes that may contribute to the survival of hypertrophied β-cells.
Troglitazone but not Metformin Restores Insulin-Stimulated Phosphoinositide 3-Kinase Activity and Increases p110β Protein Levels in Skeletal Muscle of Type 2 Diabetic Subjects
Authors: Kim Young-Bum , Ciaraldi Theodore P. , Kong Alice , Kim Dennis , Chu Neelima , Mohideen Pharis , Mudaliar Sunder , Henry Robert R. , Kahn Barbara B.
Published: March 2007

Insulin stimulation of phosphatidylinositol (PI) 3-kinase activity is defective in skeletal muscle of type 2 diabetic individuals. We studied the impact of antidiabetic therapy on this defect in type 2 diabetic subjects who failed glyburide treatment by the addition of troglitazone (600 mg/day) or metformin (2,550 mg/day) therapy for 3–4 months. Improvement in glycemic control was similar for the two groups, as indicated by changes in fasting glucose and HbA1c levels. Insulin action on whole-body glucose disposal rate (GDR) was determined before and after treatment using the hyperinsulinemic (300 mU · m−2 · min−1) euglycemic (5.0–5.5 mmol/l) clamp technique. Needle biopsies of vastus lateralis muscle were obtained before and after each 3-h insulin infusion. Troglitazone treatment resulted in a 35 ± 9% improvement in GDR (P < 0.01), which was greater than (P < 0.05) the 22 ± 13% increase (P < 0.05) after metformin treatment. Neither treatment had any effect on basal insulin receptor substrate-1 (IRS-1)-associated PI 3-kinase activity in muscle. However, insulin stimulation of PI 3-kinase activity was augmented nearly threefold after troglitazone treatment (from 67 ± 22% stimulation over basal pre-treatment to 211 ± 62% post-treatment, P < 0.05), whereas metformin had no effect. The troglitazone effect on PI 3-kinase activity was associated with a 46 ± 22% increase (P < 0.05) in the amount of the p110β catalytic subunit of PI 3-kinase. Insulin-stimulated Akt activity also increased after troglitazone treatment (from 32 ± 8 to 107 ± 32% stimulation, P < 0.05) but was unchanged after metformin therapy. Protein expression of other key insulin signaling molecules (IRS-1, the p85 subunit of PI 3-kinase, and Akt) was unaltered after either treatment. We conclude that the mechanism for the insulin-sensitizing effect of troglitazone, but not metformin, involves enhanced PI 3-kinase pathway activation in skeletal muscle of obese type 2 diabetic subjects.
Normalization of Plasma Glucose Concentration by Insulin Therapy Improves Insulin-Stimulated Glycogen Synthesis in Type 2 Diabetes
Authors: Pratipanawatr Thongchai , Cusi Kenneth , Ngo Peter , Pratipanawatr Wilailak , Mandarino Lawrence J. , DeFronzo Ralph A.
Published: March 2007

Considerable evidence suggests that skeletal muscle insulin resistance is an inherent feature of type 2 diabetes and contributes to the pathogenesis of the disease. In patients with poorly controlled diabetes, hyperglycemia is thought to produce additional insulin resistance in muscle. The magnitude and nature of hyperglycemia-induced insulin resistance is not known. The purpose of the present study was to determine the biochemical mechanisms responsible for increased insulin-stimulated glucose disposal after the achievement of tight glycemic control with a mixed-split regimen. We performed hyperinsulinemic-euglycemic clamps with indirect calorimetry and vastus lateralis muscle biopsies in eight type 2 diabetic patients who had poor glycemic control (HbA1c 10.1%) and again after 3 months of intensive insulin therapy designed to produce near-normoglycemia (HbA1c 6.6%). Improved glycemic control increased insulin-stimulated glucose disposal (5.16 ± 0.32 vs. 3.69 ± 0.33 mg · kg−1 · min−1; P < 0.01); nonoxidative glucose disposal, which primarily reflects glycogen synthesis (2.11 ± 0.26 vs. 0.90 ± 0.16 mg · kg−1 · min−1; P < 0.01); and glycogen synthase fractional velocity (0.094 ± 0.017 vs. 0.045 ± 0.007; P < 0.05). There was no improvement in insulin-stimulated glucose oxidation (3.05 ± 0.25 vs. 2.79 ± 0.20 mg · kg−1 · min−1), hexokinase II mRNA expression (increase over basal values), or hexokinase II enzymatic activity (0.51 ± 0.16 vs. 0.42 ± 0.18 pmol · min−1 · μg−1 protein). All of the increase in insulin-stimulated glucose disposal could be accounted for by increased glycogen synthesis, which is likely attributable to increased activation of glycogen synthase by insulin.