bluuebunny/crossref_metadata_2025 huggingface dataset:
Stimulated Endocrine Cell Proliferation and Differentiation in Transplanted Human Pancreatic Islets
Authors: Tyrberg Björn , Ustinov Jarkko , Otonkoski Timo , Andersson Arne
Published: March 2007

Neogenesis is crucial for the maintenance of β-cell mass in the human pancreas and possibly for the outcome of clinical islet transplantation. To date, no studies have reported a stimulation of human β-cell neogenesis in vivo. Therefore, we investigated whether human α-, β-, and duct cell growth can be stimulated when human islets are xenotransplanted to obese hyperglycemic-hyperinsulinemic ob/ob mice immuno-suppressed with anti-lymphocyte serum. Moreover, we wanted to study whether β-cell growth and duct-to-β-cell differentiation were induced in the hepatocyte growth factor (HGF)-dependent compensatory kidney growth model. For that purpose, we evaluated human islets grafted to nude (nu/nu) mice before uninephrectomy of the contralateral kidney for DNA-synthesis and duct cell expression of the β-cell-specific transcription factor Nkx 6.1 as an estimate of differentiation. Human islet grafts were well preserved after 2 weeks when transplanted to ob/ob mice during anti-lymphocyte immunosuppression. Both human β-cells (P < 0.01) and duct cells (P < 0.001) were growth stimulated when islets were transplanted to ob/ob mice. We also observed a correlation between increased duct cell proliferation and increased organ donor age (P =0.02). Moreover, duct (P < 0.05) and β-cell (P <0.05) proliferation, as well as duct cell Nkx 6.1 expression (P <0.05), were enhanced by the compensatory kidney growth after uninephrectomy. We conclude that it is possible to stimulate human β-cell neogenesis in vivo, provided that the recipient carries certain growth-stimulatory traits. Furthermore, it seems that duct cell proliferation increases with increasing organ donor age. Altogether, these data and previous results from our laboratory suggest that human β-cell neogenesis becomes more dependent on differentiation and less dependent on proliferation with increasing age.
Defective Stimulus-Secretion Coupling in Islets ofPsammomys obesus, an Animal Model for Type 2 Diabetes
Authors: Nesher Rafael , Warwar Nasim , Khan Akhtar , Efendic Suad , Cerasi Erol , Kaiser Nurit
Published: March 2007

Psammomys obesus is a model of type 2 diabetes that displays resistance to insulin and deranged β-cell response to glucose. We examined the major signaling pathways for insulin release in P. obesus islets. Islets from hyperglycemic animals utilized twice as much glucose as islets from normoglycemic diabetes-prone or diabetes-resistant controls but exhibited similar rates of glucose oxidation. Fractional oxidation of glucose was constant in control islets over a range of concentrations, whereas islets from hyperglycemic P. obesus showed a decline at high glucose. The mitochondrial substrates α-ketoisocaproate and monomethyl succinate had no effect on insulin secretion in P. obesus islets. Basal insulin release in islets from diabetes-resistant P. obesus was unaffected by glucagon-like peptide 1 (GLP-1) or forskolin, whereas that of islets of the diabetic line was augmented by the drugs. GLP-1 and forskolin potentiated the insulin response to maximal (11.1 mmol/l) glucose in islets from all groups. The phorbol ester phorbol myristic acid (PMA) potentiated basal insulin release in islets from prediabetic animals, but not those from hyperglycemic or diabetes-resistant P. obesus. At the maximal stimulatory glucose concentration, PMA potentiated insulin response in islets from normoglycemic prediabetic and diabetes-resistant P. obesus but had no effect on islets from hyperglycemic P. obesus. Maintenance of islets from hyperglycemic P. obesus for 18 h in low (3.3 mmol/l) glucose in the presence of diazoxide (375 μmol/l) dramatically improved the insulin response to glucose and restored the responsiveness to PMA. Immunohistochemical analysis indicated that hyperglycemia was associated with reduced expression ofα-protein kinase C (PKC) and diminished translocation of λ-PKC. In summary, we found that 1) P. obesus islets have low oxidative capacity, probably resulting in limited ability to generate ATP to initiate and drive the insulin secretion; 2) insulin response potentiated by cyclic AMP—dependent protein kinase is intact in P. obesus islets, and increased sensitivity to GLP-1 or forskolin in the diabetic line may be secondary to increased sensitivity to glucose; and 3) islets of hyperglycemic P. obesus display reduced expression of α-PKC and diminished translocation of λ-PKC associated with impaired response to PMA. We conclude that low β-cell oxidative capacity coupled with impaired PKC-dependent signaling may contribute to the animals' poor adaptation to a high-energy diet.
Lipotoxicity of the Pancreatic β-Cell Is Associated With Glucose-Dependent Esterification of Fatty Acids Into Neutral Lipids
Authors: Briaud Isabelle , Harmon Jamie S. , Kelpe Cynthia L. , Segu Venkatesh Babu G. , Poitout Vincent
Published: March 2007

Prolonged exposure of isolated islets to supraphysiologic concentrations of palmitate decreases insulin gene expression in the presence of elevated glucose levels. This study was designed to determine whether or not this phenomenon is associated with a glucose-dependent increase in esterification of fatty acids into neutral lipids. Gene expression of sn-glycerol-3-phosphate acyltransferase (GPAT), diacylglycerol acyltransferase (DGAT), and hormone-sensitive lipase (HSL), three key enzymes of lipid metabolism, was detected in isolated rat islets. Their levels of expression were not affected after a 72-h exposure to elevated glucose and palmitate. To determine the effects of glucose on palmitate-induced neutral lipid synthesis, isolated rat islets were cultured for 72 h with trace amounts of [14C]palmitate with or without 0.5 mmol/l unlabeled palmitate,at 2.8 or 16.7 mmol/l glucose. Glucose increased incorporation of[14C]palmitate into complex lipids. Addition of exogenous palmitate directed lipid metabolism toward neutral lipid synthesis. As a result, neutral lipid mass was increased upon prolonged incubation with elevated palmitate only in the presence of high glucose. The ability of palmitate to increase neutral lipid synthesis in the presence of high glucose was concentration-dependent in HIT cells and was inversely correlated to insulin mRNA levels. 2-Bromopalmitate, an inhibitor of fatty acid mitochondrialβ-oxidation, reproduced the inhibitory effect of palmitate on insulin mRNA levels. In contrast, palmitate methyl ester, which is not metabolized,and the medium-chain fatty acid octanoate, which is readily oxidized, did not affect insulin gene expression, suggesting that fatty-acid inhibition of insulin gene expression requires activation of the esterification pathway. These results demonstrate that inhibition of insulin gene expression upon prolonged exposure of islets to palmitate is associated with a glucose-dependent increase in esterification of fatty acids into neutral lipids.
Effects of Short-Term Improvement of Insulin Treatment and Glycemia on Hepatic Glycogen Metabolism in Type 1 Diabetes
Authors: Bischof Martin G. , Krssak Martin , Krebs Michael , Bernroider Elisabeth , Stingl Harald , Waldhäusl Werner , Roden Michael
Published: March 2007

Insufficiently treated type 1 diabetic patients exhibit inappropriate postprandial hyperglycemia and reduction in liver glycogen stores. To examine the effect of acute improvement of metabolic control on hepatic glycogen metabolism, lean young type 1 diabetic (HbA1c 8.8 ± 0.3%)and matched nondiabetic subjects (HbA1c 5.4 ± 0.1%) were studied during the course of a day with three isocaloric mixed meals. Hepatic glycogen concentrations were determined noninvasively using in vivo 13C nuclear magnetic resonance spectroscopy. Rates of net glycogen synthesis and breakdown were calculated from linear regression of the glycogen concentration time curves from 7:30-10:30 P.M. and from 10:30 P.M. to 8:00 A.M., respectively. The mean plasma glucose concentration was ∼2.4-fold higher in diabetic than in nondiabetic subjects (13.6 ± 0.4 vs. 5.8± 0.1 mmol/l, P < 0.001). Rates of net glycogen synthesis and net glycogen breakdown were reduced by ∼74% (0.11 ± 0.02 vs. 0.43 ± 0.04 mmol/l liver/min, P < 0.001) and by ∼47%(0.10 ± 0.01 vs. 0.19 ± 0.01 mmol/l liver/min, P <0.001) in diabetic patients, respectively. During short-term (24-h)intensified insulin treatment, the mean plasma glucose level was not different between diabetic and nondiabetic subjects (6.4 ± 0.1 mmol/l). Net glycogen synthesis and breakdown increased by ∼92% (0.23 ± 0.04 mmol/l liver/min, P = 0.017) and by ∼40% (0.14 ± 0.01 mmol/l liver/min, P = 0.011), respectively. In conclusion, poorly controlled type 1 diabetic patients present with marked reduction in both hepatic glycogen synthesis and breakdown. Both defects in glycogen metabolism are improved but not normalized by short-term restoration of insulinemia and glycemia.
Hyperglycemia contributes to impaired insulin response in GK rat islets.
Authors: Ling Z C , Hong-Lie C , Ostenson C G , Efendic S , Khan A
Published: March 2007

Insulin secretion and glucose metabolism were compared in pancreatic islets from type 2 diabetic GK rats treated with phlorizin or vehicle. Treatment of control and GK rats with phlorizin for 30 days did not affect body weight, islet glucose utilization, or islet glucose oxidation. In phlorizin-treated GK rats, glucose-induced insulin release was about twofold higher at 11.0 and 16.7 mmol/l glucose compared with vehicle, treated GK rats, whereas phlorizin had no effect on control Wistar rats. However, also in phlorizin-treated GK rats, the amount of insulin released by the islets was significantly less than that from control rats (5.29+/-0.33 vs. 7.50+/-1.31 pmol x min(-1) islet(-1) at 16.7 mmol/l glucose; P<0.001). Islet glucose-6-phosphatase activity was significantly higher in GK rats than in control rats; phlorizin treatment significantly decreased this activity. These findings demonstrate that hyperglycemia per se constitutes an important factor for impaired insulin release in GK rats. Correction of hyperglycemia normalizes islet glucose-6-phosphatase activity, which may be an underlying factor for the partial improvement of glucose-induced insulin release.
Endocrine pancreas in insulin receptor-deficient mouse pups.
Authors: Jackerott M , Baudry A , Lamothe B , Bucchini D , Jami J , Joshi R L
Published: March 2007

Insulin receptor (IR)-deficient pups rapidly become hyperglycemic and hyperinsulinemic and die of diabetic ketoacidosis within a few days. Immunocytochemical analysis of the endocrine pancreas revealed that IR deficiency did not alter islet morphology or the number of beta-, alpha-, delta-, and pancreatic polypeptide (PP) cells. The lack of IR did not result in major changes in the expression of islet hormone genes or of beta-cell-specific marker genes encoding pancreas duodenum homeobox-containing transcription factor-1 (PDX-1), glucokinase (GCK), and GLUT2, as shown by reverse transcriptase-polymerase chain reaction analysis. The serum glucagon levels in IR-deficient and nondiabetic littermates were comparable. Finally, total insulin content in the pancreas of IR-deficient pups was gradually depleted, indicating sustained insulin secretion, not compensated for by increased insulin biosynthesis. These findings are discussed in light of recent results suggesting a role of IR in beta-cell function.
Decreased insulin secretion in type 2 diabetes: a problem of cellular mass or function?
Authors: Clark A , Jones L C , de Koning E , Hansen B C , Matthews D R
Published: March 2007

Type 2 diabetes is characterized by diminished or inappropriate secretion of insulin, which could be a defect of either islet cell function or beta-cell mass. Quantitation of islet cell populations in postmortem pancreas demonstrates little change of beta-cell mass in type 2 diabetes. Reduction of islet cell mass (up to 30%) is associated largely with islet amyloid deposition, and the degree of amyloidosis is independent of the duration of the disease. Insulin secretory capacity is dependent on both function and mass of cells. beta-Cell secretion is heterogeneous; increasing glucose concentrations result in recruitment of beta-cells into the secretory pool, indicating a large reserve of secretory capacity that can be recruited in insulin resistant conditions. The Starling curve of islet function describes the relationship of insulin secretion to increasing levels of insulin resistance and hyperglycemia in type 2 diabetes. Longitudinal studies in Macaca mulatta monkeys show that insulin resistance is accompanied by increased islet mass and onset of diabetes is associated with deposition of amyloid and reduction of beta-cells. Increasing the function of unresponsive beta-cells rather than the mass of cells may be a more effective therapeutic target for type 2 diabetes.
Regulation of beta-cell mass by hormones and growth factors.
Authors: Nielsen J H , Galsgaard E D , Møldrup A , Friedrichsen B N , Billestrup N , Hansen J A , Lee Y C , Carlsson C
Published: March 2007

Substantial new information has accumulated on molecular mechanisms of pancreas development, regulation of beta-cell gene expression, and the role of growth factors in the differentiation, growth, and regeneration of beta-cells. The present review focuses on some recent studies on the mechanism of action of cytokines such as growth hormone (GH) and prolactin (PRL) in beta-cell proliferation and gene expression-in particular, the role of signal transducers and activators of transcription (STAT) proteins. The implication of the discovery of suppressors of cytokine signaling (SOCS) proteins for the interaction between stimulatory and inhibitory cytokines, including GH, PRL, leptin, and the proinflammatory cytokines interleukin-1 and interferon-gamma, in beta-cell survival is not yet clear. Recent studies indicate a role of cell adhesion molecules and the delta-like protein preadipocyte factor 1/fetal antigen 1 (Pref-1/FA-1) in cytokine-induced beta-cell growth and development. Surprisingly, glucagon-like peptide-1 (GLP-1) was recently found to stimulate not only insulin secretion but also beta-cell replication and differentiation, which may present a new perspective in treatment of type 2 diabetes. Together with the intriguing reports on positive effects of insulin on both beta-cell growth and function, a picture is emerging of an integrated network of signaling events acting in concert to control beta-cell mass adaptation to insulin demand.
Regulatory elements involved in human pdx-1 gene expression.
Authors: Marshak S , Ben-Shushan E , Shoshkes M , Havin L , Cerasi E , Melloul D
Published: March 2007

PDX-1 was shown to be expressed early during development in cells of both exocrine and endocrine origin; later it becomes restricted primarily to beta-cells where it regulates the expression of beta-cell-specific genes and mediates the glucose effect on insulin gene transcription. Therefore, it was important to identify the molecular mechanisms that specifically govern the expression of pdx-1 in the mature beta-cell. To address this question, we analyzed 7 kb of the 5' flanking region of the human pdx-1 gene. By transient transfections of beta- and non-beta-cell lines with different 5' and 3' deletions of that region, a strong beta-cell-specific enhancer element located between -3.71 and 3.46 kb was revealed. We also sequenced about 4.5 kb of the human 5' flanking region and compared it with that of the mouse pdx-1 gene. This comparison revealed three short conserved regions, designated PH1, PH2, and PH3. We showed that HNF-3beta can bind and stimulate the activity of the human PH1 and PH2 elements in non-beta-cells. Results reported by Wu et al. (7) and Sharma et al. (6) also indicate that expression of the mouse pdx-1 is controlled by an HNF-3-like element. Thus, it can be stated that at least some aspects of pdx-1 expression rely on the transcription factor HNF-3beta. Because HNF-3beta is not restricted to beta-cells, the selective transcription of pdx-1 is likely to rely on additional factors. Our findings that the PH1 enhancer element binds both HNF-3beta and PDX-1 and that mutations in each individual site dramatically impair its transcriptional activity suggest that these factors cooperate with one another. We therefore propose that a possible feedback mechanism might control the expression of pdx-1 at different stages during development.
Imidazoline compounds protect against interleukin 1beta-induced beta-cell apoptosis.
Authors: Zaitsev S V , Appelskog I B , Kapelioukh I L , Yang S N , Köhler M , Efendic S , Berggren P O
Published: March 2007

Imidazoline compounds have been considered for the treatment of type 2 diabetes. We have now investigated the effects of imidazolines on interleukin (IL)-1beta-induced beta-cell apoptosis and the signal transduction pathways involved. Inhibition of Ca2+ influx into beta-cells by D-600, a blocker of voltage-gated L-type Ca2+ channels, suppressed IL-1beta-induced apoptosis. Our data show that calcineurin, Ca2+/calmodulin-dependent serine/threonine protein phosphatase 2B, is responsible for the effect of Ca2+ on beta-cell apoptosis. We also demonstrate that IL-1beta-mediated apoptosis correlates with expression of inducible nitric oxide synthase (iNOS) and the increase in intracellular production of nitric oxide. An inhibitor of cGMP-dependent protein kinase (PKG), KT5823, suppressed IL-1beta-induced apoptosis, suggesting the involvement of a PKG-dependent pathway in the apoptotic process. One of the major findings in this study is that imidazoline compounds RX871024 and efaroxan, suggested as prototypes of a new generation of drugs against type 2 diabetes, can protect against IL-1beta-induced apoptosis in pancreatic beta-cells, possibly by their inhibition of the expression of iNOS, a key element in the IL-1beta-induced apoptotic pathway in pancreatic beta-cells. These data suggest that imidazoline compounds should be explored as a potential therapeutic agent for the treatment of both type 1 and type 2 diabetes.
beta-cell genes and diabetes: molecular and clinical characterization of mutations in transcription factors.
Authors: Frayling T M , Evans J C , Bulman M P , Pearson E , Allen L , Owen K , Bingham C , Hannemann M , Shepherd M , Ellard S , Hattersley A T
Published: March 2007

beta-Cell transcription factor genes are important in the pathophysiology of the beta-cell, with mutations in hepatocyte nuclear factor (HNF)-1alpha, HNF-4alpha, insulin promoter factor (IPF)-1, HNF-1beta, and NeuroD1/BETA2, all resulting in early-onset type 2 diabetes. We assessed the relative contribution of these genes to early-onset type 2 diabetes using linkage and sequencing analysis in a cohort of 101 families (95% U.K. Caucasian). The relative distribution of the 90 families fitting maturity-onset diabetes of the young (MODY) criteria was 63% HNF-1alpha, 2% HNF-4alpha, 0% IPF-1, 1% HNF-1beta, 0% NeuroD1/ BETA2, and 20% glucokinase. We report the molecular genetic and clinical characteristics of these patients including 29 new families and 8 novel HNF-1alpha gene mutations. Mutations in the transactivation domain are more likely to be protein truncating rather than result in amino acid substitutions, suggesting that a relatively severe disruption of this domain is necessary to result in diabetes. Mutations in the different transcription factors result in clinical heterogeneity. IPF-1 mutations are associated with a higher age at diagnosis (42.7 years) than HNF-1alpha (20.4 years), HNF-1beta (24.2 years), or HNF-4alpha (26.3 years) gene mutations. Subjects with HNF-1beta mutations, in contrast to the other transcription factors, frequently present with renal disease. A comparison of age at diagnosis between subjects with different types and locations of HNF-1alpha mutations did not reveal genotype-phenotype correlations. In conclusion, mutations in transcription factors expressed in the beta-cell are the major cause of MODY, and the phenotype clearly varies with the gene that is mutated. There is little evidence to indicate that different mutations within the same gene have different phenotypes.
Distinction Between Interleukin-1–Induced Necrosis and Apoptosis of Islet Cells
Authors: Hoorens Anne , Stangé Geert , Pavlovic Dejan , Pipeleers Daniel
Published: March 2007

Interleukin (IL)-1β is known to cause β-cell death in isolated rat islets. This effect has been attributed to induction of nitric oxide (NO) synthase in β-cells and subsequent generation of toxic NO levels; it was not observed, however, in dispersed rat β-cells. The present study demonstrates that IL-1β induces NO-dependent necrosis in rat β-cells cultured for 3 days at high cell density or in cell aggregates but not as single cells. Its cytotoxic condition is not explained by higher NO production rates but might result from higher intercellular NO concentrations in statically cultured cell preparations with cell-to-cell contacts; nitrite levels in collected culture medium are not a reliable index for these intercellular concentrations. Absence of IL-1–induced necrosis in rat α-cells or in human β-cells is attributed to the cytokine’s failure to generate NO in these preparations, not to their reduced sensitivity to NO: the NO donor GEA 3162 (15 min, 50–100 μmol/l) exerts a comparable necrotic effect in rat and human α- or β-cells. In preparations in which IL-1β does not cause β-cell necrosis, its combination with γ-interferon (IFN-γ) results in NO-independent apoptosis, starting after 3 days and increasing with the duration of exposure. Because IFN-γ alone was apoptotic for rat α-cells, it is proposed that IL-1β can make β-cells susceptible to this effect, conceivably through altering their phenotype. It is concluded that IL-1β can cause NO-dependent necrosis or NO-independent apoptosis of islet cells, depending on the species and on the environmental conditions. The experiments in isolated human β-cell preparations suggest that these cells may preferentially undergo apoptosis when exposed to IL-1β plus IFN-γ unless neighboring non–β-cells produce toxic NO levels.
Cellular Mechanism of Nutritionally Induced Insulin Resistance in Psammomys Obesus
Authors: Ikeda Yukio , Olsen Grith S. , Ziv Ehud , Hansen Lone L. , Busch Anna K. , Hansen Bo F. , Shafrir Eleazar , Mosthaf-Seedorf Luitgard
Published: March 2007

The sand rat (Psammomys obesus) is an animal model of nutritionally induced diabetes. We report here that several protein kinase C (PKC) isoforms (α, ε, and ζ, representing all three subclasses of PKC) are overexpressed in the skeletal muscle of diabetic animals of this species. This is most prominent for the ε isotype of PKC. Interestingly, increased expression of PKCε could already be detected in normoinsulinemic, normoglycemic (prediabetic) animals of the diabetes-prone (DP) line when compared with a diabetes-resistant (DR) line. In addition, plasma membrane (PM)–associated fractions of PKCα and PKCε were significantly increased in skeletal muscle of diabetic animals, suggesting chronic activation of these PKC isotypes in the diabetic state. The increased PM association of these PKC isotypes revealed a significant correlation with the diacylglycerol content in the muscle samples. Altered expression/activity of PKCε, in particular, may thus contribute to the development of diabetes in these animals; along with other PKC isotypes, it may be involved in the progression of the disease. This may possibly occur through inhibition of insulin receptor (IR) tyrosine kinase activity mediated by serine/threonine phosphorylation of the IR or insulin receptor substrate 1 (IRS-1). However, overexpression of PKCε also mediated downregulation of IR numbers in a cell culture model (HEK293), resulting in attenuation of insulin downstream signaling (reduced protein kinase B [PKB]/Akt activity). In accordance with this, we detected decreased 125I-labeled insulin binding, probably reflecting a downregulation of IR numbers, in skeletal muscle of Psammomys animals from the DP line. The number of IRs was inversely correlated to both the expression and PM-associated levels of PKCε. These data suggest that overexpression of PKCε may be causally related to the development of insulin resistance in these animals, possibly by increasing the degradation of IRs.
Autonomic Neuropathy in Nondiabetic Offspring of Type 2 Diabetic Subjects Is Associated With Urinary Albumin Excretion Rate and 24-h Ambulatory Blood Pressure
Authors: Hauerslev Foss C. , Vestbo E. , Frøland A. , Gjessing H.J. , Mogensen C.E. , Damsgaard E.M.
Published: March 2007

The aim of this study was to examine the impact of parental type 2 diabetes on the autonomic nervous system and to determine whether autonomic neuropathy is present and associated with changes in 24-h ambulatory blood pressure (AMBP) and urinary albumin excretion rate (UAER) in nondiabetic subjects with parental type 2 diabetes. We examined 223 nondiabetic offspring of type 2 diabetic subjects and a control group of 258 offspring of nondiabetic subjects. The autonomic nervous system was assessed by three cardiovascular reflex tests, 24-h AMBP was measured with an oscillometric recorder (90207; Spacelabs, Redmond, WA), and UAER was determined through three overnight urine samples. The subjects with parental type 2 diabetes had significantly lower heart rate variation in all three bedside tests (P < 0.01) than subjects without parental diabetes. The prevalence of autonomic neuropathy in the nondiabetic offspring with parental type 2 diabetes (6.7%) was significantly (P < 0.01) higher compared with the control group (1.6%). Autonomic neuropathy was associated with a higher fasting insulin level (P < 0.05), higher UAER (P < 0.001), higher 24-h mean AMBP (P < 0.01), and reduced diurnal blood pressure variation (P < 0.001) after adjustment for age, sex, and BMI. In conclusion, parental type 2 diabetes was found to be associated with alterations in the autonomic nervous system in nondiabetic subjects. The presence of autonomic neuropathy in subjects with parental type 2 diabetes was associated with higher UAER, fasting insulin level, and 24-h AMBP and a reduced diurnal blood pressure variation. This study indicates that parental type 2 diabetes has an impact on the cardiac autonomic function in nondiabetic subjects.
A Genome Scan for Type 2 Diabetes Susceptibility Loci in a Genetically Isolated Population
Authors: Permutt Marshall A. , Wasson Jonathon C. , Suarez Brian K. , Lin Jennifer , Thomas Jeffrey , Meyer Joanne , Lewitzky Steve , Rennich Jean S. , Parker Alex , DuPrat Laura , Maruti Sanchit , Chayen Susan , Glaser Benjamin
Published: March 2007

A total of 896 individuals of Ashkenazi Jewish descent were ascertained in Israel from 267 multiplex families, including 472 sib-pairs affected with type 2 diabetes. A genome-wide scan with average marker spacing of 9.5 cM revealed five regions on four chromosomes (4q, 8q, 14q, and 20q) that exhibited nominal evidence for linkage (P < 0.05). The highest observed nonparametric linkage Z score was 2.41 (equivalent to a logarithm of odds score of 1.26) at marker D4S1501. A maximal signal, with a Z score of 2.05, was observed on chromosome 20 near marker D20S195, and another on 20p near marker D20S103 (Z 1.80). A single marker on chromosome 8 (D8S593) and two adjacent markers on chromosome 14 (D14S749 and D14S605) also attained evidence of linkage. To explore the hypothesis that the signals on chromosomes 4 and 20 are differentially attributable to variation in BMI or age of onset, an ordered subset analysis was conducted. This analysis revealed that only when the families were ranked by BMI (in increasing order) did a subset attain nominal significance, and only for chromosome 4. The findings reported here lend credence to the hypothesis, now supported by four studies of Caucasian populations and most recently by a combined analysis of 1,852 pedigrees, that a type 2 diabetes susceptibility locus resides on chromosome 20q. This population, because of its unique genetic attributes, may facilitate identification of this and other genes contributing to type 2 diabetes.
Decreased Fasting and Oral Glucose Stimulated C-peptide in Nondiabetic Subjects With Sequence Variants in the Sulfonylurea Receptor 1 Gene
Authors: Weisnagel S. John , Rankinen Tuomo , Nadeau André , Rao D.C. , Chagnon Yvon C. , Pérusse Louis , Bouchard Claude
Published: March 2007

The high-affinity sulfonylurea receptor 1 (SUR1) plays an important role in regulating insulin secretion. In the Québec Family Study, we genotyped 731 individuals (685 nondiabetic [ND] subjects) for the SUR1 gene IVS15-3c→t and exon 18 Thr759(ACC→ACT) polymorphisms using polymerase chain reaction–restriction fragment-length polymorphism analysis. Phenotypes measured were fasting plasma glucose (GLU), fasting plasma insulin (INS), and fasting C-peptide (CPEP), as well as oral glucose tolerance test (OGTT) responses; they were adjusted for age, sex, waist circumference, and the sum of six skinfold thicknesses. In ND subjects, exon 18 Thr759(ACC→ACT) T allele carriers (T+) had lower CPEP (P = 0.022, −12.8%) and acute C-peptide responses (area above basal in first 30 min [CP30]) (P = 0.051, −12.4%) than noncarriers (T–). Also, in those with the cT/tC haplotype (from both IVS15-3c→t and exon 18 Thr759[ACC→ACT] polymorphisms), CPEP (P = 0.005, −21.2%), CP30 (P = 0.034, −19.2%), and total C-peptide responses (P = 0.016, −20.2%) were lower than that in cT– subjects. In overweight individuals (BMI >25 kg/m2), differences between carriers and noncarriers of the T or cT alleles were greater for GLU (P = 0.023–0.034), CPEP (P = 0.021–0.015), acute OGTT insulin response (P = 0.014–0.019), and CP30 (P = 0.034–0.019). These results suggest that the T and cT allele variants are associated with lower insulin secretion parameters, particularly in female and overweight subjects, adding evidence to the role of SUR1 sequence variants in decreased insulin secretion.
Circulating Ghrelin Levels Are Decreased in Human Obesity
Authors: Tschöp Matthias , Weyer Christian , Tataranni P. Antonio , Devanarayan Viswanath , Ravussin Eric , Heiman Mark L.
Published: March 2007

Ghrelin is a novel endogenous natural ligand for the growth hormone (GH) secretagogue receptor that has recently been isolated from the rat stomach. Ghrelin administration stimulates GH secretion but also causes weight gain by increasing food intake and reducing fat utilization in rodents. To investigate the possible involvement of ghrelin in the pathogenesis of human obesity, we measured body composition (by dual X-ray absorption) as well as fasting plasma ghrelin concentrations (radioimmunoassay) in 15 Caucasians (8 men and 7 women, 31 ± 9 years of age, 92 ± 24 kg body wt, and 29±10% body fat, mean ± SD) and 15 Pima Indians (8 men and 7 women, 33 ± 5 years of age, 97 ± 29 kg body wt, and 30 ± 8% body fat). Fasting plasma ghrelin was negatively correlated with percent body fat (r = –0.45; P = 0.01), fasting insulin (r = – 0.45; P = 0.01) and leptin (r = –0.38; P = 0.03) concentrations. Plasma ghrelin concentration was decreased in obese Caucasians as compared with lean Caucasians (P < 0.01). Also, fasting plasma ghrelin was lower in Pima Indians, a population with a very high prevalence of obesity, compared with Caucasians (87 ± 28 vs. 129 ± 34 fmol/ml; P < 0.01). This result did not change after adjustment for fasting plasma insulin concentration. There was no correlation between fasting plasma ghrelin and height. Prospective clinical studies are now needed to establish the role of ghrelin in the pathogenesis of human obesity.
Effects of Free Fatty Acids on Gluconeogenesis and Autoregulation of Glucose Production in Type 2 Diabetes
Authors: Boden Guenther , Chen Xinhua , Capulong Edwin , Mozzoli Maria
Published: March 2007

Effects of endogenously derived free fatty acids (FFAs) on rates of gluconeogenesis (GNG) (determined with 2H2O), glycogenolysis (GL), and endogenous glucose production (EGP) were studied in 18 type 2 diabetic patients and in 7 nondiabetic control subjects under three experimental conditions: 1) during an 8-h fast (from 16–24 h after the last meal), when plasma FFA levels increased slowly; 2) during 4 h (from 16–20 h) of nicotinic acid (NA) administration (fasting plus NA), when plasma FFAs decreased acutely; and 3) during 4 h (from 20–24 h) after discontinuation of NA (FFA rebound), when plasma FFAs increased acutely. During fasting, FFAs increased from 636 to 711 μmol/l in type 2 diabetic patients and from 462 to 573 μmol/l in control subjects (P < 0.04), but GNG did not change in diabetic patients (6.9 vs. 6.5 μmol · kg–1 · min–1, P > 0.05) or in control subjects (5.1 vs. 5.4 μmol · kg–1 · min–1, P > 0.05). During fasting plus NA, FFAs decreased in diabetic patients and control subjects (from 593 to 193 and from 460 to 162 μmol/l, respectively); GNG decreased (from 6.1 to 4.2 and from 4.7 to 3.5 μmol · kg–1 · min−1), whereas GL decreased in diabetic patients (from 5.3 to 4.4 μmol · kg−1 · min−1) but increased in control subjects (from 5.4 to 7.2 μmol · kg−1 min−1). During the FFA rebound, FFAs increased in diabetic patients and control subjects (from 193 to 1,239 and from 162 to 1,491 μmol/l, respectively); GNG increased (from 4.2 to 5.4 and from 3.4 to 5.3 μmol · kg–1 · min–1 respectively), and GL decreased (from 4.4 to 3.4 and from 7.3 to 4.3 μmol · kg−1 · min−1, respectively). In summary, during an extended overnight fast, increasing plasma FFA levels stimulated GNG, whereas decreasing FFA levels inhibited GNG in both diabetic and control subjects; 20 h after the last meal, approximately one-third of GNG in both diabetic and control subjects was dependent on FFAs; and autoregulation of EGP by GL in response to decreasing GNG was impaired in diabetic patients.
Pro12Ala Polymorphism in the Peroxisome Proliferator–Activated Receptor-γ2 Gene Is Associated With Increased Antilipolytic Insulin Sensitivity
Authors: Stumvoll Michael , Wahl Hans Günther , Löblein Klaus , Becker Regine , Machicao Fausto , Jacob Stephan , Häring Hans
Published: March 2007

The Pro12Ala polymorphism of the peroxisome proliferator–activated receptor (PPAR)-γ2 is associated with reduced transcriptional activity in vitro and increased insulin sensitivity in humans in vivo. The mechanism by which this polymorphism influences insulin sensitivity in humans is unclear. PPAR-γ2 is mainly expressed in adipocytes, and free fatty acids released from adipose tissue are key mediators of peripheral insulin resistance. Therefore, we examined insulin suppression of lipolysis in 51 subjects without (Pro/Pro) and 17 subjects with the polymorphism (X/Ala). Both groups were lean (BMI <27.0 kg/m2) and matched for age, BMI, waist-to-hip ratio, and sex. The isotopically (infusion of d5 glycerol) determined glycerol rate of appearance was used as an index of lipolysis. Insulin sensitivity of lipolysis was expressed as the insulin concentration resulting in half-maximal suppression (EC50). This was directly determined during a three-step hyperinsulinemic-euglycemic clamp (n = 21) or estimated indirectly during a standard hyperinsulinemic-euglycemic clamp (n = 47). The insulin sensitivity index (ISI) of glucose disposal was 0.095 ± 0.006 μmol · kg−1 · min−1 · pmol−1 · l–1 in the control group and 0.129 ± 0.008 μmol · kg−1 · min−1 · pmol−1 · l–1 in the X/Ala group (P = 0.003). The EC50 was 56 ± 2 pmol/l in the control group and 44 ± 3 pmol/l in the X/Ala group (P = 0.001). The EC50 of lipolysis and ISI was significantly correlated (r = 0.42, P = 0.002). In conclusion, in lean subjects, the Pro12Ala polymorphism is associated with increased insulin sensitivity of glucose disposal and suppression of lipolysis. This result suggests that an altered transcriptional activity of PPAR-γ2 in X/Ala subjects either causes a more efficient suppression of lipolysis in adipose tissue, which in turn results in improved insulin-stimulated glucose disposal in muscle, or, alternatively, beneficially affects insulin signaling in both tissues independently of one another.
The Gly972Arg Polymorphism in the Insulin Receptor Substrate-1 Gene Contributes to the Variation in insulin Secretion in Normal Glucose-Tolerant Humans
Authors: Stumvoll Michael , Fritsche Andreas , Volk Annette , Stefan Norbert , Madaus Alexander , Maerker Elke , Teigeler Anna , Koch Matthias , Machicao Fausto , Häring Hans
Published: March 2007

The Gly972Arg polymorphism in the insulin receptor substrate (IRS)-1 was found in some studies to have a higher prevalence in type 2 diabetic subjects than in control subjects. Previously, transfection of IRS-1 with this polymorphism into insulin-secreting cells resulted in a marked reduction of glucose-stimulated insulin secretion compared with the wild-type transfected cells. In the present study, we compared insulin secretion in well-matched normal glucose-tolerant subjects with and without this polymorphism. Several validated indexes of β-cell function from the oral glucose tolerance test were significantly lower in X/Arg (n = 31) compared with Gly/Gly (n = 181) (P between 0.002 and 0.05), whereas insulin sensitivity (measured with a euglycemic clamp) was not different. During a modified hyperglycemic clamp, insulin secretion rates were significantly lower in Gly/Arg (n = 8) compared with Gly/Gly (n = 36) during the first phase (1,711 ± 142 vs. 3,014 ± 328 pmol/min, P = 0.05) and after maximal stimulation with arginine (5,340 ± 639 vs. 9,075 ± 722 pmol/min, P = 0.03). In summary, our results suggest that the Gly972Arg polymorphism in IRS-1 is associated with decreased insulin secretion in response to glucose but not with insulin sensitivity. It is possible that this polymorphism causes insulin resistance at the level of the β-cell and contributes to the polygenic etiology of type 2 diabetes.
Tissue Inhibitor of Metalloproteinase-1 Prevents Cytokine-Mediated Dysfunction and Cytotoxicity in Pancreatic Islets and β-cells
Authors: Han Xiao , Sun Yujie , Scott Stephen , Bleich David
Published: March 2007

In addition to inhibiting matrix metalloproteinase-2 and matrix metalloproteinase-9 activity, recent studies suggest that tissue inhibitor of metalloproteinase (TIMP)-1 may inhibit apoptosis in various cell lines. To address this question in pancreatic islets and β-cells, we treated rat pancreatic islets and INS-1 cells with a high-dose combination of the cytokines interleukin (IL)-1β, tumor necrosis factor-α, and interferon-γ with or without the addition of TIMP-1 and TIMP-2 protein. Using flow cytometry, we quantitated DNA fragmentation to assess cellular apoptosis and confirmed these observations with DNA laddering experiments. Next, we transfected the mouse TIMP-1 gene into INS-1 cells and performed Western immunoblotting to demonstrate expression of TIMP-1 protein. We treated TIMP-1–expressing INS-1 cells with high-dose cytokines and again used flow cytometry to assess DNA fragmentation. We also evaluated the effect of TIMP-1 on IL-1β–induced inhibition of glucose-stimulated insulin secretion (GSIS) in freshly isolated rat pancreatic islets. Finally, we evaluated the effect of TIMP-1 on inducible nitric oxide synthase (iNOS) gene expression and nuclear factor (NF)-κB activity in INS-1 cells stimulated with high-dose cytokines. TIMP-1 but not TIMP-2 prevented cytokine-induced apoptosis and cytokine-mediated inhibition of GSIS in rat islets and β-cells. TIMP-1 mediated these effects by inhibiting cytokine activation of NF-κB, but it did not affect nitric oxide production or iNOS gene expression. Therefore, TIMP-1 may be an ideal gene to prevent cytokine-mediated β-cell destruction and dysfunction in models of type 1 diabetes and islet transplantation rejection.
Pioglitazone Ameliorates Tumor Necrosis Factor-α–Induced Insulin Resistance by a Mechanism Independent of Adipogenic Activity of Peroxisome Proliferator–Activated Receptor-γ
Authors: Iwata Minoru , Haruta Tetsuro , Usui Isao , Takata Yasumitsu , Takano Atsuko , Uno Tatsuhito , Kawahara Junko , Ueno Eiichi , Sasaoka Toshiyasu , Ishibashi Osamu , Kobayashi Masashi
Published: March 2007

Tumor necrosis factor (TNF)-α is one of the candidate mediators of insulin resistance associated with obesity, a major risk factor for the development of type 2 diabetes. The insulin resistance induced by TNF-α is antagonized by thiazolidinediones (TZDs), a new class of insulin-sensitizing drugs. The aim of the current study was to dissect the mechanism whereby pioglitazone, one of the TZDs, ameliorates TNF-α–induced insulin resistance in 3T3-L1 adipocytes. Pioglitazone restored insulin-stimulated 2-deoxyglucose (DOG) uptake, which was reduced by TNF-α, with concomitant restorations in tyrosine phosphorylation and protein levels of insulin receptor (IR) and insulin receptor substrate (IRS)-1, as well as association of the p85 regulatory subunit of phosphatidylinositol (PI) 3-kinase with IRS-1 and PI 3-kinase activity. Adenovirus-mediated gene transfer of either wild-type human peroxisome proliferator–activated receptor (PPAR)-γ2 or a mutant carrying a replacement at the consensus mitogen-activated protein kinase phosphorylation site (hPPAR-γ2-S112A) promoted adipogenesis of 3T3-L1 fibroblasts and restored TNF-α–induced decrease of triglyceride in adipocytes as effectively as pioglitazone. Overexpression of the PPAR-γ proteins in TNF-α–treated adipocytes restored protein levels of IR/IRS-1, but did not improve insulin-stimulated tyrosine phosphorylation of IR/IRS-1 or insulin-stimulated 2-DOG uptake. These results indicate that the ability of pioglitazone to restore insulin-stimulated tyrosine phosphorylation of IR/IRS-1, which is necessary for amelioration of TNF-α–induced insulin resistance, may be independent of the adipogenic activity of PPAR-γ that regulates protein levels of IR/IRS-1.
Troglitazone Induces GLUT4 Translocation in L6 Myotubes
Authors: Yonemitsu Shin , Nishimura Haruo , Shintani Mitsuyo , Inoue Ryou , Yamamoto Yuji , Masuzaki Hiroaki , Ogawa Yoshihiro , Hosoda Kiminori , Inoue Gen , Hayashi Tatsuya , Nakao Kazuwa
Published: March 2007

A number of studies have demonstrated that insulin resistance in the skeletal muscle plays a pivotal role in the insulin resistance associated with obesity and type 2 diabetes. A decrease in GLUT4 translocation from the intracellular pool to the plasma membranes in skeletal muscles has been implicated as a possible cause of insulin resistance. Herein, we examined the effects of an insulin-sensitizing drug, troglitazone (TGZ), on glucose uptake and the translocation of GLUT4 in L6 myotubes. The prolonged exposure (24 h) of L6 myotubes to TGZ (10−5 mol/l) caused a substantial increase in the 2-deoxy-[3H]d-glucose (2-DG) uptake without changing the total amount of the glucose transporters GLUT4, GLUT1, and GLUT3. The TGZ-induced 2-DG uptake was completely abolished by cytochalasin-B (10 μmol/l). The ability of TGZ to translocate GLUT4 from light microsomes to the crude plasma membranes was greater than that of insulin. Both cycloheximide treatment (3.5 × 10−6 mol/l) and the removal of TGZ by washing reversed the 2-DG uptake to the basal level. Moreover, insulin did not enhance the TGZ-induced 2-DG uptake additively. The TGZ-induced 2-DG uptake was only partially reversed by wortmannin to 80%, and TGZ did not change the expression and the phosphorylation of protein kinase B; the expression of protein kinase C (PKC)-λ, PKC-β2, and PKC-ζ; or 5′AMP-activated protein kinase activity. α-Tocopherol, which has a molecular structure similar to that of TGZ, did not increase 2-DG uptake. We conclude that the glucose transport in L6 myotubes exposed to TGZ for 24 h is the result of an increased translocation of GLUT4. The present results imply that the effects of troglitazone on GLUT4 translocation may include a new mechanism for improving glucose transport in skeletal muscle.
Liver-Specific igf-1 Gene Deletion Leads to Muscle Insulin Insensitivity
Authors: Yakar Shoshana , Liu Jun-Li , Fernandez Ana M. , Wu Yiping , Schally Andrew V. , Frystyk Jan , Chernausek Steve D. , Mejia Wilson , Le Roith Derek
Published: March 2007

Insulin and insulin-like growth factors (IGFs) mediate a variety of signals involved in mammalian development and metabolism. To study the metabolic consequences of IGF-I deficiency, we used the liver IGF-I–deficient (LID) mouse model. The LID mice show a marked reduction (∼75%) in circulating IGF-I and elevated growth hormone (GH) levels. Interestingly, LID mice show a fourfold increase in serum insulin levels (2.2 vs. 0.6 ng/ml in control mice) and abnormal glucose clearance after insulin injection. Fasting blood glucose levels and those after a glucose tolerance test were similar between the LID mice and their control littermates. Thus, the high levels of circulating insulin enable the LID mice to maintain normoglycemia in the presence of apparent insulin insensitivity. Insulin-induced autophosphorylation of the insulin receptor and tyrosine phosphorylation of insulin receptor substrate (IRS)-1 were absent in muscle, but were normal in liver and white adipose tissue of the LID mice. In contrast, IGF-I–induced autophosphorylation of its cognate receptor and phosphorylation of IRS-1 were normal in muscle of LID mice. Thus, the insulin insensitivity seen in the LID mice is muscle specific. Recombinant human IGF-I treatment of the LID mice caused a reduction in insulin levels and an increase in insulin sensitivity. Treatment of the LID mice with GH-releasing hormone antagonist, which reduces GH levels, also increased insulin sensitivity. These data provide evidence of the role of circulating IGF-I as an important component of overall insulin action in peripheral tissues.
Upregulation of Glucose Metabolism During Intimal Lesion Formation Is Coupled to the Inhibition of Vascular Smooth Muscle Cell Apoptosis
Authors: Hall Jennifer L. , Chatham John C. , Eldar-Finkelman Hagit , Gibbons Gary H.
Published: March 2007

The purpose of this study was to define the role of metabolic regulatory genes in the pathogenesis of vascular lesions. The glucose transporter isoform, GLUT1, was significantly increased in the neointima after balloon injury. To define the role of GLUT1 in vascular biology, we established cultured vascular smooth muscle cells (VSMCs) with constitutive upregulation of GLUT1, which led to a threefold increase in glucose uptake as well as significant increases in both nonoxidative and oxidative glucose metabolism as assessed by 13C–nuclear magnetic resonance spectroscopy. We hypothesized that the differential enhancement of glucose metabolism in the neointima contributed to formation of lesions by increasing the resistance of VSMCs to apoptosis. Indeed, upregulation of GLUT1 significantly inhibited apoptosis induced by serum withdrawal (control 20 ± 1% vs. GLUT1 11 ± 1%, P < 0.0005) as well as Fas-ligand (control 12 ± 1% vs. GLUT1 6 ± 1.0%, P < 0.0005). Provocatively, the enhanced glucose metabolism in GLUT1 overexpressing VSMC as well as neointimal tissue correlated with the inactivation of the proapoptotic kinase, glycogen synthase kinase 3β (GSK3β). Transient overexpression of GSK3β was sufficient to induce apoptosis (control 7 ± 1% vs. GSK3β 28 ± 2%, P < 0.0001). GSK3β-induced apoptosis was significantly attenuated by GLUT1 overexpression (GSK3β 29 ± 3% vs. GLUT1 + GSK3β 6 ± 1%, n = 12, P < 0.001), suggesting that the antiapoptotic effect of enhanced glucose metabolism is linked to the inactivation of GSK3β. Taken together, upregulation of glucose metabolism during intimal lesion formation promotes an antiapoptotic signaling pathway that is linked to the inactivation of GSK3β.