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Sample records for insulin responses cell

  1. Heterogeneous response of isolated adult rat heart cells to insulin

    International Nuclear Information System (INIS)

    Haworth, R.A.; Hunter, D.R.; Berkoff, H.A.

    1984-01-01

    3-O-Methylglucose uptake by Ca2+-resistant adult rat heart cells in suspension was measured, free of artifactual inhibitor-insensitive uptake, and with an accuracy of +/- 1.9% pellet water. (Ca2+-resistant cells are cells which retain their original rod-shaped morphology in the presence of physiological levels of Ca2+.) High levels of insulin (10(-6) M) stimulated the rate of 3-O-methylglucose uptake approximately 10-fold. In the presence of low levels of insulin (3 X 10(-11) M, 10(-10) M) uptake was biphasic; it could not be described by a single exponential function within experimental error, but required the sum of two exponentials. Deviation from a single exponential function was not so great with high levels of insulin (10(-6) M) or no insulin. Cell sugar uptake was also investigated using autoradiography of cells which had accumulated [2-14C]deoxyglucose under similar conditions. This showed considerable heterogeneity of 2-deoxyglucose uptake by cells treated with low levels of insulin, but significantly less heterogeneity of 2-deoxyglucose uptake by cells treated with high levels of insulin. It is concluded that the deviation of 3-O-methylglucose uptake from a single exponential observed at low insulin levels can be accounted for in terms of a heterogeneous response of cells to insulin

  2. Variations in insulin responsiveness in rat fat cells are due to metabolic differences rather than insulin binding

    DEFF Research Database (Denmark)

    Hansen, Finn Mølgård; Nilsson, Poul; Sonne, Ole

    1983-01-01

    -insulin to fat cells. Insulin binding was not correlated to the plasma insulin level which however was reflected in the lipoprotein lipase activity in the adipose tissue. In conclusion, these results indicate that variations in insulin responsiveness in fat cells are due to alterations in cellular metabolism...

  3. Measuring phospholipase D activity in insulin-secreting pancreatic beta-cells and insulin-responsive muscle cells and adipocytes.

    Science.gov (United States)

    Cazzolli, Rosanna; Huang, Ping; Teng, Shuzhi; Hughes, William E

    2009-01-01

    Phospholipase D (PLD) is an enzyme producing phosphatidic acid and choline through hydrolysis of phosphatidylcholine. The enzyme has been identified as a member of a variety of signal transduction cascades and as a key regulator of numerous intracellular vesicle trafficking processes. A role for PLD in regulating glucose homeostasis is emerging as the enzyme has recently been identified in events regulating exocytosis of insulin from pancreatic beta-cells and also in insulin-stimulated glucose uptake through controlling GLUT4 vesicle exocytosis in muscle and adipose tissue. We present methodologies for assessing cellular PLD activity in secretagogue-stimulated insulin-secreting pancreatic beta-cells and also insulin-stimulated adipocyte and muscle cells, two of the principal insulin-responsive cell types controlling blood glucose levels.

  4. MCF-7 human mammary adenocarcinoma cells exhibit augmented responses to human insulin on a collagen IV surface

    DEFF Research Database (Denmark)

    Listov-Saabye, Nicolai; Jensen, Marianne Blirup; Kiehr, Benedicte

    2009-01-01

    Human mammary cell lines are extensively used for preclinical safety assessment of insulin analogs. However, it is essentially unknown how mitogenic responses can be optimized in mammary cell-based systems. We developed an insulin mitogenicity assay in MCF-7 human mammary adenocarcinoma cells......, under low serum (0.1% FCS) and phenol red-free conditions, with 3H thymidine incorporation as endpoint. Based on EC50 values determined from 10-fold dilution series, beta-estradiol was the most potent mitogen, followed by human IGF-1, human AspB10 insulin and native human insulin. AspB10 insulin...... was significantly more mitogenic than native insulin, validating the ability of the assay to identify hypermitogenic human insulin analogs. With MCF-7 cells on a collagen IV surface, the ranking of mitogens was maintained, but fold mitogenic responses and dynamic range and steepness of dose-response curves were...

  5. Human adipose cells in vitro are either refractory or responsive to insulin, reflecting host metabolic state.

    Directory of Open Access Journals (Sweden)

    Vladimir A Lizunov

    Full Text Available While intercellular communication processes are frequently characterized by switch-like transitions, the endocrine system, including the adipose tissue response to insulin, has been characterized by graded responses. Yet here individual cells from adipose tissue biopsies are best described by a switch-like transition between the basal and insulin-stimulated states for the trafficking of the glucose transporter GLUT4. Two statistically-defined populations best describe the observed cellular heterogeneity, representing the fractions of refractive and responsive adipose cells. Furthermore, subjects exhibiting high systemic insulin sensitivity indices (SI have high fractions of responsive adipose cells in vitro, while subjects exhibiting decreasing SI have increasing fractions of refractory cells in vitro. Thus, a two-component model best describes the relationship between cellular refractory fraction and subject SI. Since isolated cells exhibit these different response characteristics in the presence of constant culture conditions and milieu, we suggest that a physiological switching mechanism at the adipose cellular level ultimately drives systemic SI.

  6. Insulin-induced translocation of IR to the nucleus in insulin responsive cells requires a nuclear translocation sequence.

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    Kesten, Dov; Horovitz-Fried, Miriam; Brutman-Barazani, Tamar; Sampson, Sanford R

    2018-04-01

    Insulin binding to its cell surface receptor (IR) activates a cascade of events leading to its biological effects. The Insulin-IR complex is rapidly internalized and then is either recycled back to the plasma membrane or sent to lysosomes for degradation. Although most of the receptor is recycled or degraded, a small amount may escape this pathway and migrate to the nucleus of the cell where it might be important in promulgation of receptor signals. In this study we explored the mechanism by which insulin induces IR translocation to the cell nucleus. Experiments were performed cultured L6 myoblasts, AML liver cells and 3T3-L1 adipocytes. Insulin treatment induced a rapid increase in nuclear IR protein levels within 2 to 5 min. Treatment with WGA, an inhibitor of nuclear import, reduced insulin-induced increases nuclear IR protein; IR was, however, translocated to a perinuclear location. Bioinformatics tools predicted a potential nuclear localization sequence (NLS) on IR. Immunofluorescence staining showed that a point mutation on the predicted NLS blocked insulin-induced IR nuclear translocation. In addition, blockade of nuclear IR activation in isolated nuclei by an IR blocking antibody abrogated insulin-induced increases in IR tyrosine phosphorylation and nuclear PKCδ levels. Furthermore, over expression of mutated IR reduced insulin-induced glucose uptake and PKB phosphorylation. When added to isolated nuclei, insulin induced IR phosphorylation but had no effect on nuclear IR protein levels. These results raise questions regarding the possible role of nuclear IR in IR signaling and insulin resistance. Copyright © 2018 Elsevier B.V. All rights reserved.

  7. Subcellular localization, mobility, and kinetic activity of glucokinase in glucose-responsive insulin-secreting cells.

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    Stubbs, M; Aiston, S; Agius, L

    2000-12-01

    We investigated the subcellular localization, mobility, and activity of glucokinase in MIN6 cells, a glucose-responsive insulin-secreting beta-cell line. Glucokinase is present in the cytoplasm and a vesicular/granule compartment that is partially colocalized with insulin granules. The granular staining of glucokinase is preserved after permeabilization of the cells with digitonin. There was no evidence for changes in distribution of glucokinase between the cytoplasm and the granule compartment during incubation of the cells with glucose. The rate of release of glucokinase and of phosphoglucoisomerase from digitonin-permeabilized cells was slower when cells were incubated at an elevated glucose concentration (S0.5 approximately 15 mmol/l). This effect of glucose was counteracted by competitive inhibitors of glucokinase (5-thioglucose and mannoheptulose) but was unaffected by fructose analogs and may be due to changes in cell shape or conformation of the cytoskeleton that are secondary to glucose metabolism. Based on the similar release of glucokinase and phosphoglucoisomerase, we found no evidence for specific binding of cytoplasmic digitonin-extractable glucokinase. The affinity of beta-cells for glucose is slightly lower than that in cell extracts and, unlike that in hepatocytes, is unaffected by fructose, tagatose, or a high-K+ medium, which is consistent with the lack of change in glucokinase distribution or release. We conclude that glucokinase is present in two locations, cytoplasm and the granular compartment, and that it does not translocate between them. This conclusion is consistent with the lack of adaptive changes in the glucose phosphorylation affinity. The glucokinase activity associated with the insulin granules may have a role in either direct or indirect coupling between glucose phosphorylation and insulin secretion.

  8. Lipid content and response to insulin are not invariably linked in human muscle cells

    OpenAIRE

    Aguer , Céline; Mercier , Jacques; Kitzmann , Magali

    2009-01-01

    Abstract In type 2 diabetes, a strong correlation between intramyocellular lipid accumulation and insulin resistance exists but whether intramyocellular accumulation is a cause or a consequence of insulin resistance is not clear. Lipid accumulation and response to insulin were evaluated in primary human myotubes derived from non-diabetic subjects and type 2 diabetic patients. Myotubes derived from type 2 diabetic patients had a defective response to insulin without showing a signif...

  9. Generation of glucose-responsive, insulin-producing cells from human umbilical cord blood-derived mesenchymal stem cells.

    Science.gov (United States)

    Prabakar, Kamalaveni R; Domínguez-Bendala, Juan; Molano, R Damaris; Pileggi, Antonello; Villate, Susana; Ricordi, Camillo; Inverardi, Luca

    2012-01-01

    We sought to assess the potential of human cord blood-derived mesenchymal stem cells (CB-MSCs) to derive insulin-producing, glucose-responsive cells. We show here that differentiation protocols based on stepwise culture conditions initially described for human embryonic stem cells (hESCs) lead to differentiation of cord blood-derived precursors towards a pancreatic endocrine phenotype, as assessed by marker expression and in vitro glucose-regulated insulin secretion. Transplantation of these cells in immune-deficient animals shows human C-peptide production in response to a glucose challenge. These data suggest that human cord blood may be a promising source for regenerative medicine approaches for the treatment of diabetes mellitus.

  10. Activated α2-macroglobulin binding to human prostate cancer cells triggers insulin-like responses.

    Science.gov (United States)

    Misra, Uma Kant; Pizzo, Salvatore Vincent

    2015-04-10

    Ligation of cell surface GRP78 by activated α2-macroglobulin (α2M*) promotes cell proliferation and suppresses apoptosis. α2M*-treated human prostate cancer cells exhibit a 2-3-fold increase in glucose uptake and lactate secretion, an effect similar to insulin treatment. In both α2M* and insulin-treated cells, the mRNA levels of SREBP1-c, SREBP2, fatty-acid synthase, acetyl-CoA carboxylase, ATP citrate lyase, and Glut-1 were significantly increased together with their protein levels, except for SREBP2. Pretreatment of cells with α2M* antagonist antibody directed against the carboxyl-terminal domain of GRP78 blocks these α2M*-mediated effects, and silencing GRP78 expression by RNAi inhibits up-regulation of ATP citrate lyase and fatty-acid synthase. α2M* induces a 2-3-fold increase in lipogenesis as determined by 6-[(14)C]glucose or 1-[(14)C]acetate incorporation into free cholesterol, cholesterol esters, triglycerides, free fatty acids, and phosphatidylcholine, which is blocked by inhibitors of fatty-acid synthase, PI 3-kinase, mTORC, or an antibody against the carboxyl-terminal domain of GRP78. We also assessed the incorporation of [(14)CH3]choline into phosphatidylcholine and observed similar effects. Lipogenesis is significantly affected by pretreatment of prostate cancer cells with fatostatin A, which blocks sterol regulatory element-binding protein proteolytic cleavage and activation. This study demonstrates that α2M* functions as a growth factor, leading to proliferation of prostate cancer cells by promoting insulin-like responses. An antibody against the carboxyl-terminal domain of GRP78 may have important applications in prostate cancer therapy. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  11. The beta-cell response to glucagon and mixed meal stimulation in non-insulin dependent diabetes

    DEFF Research Database (Denmark)

    Gjessing, H J; Damsgaard, E M; Matzen, L E

    1988-01-01

    The aim of this study was to evaluate the correlations of the C-peptide and insulin responses after stimulation with glucagon intravenously as well as the 24-h urinary excretion of C-peptide to the C-peptide response to a standard mixed meal in 30 patients with non-insulin dependent diabetes...... plasma C-peptide (r = 0.55, p less than 0.01). The C-peptide and insulin responses after meal stimulation correlated modestly inversely with HbA1. In conclusion, measurement of C-peptide in fasting state, as well as measurements of C-peptide and insulin after glucagon stimulation, only modestly predict...... the C-peptide response to physiologic stimulation in NIDDM. Twenty-four-hour urinary C-peptide excretion does not predict this response. Patients with NIDDM seem to show a better metabolic control if they have a more pronounced beta-cell response to physiologic stimulation....

  12. Deletion of glutamate dehydrogenase in beta-cells abolishes part of the insulin secretory response not required for glucose homeostasis

    DEFF Research Database (Denmark)

    Carobbio, Stefania; Frigerio, Francesca; Rubi, Blanca

    2009-01-01

    Insulin exocytosis is regulated in pancreatic ss-cells by a cascade of intracellular signals translating glucose levels into corresponding secretory responses. The mitochondrial enzyme glutamate dehydrogenase (GDH) is regarded as a major player in this process, although its abrogation has not been...... tested yet in animal models. Here, we generated transgenic mice, named betaGlud1(-/-), with ss-cell-specific GDH deletion. Our results show that GDH plays an essential role in the full development of the insulin secretory response. In situ pancreatic perfusion revealed that glucose-stimulated insulin...... secretion was reduced by 37% in betaGlud1(-/-). Furthermore, isolated islets with either constitutive or acute adenovirus-mediated knock-out of GDH showed a 49 and 38% reduction in glucose-induced insulin release, respectively. Adenovirus-mediated re-expression of GDH in betaGlud1(-/-) islets fully restored...

  13. Insulin-induced enhancement of MCF-7 breast cancer cell response to 5-fluorouracil and cyclophosphamide.

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    Agrawal, Siddarth; Łuc, Mateusz; Ziółkowski, Piotr; Agrawal, Anil Kumar; Pielka, Ewa; Walaszek, Kinga; Zduniak, Krzysztof; Woźniak, Marta

    2017-06-01

    The study was designed to evaluate the potential use of insulin for cancer-specific treatment. Insulin-induced sensitivity of MCF-7 breast cancer cells to chemotherapeutic agents 5-fluorouracil and cyclophosphamide was evaluated. To investigate and establish the possible mechanisms of this phenomenon, we assessed cell proliferation, induction of apoptosis, activation of apoptotic and autophagic pathways, expression of glucose transporters 1 and 3, formation of reactive oxygen species, and wound-healing assay. Additionally, we reviewed the literature regarding theuse of insulin in cancer-specific treatment. We found that insulin increases the cytotoxic effect of 5-fluorouracil and cyclophosphamide in vitro up to two-fold. The effect was linked to enhancement of apoptosis, activation of apoptotic and autophagic pathways, and overexpression of glucose transporters 1 and 3 as well as inhibition of cell proliferation and motility. We propose a model for insulin-induced sensitization process. Insulin acts as a sensitizer of cancer cells to cytotoxic therapy through various mechanisms opening a possibility for metronomic insulin-based treatments.

  14. Prolonged Growth Hormone/Insulin/Insulin-like Growth Factor Nutrient Response Signaling Pathway as a Silent Killer of Stem Cells and a Culprit in Aging.

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    Ratajczak, Mariusz Z; Bartke, Andrzej; Darzynkiewicz, Zbigniew

    2017-08-01

    The dream of slowing down the aging process has always inspired mankind. Since stem cells are responsible for tissue and organ rejuvenation, it is logical that we should search for encoded mechanisms affecting life span in these cells. However, in adult life the hierarchy within the stem cell compartment is still not very well defined, and evidence has accumulated that adult tissues contain rare stem cells that possess a broad trans-germ layer differentiation potential. These most-primitive stem cells-those endowed with pluripotent or multipotent differentiation ability and that give rise to other cells more restricted in differentiation, known as tissue-committed stem cells (TCSCs) - are of particular interest. In this review we present the concept supported by accumulating evidence that a population of so-called very small embryonic-like stem cells (VSELs) residing in adult tissues positively impacts the overall survival of mammals, including humans. These unique cells are prevented in vertebrates from premature depletion by decreased sensitivity to growth hormone (GH), insulin (INS), and insulin-like growth factor (IGF) signaling, due to epigenetic changes in paternally imprinted genes that regulate their resistance to these factors. In this context, we can envision nutrient response GH/INS/IGF signaling pathway as a lethal factor for these most primitive stem cells and an important culprit in aging.

  15. Pancreatic Endoderm-Derived From Diabetic Patient-Specific Induced Pluripotent Stem Cell Generates Glucose-Responsive Insulin-Secreting Cells.

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    Rajaei, Bahareh; Shamsara, Mehdi; Amirabad, Leila Mohammadi; Massumi, Mohammad; Sanati, Mohammad Hossein

    2017-10-01

    Human-induced pluripotent stem cells (hiPSCs) can potentially serve as an invaluable source for cell replacement therapy and allow the creation of patient- and disease-specific stem cells without the controversial use of embryos and avoids any immunological incompatibility. The generation of insulin-producing pancreatic β-cells from pluripotent stem cells in vitro provides an unprecedented cell source for personal drug discovery and cell transplantation therapy in diabetes. A new five-step protocol was introduced in this study, effectively induced hiPSCs to differentiate into glucose-responsive insulin-producing cells. This process mimics in vivo pancreatic organogenesis by directing cells through stages resembling definitive endoderm, primitive gut-tube endoderm, posterior foregut, pancreatic endoderm, and endocrine precursor. Each stage of differentiation were characterized by stage-specific markers. The produced cells exhibited many properties of functional β-cells, including expression of critical β-cells transcription factors, the potency to secrete C-peptide in response to high levels of glucose and the presence of mature endocrine secretory granules. This high efficient differentiation protocol, established in this study, yielded 79.18% insulin-secreting cells which were responsive to glucose five times higher than the basal level. These hiPSCs-derived glucose-responsive insulin-secreting cells might provide a promising approach for the treatment of type I diabetes mellitus. J. Cell. Physiol. 232: 2616-2625, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  16. Insulin receptor internalization defect in an insulin-resistant mouse melanoma cell line

    International Nuclear Information System (INIS)

    Androlewicz, M.J.; Straus, D.S.; Brandenburg, D.F.

    1989-01-01

    Previous studies from this laboratory demonstrated that the PG19 mouse melanoma cell line does not exhibit a biological response to insulin, whereas melanoma x mouse embryo fibroblast hybrids do respond to insulin. To investigate the molecular basis of the insulin resistance of the PG19 melanoma cells, insulin receptors from the insulin-resistant melanoma cells and insulin-sensitive fibroblast x melanoma hybrid cells were analyzed by the technique of photoaffinity labeling using the photoprobe 125 I-NAPA-DP-insulin. Photolabeled insulin receptors from the two cell types have identical molecular weights as determined by SDS gel electrophoresis under reducing and nonreducing conditions, indicating that the receptors on the two cell lines are structurally similar. Insulin receptor internalization studies revealed that the hybrid cells internalize receptors to a high degree at 37 degree C, whereas the melanoma cells internalize receptors to a very low degree or not at all. The correlation between ability to internalize insulin receptors and sensitivity to insulin action in this system suggests that uptake of the insulin-receptor complex may be required for insulin action in these cells. Insulin receptors from the two cell lines autophosphorylate in a similar insulin-dependent manner both in vitro and in intact cells, indicating that insulin receptors on the melanoma and hybrid cells have functional tyrosine protein kinase activity. Therefore, the block in insulin action in the PG19 melanoma cells appears to reside at a step beyond insulin-stimulated receptor autophosphorylation

  17. Insulin and insulin-like growth factor receptors and responses

    International Nuclear Information System (INIS)

    Roth, R.A.; Steele-Perkins, G.; Hari, J.; Stover, C.; Pierce, S.; Turner, J.; Edman, J.C.; Rutter, W.J.

    1988-01-01

    Insulin is a member of a family of structurally related hormones with diverse physiological functions. In humans, the best-characterized members of this family include insulin, insulin-like growth factor (IGF)-I, and IGF-II. Each of these three polypeptide hormones has its own distinct receptor. The structures of each of these receptors have now been deduced from analyses of isolated cDNA clones. To study further the responses mediated through these three different receptors, the authors have been studying cells expressing the proteins encoded by these three cDNAs. The isolated cDNAs have been transfected into Chinese hamster ovary (CHO) cells, and the resulting transfected cell lines have been characterized as to the ligand-binding activities and signal-transducing activities of the expressed proteins

  18. Relationship Between β-cell Response and Insulin Sensitivity in Horses based on the Oral Sugar Test and the Euglycemic Hyperinsulinemic Clamp.

    Science.gov (United States)

    Lindåse, S; Nostell, K; Söder, J; Bröjer, J

    2017-09-01

    A hyperbolic relationship between β-cell response and insulin sensitivity (IS) has been described in several species including rodents, dogs, and humans. This relationship has not been elucidated in the horse. To determine whether the hyperbolic relationship between β-cell response and IS exists in horses by using indices of β-cell response from the oral sugar test (OST) and IS measurements from the euglycemic hyperinsulinemic clamp (EHC). A second aim was to compare how well IS estimates from the OST and EHC correlate. Forty-nine horses with different degrees of insulin regulation (normal-to-severe insulin dysregulation). Cross-sectional study. Horses were examined with an OST and an EHC. Decreased IS was associated with increased β-cell response in the horses. Nine of 12 comparisons between indices of β-cell response and IS measures fulfilled the criteria for a hyperbolic relationship. Indices of IS calculated from the OST correlated highly with the insulin-dependent glucose disposal rate (M) and the insulin-dependent glucose disposal rate per unit of insulin (M/I) determined from the EHC (r = 0.81-0.87). A hyperbolic relationship between β-cell response and IS exists in horses, which suggest that horses with insulin dysregulation respond not only with postprandial hyperinsulinemia but are also insulin resistant. The OST is primarily a test for β-cell response rather than a test for IS, but calculated indices of IS from the OST may be useful to estimate IS in horses, especially when the horse is insulin resistant. Copyright © 2017 The Authors. Journal of Veterinary Internal Medicine published by Wiley Periodicals, Inc. on behalf of the American College of Veterinary Internal Medicine.

  19. Do we really need to differentiate mesenchymal stem cells into insulin-producing cells for attenuation of the autoimmune responses in type 1 diabetes: immunoprophylactic effects of precursors to insulin-producing cells.

    Science.gov (United States)

    Sharma, Anshu; Rani, Rajni

    2017-07-12

    Type 1 diabetes (T1D) is a multifactorial autoimmune disorder where pancreatic beta cells are lost before the clinical manifestations of the disease. Administration of mesenchymal stem cells (MSCs) or MSCs differentiated into insulin-producing cells (IPCs) have yielded limited success when used therapeutically. We have evaluated the immunoprophylactic potentials of precursors to insulin-producing cells (pIPCs) and IPCs in nonobese diabetic (NOD) mice to ask a basic question: do we need to differentiate MSCs into IPCs or will pIPCs suffice to attenuate autoimmune responses in T1D? Bone marrow-derived MSCs from Balb/c mice were characterized following the International Society for Cellular Therapy (ISCT) guidelines. MSCs cultured in high-glucose media for 11 to 13 passages were characterized for the expression of pancreatic lineage genes using real-time polymerase chain reaction. Expression of the PDX1 gene in pIPCs was assessed using Western blot and fluorescence-activated cell sorting (FACS). Triple-positive MSCs were differentiated into IPCs using a three-step protocol after sorting them for cell surface markers, i.e. CD29, CD44, and SCA-1. Nonobese diabetic mice were administered pIPCs, IPCs, or phosphate-buffered saline (PBS) into the tail vein at weeks 9 or 10 and followed-up for 29-30 weeks for fasting blood glucose levels. Two consecutive blood sugar levels of more than 250 mg/dl were considered diabetic. MSCs grown in high-glucose media for 11 to 13 passages expressed genes of the pancreatic lineage such as PDX1, beta2, neurogenin, PAX4, Insulin, and glucagon. Furthermore, Western blot and FACS analysis for PDX-1, a transcription factor necessary for beta cell maturation, confirmed that these cells were precursors of insulin-producing cells (pIPCs). NOD mice administered with pIPCs were better protected from developing diabetes with a protective efficacy of 78.4% (p cells seem to have better potential to arrest autoimmune response in type 1 diabetes when

  20. GLUT3 is present in Clone 9 liver cells and translocates to the plasma membrane in response to insulin.

    Science.gov (United States)

    Defries, Danielle M; Taylor, Carla G; Zahradka, Peter

    2016-08-26

    Clone 9 cells have been reported to express only the GLUT1 facilitative glucose transporter; however, previous studies have not examined Clone 9 cells for GLUT3 content. The current study sought to profile the presence of glucose transporters in Clone 9 cells, H4IIE hepatoma cells, and L6 myoblasts and myotubes. While the other cell types contained the expected complement of transporters, Clone 9 cells had GLUT3 which was previously not reported. Interestingly, both GLUT3 mRNA and protein were detected in Clone 9 cells, but only mRNA for GLUT1 was detected. Glucose transport in Clone 9 cells was insulin-sensitive in a concentration-dependent manner, concomitant with the presence of GLUT3 in the plasma membrane after insulin treatment. Although basal glucose uptake was unaffected, insulin-stimulated glucose uptake was abolished with siRNA-mediated GLUT3 knockdown. These results contradict previous reports that Clone 9 cells exclusively express GLUT1 and suggest GLUT3 is a key insulin-sensitive glucose transporter required for insulin-stimulated glucose uptake by Clone 9 cells. Copyright © 2016 Elsevier Inc. All rights reserved.

  1. Effect of metformin combined with clomiphene on insulin resistance, oxidative stress response and T cell immune response in patients with PCOS

    Directory of Open Access Journals (Sweden)

    Xiao-Lian Zhang

    2017-08-01

    Full Text Available Objective: To study the effect of metformin combined with clomiphene on insulin resistance, oxidative stress response and T cell immune response in patients with polycystic ovarian syndrome (PCOS. Methods: A total of 94 patients who were diagnosed with PCOS in Jingzhou Second People’s Hospital between September 2014 and October 2016 were selected and randomly divided into the combined group who received the metformin combined with clomiphene therapy and the control group who received clomiphene therapy. The insulin resistance, oxidative stress response and T cell immune response were evaluated before treatment and 3 menstrual cycles after treatment. Results: 3 menstrual cycles after treatment, HOMA-IR level, serum F-Ins, F-CP, TOS, MDA, AOPP and IL-17 contents as well as peripheral blood RORγt mRNA expression of combined group were significantly lower than those before treatment while HOMA-β level, serum TAS, SOD, GSH-Px, VitC, VitE, IL-10 and TGF-β1 contents as well as peripheral blood Foxp3 mRNA expression were significantly higher than those before treatment; HOMA-IR and HOMA-β levels, serum F-Ins, F-CP, TOS, MDA, AOPP, IL-17, TAS, SOD, GSH-Px, VitC, VitE, IL-10 and TGF-β1 contents as well as peripheral blood Foxp3 and RORγt mRNA expression of control group were not different from those before treatment. Conclusion: Metformin combined with clomiphene can significantly improve the insulin resistance, oxidative stress response and T cell immune response in patients with PCOS.

  2. Hormonal, metabolic and cardiovascular responses to hypoglycaemia in Type 1 (insulin-dependent) diabetes with and without residual B cell function

    DEFF Research Database (Denmark)

    Madsbad, S; Hilsted, J; Krarup, T

    1982-01-01

    Hormonal, metabolic and cardiovascular responses to insulin induced hypoglycaemia were investigated in seven Type 1 (insulin-dependent) diabetic patients with residual B cell function, eight Type 1 diabetic patients without B cell function and six healthy subjects. No differences were found between...... the diabetic groups regarding nadir of glucose and rate of recovery to normoglycaemia. The patients with residual B cell function had a glucagon response to hypoglycaemia which was close to that of normal subjects. In patients without B cell function, the glucagon response to hypoglycaemia was present, albeit...... significantly smaller than in the patients with preserved B cell function (0.025 ng/ml, range 0.007-0.042 versus 0.054 ng/ml, range 0.029-0.087). The group without B cell function had signs of an exaggerated rate of lipolysis and ketogenesis compared with the patients with B cell function and the normal...

  3. Intracellular insulin-receptor dissociation and segregation in a rat fibroblast cell line transfected with a human insulin receptor gene

    International Nuclear Information System (INIS)

    Levy, J.R.; Olefsky, J.M.

    1988-01-01

    The cellular processing of insulin and insulin receptors was studied using a rat fibroblast cell line that had been transfected with a normal human insulin receptor gene, expressing approximately 500 times the normal number of native fibroblasts insulin receptors. These cells bind and internalize insulin normally. Biochemically assays based on the selective precipitation by polyethylene glycol of intact insulin-receptor complexes but not of free intracellular insulin were developed to study the time course of intracellular insulin-receptor dissociation. Fibroblasts were incubated with radiolabeled insulin at 4 0 C, and internalization of insulin-receptor complexes was initiated by warming the cells to 37 0 C. Within 2 min, 90% of the internalized radioactivity was composed of intact insulin-receptor complexes. The dissociation of insulin from internalized insulin-receptor complexes was markedly inhibited by monensin and chloroquine. Furthermore, chloroquine markedly increased the number of cross-linkable intracellular insulin-receptor complexes, as analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis autoradiography. These findings suggest that acidification of intracellular vesicles is responsible for insulin-receptor dissociation. Physical segregation of dissociated intracellular insulin from its receptor was monitored. The results are consistent with the view that segregation of insulin and receptor occurs 5-10 min after initiation of dissociation. These studies demonstrate the intracellular itinerary of insulin-receptor complexes, including internalization, dissociation of insulin from the internalized receptor within an acidified compartment, segregation of insulin from the receptor, and subsequent ligand degradation

  4. Development of the insulin secretion mechanism in fetal and neonatal rat pancreatic B-cells: response to glucose, K+, theophylline, and carbamylcholine

    Directory of Open Access Journals (Sweden)

    A.C. Mendonça

    1998-06-01

    Full Text Available We studied the development of the insulin secretion mechanism in the pancreas of fetal (19- and 21-day-old, neonatal (3-day-old, and adult (90-day-old rats in response to stimulation with 8.3 or 16.7 mM glucose, 30 mM K+, 5 mM theophylline (Theo and 200 µM carbamylcholine (Cch. No effect of glucose or high K+ was observed on the pancreas from 19-day-old fetuses, whereas Theo and Cch significantly increased insulin secretion at this age (82 and 127% above basal levels, respectively. High K+ also failed to alter the insulin secretion in the pancreas from 21-day-old fetuses, whereas 8.3 mM and 16.7 mM glucose significantly stimulated insulin release by 41 and 54% above basal levels, respectively. Similar results were obtained with Theo and Cch. A more marked effect of glucose on insulin secretion was observed in the pancreas of 3-day-old rats, reaching 84 and 179% above basal levels with 8.3 mM and 16.7 mM glucose, respectively. At this age, both Theo and Cch increased insulin secretion to close to two-times basal levels. In islets from adult rats, 8.3 mM and 16.7 mM glucose, Theo, and Cch increased the insulin release by 104, 193, 318 and 396% above basal levels, respectively. These data indicate that pancreatic B-cells from 19-day-old fetuses were already sensitive to stimuli that use either cAMP or IP3 and DAG as second messengers, but insensitive to stimuli such as glucose and high K+ that induce membrane depolarization. The greater effect of glucose on insulin secretion during the neonatal period indicates that this period is crucial for the maturation of the glucose-sensing mechanism in B-cells.

  5. Introduction of exogenous growth hormone receptors augments growth hormone-responsive insulin biosynthesis in rat insulinoma cells

    DEFF Research Database (Denmark)

    Billestrup, N; Møldrup, A; Serup, P

    1990-01-01

    The stimulation of insulin biosynthesis in the pancreatic insulinoma cell line RIN5-AH by growth hormone (GH) is initiated by GH binding to specific receptors. To determine whether the recently cloned rat hepatic GH receptor is able to mediate the insulinotropic effect of GH, we have transfected ...

  6. Chronic Insulin Exposure Induces ER Stress and Lipid Body Accumulation in Mast Cells at the Expense of Their Secretory Degranulation Response.

    Directory of Open Access Journals (Sweden)

    William E Greineisen

    Full Text Available Lipid bodies (LB are reservoirs of precursors to inflammatory lipid mediators in immunocytes, including mast cells. LB numbers are dynamic, increasing dramatically under conditions of immunological challenge. We have previously shown in vitro that insulin-influenced lipogenic pathways induce LB biogenesis in mast cells, with their numbers attaining steatosis-like levels. Here, we demonstrate that in vivo hyperinsulinemia resulting from high fat diet is associated with LB accumulation in murine mast cells and basophils. We characterize the lipidome of purified insulin-induced LB, and the shifts in the whole cell lipid landscape in LB that are associated with their accumulation, in both model (RBL2H3 and primary mast cells. Lipidomic analysis suggests a gain of function associated with LB accumulation, in terms of elevated levels of eicosanoid precursors that translate to enhanced antigen-induced LTC4 release. Loss-of-function in terms of a suppressed degranulation response was also associated with LB accumulation, as were ER reprogramming and ER stress, analogous to observations in the obese hepatocyte and adipocyte. Taken together, these data suggest that chronic insulin elevation drives mast cell LB enrichment in vitro and in vivo, with associated effects on the cellular lipidome, ER status and pro-inflammatory responses.

  7. Development of glucose-responsive 'smart' insulin systems.

    Science.gov (United States)

    Rege, Nischay K; Phillips, Nelson F B; Weiss, Michael A

    2017-08-01

    The complexity of modern insulin-based therapy for type I and type II diabetes mellitus and the risks associated with excursions in blood-glucose concentration (hyperglycemia and hypoglycemia) have motivated the development of 'smart insulin' technologies (glucose-responsive insulin, GRI). Such analogs or delivery systems are entities that provide insulin activity proportional to the glycemic state of the patient without external monitoring by the patient or healthcare provider. The present review describes the relevant historical background to modern GRI technologies and highlights three distinct approaches: coupling of continuous glucose monitoring (CGM) to deliver devices (algorithm-based 'closed-loop' systems), glucose-responsive polymer encapsulation of insulin, and molecular modification of insulin itself. Recent advances in GRI research utilizing each of the three approaches are illustrated; these include newly developed algorithms for CGM-based insulin delivery systems, glucose-sensitive modifications of existing clinical analogs, newly developed hypoxia-sensitive polymer matrices, and polymer-encapsulated, stem-cell-derived pancreatic β cells. Although GRI technologies have yet to be perfected, the recent advances across several scientific disciplines that are described in this review have provided a path towards their clinical implementation.

  8. Induction of insulin secretion in engineered liver cells by nitric oxide

    Directory of Open Access Journals (Sweden)

    Özcan Sabire

    2007-10-01

    Full Text Available Abstract Background Type 1 Diabetes Mellitus results from an autoimmune destruction of the pancreatic beta cells, which produce insulin. The lack of insulin leads to chronic hyperglycemia and secondary complications, such as cardiovascular disease. The currently approved clinical treatments for diabetes mellitus often fail to achieve sustained and optimal glycemic control. Therefore, there is a great interest in the development of surrogate beta cells as a treatment for type 1 diabetes. Normally, pancreatic beta cells produce and secrete insulin only in response to increased blood glucose levels. However in many cases, insulin secretion from non-beta cells engineered to produce insulin occurs in a glucose-independent manner. In the present study we engineered liver cells to produce and secrete insulin and insulin secretion can be stimulated via the nitric oxide pathway. Results Expression of either human insulin or the beta cell specific transcription factors PDX-1, NeuroD1 and MafA in the Hepa1-6 cell line or primary liver cells via adenoviral gene transfer, results in production and secretion of insulin. Although, the secretion of insulin is not significantly increased in response to high glucose, treatment of these engineered liver cells with L-arginine stimulates insulin secretion up to three-fold. This L-arginine-mediated insulin release is dependent on the production of nitric oxide. Conclusion Liver cells can be engineered to produce insulin and insulin secretion can be induced by treatment with L-arginine via the production of nitric oxide.

  9. SAD-A potentiates glucose-stimulated insulin secretion as a mediator of glucagon-like peptide 1 response in pancreatic β cells.

    Science.gov (United States)

    Nie, Jia; Lilley, Brendan N; Pan, Y Albert; Faruque, Omar; Liu, Xiaolei; Zhang, Weiping; Sanes, Joshua R; Han, Xiao; Shi, Yuguang

    2013-07-01

    Type 2 diabetes is characterized by defective glucose-stimulated insulin secretion (GSIS) from pancreatic β cells, which can be restored by glucagon-like peptide 1 (GLP-1), an incretin hormone commonly used for the treatment of type 2 diabetes. However, molecular mechanisms by which GLP-1 affects glucose responsiveness in islet β cells remain poorly understood. Here we investigated a role of SAD-A, an AMP-activated protein kinase (AMPK)-related kinase, in regulating GSIS in mice with conditional SAD-A deletion. We show that selective deletion of SAD-A in pancreas impaired incretin's effect on GSIS, leading to glucose intolerance. Conversely, overexpression of SAD-A significantly enhanced GSIS and further potentiated GLP-1's effect on GSIS from isolated mouse islets. In support of SAD-A as a mediator of incretin response, SAD-A is expressed exclusively in pancreas and brain, the primary targeting tissues of GLP-1 action. Additionally, SAD-A kinase is activated in response to stimulation by GLP-1 through cyclic AMP (cAMP)/Ca(2+)-dependent signaling pathways in islet β cells. Furthermore, we identified Thr443 as a key autoinhibitory phosphorylation site which mediates SAD-A's effect on incretin response in islet β cells. Consequently, ablation of Thr443 significantly enhanced GLP-1's effect on GSIS from isolated mouse islets. Together, these findings identified SAD-A kinase as a pancreas-specific mediator of incretin response in islet β cells.

  10. Differential responsiveness of luteinized human granulosa cells to gonadotropins and insulin-like growth factor I for induction of aromatase activity

    International Nuclear Information System (INIS)

    Christman, G.M.; Randolph, J.F. Jr.; Peegel, H.; Menon, K.M.

    1991-01-01

    The objective of this study was to examine the in vitro responsiveness of cultured luteinized human granulosa cells over time to insulin-like growth factor 1 (IGF-1), human follicle-stimulating hormone (FSH), and human chorionic gonadotropin (hCG) for the induction of aromatase activity. Granulosa cells were retrieved from preovulatory follicles in patients undergoing in vitro fertilization. Cells were cultured for a period of 72 hours or 10 days. The ability of hCG, human FSH, and/or IGF-I to induce aromatase activity was assayed by the stereospecific release of tritium from [1B-3H]androstenedione. Short-term cultures (72 hours) demonstrated a marked rise in aromatase activity in response to human FSH and IGF-I, whereas a smaller response to hCG was observed. In contrast, 10-day cultures demonstrated responsiveness predominantly to hCG rather than human FSH for the induction of aromatase activity with no remarkable effect of IGF-I. Luteinized human granulosa cells undergo a transformation from an initial human FSH and IGF-I responsive state to an hCG responsive state in long-term cultures

  11. Insulin signaling in Caenorhabditis elegans regulates both endocrine-like and cell-autonomous outputs.

    Science.gov (United States)

    Iser, Wendy B; Gami, Minaxi S; Wolkow, Catherine A

    2007-03-15

    In C. elegans, insulin signaling affects development, lifespan and stress resistance. Several studies have shown that insulin signaling affects lifespan in an endocrine-like manner from different cells, while the major downstream target of insulin, the FOXO transcription factor encoded by daf-16, may act preferentially in intestinal cells to prolong lifespan. This discrepancy raised the possibility that insulin may have both endocrine and cell-intrinsic outputs. Here, we further investigated the types of cells capable of producing endocrine outputs of insulin and also identified a new cell-intrinsic insulin output. We found that insulin signaling within groups of neurons promoted wildtype lifespan, showing that the endocrine outputs of insulin were not restricted to specific cells. In contrast, DAF-16 appeared to have a greater effect on lifespan when expressed in a combination of tissues. These results suggest that insulin signaling may regulate DAF-16 through cell-intrinsic and endocrine pathways. We also found that an insulin-dependent response to fasting in intestinal cells was preferentially regulated by intestinal insulin signaling and was less responsive to insulin signaling from non-intestinal cells. Together, these results show that C. elegans insulin signaling has endocrine as well as tissue-specific outputs which could influence lifespan in a combinatorial fashion.

  12. Changes in Gastrointestinal Hormone Responses, Insulin Sensitivity, and Beta-Cell Function Within 2 Weeks After Gastric Bypass in Non-diabetic Subjects

    DEFF Research Database (Denmark)

    Jacobsen, Siv Hesse; Olesen, S C; Dirksen, C

    2012-01-01

    measured fasting and postprandial glucose, insulin, C-peptide, glucagon, total and intact glucagon-like peptide-1 (GLP-1), glucose-dependent insulinotropic polypeptide (GIP), glucagon-like peptide-2 (GLP-2), peptide YY(3-36) (PYY), cholecystokinin (CCK), total and active ghrelin, gastrin, somatostatin......, pancreatic polypeptide (PP), amylin, leptin, free fatty acids (FFA), and registered postprandial dumping. Insulin sensitivity was measured by homeostasis model assessment of insulin resistance. RESULTS: Fasting glucose, insulin, ghrelin, and PYY were significantly decreased and FFA was elevated...... postoperatively. Insulin sensitivity increased after surgery. The postprandial response increased for C-peptide, GLP-1, GLP-2, PYY, CCK, and glucagon (in response to the mixed meal) and decreased for total and active ghrelin, leptin, and gastrin, but were unchanged for GIP, amylin, PP, and somatostatin after...

  13. Identified peptidergic neurons in the Drosophila brain regulate insulin-producing cells, stress responses and metabolism by coexpressed short neuropeptide F and corazonin.

    Science.gov (United States)

    Kapan, Neval; Lushchak, Oleh V; Luo, Jiangnan; Nässel, Dick R

    2012-12-01

    Insulin/IGF-like signaling regulates the development, growth, fecundity, metabolic homeostasis, stress resistance and lifespan in worms, flies and mammals. Eight insulin-like peptides (DILP1-8) are found in Drosophila. Three of these (DILP2, 3 and 5) are produced by a set of median neurosecretory cells (insulin-producing cells, IPCs) in the brain. Activity in the IPCs of adult flies is regulated by glucose and several neurotransmitters and neuropeptides. One of these, short neuropeptide F (sNPF), regulates food intake, growth and Dilp transcript levels in IPCs via the sNPF receptor (sNPFR1) expressed on IPCs. Here we identify a set of brain neurons that utilizes sNPF to activate the IPCs. These sNPF-expressing neurons (dorsal lateral peptidergic neurons, DLPs) also produce the neuropeptide corazonin (CRZ) and have axon terminations impinging on IPCs. Knockdown of either sNPF or CRZ in DLPs extends survival in flies exposed to starvation and alters carbohydrate and lipid metabolism. Expression of sNPF in DLPs in the sNPF mutant background is sufficient to rescue wild-type metabolism and response to starvation. Since CRZ receptor RNAi in IPCs affects starvation resistance and metabolism, similar to peptide knockdown in DLPs, it is likely that also CRZ targets the IPCs. Knockdown of sNPF, but not CRZ in DLPs decreases transcription of Dilp2 and 5 in the brain, suggesting different mechanisms of action on IPCs of the two co-released peptides. Our findings indicate that sNPF and CRZ co-released from a small set of neurons regulate IPCs, stress resistance and metabolism in adult Drosophila.

  14. Studies on binding and mitogenic effect of insulin and insulin-like growth factor I in glomerular mesangial cells

    International Nuclear Information System (INIS)

    Conti, F.G.; Striker, L.J.; Lesniak, M.A.; MacKay, K.; Roth, J.; Striker, G.E.

    1988-01-01

    The mesangial cells are actively involved in regulating glomerular hemodynamics. Their overlying endothelium is fenestrated; therefore, these cells are directly exposed to plasma substances, including hormones such as insulin and insulin-like growth factor I (IGF-I). These peptides may contribute to the mesangial sclerosis and cellular hyperplasia that characterize diabetic glomerulopathy. We report herein the characterization of the receptors and the mitogenic effects of IGF-I and insulin on mouse glomerular mesangial cells in culture. The IGF-I receptor was characterized on intact cells. The Kd of the IGF-I receptor was 1.47 X 10(-9) M, and the estimated number of sites was 64,000 receptors/cell. The binding was time, temperature, and pH dependent, and the receptor showed down-regulation after exposure to serum. The expression of the receptor did not change on cells at different densities. The specific binding for insulin was too low to allow characterization of the insulin receptor on intact cells. However, it was possible to identify the insulin receptor in a wheat germ agglutinin-purified preparation of solubilized mesangial cells. This receptor showed the characteristic features of the insulin receptor, including pH dependence of binding and a curvilinear Scatchard plot. The mitogenic effects of insulin and IGF-I on mesangial cells were measured by the incorporation of [3H]thymidine into DNA. IGF-I was more potent than insulin. The half-maximal response to IGF-I stimulation occurred at 1.3 X 10(-10) M, and a similar increase with insulin was observed at concentrations in the range of 10(-7) M, suggesting that this insulin action was mediated through the IGF-I receptor. These data show that the mouse microvascular smooth muscle cells of the glomerulus express a cell surface receptor for IGF-I in vitro and that this peptide is a potent mitogen for these mesangial cells

  15. A method to determine insulin responsiveness in synaptosomes isolated from frozen brain tissue.

    Science.gov (United States)

    Franklin, Whitney; Taglialatela, Giulio

    2016-03-01

    Studying the insulin signaling response at the synapse is an important approach to understand molecular mechanisms involved in disease-related neurodegenerative processes. We developed a method for studying the insulin responsiveness at the synaptic level by isolating functional synaptosomes from fresh or frozen tissue and exposing them to insulin in the presence of ATP (a critical step) to detect insulin receptor (IR) activation. We performed an ATP dose-response curve, insulin dose-response curve, and insulin response time course to optimize this method. We also demonstrated that our protocol reflects the degree of insulin responsiveness in vivo by using an animal model of known insulin resistance, AtENPP1-Tg mice. This method is advantageous over other methods detecting IR in total brain homogenates due to the ability to detect IR response without confounding contributions from other cell areas and cell types also expressing IR. Furthermore, ex vivo insulin stimulation can be compared to baseline synaptosomes obtained from the same animal which improves reliability and statistical power while decreasing the number of animals required to perform individual experiments. We have developed a reliable, efficient method to measure insulin-driven ex vivo phosphorylation of the synaptosomal insulin receptor that can reliably reflect the pre-existing insulin responsiveness status in the CNS of the animal. To the best of our knowledge, this is the first evidence of stimulation of isolated synaptosomes with insulin and a promising new technique to study the synaptic CNS insulin responsiveness under physiological or disease conditions. Copyright © 2016 Elsevier B.V. All rights reserved.

  16. Adiposity in Children Born Small for Gestational Age Is Associated With β-Cell Function, Genetic Variants for Insulin Resistance, and Response to Growth Hormone Treatment

    DEFF Research Database (Denmark)

    Thankamony, Ajay; Jensen, Rikke Beck; O'Connell, Susan M

    2016-01-01

    -ray absorptiometry. The main outcome measures were treatment-related changes in height, IGF-1 standard deviation score, insulin sensitivity, insulin secretion, and disposition index. Combined multiallele gene scores based on single nucleotide polymorphisms with known associations with lower insulin sensitivity (gene...... (baseline vs 1-year z-scores, -0.26 ± 1.2 vs -1.23 ± 1.54; P IGF-1 responses (p-trends = .042), first-year height gains (B [95% confidence interval], 0.61 cm/y [0.28,0.95]; P

  17. Stress Hyperglycemia, Insulin Treatment, and Innate Immune Cells

    Directory of Open Access Journals (Sweden)

    Fangming Xiu

    2014-01-01

    Full Text Available Hyperglycemia (HG and insulin resistance are the hallmarks of a profoundly altered metabolism in critical illness resulting from the release of cortisol, catecholamines, and cytokines, as well as glucagon and growth hormone. Recent studies have proposed a fundamental role of the immune system towards the development of insulin resistance in traumatic patients. A comprehensive review of published literatures on the effects of hyperglycemia and insulin on innate immunity in critical illness was conducted. This review explored the interaction between the innate immune system and trauma-induced hypermetabolism, while providing greater insight into unraveling the relationship between innate immune cells and hyperglycemia. Critical illness substantially disturbs glucose metabolism resulting in a state of hyperglycemia. Alterations in glucose and insulin regulation affect the immune function of cellular components comprising the innate immunity system. Innate immune system dysfunction via hyperglycemia is associated with a higher morbidity and mortality in critical illness. Along with others, we hypothesize that reduction in morbidity and mortality observed in patients receiving insulin treatment is partially due to its effect on the attenuation of the immune response. However, there still remains substantial controversy regarding moderate versus intensive insulin treatment. Future studies need to determine the integrated effects of HG and insulin on the regulation of innate immunity in order to provide more effective insulin treatment regimen for these patients.

  18. Nicotinamide induces differentiation of embryonic stem cells into insulin-secreting cells

    International Nuclear Information System (INIS)

    Vaca, Pilar; Berna, Genoveva; Araujo, Raquel; Carneiro, Everardo M.; Bedoya, Francisco J.; Soria, Bernat; Martin, Franz

    2008-01-01

    The poly(ADP-ribose) polymerase (PARP) inhibitor, nicotinamide, induces differentiation and maturation of fetal pancreatic cells. In addition, we have previously reported evidence that nicotinamide increases the insulin content of cells differentiated from embryonic stem (ES) cells, but the possibility of nicotinamide acting as a differentiating agent on its own has never been completely explored. Islet cell differentiation was studied by: (i) X-gal staining after neomycin selection; (ii) BrdU studies; (iii) single and double immunohistochemistry for insulin, C-peptide and Glut-2; (iv) insulin and C-peptide content and secretion assays; and (v) transplantation of differentiated cells, under the kidney capsule, into streptozotocin (STZ)-diabetic mice. Here we show that undifferentiated mouse ES cells treated with nicotinamide: (i) showed an 80% decrease in cell proliferation; (ii) co-expressed insulin, C-peptide and Glut-2; (iii) had values of insulin and C-peptide corresponding to 10% of normal mouse islets; (iv) released insulin and C-peptide in response to stimulatory glucose concentrations; and (v) after transplantation into diabetic mice, normalized blood glucose levels over 7 weeks. Our data indicate that nicotinamide decreases ES cell proliferation and induces differentiation into insulin-secreting cells. Both aspects are very important when thinking about cell therapy for the treatment of diabetes based on ES cells

  19. Targeting development of incretin-producing cells increases insulin secretion

    DEFF Research Database (Denmark)

    Petersen, Natalia; Reimann, Frank; van Es, Johan H

    2015-01-01

    the number of intestinal L cells, which produce GLP-1, is an alternative strategy to augment insulin responses and improve glucose tolerance. Blocking the NOTCH signaling pathway with the γ-secretase inhibitor dibenzazepine increased the number of L cells in intestinal organoid-based mouse and human culture...... of the development of incretin-producing cells in the intestine has potential as a therapeutic strategy to improve glycemic control....

  20. Globular adiponectin ameliorates metabolic insulin resistance via AMPK-mediated restoration of microvascular insulin responses

    Science.gov (United States)

    Zhao, Lina; Fu, Zhuo; Wu, Jing; Aylor, Kevin W; Barrett, Eugene J; Cao, Wenhong; Liu, Zhenqi

    2015-01-01

    Abstract Hypoadiponectinaemia is closely associated with endothelial dysfunction and insulin resistance, and microvasculature plays a critical role in the regulation of insulin action in muscle. Here we tested whether adiponectin replenishment could improve metabolic insulin sensitivity in male rats fed a high-fat diet (HFD) via the modulation of microvascular insulin responses. Male Sprague–Dawley rats were fed either a HFD or low-fat diet (LFD) for 4 weeks. Small resistance artery myograph changes in tension, muscle microvascular recruitment and metabolic response to insulin were determined. Compared with rats fed a LFD, HFD feeding abolished the vasodilatory actions of globular adiponectin (gAd) and insulin on pre-constricted distal saphenous arteries. Pretreatment with gAd improved insulin responses in arterioles isolated from HFD rats, which was blocked by AMP-activated protein kinase (AMPK) inhibition. Similarly, HFD abolished microvascular responses to either gAd or insulin and decreased insulin-stimulated glucose disposal by ∼60%. However, supplementing gAd fully rescued insulin’s microvascular action and significantly improved the metabolic responses to insulin in HFD male rats and these actions were abolished by inhibition of either AMPK or nitric oxide production. We conclude that HFD induces vascular adiponectin and insulin resistance but gAd administration can restore vascular insulin responses and improve insulin’s metabolic action via an AMPK- and nitric oxide-dependent mechanism in male rats. Key points Adiponectin is an adipokine with anti-inflammatory and anti-diabetic properties. Hypoadiponectinaemia is closely associated with endothelial dysfunction and insulin resistance in obesity and diabetes. Insulin resistance is present in muscle microvasculature and this may contribute to decreased insulin delivery to, and action in, muscle. In this study we examined whether adiponectin ameliorates metabolic insulin resistance by affecting muscle

  1. In vivo response of Mesocestoides vogae to human insulin.

    Science.gov (United States)

    Canclini, L; Esteves, A

    2009-02-01

    Successful host invasion by parasitic helminths involves detection and appropriate response to a range of host-derived signals. Insulin signal response pathways are ancient and highly-conserved throughout the metazoans. However, very little is known about helminth insulin signalling and the potential role it may play in host-parasite interactions. The response of Mesocestoides vogae (Cestoda: Cyclophyllidea) larvae to human insulin was investigated, focusing on tyrosine-phosphorylation status, glucose content, survival and asexual reproduction rate. Parasite larvae were challenged with different levels of insulin for variable periods. The parameters tested were influenced by human insulin, and suggested a host-parasite molecular dialogue.

  2. Evaluation of insulin expression and secretion in genetically engineered gut K and L-cells

    Directory of Open Access Journals (Sweden)

    Ahmad Zalinah

    2012-09-01

    Full Text Available Abstract Background Gene therapy could provide an effective treatment of diabetes. Previous studies have investigated the potential for several cell and tissue types to produce mature and active insulin. Gut K and L-cells could be potential candidate hosts for gene therapy because of their special features. Results In this study, we isolated gut K and L-cells to compare the potential of both cell types to produce insulin when exposed to similar conditions. The isolated pure K and L-cells were transfected with recombinant plasmids encoding insulin and with specific promoters for K or L-cells. Insulin expression was studied in response to glucose or meat hydrolysate. We found that glucose and meat hydrolysate efficiently induced insulin secretion from K and L-cells. However, the effects of meat hydrolysate on insulin secretion were more potent in both cells compared with glucose. Results of enzyme-linked immunosorbent assays showed that L-cells secreted more insulin compared with K-cells regardless of the stimulator, although this difference was not statistically significant. Conclusion The responses of K and L-cells to stimulation with glucose or meat hydrolysate were generally comparable. Therefore, both K and L-cells show similar potential to be used as surrogate cells for insulin gene expression in vitro. The potential use of these cells for diabetic gene therapy warrants further investigation.

  3. Chronic Insulin Exposure Induces ER Stress and Lipid Body Accumulation in Mast Cells at the Expense of Their Secretory Degranulation Response

    OpenAIRE

    Greineisen, William E.; Maaetoft-Udsen, Kristina; Speck, Mark; Balajadia, Januaria; Shimoda, Lori M. N.; Sung, Carl; Turner, Helen

    2015-01-01

    Lipid bodies (LB) are reservoirs of precursors to inflammatory lipid mediators in immunocytes, including mast cells. LB numbers are dynamic, increasing dramatically under conditions of immunological challenge. We have previously shown in vitro that insulin-influenced lipogenic pathways induce LB biogenesis in mast cells, with their numbers attaining steatosis-like levels. Here, we demonstrate that in vivo hyperinsulinemia resulting from high fat diet is associated with LB accumulation in muri...

  4. Insulin regulation of Na/K pump activity in rat hepatoma cells

    International Nuclear Information System (INIS)

    Gelehrter, T.D.; Shreve, P.D.; Dilworth, V.M.

    1984-01-01

    Insulin rapidly increases Na/K pump activity in HTC rat hepatoma cells in tissue culture, as measured by the ouabain-sensitive influx of the potassium analogue 86Rb+. Increased influx is observed within minutes and is maximal (70% above control) within 1-2 h. The effect appears to be mediated by the insulin receptors, as: the concentration dependence on insulin is identical to that for insulin induction of tyrosine aminotransferase and stimulation of 2-aminoisobutyric acid transport, proinsulin is 6% as potent as insulin, and the effect is blocked by anti-receptor antibodies. The early stimulation of potassium influx is not blocked by cycloheximide and is not associated with an increased number of pump sites as measured by 3 H-ouabain binding. The insulin effect is blocked by amiloride, which blocks sodium influx, and is mimicked by the sodium ionophore monensin, which increases sodium influx and intracellular accumulation. Insulin also rapidly increases the initial rate of 22 Na+ influx, suggesting that insulin may enhance Na/K pump activity, in part, by increasing intracellular sodium concentration. Incubation of HTC cells with insulin for 24 h causes complete unresponsiveness to the insulin induction of transaminase and stimulation of amino acid transport, a phenomenon mediated by postbinding mechanisms. In contrast, similar incubation with insulin does not cause unresponsiveness to the insulin stimulation of Na/K pump activity. Therefore, the site of regulation of responsiveness to insulin must be distal to, or separate from, those events causing stimulation of ion fluxes

  5. Characterisation of insulin-producing cells differentiated from tonsil derived mesenchymal stem cells.

    Science.gov (United States)

    Kim, So-Yeon; Kim, Ye-Ryung; Park, Woo-Jae; Kim, Han Su; Jung, Sung-Chul; Woo, So-Youn; Jo, Inho; Ryu, Kyung-Ha; Park, Joo-Won

    2015-01-01

    Tonsil-derived (T-) mesenchymal stem cells (MSCs) display mutilineage differentiation potential and self-renewal capacity and have potential as a banking source. Diabetes mellitus is a prevalent disease in modern society, and the transplantation of pancreatic progenitor cells or various stem cell-derived insulin-secreting cells has been suggested as a novel therapy for diabetes. The potential of T-MSCs to trans-differentiate into pancreatic progenitor cells or insulin-secreting cells has not yet been investigated. We examined the potential of human T-MSCs to trans-differentiate into pancreatic islet cells using two different methods based on β-mercaptoethanol and insulin-transferin-selenium, respectively. First, we compared the efficacy of the two methods for inducing differentiation into insulin-producing cells. We demonstrated that the insulin-transferin-selenium method is more efficient for inducing differentiation into insulin-secreting cells regardless of the source of the MSCs. Second, we compared the differentiation potential of two different MSC types: T-MSCs and adipose-derived MSCs (A-MSCs). T-MSCs had a differentiation capacity similar to that of A-MSCs and were capable of secreting insulin in response to glucose concentration. Islet-like clusters differentiated from T-MSCs had lower synaptotagmin-3, -5, -7, and -8 levels, and consequently lower secreted insulin levels than cells differentiated from A-MSCs. These results imply that T-MSCs can differentiate into functional pancreatic islet-like cells and could provide a novel, alternative cell therapy for diabetes mellitus. Copyright © 2015 International Society of Differentiation. Published by Elsevier B.V. All rights reserved.

  6. IRS-1: essential for insulin- and IL-4-stimulated mitogenesis in hematopoietic cells.

    Science.gov (United States)

    Wang, L M; Myers, M G; Sun, X J; Aaronson, S A; White, M; Pierce, J H

    1993-09-17

    Although several interleukin-3 (IL-3)-dependent cell lines proliferate in response to IL-4 or insulin, the 32D line does not. Insulin and IL-4 sensitivity was restored to 32D cells by expression of IRS-1, the principal substrate of the insulin receptor. Although 32D cells possessed receptors for both factors, they lacked the IRS-1--related protein, 4PS, which becomes phosphorylated by tyrosine in insulin- or IL-4--responsive lines after stimulation. These results indicate that factors that bind unrelated receptors can use similar mitogenic signaling pathways in hematopoietic cells and that 4PS and IRS-1 are functionally similar proteins that are essential for insulin- and IL-4--induced proliferation.

  7. Internalization and localization of basal insulin peglispro in cells.

    Science.gov (United States)

    Moyers, Julie S; Volk, Catherine B; Cao, Julia X C; Zhang, Chen; Ding, Liyun; Kiselyov, Vladislav V; Michael, M Dodson

    2017-10-15

    Basal insulin peglispro (BIL) is a novel, PEGylated insulin lispro that has a large hydrodynamic size compared with insulin lispro. It has a prolonged duration of action, which is related to a delay in insulin absorption and a reduction in clearance. Given the different physical properties of BIL compared with native insulin and insulin lispro, it is important to assess the cellular internalization characteristics of the molecule. Using immunofluorescent confocal imaging, we compared the cellular internalization and localization patterns of BIL, biosynthetic human insulin, and insulin lispro. We assessed the effects of BIL on internalization of the insulin receptor (IR) and studied cellular clearance of BIL. Co-localization studies using antibodies to either insulin or PEG, and the early endosomal marker EEA1 showed that the overall internalization and subcellular localization pattern of BIL was similar to that of human insulin and insulin lispro; all were rapidly internalized and co-localized with EEA1. During ligand washout for 4 h, concomitant loss of insulin, PEG methoxy group, and PEG backbone immunostaining was observed for BIL, similar to the loss of insulin immunostaining observed for insulin lispro and human insulin. Co-localization studies using an antibody to the lysosomal marker LAMP1 did not reveal evidence of lysosomal localization for insulin lispro, human insulin, BIL, or PEG using either insulin or PEG immunostaining reagents. BIL and human insulin both induced rapid phosphorylation and internalization of human IR. Our findings show that treatment of cells with BIL stimulates internalization and localization of IR to early endosomes. Both the insulin and PEG moieties of BIL undergo a dynamic cellular process of rapid internalization and transport to early endosomes followed by loss of cellular immunostaining in a manner similar to that of insulin lispro and human insulin. The rate of clearance for the insulin lispro portion of BIL was slower than

  8. The adipose transcriptional response to insulin is determined by obesity, not insulin sensitivity

    DEFF Research Database (Denmark)

    Rydén, Mikael; Hrydziuszko, Olga; Mileti, Enrichetta

    2016-01-01

    Metabolically healthy obese subjects display preserved insulin sensitivity and a beneficial white adipose tissue gene expression pattern. However, this observation stems from fasting studies when insulin levels are low. We investigated adipose gene expression by 5'Cap-mRNA sequencing in 17 healthy...... non-obese (NO), 21 insulin-sensitive severely obese (ISO), and 30 insulin-resistant severely obese (IRO) subjects, before and 2 hr into a hyperinsulinemic euglycemic clamp. ISO and IRO subjects displayed a clear but globally similar transcriptional response to insulin, which differed from the small...... effects observed in NO subjects. In the obese, 231 genes were altered; 71 were enriched in ISO subjects (e.g., phosphorylation processes), and 52 were enriched in IRO subjects (e.g., cellular stimuli). Common cardio-metabolic risk factors and gender do not influence these findings. This study demonstrates...

  9. Insulin resistance, insulin response, and obesity as indicators of metabolic risk

    DEFF Research Database (Denmark)

    Ferrannini, Ele; Balkau, Beverley; Coppack, Simon W

    2007-01-01

    CONTEXT: Insulin resistance (IR) and obesity, especially abdominal obesity, are regarded as central pathophysiological features of a cluster of cardiovascular risk factors (CVRFs), but their relative roles remain undefined. Moreover, the differential impact of IR viz. insulin response has not been...... evaluated. OBJECTIVE: The objective of this study was to dissect out the impact of obesity, abdominal obesity, and IR/insulin response on CVRF. DESIGN: This was a cross-sectional study. SETTING: The study was conducted at 21 research centers in Europe. SUBJECTS: The study included a cohort of 1308......-cholesterol, and lower high-density lipoprotein-cholesterol, and insulin response to higher heart rate, blood pressure and fasting glucose, and the same dyslipidemic profile as IR (P

  10. Insulin-producing cells generated from dedifferentiated human pancreatic beta cells expanded in vitro.

    Directory of Open Access Journals (Sweden)

    Holger A Russ

    Full Text Available Expansion of beta cells from the limited number of adult human islet donors is an attractive prospect for increasing cell availability for cell therapy of diabetes. However, attempts at expanding human islet cells in tissue culture result in loss of beta-cell phenotype. Using a lineage-tracing approach we provided evidence for massive proliferation of beta-cell-derived (BCD cells within these cultures. Expansion involves dedifferentiation resembling epithelial-mesenchymal transition (EMT. Epigenetic analyses indicate that key beta-cell genes maintain open chromatin structure in expanded BCD cells, although they are not transcribed. Here we investigated whether BCD cells can be redifferentiated into beta-like cells.Redifferentiation conditions were screened by following activation of an insulin-DsRed2 reporter gene. Redifferentiated cells were characterized for gene expression, insulin content and secretion assays, and presence of secretory vesicles by electron microscopy. BCD cells were induced to redifferentiate by a combination of soluble factors. The redifferentiated cells expressed beta-cell genes, stored insulin in typical secretory vesicles, and released it in response to glucose. The redifferentiation process involved mesenchymal-epithelial transition, as judged by changes in gene expression. Moreover, inhibition of the EMT effector SLUG (SNAI2 using shRNA resulted in stimulation of redifferentiation. Lineage-traced cells also gave rise at a low rate to cells expressing other islet hormones, suggesting transition of BCD cells through an islet progenitor-like stage during redifferentiation.These findings demonstrate for the first time that expanded dedifferentiated beta cells can be induced to redifferentiate in culture. The findings suggest that ex-vivo expansion of adult human islet cells is a promising approach for generation of insulin-producing cells for transplantation, as well as basic research, toxicology studies, and drug

  11. Effects of turtle oil on insulin sensitivity and glucose metabolism in insulin resistant cell model

    International Nuclear Information System (INIS)

    Bai Jing; Tian Yaping; Guo Duo

    2007-01-01

    To evaluate the effects of turtle oil on insulin sensitivity and glucose metabolism in an insulin-resistant (IR) cell model which was established by the way of high concentration of insulin induction with HepG 2 cell in vitro culture. The IR cells were treated by turtle oil, the glucose consumption and 3 H-D-glucose incorporation rate in IR cells were detected by the way of glucose oxidase and 3 H-D-glucose incorporation assay respectively. The state of cell proliferation was tested by MTT method. The results showed that the incorporation rate of 3 H-D-glucose in IR cells was significantly lower than that in the control cells(P 3 H-D-glucose incorporation rate in either IR cells or control cells was increased with the increase of insulin concentration. Moreover, the 3 H-D-glucose incorporation rate of IR cells increased slower than that of control cells. The MTT assay showed that turtle oil can promote the proliferation of IR cell and control cell. The glucose uptake and glucose consumption in IR cell which treated with turtle oil was significantly increase than that in the control cells (P<0.05). Turtle oil can improve the insulin sensitivity and glucose metabolism in the IR cell model. (authors)

  12. Adverse effects of reduced oxygen tension on the proliferative capacity of rat kidney and insulin-secreting cell lines involve DNA damage and stress responses

    International Nuclear Information System (INIS)

    Chen Jianhua; Jones, R. Huw; Tarry-Adkins, Jane; Smith, Noel H.; Ozanne, Susan E.

    2008-01-01

    Standard cell culture conditions do not reflect the physiological environment in terms of oxygen tension (20% vs 3%). The effects of lowering oxygen tension on cell proliferation in culture can be beneficial as well as detrimental depending on the cell line studied, but the molecular mechanism underlying such effects is not fully understood. We observed that the proliferative capacity of the rat cell lines NRK and INS-1 was inhibited when cultured under 3% oxygen as compared to 20% oxygen. Suppression of proliferation in NRK cells was accompanied by induction of DNA double strand breaks whereas in INS-1 cells it was accompanied by up-regulation of p53 and p27. Although Sirt1 was up-regulated in both cell lines by 3% oxygen the effects on antioxidant enzymes (MnSOD, CuZnSOD and catalase) were cell line specific. Marked up-regulation of heme oxygenase-1 (HO-1) was detected in both NRK and INS-1 cells when cultured in 3% oxygen. HO-1 expression can be readily induced by exposure to hydrogen peroxide in culture. These results suggest that reduced oxygen tension suppresses the proliferative capacity of these two cell lines through a stress response that is similar to an oxidative stress response but the molecular events that lead to the reduced cell proliferation are cell line specific

  13. Insulin resistance in vascular endothelial cells promotes intestinal tumour formation

    DEFF Research Database (Denmark)

    Wang, X; Häring, M-F; Rathjen, Thomas

    2017-01-01

    in vascular endothelial cells. Strikingly, these mice had 42% more intestinal tumours than controls, no change in tumour angiogenesis, but increased expression of vascular cell adhesion molecule-1 (VCAM-1) in primary culture of tumour endothelial cells. Insulin decreased VCAM-1 expression and leukocyte...... adhesion in quiescent tumour endothelial cells with intact insulin receptors and partly prevented increases in VCAM-1 and leukocyte adhesion after treatment with tumour necrosis factor-α. Knockout of insulin receptors in endothelial cells also increased leukocyte adhesion in mesenteric venules...

  14. Liver-derived systemic factors drive β-cell hyperplasia in insulin resistant states

    Energy Technology Data Exchange (ETDEWEB)

    El Ouaamari, Abdelfattah; Kawamori, Dan; Dirice, Ercument; Liew, Chong Wee; Shadrach, Jennifer L.; Hu, Jiang; Katsuta, Hitoshi; Hollister-Lock, Jennifer; Qian, Weijun; Wagers, Amy J.; Kulkarni, Rohit N.

    2013-02-21

    Integrative organ cross-talk regulates key aspects of energy homeostasis and its dysregulation may underlie metabolic disorders such as obesity and diabetes. To test the hypothesis that cross-talk between the liver and pancreatic islets modulates β-cell growth in response to insulin resistance, we used the Liver-specific Insulin Receptor Knockout (LIRKO) mouse, a unique model that exhibits dramatic islet hyperplasia. Using complementary in vivo parabiosis and transplantation assays, and in vitro islet culture approaches, we demonstrate that humoral, non-neural, non-cell autonomous factor(s) induce β-cell proliferation in LIRKO mice. Furthermore, we report that a hepatocyte-derived factor(s) stimulates mouse and human β-cell proliferation in ex vivo assays, independent of ambient glucose and insulin levels. These data implicate the liver as a critical source of β-cell growth factors in insulin resistant states.

  15. From Human Mesenchymal Stem Cells to Insulin-Producing Cells: Comparison between Bone Marrow- and Adipose Tissue-Derived Cells.

    Science.gov (United States)

    Gabr, Mahmoud M; Zakaria, Mahmoud M; Refaie, Ayman F; Abdel-Rahman, Engy A; Reda, Asmaa M; Ali, Sameh S; Khater, Sherry M; Ashamallah, Sylvia A; Ismail, Amani M; Ismail, Hossam El-Din A; El-Badri, Nagwa; Ghoneim, Mohamed A

    2017-01-01

    The aim of this study is to compare human bone marrow-derived mesenchymal stem cells (BM-MSCs) and adipose tissue-derived mesenchymal stem cells (AT-MSCs), for their differentiation potentials to form insulin-producing cells. BM-MSCs were obtained during elective orthotopic surgery and AT-MSCs from fatty aspirates during elective cosmetics procedures. Following their expansion, cells were characterized by phenotyping, trilineage differentiation ability, and basal gene expression of pluripotency genes and for their metabolic characteristics. Cells were differentiated according to a Trichostatin-A based protocol. The differentiated cells were evaluated by immunocytochemistry staining for insulin and c-peptide. In addition the expression of relevant pancreatic endocrine genes was determined. The release of insulin and c-peptide in response to a glucose challenge was also quantitated. There were some differences in basal gene expression and metabolic characteristics. After differentiation the proportion of the resulting insulin-producing cells (IPCs), was comparable among both cell sources. Again, there were no differences neither in the levels of gene expression nor in the amounts of insulin and c-peptide release as a function of glucose challenge. The properties, availability, and abundance of AT-MSCs render them well-suited for applications in regenerative medicine. Conclusion . BM-MSCs and AT-MSCs are comparable regarding their differential potential to form IPCs. The availability and properties of AT-MSCs render them well-suited for applications in regenerative medicine.

  16. From Human Mesenchymal Stem Cells to Insulin-Producing Cells: Comparison between Bone Marrow- and Adipose Tissue-Derived Cells

    Directory of Open Access Journals (Sweden)

    Mahmoud M. Gabr

    2017-01-01

    Full Text Available The aim of this study is to compare human bone marrow-derived mesenchymal stem cells (BM-MSCs and adipose tissue-derived mesenchymal stem cells (AT-MSCs, for their differentiation potentials to form insulin-producing cells. BM-MSCs were obtained during elective orthotopic surgery and AT-MSCs from fatty aspirates during elective cosmetics procedures. Following their expansion, cells were characterized by phenotyping, trilineage differentiation ability, and basal gene expression of pluripotency genes and for their metabolic characteristics. Cells were differentiated according to a Trichostatin-A based protocol. The differentiated cells were evaluated by immunocytochemistry staining for insulin and c-peptide. In addition the expression of relevant pancreatic endocrine genes was determined. The release of insulin and c-peptide in response to a glucose challenge was also quantitated. There were some differences in basal gene expression and metabolic characteristics. After differentiation the proportion of the resulting insulin-producing cells (IPCs, was comparable among both cell sources. Again, there were no differences neither in the levels of gene expression nor in the amounts of insulin and c-peptide release as a function of glucose challenge. The properties, availability, and abundance of AT-MSCs render them well-suited for applications in regenerative medicine. Conclusion. BM-MSCs and AT-MSCs are comparable regarding their differential potential to form IPCs. The availability and properties of AT-MSCs render them well-suited for applications in regenerative medicine.

  17. Rapid internalization of the insulin receptor in rat hepatoma cells

    International Nuclear Information System (INIS)

    Backer, J.M.; White, M.F.; Kahn, C.R.

    1987-01-01

    The authors have studied the internalization of the insulin receptor (IR) in rat hepatoma cells (Fao). The cells were surface-iodinated at 4 0 C, stimulated with insulin at 37 0 C, and then cooled rapidly, trypsinized at 4 0 C and solubilized. The IR was immunoprecipitated with a specific antibody, and internalization of the IR was assessed by the appearance of trypsin-resistant bands on SDS-PAGE. Insulin induced the internalization of surface receptors with a t 1/2 of 9-10 mins; cells not exposed to insulin internalized less than 20% of the IR during 1 h at 37 0 C. Further experiments demonstrated that the accumulation of trypsin-resistant IR paralleled a loss of receptor from the cell surface. Insulin-stimulated cells were chilled and iodinated at 4 0 C, followed by solubilization, immunoprecipitation and SDS-PAGE; alternatively, insulin-stimulated cells were chilled, surface-bound ligand removed by washing the cells at pH 4.2, and specific [ 125 I]insulin binding measured at 4 0 C. Both techniques confirmed the disappearance of IR from the cell surface at rates comparable to the insulin-stimulated internalization described above. The total amount of phosphotyrosine-containing IR, as assessed by immunoprecipitation with an anti-phosphotyrosine antibody, remained constant during this time interval, suggesting that active kinase is translocated into the cell. In summary, the authors data indicate that insulin binding increases the rate of IR internalization of Fao cells. This relocation may facilitate the interaction of the activated tyrosine kinase in the IR with intracellular substrates, thus transmitting the insulin signal to metabolic pathways

  18. Identification and characterization of insulin receptors on foetal-mouse brain-cortical cells.

    OpenAIRE

    Van Schravendijk, C F; Hooghe-Peters, E L; De Meyts, P; Pipeleers, D G

    1984-01-01

    The occurrence of insulin receptors was investigated in freshly dissociated brain-cortical cells from mouse embryos. By analogy with classical insulin-binding cell types, binding of 125I-insulin to foetal brain-cortical cells was time- and pH-dependent, only partially reversible, and competed for by unlabelled insulin and closely related peptides. Desalanine-desasparagine-insulin, pig proinsulin, hagfish insulin and turkey insulin were respectively 2%, 4%, 2% and 200% as potent as bovine insu...

  19. Aβ-Induced Insulin Resistance and the Effects of Insulin on the Cholesterol Synthesis Pathway and Aβ Secretion in Neural Cells.

    Science.gov (United States)

    Najem, Dema; Bamji-Mirza, Michelle; Yang, Ze; Zhang, Wandong

    2016-06-01

    Alzheimer's disease (AD) is characterized by amyloid-β (Aβ) toxicity, tau pathology, insulin resistance, neuroinflammation, and dysregulation of cholesterol homeostasis, all of which play roles in neurodegeneration. Insulin has polytrophic effects on neurons and may be at the center of these pathophysiological changes. In this study, we investigated possible relationships among insulin signaling and cholesterol biosynthesis, along with the effects of Aβ42 on these pathways in vitro. We found that neuroblastoma 2a (N2a) cells transfected with the human gene encoding amyloid-β protein precursor (AβPP) (N2a-AβPP) produced Aβ and exhibited insulin resistance by reduced p-Akt and a suppressed cholesterol-synthesis pathway following insulin treatment, and by increased phosphorylation of insulin receptor subunit-1 at serine 612 (p-IRS-S612) as compared to parental N2a cells. Treatment of human neuroblastoma SH-SY5Y cells with Aβ42 also increased p-IRS-S612, suggesting that Aβ42 is responsible for insulin resistance. The insulin resistance was alleviated when N2a-AβPP cells were treated with higher insulin concentrations. Insulin increased Aβ release from N2a-AβPP cells, by which it may promote Aβ clearance. Insulin increased cholesterol-synthesis gene expression in SH-SY5Y and N2a cells, including 24-dehydrocholesterol reductase (DHCR24) and 3-hydroxy-3-methyl-glutaryl-CoA reductase (HMGCR) through sterol-regulatory element-binding protein-2 (SREBP2). While Aβ42-treated SH-SY5Y cells exhibited increased HMGCR expression and c-Jun phosphorylation as pro-inflammatory responses, they also showed down-regulation of neuro-protective/anti-inflammatory DHCR24. These results suggest that Aβ42 may cause insulin resistance, activate JNK for c-Jun phosphorylation, and lead to dysregulation of cholesterol homeostasis, and that enhancing insulin signaling may relieve the insulin-resistant phenotype and the dysregulated cholesterol-synthesis pathway to promote A

  20. Clinical study on insulin receptors of mononuclear cells in diabetes

    Energy Technology Data Exchange (ETDEWEB)

    Dalimunthe, D [Hiroshima Univ. (Japan). School of Medicine

    1980-12-01

    /sup 125/I-insulin binding activity to mononuclear cells was studied in 75 noninsulin-dependent diabetic subjects and 31 normal subjects and the following results were obtained. 1. /sup 125/I-insulin binding is directly proportional to the mononuclear cell concentrations. There is a linear increase of specific /sup 125/I-insulin binding. 2. The binding of /sup 125/I-insulin to mononuclear cells is displaced by the increasing concentration of native insulin. 3. The /sup 125/I-insulin degradation in the incubation medium after incubation of mononuclear cells for 24 hours at 4/sup 0/C was almost 5% in this study. 4. The insulin binding activity in diabetic subjects was lower than that in normal subjects (P < 0.001) without any significant difference in affinity constant. 5. The relationship of binding activity to age of diabetics (r = 0.06, N.S), relative body weitht (r = 0.06, N.S) and duration of diabetes from onset was not significant. 6. In untreated noninsulin-dependent diabetics the insulin binding activity was inversely correlated to fasting blood glucose level (r = 0.78, P < 0.001) and slightly inversely correlated to serum insulin level (r = 0.47, P < 0.01). A slight inverse correlation was also observed in serum triglyceride level (r = 0.53, P < 0.01) and in total cholesterol level (r = 0.29, P < 0.05). 7. No significant difference between the binding activity was observed by grade of diabetic retinopathy. 8. After treatment with diet and/or sulfonylurea, the diabetics exhibited a significant increase in insulin binding activity (P < 0.005) but no significant difference in plasma insulin level, body weight and plasma lipid levels was observed.

  1. Insulin promotes cell migration by regulating PSA-NCAM

    Energy Technology Data Exchange (ETDEWEB)

    Monzo, Hector J.; Coppieters, Natacha [Centre for Brain Research, Faculty of Medical and Health Sciences, The University of Auckland, Private Bag, 92019, Auckland (New Zealand); Department of Anatomy and Medical Imaging, Faculty of Medical and Health Sciences, The University of Auckland, Private Bag, 92019, Auckland (New Zealand); Park, Thomas I.H. [Centre for Brain Research, Faculty of Medical and Health Sciences, The University of Auckland, Private Bag, 92019, Auckland (New Zealand); Department of Pharmacology, Faculty of Medical and Health Sciences, The University of Auckland, Private Bag, 92019, Auckland (New Zealand); Dieriks, Birger V.; Faull, Richard L.M. [Centre for Brain Research, Faculty of Medical and Health Sciences, The University of Auckland, Private Bag, 92019, Auckland (New Zealand); Department of Anatomy and Medical Imaging, Faculty of Medical and Health Sciences, The University of Auckland, Private Bag, 92019, Auckland (New Zealand); Dragunow, Mike [Centre for Brain Research, Faculty of Medical and Health Sciences, The University of Auckland, Private Bag, 92019, Auckland (New Zealand); Department of Pharmacology, Faculty of Medical and Health Sciences, The University of Auckland, Private Bag, 92019, Auckland (New Zealand); Curtis, Maurice A., E-mail: m.curtis@auckland.ac.nz [Centre for Brain Research, Faculty of Medical and Health Sciences, The University of Auckland, Private Bag, 92019, Auckland (New Zealand); Department of Anatomy and Medical Imaging, Faculty of Medical and Health Sciences, The University of Auckland, Private Bag, 92019, Auckland (New Zealand)

    2017-06-01

    Cellular interactions with the extracellular environment are modulated by cell surface polysialic acid (PSA) carried by the neural cell adhesion molecule (NCAM). PSA-NCAM is involved in cellular processes such as differentiation, plasticity, and migration, and is elevated in Alzheimer's disease as well as in metastatic tumour cells. Our previous work demonstrated that insulin enhances the abundance of cell surface PSA by inhibiting PSA-NCAM endocytosis. In the present study we have identified a mechanism for insulin-dependent inhibition of PSA-NCAM turnover affecting cell migration. Insulin enhanced the phosphorylation of the focal adhesion kinase leading to dissociation of αv-integrin/PSA-NCAM clusters, and promoted cell migration. Our results show that αv-integrin plays a key role in the PSA-NCAM turnover process. αv-integrin knockdown stopped PSA-NCAM from being endocytosed, and αv-integrin/PSA-NCAM clusters co-labelled intracellularly with Rab5, altogether indicating a role for αv-integrin as a carrier for PSA-NCAM during internalisation. Furthermore, inhibition of p-FAK caused dissociation of αv-integrin/PSA-NCAM clusters and counteracted the insulin-induced accumulation of PSA at the cell surface and cell migration was impaired. Our data reveal a functional association between the insulin/p-FAK-dependent regulation of PSA-NCAM turnover and cell migration through the extracellular matrix. Most importantly, they identify a novel mechanism for insulin-stimulated cell migration. - Highlights: • Insulin modulates PSA-NCAM turnover through upregulation of p-FAK. • P-FAK modulates αv-integrin/PSA-NCAM clustering. • αv-integrin acts as a carrier for PSA-NCAM endocytosis. • Cell migration is promoted by cell surface PSA. • Insulin promotes PSA-dependent migration in vitro.

  2. Insulin promotes cell migration by regulating PSA-NCAM

    International Nuclear Information System (INIS)

    Monzo, Hector J.; Coppieters, Natacha; Park, Thomas I.H.; Dieriks, Birger V.; Faull, Richard L.M.; Dragunow, Mike; Curtis, Maurice A.

    2017-01-01

    Cellular interactions with the extracellular environment are modulated by cell surface polysialic acid (PSA) carried by the neural cell adhesion molecule (NCAM). PSA-NCAM is involved in cellular processes such as differentiation, plasticity, and migration, and is elevated in Alzheimer's disease as well as in metastatic tumour cells. Our previous work demonstrated that insulin enhances the abundance of cell surface PSA by inhibiting PSA-NCAM endocytosis. In the present study we have identified a mechanism for insulin-dependent inhibition of PSA-NCAM turnover affecting cell migration. Insulin enhanced the phosphorylation of the focal adhesion kinase leading to dissociation of αv-integrin/PSA-NCAM clusters, and promoted cell migration. Our results show that αv-integrin plays a key role in the PSA-NCAM turnover process. αv-integrin knockdown stopped PSA-NCAM from being endocytosed, and αv-integrin/PSA-NCAM clusters co-labelled intracellularly with Rab5, altogether indicating a role for αv-integrin as a carrier for PSA-NCAM during internalisation. Furthermore, inhibition of p-FAK caused dissociation of αv-integrin/PSA-NCAM clusters and counteracted the insulin-induced accumulation of PSA at the cell surface and cell migration was impaired. Our data reveal a functional association between the insulin/p-FAK-dependent regulation of PSA-NCAM turnover and cell migration through the extracellular matrix. Most importantly, they identify a novel mechanism for insulin-stimulated cell migration. - Highlights: • Insulin modulates PSA-NCAM turnover through upregulation of p-FAK. • P-FAK modulates αv-integrin/PSA-NCAM clustering. • αv-integrin acts as a carrier for PSA-NCAM endocytosis. • Cell migration is promoted by cell surface PSA. • Insulin promotes PSA-dependent migration in vitro.

  3. Characterization of insulin-like growth factor I and insulin receptors on cultured bovine adrenal fasciculata cells. Role of these peptides on adrenal cell function

    International Nuclear Information System (INIS)

    Penhoat, A.; Chatelain, P.G.; Jaillard, C.; Saez, J.M.

    1988-01-01

    We have characterized insulin-like growth factor I (IGF-I) and insulin receptors in cultured bovine adrenal cells by binding and cross-linking affinity experiments. At equilibrium the dissociation constant and the number of binding sites per cell for IGF-I were 1.4 +/- (SE) 0.3 x 10(-9) M and 19,200 +/- 2,100, respectively. Under reduction conditions, disuccinimidyl suberate cross-linked [ 125 I]iodo-IGF-I to one receptor complex with an Mr of 125,000. Adrenal cells also contain specific insulin receptors with an apparent dissociation constant (Kd) of 10(-9) M. Under reduction conditions [ 125 I]iodo-insulin binds to one band with an approximate Mr of 125,000. IGF-I and insulin at micromolar concentrations, but not at nanomolar concentrations, slightly stimulated DNA synthesis, but markedly potentiated the mitogenic action of fibroblast growth factor. Adrenal cells cultured in a serum-free medium containing transferrin, ascorbic acid, and insulin (5 micrograms/ml) maintained fairly constant angiotensin-II (A-II) receptor concentration per cell and increased cAMP release on response to ACTH and their steroidogenic response to both ACTH and A-II. When the cells were cultured in the same medium without insulin, the number of A-II receptors significantly decreased to 65% and the increased responsiveness was blunted. Treatment of such cells for 3 days with increasing concentrations of IGF-I (1-100 ng/ml) produced a 2- to 3-fold increase in A-II receptors and enhanced the cAMP response (3- to 4-fold) to ACTH and the steroidogenic response (4- to 6-fold) to ACTH and A-II. These effects were time and dose dependent (ED50 approximately equal to 10(-9) M). Insulin at micromolar concentrations produced an effect similar to that of IGF-I, but at nanomolar concentrations the effect was far less

  4. [Molecular mechanism for ET-1-induced insulin resistance in skeletal muscle cells].

    Science.gov (United States)

    Horinouchi, Takahiro; Mazaki, Yuichi; Terada, Koji; Miwa, Soichi

    2018-01-01

    Insulin resistance is a condition where the sensitivity to insulin of the tissues expressing insulin receptor (InsR) is decreased due to a functional disturbance of InsR-mediated intracellular signaling. Insulin promotes the entry of glucose into the tissues and skeletal muscle is the most important tissue responsible for the insulin's action of decreasing blood glucose levels. Endothelin-1 (ET-1), a potent vasoconstrictor and pro-inflammatory peptide, induces insulin resistance through a direct action on skeletal muscle. However, the signaling pathways of ET-1-induced insulin resistance in skeletal muscle remain unclear. Here we show molecular mechanism underlying the inhibitory effect of ET-1 on insulin-stimulated Akt phosphorylation and glucose uptake in myotubes of rat L6 skeletal muscle cell line. mRNA expression levels of differentiation marker genes, MyoD and myogenin, were increased during L6 myoblasts differentiation into myotubes. Some of myotubes possessed the ability to spontaneously contract. In myotubes, insulin promoted Akt phosphorylation at Thr 308 and Ser 473 , and [ 3 H]-labelled 2-deoxy-D-glucose ([ 3 H]2-DG) uptake. The insulin-facilitated Akt phosphorylation and [ 3 H]2-DG uptake were inhibited by ET-1. The inhibitory effect of ET-1 was counteracted by blockade of ET type A receptor (ET A R), inhibition of G q/11 protein, and siRNA knockdown of G protein-coupled receptor kinase 2 (GRK2). The exogenously overexpressed GRK2 directly bound to endogenous Akt and their association was facilitated by ET-1. In summary, activation of ET A R with ET-1 inhibits insulin-induced Akt phosphorylation and [ 3 H]2-DG uptake in a G q/11 protein- and GRK2-dependent manner in skeletal muscle. These findings indicate that ET A R and GRK2 are potential targets for insulin resistance.

  5. ADAMTS13 expression in human chondrosarcoma cells induced by insulin

    Directory of Open Access Journals (Sweden)

    Rıdvan Fırat

    2014-06-01

    Full Text Available Objectives: A Disintegrin-like Metalloproteinase with Thrombospondin Motifs (ADAMTS proteins is a proteinase enzyme group that primarily located in the extracellular matrix (ECM. Insulin has been known to stimulate proteoglycan biosynthesis in chondrosarcoma chondrocytes and thereby the levels of ADAMTS proteins. The aim of this study is to evaluate the time-dependent effects of insulin on the ADAMTS13 expression in OUMS-27 human chondrosarcoma cell line to test the hypothesis that insulin diminishes ADAMTS13 expression because of its anabolic effects. Methods: To test this hypothesis OUMS-27 cells were cultured in Dulbecco’s modified Eagle’ medium (DMEM containing 10μg/mL insulin. The medium containing insulin was changed every other day up to 11th day. Cells were harvested at 1, 3, 7, and 11th days and protein and RNA isolations were performed at the proper times. The levels of RNA expression of ADAMTS13 was quantified by qRT-PCR using appropriate primers while protein levels was detected by Western blot technique using anti-ADAMTS13 antibody. Results: Although there was a decrease in both RNA and protein levels in insulin-applied groups compared to the control cells, it was not statistically significant. Conclusion: Under the light of our findings, it is suggested that insulin does not participate in regulation of ADAMTS13 in OUMS-27 chondrosarcoma cells. J Clin Exp Invest 2014; 5 (2: 226-232

  6. Reversal of hyperglycemia in mice by using human expandable insulin-producing cells differentiated from fetal liver progenitor cells

    Science.gov (United States)

    Zalzman, Michal; Gupta, Sanjeev; Giri, Ranjit K.; Berkovich, Irina; Sappal, Baljit S.; Karnieli, Ohad; Zern, Mark A.; Fleischer, Norman; Efrat, Shimon

    2003-06-01

    Beta-cell replacement is considered to be the most promising approach for treatment of type 1 diabetes. Its application on a large scale is hindered by a shortage of cells for transplantation. Activation of insulin expression, storage, and regulated secretion in stem/progenitor cells offers novel ways to overcome this shortage. We explored whether fetal human progenitor liver cells (FH) could be induced to differentiate into insulin-producing cells after expression of the pancreatic duodenal homeobox 1 (Pdx1) gene, which is a key regulator of pancreatic development and insulin expression in beta cells. FH cells possess a considerable replication capacity, and this was further extended by introduction of the gene for the catalytic subunit of human telomerase. Immortalized FH cells expressing Pdx1 activated multiple beta-cell genes, produced and stored considerable amounts of insulin, and released insulin in a regulated manner in response to glucose. When transplanted into hyperglycemic immunodeficient mice, the cells restored and maintained euglycemia for prolonged periods. Quantitation of human C-peptide in the mouse serum confirmed that the glycemia was normalized by the transplanted human cells. This approach offers the potential of a novel source of cells for transplantation into patients with type 1 diabetes.

  7. Extrinsic factors promoting insulin producing cell-differentiation and insulin expression enhancement-hope for diabetics.

    Science.gov (United States)

    Dave, Shruti

    2013-11-01

    Diabetes mellitus (DM) is considered to be an autoimmune disorder leading to destruction of beta-cells resulting in to a loss of blood sugar control. Attempts using many pharmacological compositions including exogenous insulin have failed to show tight control of glycemia and associated manifestations. Stem cells are considered a potential tool for the supply of insulin-producing cells (IPC) generation in vitro. Stem cell differentiation in to pancreatic lineages requires influence of both intrinsic and extrinsic factors. Application of islet growth factors is considered to be potential for enhancement of beta-cell replication, function and survival. Use of certain extrinsic factors is known to facilitate expression of transcription factors known to be important for beta-cell differentiation and production of insulin enabling IPC generation. Hierarchies of secreted signals and transcription factors have been identified by studies from several laboratories that guide cell differentiation in to IPC. This knowledge provides insights for in vitro IPC differentiation from stem cells. Current advancement in medical knowledge promises an insulin independency for DM patients. The review sheds light on few specific extrinsic factors which facilitate differentiation of stem cells in to IPC in vitro have been discussed; which can be proven as a potential therapeutic option for treatment of DM and associated diseases.

  8. Counter-regulatory hormone responses to spontaneous hypoglycaemia during treatment with insulin Aspart or human soluble insulin

    DEFF Research Database (Denmark)

    Brock Jacobsen, I; Vind, B F; Korsholm, Lars

    2011-01-01

    examined in a randomized, double-blinded cross-over study for two periods of 8 weeks. Sixteen patients with type 1 diabetes were subjected to three daily injections of human soluble insulin or Aspart in addition to Neutral Protamine Hagedorn (NPH) insulin twice daily. Each intervention period was followed......-regulatory responses regarding growth hormone, glucagon and ghrelin whereas no differences were found in relation to free fatty acid, cortisol, insulin-like growth factor (IGF)-I, IGF-II and IGF-binding proteins 1 and 2. Treatment with insulin Aspart resulted in well-defined peaks in serum insulin concentrations...... elicited a slightly different physiological response to spontaneous hypoglycaemia compared with human insulin. Keywords hypoglycaemia counter-regulation, insulin Aspart, type 1 diabetes....

  9. Immunocytochemical detection of glucagon and insulin cells in ...

    Indian Academy of Sciences (India)

    Madhu urs

    throughout the pancreas during the reproductive cycle, while insulin-IR cells were found to be pulsating in their secretion. Mean size ..... The interaction effect between the species .... Singh D P 1974 Analysis of environmental factors regulating.

  10. Quercetin suppresses insulin receptor signaling through inhibition of the insulin ligand–receptor binding and therefore impairs cancer cell proliferation

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Feng [Department of Gastroenterology, The Tenth People’s Hospital of Shanghai, Tongji University, Shanghai 200072 (China); Department of Nanomedicine, Houston Methodist Research Institute, Houston, TX 77030 (United States); Yang, Yong, E-mail: yyang@houstonmethodist.org [Department of Nanomedicine, Houston Methodist Research Institute, Houston, TX 77030 (United States); Department of Medicine, Weill Cornell Medical College, New York, NY 10065 (United States)

    2014-10-03

    Graphical abstract: - Highlights: • Quercetin inhibits insulin ligand–receptor interactions. • Quercetin reduces downstream insulin receptor signaling. • Quercetin blocks insulin induced glucose uptake. • Quercetin suppresses insulin stimulated cancer cell proliferation and tumor growth. - Abstract: Although the flavonoid quercetin is known to inhibit activation of insulin receptor signaling, the inhibitory mechanism is largely unknown. In this study, we demonstrate that quercetin suppresses insulin induced dimerization of the insulin receptor (IR) through interfering with ligand–receptor interactions, which reduces the phosphorylation of IR and Akt. This inhibitory effect further inhibits insulin stimulated glucose uptake due to decreased cell membrane translocation of glucose transporter 4 (GLUT4), resulting in impaired cancer cell proliferation. The effect of quercetin in inhibiting tumor growth was also evident in an in vivo model, indicating a potential future application for quercetin in the treatment of cancers.

  11. Quercetin suppresses insulin receptor signaling through inhibition of the insulin ligand–receptor binding and therefore impairs cancer cell proliferation

    International Nuclear Information System (INIS)

    Wang, Feng; Yang, Yong

    2014-01-01

    Graphical abstract: - Highlights: • Quercetin inhibits insulin ligand–receptor interactions. • Quercetin reduces downstream insulin receptor signaling. • Quercetin blocks insulin induced glucose uptake. • Quercetin suppresses insulin stimulated cancer cell proliferation and tumor growth. - Abstract: Although the flavonoid quercetin is known to inhibit activation of insulin receptor signaling, the inhibitory mechanism is largely unknown. In this study, we demonstrate that quercetin suppresses insulin induced dimerization of the insulin receptor (IR) through interfering with ligand–receptor interactions, which reduces the phosphorylation of IR and Akt. This inhibitory effect further inhibits insulin stimulated glucose uptake due to decreased cell membrane translocation of glucose transporter 4 (GLUT4), resulting in impaired cancer cell proliferation. The effect of quercetin in inhibiting tumor growth was also evident in an in vivo model, indicating a potential future application for quercetin in the treatment of cancers

  12. Transfer plate radioassay using adsorbed anti-insulin antibody to detect insulin secreted by islet cell cultures

    International Nuclear Information System (INIS)

    Scearce, R.M.; Oie, H.K.; Gazdar, A.F.; Chick, W.L.; Eisenbarth, G.S.

    1981-01-01

    A solid-phase radioimmunoassay for detection of insulin synthesized by islet cell clones is described. This assay employs anti-insulin antibody adsorbed onto fenestrated polyvinyl chloride 96-well plates ('transfer plates'). The calibrated aperture in the bottom of each transfer plate well permits fluid to enter the wells when transfer plates are lowered into microculture wells containing insulin. With this assay it is possible to rapidly screen hundreds of islet cell cultures for insulin production. The authors have used this assay to facilitate cloning of the RIN rat insulinoma cell line. The assay readily detects insulin synthesis by RIN cells and [ 125 I]insulin is not displaced by culture medium from cells which do not produce insulin. The transfer plate format should be applicable to semiautomate other radioimmunoassays. (Auth.)

  13. Common elements in interleukin 4 and insulin signaling pathways in factor-dependent hematopoietic cells.

    Science.gov (United States)

    Wang, L M; Keegan, A D; Li, W; Lienhard, G E; Pacini, S; Gutkind, J S; Myers, M G; Sun, X J; White, M F; Aaronson, S A

    1993-05-01

    Interleukin 4 (IL-4), insulin, and insulin-like growth factor I (IGF-I) efficiently induced DNA synthesis in the IL-3-dependent murine myeloid cell lines FDC-P1 and FDC-P2. Although these factors could not individually sustain long-term growth of these lines, a combination of IL-4 with either insulin or IGF-I did support continuous growth. The principal tyrosine-phosphorylated substrate observed in FDC cells stimulated with IL-4, previously designated 4PS, was of the same size (170 kDa) as the major substrate phosphorylated in response to insulin or IGF-I. These substrates had phosphopeptides of the same size when analyzed by digestion with Staphylococcus aureus V8 protease, and each tightly associated with the 85-kDa component of phosphatidylinositol 3-kinase after factor stimulation. IRS-1, the principal substrate phosphorylated in response to insulin or IGF-I stimulation in nonhematopoietic cells, is similar in size to 4PS. However, anti-IRS-1 antibodies failed to efficiently precipitate 4PS, and some phosphopeptides generated by V8 protease digestion of IRS-1 were distinct in size from the phosphopeptides of 4PS. Nevertheless, IL-4, insulin, and IGF-I were capable of stimulating tyrosine phosphorylation of IRS-1 in FDC cells that expressed this substrate as a result of transfection. These findings indicate that (i) IL-4, insulin, and IGF-I use signal transduction pathways in FDC lines that have at least one major feature in common, the rapid tyrosine phosphorylation of 4PS, and (ii) insulin and IGF-I stimulation of hematopoietic cell lines leads to the phosphorylation of a substrate that may be related to but is not identical to IRS-1.

  14. Detection of transketolase in bone marrow-derived insulin-producing cells: benfotiamine enhances insulin synthesis and glucose metabolism.

    Science.gov (United States)

    Oh, Seh-Hoon; Witek, Rafal P; Bae, Si-Hyun; Darwiche, Houda; Jung, Youngmi; Pi, Liya; Brown, Alicia; Petersen, Bryon E

    2009-01-01

    Adult bone marrow (BM)-derived insulin-producing cells (IPCs) are capable of regulating blood glucose levels in chemically induced hyperglycemic mice. Using cell transplantation therapy, fully functional BM-derived IPCs help to mediate treatment of diabetes mellitus. Here, we demonstrate the detection of the pentose phosphate pathway enzyme, transketolase (TK), in BM-derived IPCs cultured under high-glucose conditions. Benfotiamine, a known activator of TK, was not shown to affect the proliferation of insulinoma cell line, INS-1; however, when INS-1 cells were cultured with oxythiamine, an inhibitor of TK, cell proliferation was suppressed. Treatment with benfotiamine activated glucose metabolism in INS-1 cells in high-glucose culture conditions, and appeared to maximize the BM-derived IPCs ability to synthesize insulin. Benfotiamine was not shown to induce the glucose receptor Glut-2, however it was shown to activate glucokinase, the enzyme responsible for conversion of glucose to glucose-6-phosphate. Furthermore, benfotiamine-treated groups showed upregulation of the downstream glycolytic enzyme, glyceraldehyde phosphate dehydrogenase (GAPDH). However, in cells where the pentose phosphate pathway was blocked by oxythiamine treatment, there was a clear downregulation of Glut-2, glucokinase, insulin, and GAPDH. When benfotiamine was used to treat mice transplanted with BM-derived IPCs transplanted, their glucose level was brought to a normal range. The glucose challenge of normal mice treated with benfotiamine lead to rapidly normalized blood glucose levels. These results indicate that benfotiamine activates glucose metabolism and insulin synthesis to prevent glucose toxicity caused by high concentrations of blood glucose in diabetes mellitus.

  15. Detection of Transketolase in Bone Marrow—Derived Insulin-Producing Cells: Benfotiamine Enhances Insulin Synthesis and Glucose Metabolism

    Science.gov (United States)

    Witek, Rafal P.; Bae, Si-Hyun; Darwiche, Houda; Jung, Youngmi; Pi, Liya; Brown, Alicia; Petersen, Bryon E.

    2009-01-01

    Adult bone marrow (BM)-derived insulin-producing cells (IPCs) are capable of regulating blood glucose levels in chemically induced hyperglycemic mice. Using cell transplantation therapy, fully functional BM-derived IPCs help to mediate treatment of diabetes mellitus. Here, we demonstrate the detection of the pentose phosphate pathway enzyme, transketolase (TK), in BM-derived IPCs cultured under high-glucose conditions. Benfotiamine, a known activator of TK, was not shown to affect the proliferation of insulinoma cell line, INS-1; however, when INS-1 cells were cultured with oxythiamine, an inhibitor of TK, cell proliferation was suppressed. Treatment with benfotiamine activated glucose metabolism in INS-1 cells in high-glucose culture conditions, and appeared to maximize the BM-derived IPCs ability to synthesize insulin. Benfotiamine was not shown to induce the glucose receptor Glut-2, however it was shown to activate glucokinase, the enzyme responsible for conversion of glucose to glucose-6-phosphate. Furthermore, benfotiamine-treated groups showed upregulation of the downstream glycolytic enzyme, glyceraldehyde phosphate dehydrogenase (GAPDH). However, in cells where the pentose phosphate pathway was blocked by oxythiamine treatment, there was a clear downregulation of Glut-2, glucokinase, insulin, and GAPDH. When benfotiamine was used to treat mice transplanted with BM-derived IPCs transplanted, their glucose level was brought to a normal range. The glucose challenge of normal mice treated with benfotiamine lead to rapidly normalized blood glucose levels. These results indicate that benfotiamine activates glucose metabolism and insulin synthesis to prevent glucose toxicity caused by high concentrations of blood glucose in diabetes mellitus. PMID:18393672

  16. Expansion and conversion of human pancreatic ductal cells into insulin-secreting endocrine cells.

    Science.gov (United States)

    Lee, Jonghyeob; Sugiyama, Takuya; Liu, Yinghua; Wang, Jing; Gu, Xueying; Lei, Ji; Markmann, James F; Miyazaki, Satsuki; Miyazaki, Jun-Ichi; Szot, Gregory L; Bottino, Rita; Kim, Seung K

    2013-11-19

    Pancreatic islet β-cell insufficiency underlies pathogenesis of diabetes mellitus; thus, functional β-cell replacement from renewable sources is the focus of intensive worldwide effort. However, in vitro production of progeny that secrete insulin in response to physiological cues from primary human cells has proven elusive. Here we describe fractionation, expansion and conversion of primary adult human pancreatic ductal cells into progeny resembling native β-cells. FACS-sorted adult human ductal cells clonally expanded as spheres in culture, while retaining ductal characteristics. Expression of the cardinal islet developmental regulators Neurog3, MafA, Pdx1 and Pax6 converted exocrine duct cells into endocrine progeny with hallmark β-cell properties, including the ability to synthesize, process and store insulin, and secrete it in response to glucose or other depolarizing stimuli. These studies provide evidence that genetic reprogramming of expandable human pancreatic cells with defined factors may serve as a general strategy for islet replacement in diabetes. DOI: http://dx.doi.org/10.7554/eLife.00940.001.

  17. Specific insulin binding in bovine chromaffin cells; demonstration of preferential binding to adrenalin-storing cells

    International Nuclear Information System (INIS)

    Serck-Hanssen, G.; Soevik, O.

    1987-01-01

    Insulin binding was studied in subpopulations of bovine chromaffin cells enriched in adrenalin-producing cells (A-cells) or noradrenalin-producing cells (NA-cells). Binding of 125 I-insulin was carried out at 15 0 C for 3 hrs in the absence or presence of excess unlabeled hormone. Four fractions of cells were obtained by centrifugation on a stepwise bovine serum albumin gradient. The four fractions were all shown to bind insulin in a specific manner and the highest binding was measured in the cell layers of higher densities, containing mainly A-cells. The difference in binding of insulin to the four subpopulations of chromaffin cells seemed to be related to differences in numbers of receptors as opposed to receptor affinities. The authors conclude that bovine chromaffin cells possess high affinity binding sites for insulin and that these binding sites are mainly confined to A-cells. 24 references, 2 figures, 1 table

  18. Differentiation of insulin-producing cells from human neural progenitor cells.

    Directory of Open Access Journals (Sweden)

    Yuichi Hori

    2005-04-01

    Full Text Available BACKGROUND: Success in islet-transplantation-based therapies for type 1 diabetes, coupled with a worldwide shortage of transplant-ready islets, has motivated efforts to develop renewable sources of islet-replacement tissue. Islets and neurons share features, including common developmental programs, and in some species brain neurons are the principal source of systemic insulin. METHODS AND FINDINGS: Here we show that brain-derived human neural progenitor cells, exposed to a series of signals that regulate in vivo pancreatic islet development, form clusters of glucose-responsive insulin-producing cells (IPCs. During in vitro differentiation of neural progenitor cells with this novel method, genes encoding essential known in vivo regulators of pancreatic islet development were expressed. Following transplantation into immunocompromised mice, IPCs released insulin C-peptide upon glucose challenge, remained differentiated, and did not form detectable tumors. CONCLUSION: Production of IPCs solely through extracellular factor modulation in the absence of genetic manipulations may promote strategies to derive transplantable islet-replacement tissues from human neural progenitor cells and other types of multipotent human stem cells.

  19. Sodium arsenite impairs insulin secretion and transcription in pancreatic β-cells

    International Nuclear Information System (INIS)

    Diaz-Villasenor, Andrea; Sanchez-Soto, M. Carmen; Cebrian, Mariano E.; Ostrosky-Wegman, Patricia; Hiriart, Marcia

    2006-01-01

    Human studies have shown that chronic inorganic arsenic (iAs) exposure is associated with a high prevalence and incidence of type 2 diabetes. However, the mechanism(s) underlying this effect are not well understood, and practically, there is no information available on the effects of arsenic on pancreatic β-cells functions. Thus, since insulin secreted by the pancreas plays a crucial role in maintaining glucose homeostasis, our aim was to determine if sodium arsenite impairs insulin secretion and mRNA expression in single adult rat pancreatic β-cells. Cells were treated with 0.5, 1, 2, 5 and 10 μM sodium arsenite and incubated for 72 and 144 h. The highest dose tested (10 μM) decreased β-cell viability, by 33% and 83%, respectively. Insulin secretion and mRNA expression were evaluated in the presence of 1 and 5 μM sodium arsenite. Basal insulin secretion, in 5.6 mM glucose, was not significantly affected by 1 or 5 μM treatment for 72 h, but basal secretion was reduced when cells were exposed to 5 μM sodium arsenite for 144 h. On the other hand, insulin secretion in response to 15.6 mM glucose decreased with sodium arsenite in a dose-dependent manner in such a way that cells were no longer able to distinguish between different glucose concentrations. We also showed a significant decrease in insulin mRNA expression of cells exposed to 5 μM sodium arsenite during 72 h. Our data suggest that arsenic may contribute to the development of diabetes mellitus by impairing pancreatic β-cell functions, particularly insulin synthesis and secretion

  20. Somatic insulin signaling regulates a germline starvation response in Drosophila egg chambers

    Science.gov (United States)

    Burn, K. Mahala; Shimada, Yuko; Ayers, Kathleen; Lu, Feiyue; Hudson, Andrew M.; Cooley, Lynn

    2014-01-01

    Egg chambers from starved Drosophila females contain large aggregates of processing (P) bodies and cortically enriched microtubules. As this response to starvation is rapidly reversed upon re-feeding females or culturing egg chambers with exogenous bovine insulin, we examined the role of endogenous insulin signaling in mediating the starvation response. We found that systemic Drosophila insulin-like peptides (dILPs) activate the insulin pathway in follicle cells, which then regulate both microtubule and P body organization in the underlying germline cells. This organization is modulated by the motor proteins Dynein and Kinesin. Dynein activity is required for microtubule and P body organization during starvation, while Kinesin activity is required during nutrient-rich conditions. Blocking the ability of egg chambers to form P body aggregates in response to starvation correlated with reduced progeny survival. These data suggest a potential mechanism to maximize fecundity even during periods of poor nutrient availability, by mounting a protective response in immature egg chambers. PMID:25481758

  1. Investigations on the insulin receptor of isolated fat cells

    International Nuclear Information System (INIS)

    Eichler, W.

    1980-01-01

    Fat cells, isolated from the epididymal adipose tissue of rats, were incubed with iodine 125 insulin after previous incubation with various antagonists. By varying the antagonist concentration, it was possible to determine the effect these substances have on the insulin receptor, i.e. the insulin similarity. By varying the preincubation time, toxicity of the test substances could be detected, which pretended repression effects; and by finally verying the incubation time the effects on the receptor via the membrane could be distinguished from direct receptor bindings of the antagonist. (orig./MG) [de

  2. High passage MIN6 cells have impaired insulin secretion with impaired glucose and lipid oxidation.

    Directory of Open Access Journals (Sweden)

    Kim Cheng

    Full Text Available Type 2 diabetes is a metabolic disorder characterized by the inability of beta-cells to secrete enough insulin to maintain glucose homeostasis. MIN6 cells secrete insulin in response to glucose and other secretagogues, but high passage (HP MIN6 cells lose their ability to secrete insulin in response to glucose. We hypothesized that metabolism of glucose and lipids were defective in HP MIN6 cells causing impaired glucose stimulated insulin secretion (GSIS. HP MIN6 cells had no first phase and impaired second phase GSIS indicative of global functional impairment. This was coupled with a markedly reduced ATP content at basal and glucose stimulated states. Glucose uptake and oxidation were higher at basal glucose but ATP content failed to increase with glucose. HP MIN6 cells had decreased basal lipid oxidation. This was accompanied by reduced expressions of Glut1, Gck, Pfk, Srebp1c, Ucp2, Sirt3, Nampt. MIN6 cells represent an important model of beta cells which, as passage numbers increased lost first phase but retained partial second phase GSIS, similar to patients early in type 2 diabetes onset. We believe a number of gene expression changes occurred to produce this defect, with emphasis on Sirt3 and Nampt, two genes that have been implicated in maintenance of glucose homeostasis.

  3. Generation of insulin-producing cells from human bone marrow-derived mesenchymal stem cells: comparison of three differentiation protocols.

    Science.gov (United States)

    Gabr, Mahmoud M; Zakaria, Mahmoud M; Refaie, Ayman F; Khater, Sherry M; Ashamallah, Sylvia A; Ismail, Amani M; El-Badri, Nagwa; Ghoneim, Mohamed A

    2014-01-01

    Many protocols were utilized for directed differentiation of mesenchymal stem cells (MSCs) to form insulin-producing cells (IPCs). We compared the relative efficiency of three differentiation protocols. Human bone marrow-derived MSCs (HBM-MSCs) were obtained from three insulin-dependent type 2 diabetic patients. Differentiation into IPCs was carried out by three protocols: conophylline-based (one-step protocol), trichostatin-A-based (two-step protocol), and β -mercaptoethanol-based (three-step protocol). At the end of differentiation, cells were evaluated by immunolabeling for insulin production, expression of pancreatic endocrine genes, and release of insulin and c-peptide in response to increasing glucose concentrations. By immunolabeling, the proportion of generated IPCs was modest ( ≃ 3%) in all the three protocols. All relevant pancreatic endocrine genes, insulin, glucagon, and somatostatin, were expressed. There was a stepwise increase in insulin and c-peptide release in response to glucose challenge, but the released amounts were low when compared with those of pancreatic islets. The yield of functional IPCs following directed differentiation of HBM-MSCs was modest and was comparable among the three tested protocols. Protocols for directed differentiation of MSCs need further optimization in order to be clinically meaningful. To this end, addition of an extracellular matrix and/or a suitable template should be attempted.

  4. Generation of Insulin-Producing Cells from Human Bone Marrow-Derived Mesenchymal Stem Cells: Comparison of Three Differentiation Protocols

    Directory of Open Access Journals (Sweden)

    Mahmoud M. Gabr

    2014-01-01

    Full Text Available Introduction. Many protocols were utilized for directed differentiation of mesenchymal stem cells (MSCs to form insulin-producing cells (IPCs. We compared the relative efficiency of three differentiation protocols. Methods. Human bone marrow-derived MSCs (HBM-MSCs were obtained from three insulin-dependent type 2 diabetic patients. Differentiation into IPCs was carried out by three protocols: conophylline-based (one-step protocol, trichostatin-A-based (two-step protocol, and β-mercaptoethanol-based (three-step protocol. At the end of differentiation, cells were evaluated by immunolabeling for insulin production, expression of pancreatic endocrine genes, and release of insulin and c-peptide in response to increasing glucose concentrations. Results. By immunolabeling, the proportion of generated IPCs was modest (≃3% in all the three protocols. All relevant pancreatic endocrine genes, insulin, glucagon, and somatostatin, were expressed. There was a stepwise increase in insulin and c-peptide release in response to glucose challenge, but the released amounts were low when compared with those of pancreatic islets. Conclusion. The yield of functional IPCs following directed differentiation of HBM-MSCs was modest and was comparable among the three tested protocols. Protocols for directed differentiation of MSCs need further optimization in order to be clinically meaningful. To this end, addition of an extracellular matrix and/or a suitable template should be attempted.

  5. Fructose downregulates miR-330 to induce renal inflammatory response and insulin signaling impairment: Attenuation by morin.

    Science.gov (United States)

    Gu, Ting-Ting; Song, Lin; Chen, Tian-Yu; Wang, Xing; Zhao, Xiao-Juan; Ding, Xiao-Qin; Yang, Yan-Zi; Pan, Ying; Zhang, Dong-Mei; Kong, Ling-Dong

    2017-08-01

    Fructose induces insulin resistance with kidney inflammation and injury. MicroRNAs are emerged as key regulators of insulin signaling. Morin has insulin-mimetic effect with the improvement of insulin resistance and kidney injury. This study investigated the protective mechanisms of morin against fructose-induced kidney injury, with particular focus on miR-330 expression change, inflammatory response, and insulin signaling impairment. miR-330, sphingosine kinase 1 (SphK1)/sphingosine-1-phosphate (S1P)/S1P receptor (S1PR)1/3 signaling, nuclear factor-κB (NF-κB)/NOD-like receptor family, pyrin domain containing 3 (NLRP3) inflammasome, and insulin signaling were detected in kidney cortex of fructose-fed rats and fructose-exposed HK-2 cells, respectively. Whether miR-330 mediated inflammatory response to affect insulin signaling was examined using SphK1 inhibitor, S1PR1/3 short interfering RNA, or miR-330 mimic/inhibitor, respectively. Fructose was found to downregulate miR-330 expression to increase SphK1/S1P/S1PR1/3 signaling, and then activate NF-κB/NLRP3 inflammasome to produce IL-1β, causing insulin signaling impairment. Moreover, morin upregulated miR-330 and partly attenuated inflammatory response and insulin signaling impairment to alleviate kidney injury. These findings suggest that morin protects against fructose-induced kidney insulin signaling impairment by upregulating miR-330 to reduce inflammatory response. Morin may be a potential therapeutic agent for the treatment of kidney injury associated with fructose-induced inflammation and insulin signaling impairment. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  6. Vasorelaxation responses to insulin in laminar vessel rings from healthy, lean horses.

    Science.gov (United States)

    Wooldridge, A A; Waguespack, R W; Schwartz, D D; Venugopal, C S; Eades, S C; Beadle, R E

    2014-10-01

    Hyperinsulinemia causes laminitis experimentally and is a risk factor for naturally occurring laminitis. The aim of this study was to investigate the effects of insulin on laminar vascular relaxation and to induce insulin-associated vascular dysfunction in vitro. Relaxation responses of isolated laminar arterial and venous rings to acetylcholine and insulin were evaluated. To alter vascular function in response to insulin, all vessel rings were incubated with insulin or vehicle, submaximally contracted, administered insulin again and relaxation responses recorded. Laminar arteries were also incubated with the mitogen-activated protein kinase (MAPK) inhibitor, PD-98059. Relaxation in response to acetylcholine was not different between arteries and veins, but veins relaxed less in response to insulin than arteries. In arteries incubated with insulin, the subsequent relaxation response to insulin was blunted. Veins had minimal relaxation to insulin regardless of incubation. Arteries incubated with PD-98059 relaxed more in response to insulin than arteries not exposed to PD-98059, indicating that MAPK plays a role in maintenance of basal tone in laminar arteries. A differing response of laminar veins and arteries to insulin-induced relaxation may be important in understanding the link between hyperinsulinemia and laminitis. In vitro induction of vascular dysfunction in response to insulin in laminar arteries may be useful for testing therapeutic interventions and for understanding the pathophysiology of laminitis. Copyright © 2014 Elsevier Ltd. All rights reserved.

  7. miR-375 induces human decidua basalis-derived stromal cells to become insulin-producing cells.

    Science.gov (United States)

    Shaer, Anahita; Azarpira, Negar; Vahdati, Akbar; Karimi, Mohammad Hosein; Shariati, Mehrdad

    2014-09-01

    This paper focuses on the development of renewable sources of isletreplacement tissue for the treatment of type I diabetes mellitus. Placental tissue-derived mesenchymal stem cells (MSCs) are a promising source for regenerative medicine due to their plasticity and easy availability. They have the potential to differentiate into insulin-producing cells. miR-375 is a micro RNA that is expressed in the pancreas and involved in islet development. Human placental decidua basalis MSCs (PDB-MSCs) were cultured from full-term human placenta. The immunophenotype of the isolated cells was checked for CD90, CD105, CD44, CD133 and CD34 markers. The MSCs (P3) were chemically transfected with hsa-miR-375. Total RNA was extracted 4 and 6 days after transfection. The expressions of insulin, NGN3, GLUT2, PAX4, PAX6, KIR6.2, NKX6.1, PDX1, and glucagon genes were evaluated using real-time qPCR. On day 6, we tested the potency of the clusters in response to the high glucose challenge and assessed the presence of insulin and NGN3 proteins via immunocytochemistry. Flow cytometry analysis confirmed that more than 90% of the cells were positive for CD90, CD105 and CD44 and negative for CD133 and CD34. Morphological changes were followed from day 2. Cell clusters formed during day 6. Insulin-producing clusters showed a deep red color with DTZ. The expression of pancreatic-specific transcription factors increased remarkably during the four days after transfection and significantly increased on day 7. The clusters were positive for insulin and NGN3 proteins, and C-peptide and insulin secretion increased in response to changes in the glucose concentration (2.8 mM and 16.7 mM). In conclusion, the MSCs could be programmed into functional insulin-producing cells by transfection of miR-375.

  8. Intermittent Hypoxia Disrupts Glucose Homeostasis in Liver Cells in an Insulin-Dependent and Independent Manner

    Directory of Open Access Journals (Sweden)

    Chen Juan Gu

    2018-05-01

    Full Text Available Background/Aims: Obstructive sleep apnea is associated with diabetes and insulin resistance, but the underlying mechanisms remain unclear. The purpose of the current study was to determine the molecular effects of intermittent hypoxia (IH on hepatic insulin signaling and glucose homeostasis, and whether c-Jun NH2-terminal-kinase (JNK contributed to metabolic responses to IH in liver cells. Methods: The human HepG2 cells and rat FAO cells were exposed to 10, 30, 120, 240 or 360 cycles of IH (1% O2 for 60 s followed by 21% O2 for 60s, 7.5 cycles per hour or normoxia as a control. In a subgroup, we exposed cells to 360 cycles of IH with the JNK inhibitor SP600125. After IH exposure, cell glycogen content and glucose output were measured using colorimetric assay kits. Canonical insulin signaling and gluconeogenic genes were measured by western blot and quantitative polymerase chain reaction. Results: IH decreased insulin-stimulated protein kinase B (AKT/glycogen synthase kinase-3β (GSK-3β phosphorylation in a time-dependent manner, while inhibiting forkhead box protein O1 (FOXO1 expression and phosphoenolpyruvate carboxykinase (PEPCK transcription independent of insulin signaling. JNK inhibitor SP600125 partially restored AKT/ GSK-3β phosphorylation and glycogen synthesis, but did not affect other IH-induced glucose metabolic changes. Conclusion: IH in vitro impaired insulin signal transduction in liver cells as assessed by inhibited AKT/GSK-3β phosphorylation via JNK activation. IH inhibited FOXO1 and gluconeogenesis in an insulin-independent manner.

  9. The glucose-dependent insulinotropic polypeptide and glucose-stimulated insulin response to exercise training and diet in obesity

    DEFF Research Database (Denmark)

    Kelly, Karen R; Brooks, Latina M; Solomon, Thomas

    2009-01-01

    the incretin effect of GIP. The purpose of this study was to assess the effects of a 12-wk exercise training intervention (5 days/wk, 60 min/day, 75% Vo(2 max)) combined with a eucaloric (EX, n = 10) or hypocaloric (EX-HYPO, pre: 1,945 +/- 190, post: 1,269 +/- 70, kcal/day; n = 9) diet on the GIP response......Aging and obesity are characterized by decreased beta-cell sensitivity and defects in the potentiation of nutrient-stimulated insulin secretion by GIP. Exercise and diet are known to improve glucose metabolism and the pancreatic insulin response to glucose, and this effect may be mediated through...... to ingested glucose, 2) GIP may mediate the attenuated glucose-stimulated insulin response after exercise/diet interventions, and 3) the increased PYY(3-36) response represents an improved capacity to regulate satiety and potentially body weight in older, obese, insulin-resistant adults....

  10. Insulin-Producing Endocrine Cells Differentiated In Vitro From Human Embryonic Stem Cells Function in Macroencapsulation Devices In Vivo.

    Science.gov (United States)

    Agulnick, Alan D; Ambruzs, Dana M; Moorman, Mark A; Bhoumik, Anindita; Cesario, Rosemary M; Payne, Janice K; Kelly, Jonathan R; Haakmeester, Carl; Srijemac, Robert; Wilson, Alistair Z; Kerr, Justin; Frazier, Mauro A; Kroon, Evert J; D'Amour, Kevin A

    2015-10-01

    The PEC-01 cell population, differentiated from human embryonic stem cells (hESCs), contains pancreatic progenitors (PPs) that, when loaded into macroencapsulation devices (to produce the VC-01 candidate product) and transplanted into mice, can mature into glucose-responsive insulin-secreting cells and other pancreatic endocrine cells involved in glucose metabolism. We modified the protocol for making PEC-01 cells such that 73%-80% of the cell population consisted of PDX1-positive (PDX1+) and NKX6.1+ PPs. The PPs were further differentiated to islet-like cells (ICs) that reproducibly contained 73%-89% endocrine cells, of which approximately 40%-50% expressed insulin. A large fraction of these insulin-positive cells were single hormone-positive and expressed the transcription factors PDX1 and NKX6.1. To preclude a significant contribution of progenitors to the in vivo function of ICs, we used a simple enrichment process to remove remaining PPs, yielding aggregates that contained 93%-98% endocrine cells and 1%-3% progenitors. Enriched ICs, when encapsulated and implanted into mice, functioned similarly to the VC-01 candidate product, demonstrating conclusively that in vitro-produced hESC-derived insulin-producing cells can mature and function in vivo in devices. A scaled version of our suspension culture was used, and the endocrine aggregates could be cryopreserved and retain functionality. Although ICs expressed multiple important β cell genes, the cells contained relatively low levels of several maturity-associated markers. Correlating with this, the time to function of ICs was similar to PEC-01 cells, indicating that ICs required cell-autonomous maturation after delivery in vivo, which would occur concurrently with graft integration into the host. Type 1 diabetes (T1D) affects approximately 1.25 million people in the U.S. alone and is deadly if not managed with insulin injections. This paper describes the production of insulin-producing cells in vitro and a new

  11. Hydrogen peroxide production regulates the mitochondrial function in insulin resistant muscle cells: effect of catalase overexpression.

    Science.gov (United States)

    Barbosa, Marina R; Sampaio, Igor H; Teodoro, Bruno G; Sousa, Thais A; Zoppi, Claudio C; Queiroz, André L; Passos, Madla A; Alberici, Luciane C; Teixeira, Felipe R; Manfiolli, Adriana O; Batista, Thiago M; Cappelli, Ana Paula Gameiro; Reis, Rosana I; Frasson, Danúbia; Kettelhut, Isis C; Parreiras-e-Silva, Lucas T; Costa-Neto, Claudio M; Carneiro, Everardo M; Curi, Rui; Silveira, Leonardo R

    2013-10-01

    The mitochondrial redox state plays a central role in the link between mitochondrial overloading and insulin resistance. However, the mechanism by which the ROS induce insulin resistance in skeletal muscle cells is not completely understood. We examined the association between mitochondrial function and H2O2 production in insulin resistant cells. Our hypothesis is that the low mitochondrial oxygen consumption leads to elevated ROS production by a mechanism associated with reduced PGC1α transcription and low content of phosphorylated CREB. The cells were transfected with either the encoded sequence for catalase overexpression or the specific siRNA for catalase inhibition. After transfection, myotubes were incubated with palmitic acid (500μM) and the insulin response, as well as mitochondrial function and fatty acid metabolism, was determined. The low mitochondrial oxygen consumption led to elevated ROS production by a mechanism associated with β-oxidation of fatty acids. Rotenone was observed to reduce the ratio of ROS production. The elevated H2O2 production markedly decreased the PGC1α transcription, an effect that was accompanied by a reduced phosphorylation of Akt and CREB. The catalase transfection prevented the reduction in the phosphorylated level of Akt and upregulated the levels of phosphorylated CREB. The mitochondrial function was elevated and H2O2 production reduced, thus increasing the insulin sensitivity. The catalase overexpression improved mitochondrial respiration protecting the cells from fatty acid-induced, insulin resistance. This effect indicates that control of hydrogen peroxide production regulates the mitochondrial respiration preventing the insulin resistance in skeletal muscle cells by a mechanism associated with CREB phosphorylation and β-oxidation of fatty acids. Copyright © 2013 Elsevier B.V. All rights reserved.

  12. Changes in phosphatidylcholine fatty acid composition are associated with altered skeletal muscle insulin responsiveness in normal man.

    Science.gov (United States)

    Clore, J N; Harris, P A; Li, J; Azzam, A; Gill, R; Zuelzer, W; Rizzo, W B; Blackard, W G

    2000-02-01

    The fatty acid composition of skeletal muscle cell membrane phospholipids (PLs) is known to influence insulin responsiveness in man. We have recently shown that the fatty acid composition of phosphatidylcholine (PC), and not phosphatidylethanolamine (PE), from skeletal muscle membranes is of particular importance in this relationship. Efforts to alter the PL fatty acid composition in animal models have demonstrated induction of insulin resistance. However, it has been more difficult to determine if changes in insulin sensitivity are associated with changes in the skeletal muscle membrane fatty acid composition of PL in man. Using nicotinic acid (NA), an agent known to induce insulin resistance in man, 9 normal subjects were studied before and after treatment for 1 month. Skeletal muscle membrane fatty acid composition of PC and PE from biopsies of vastus lateralis was correlated with insulin responsiveness using a 3-step hyperinsulinemic-euglycemic clamp. Treatment with NA was associated with a 25% increase in the half-maximal insulin concentration ([ED50] 52.0 +/- 7.5 to 64.6 +/- 9.0 microU/mL, P insulin sensitivity. Significant changes in the fatty acid composition of PC, but not PE, were also observed after NA administration. An increase in the percentage of 16:0 (21% +/- 0.3% to 21.7% +/- 0.4%, P insulin resistance with NA is associated with changes in the fatty acid composition of PC in man.

  13. Co-culture of clonal beta cells with GLP-1 and glucagon-secreting cell line impacts on beta cell insulin secretion, proliferation and susceptibility to cytotoxins.

    Science.gov (United States)

    Green, Alastair D; Vasu, Srividya; Moffett, R Charlotte; Flatt, Peter R

    2016-06-01

    We investigated the direct effects on insulin releasing MIN6 cells of chronic exposure to GLP-1, glucagon or a combination of both peptides secreted from GLUTag L-cell and αTC1.9 alpha-cell lines in co-culture. MIN6, GLUTag and αTC1.9 cell lines exhibited high cellular hormone content and release of insulin, GLP-1 and glucagon, respectively. Co-culture of MIN6 cells with GLUTag cells significantly increased cellular insulin content, beta-cell proliferation, insulin secretory responses to a range of established secretogogues and afforded protection against exposure cytotoxic concentrations of glucose, lipid, streptozotocin or cytokines. Benefits of co-culture of MIN6 cells with αTC1.9 alphacells were limited to enhanced beta-cell proliferation with marginal positive actions on both insulin secretion and cellular protection. In contrast, co-culture of MIN6 with GLUTag cells plus αTC1.9 cells, markedly enhanced both insulin secretory responses and protection against beta-cell toxins compared with co-culture with GLUTag cells alone. These data indicate important long-term effects of conjoint GLP-1 and glucagon exposure on beta-cell function. This illustrates the possible functional significance of alpha-cell GLP-1 production as well as direct beneficial effects of dual agonism at beta-cell GLP-1 and glucagon receptors. Copyright © 2016 Elsevier B.V. and Société française de biochimie et biologie Moléculaire (SFBBM). All rights reserved.

  14. Increased CD19+CD24+CD27+ B regulatory cells are associated with insulin resistance in patients with type I Hashimoto's thyroiditis.

    Science.gov (United States)

    Yang, Min; Du, Changji; Wang, Yinping; Liu, Jun

    2017-06-01

    Hashimoto's thyroiditis (HT) is characterized by dysregulated immune responses and is commonly associated with insulin resistance. However, the mechanism of insulin resistance in HT remains to be fully elucidated. The aim of the present study was to investigate the correlation between the percentage of B regulatory lymphocytes (Bregs) and insulin resistance in patients with HT but with normal thyroid function (type I). A total of 59 patients with type I HT and 38 healthy volunteers were enrolled in the study. An oral glucose tolerance test was performed to measure insulin secretion and assess β‑cell functions. Flow cytometry was performed to examine the percentages of lymphocyte populations. The patients with HT exhibited normal fasting and postprandial glucose and fasting insulin secretion, but increased secretion of early‑phase and total insulin. The patients with HT also had insufficient β‑cell compensation for insulin resistance, indicated by a reduced disposition index, in the fasting state. An elevation in the percentage of CD19+CD24+CD27+ Bregs was also observed, which correlated positively with insulin secretion and insulin resistance in the fasting state. The patients with type I HT had postprandial insulin resistance and insufficient β‑cell compensation for fasting insulin resistance. Therefore, the increase in CD19+CD24+CD27+ Bregs was closely associated with fasting insulin secretion. These results provide novel insight into the mechanism of insulin resistance in HT.

  15. Potential role of insulin signaling on vascular smooth muscle cell migration, proliferation, and inflammation pathways.

    Science.gov (United States)

    Cersosimo, Eugenio; Xu, Xiaojing; Musi, Nicolas

    2012-02-15

    To investigate the role of insulin signaling pathways in migration, proliferation, and inflammation of vascular smooth muscle cells (VSMCs), we examined the expression of active components of the phosphatidyl inositol 3 (PI-3) kinase (p-Akt) and mitogen-activated protein kinase (MAPK) (p-Erk) in primary cultures of VSMCs from human coronary arteries. VSMCs were treated in a dose-response manner with insulin (0, 1, 10, and 100 nM) for 20 min, and Akt and Erk phosphorylation were measured by Western blot analysis. In separate experiments, we evaluated the effect of 200 μM palmitate, in the presence and absence of 8 μM pioglitazone, on insulin-stimulated (100 nM for 20 min) Akt and Erk phosphorylation. The phosphorylation of Akt and Erk in VSMCs exhibited a dose dependency with a three- to fourfold increase, respectively, at the highest dose (100 nM). In the presence of palmitate, insulin-induced Akt phosphorylation was completely abolished, and there was a threefold increase in p-Erk. With addition of pioglitazone, the phosphorylation of Akt by insulin remained unchanged, whereas insulin-stimulated Erk phosphorylation was reduced by pioglitazone. These data in VSMCs indicate that high palmitate decreases insulin-stimulated Akt phosphorylation and stimulates MAPK, whereas preexposure peroxisome proliferator-activated receptor-γ agonist pioglitazone preserves Akt phosphorylation and simultaneously attenuates MAPK signaling. Our results suggest that metabolic and mitogenic insulin signals have different sensitivity, are independently regulated, and may play a role in arterial smooth muscle cells migration, proliferation, and inflammation in conditions of acute hyperinsulinemia.

  16. p53- and ERK7-dependent ribosome surveillance response regulates Drosophila insulin-like peptide secretion.

    Directory of Open Access Journals (Sweden)

    Kiran Hasygar

    2014-11-01

    Full Text Available Insulin-like signalling is a conserved mechanism that coordinates animal growth and metabolism with nutrient status. In Drosophila, insulin-producing median neurosecretory cells (IPCs regulate larval growth by secreting insulin-like peptides (dILPs in a diet-dependent manner. Previous studies have shown that nutrition affects dILP secretion through humoral signals derived from the fat body. Here we uncover a novel mechanism that operates cell autonomously in the IPCs to regulate dILP secretion. We observed that impairment of ribosome biogenesis specifically in the IPCs strongly inhibits dILP secretion, which consequently leads to reduced body size and a delay in larval development. This response is dependent on p53, a known surveillance factor for ribosome biogenesis. A downstream effector of this growth inhibitory response is an atypical MAP kinase ERK7 (ERK8/MAPK15, which is upregulated in the IPCs following impaired ribosome biogenesis as well as starvation. We show that ERK7 is sufficient and essential to inhibit dILP secretion upon impaired ribosome biogenesis, and it acts epistatically to p53. Moreover, we provide evidence that p53 and ERK7 contribute to the inhibition of dILP secretion upon starvation. Thus, we conclude that a cell autonomous ribosome surveillance response, which leads to upregulation of ERK7, inhibits dILP secretion to impede tissue growth under limiting dietary conditions.

  17. Possible contribution of taurine to distorted glucagon secretion in intra-islet insulin deficiency: a metabolome analysis using a novel α-cell model of insulin-deficient diabetes.

    Directory of Open Access Journals (Sweden)

    Megumi Bessho

    Full Text Available Glycemic instability is a serious problem in patients with insulin-deficient diabetes, and it may be due in part to abnormal endogenous glucagon secretion. However, the intracellular metabolic mechanism(s involved in the aberrant glucagon response under the condition of insulin deficiency has not yet been elucidated. To investigate the metabolic traits that underlie the distortion of glucagon secretion under insulin deficient conditions, we generated an αTC1-6 cell line with stable knockdown of the insulin receptor (IRKD, i.e., an in vitro α-cell model for insulin-deficient diabetes, which exhibits an abnormal glucagon response to glucose. A comprehensive metabolomic analysis of the IRKD αTC1-6 cells (IRKD cells revealed some candidate metabolites whose levels differed markedly compared to those in control αTC1-6 cells, but also which could affect the glucagon release in IRKD cells. Of these candidates, taurine was remarkably increased in the IRKD cells and was identified as a stimulator of glucagon in αTC1-6 cells. Taurine also paradoxically exaggerated the glucagon secretion at a high glucose concentration in IRKD cells and islets with IRKD. These results indicate that the metabolic alterations induced by IRKD in α-cells, especially the increase of taurine, may lead to the distorted glucagon response in IRKD cells, suggesting the importance of taurine in the paradoxical glucagon response and the resultant glucose instability in insulin-deficient diabetes.

  18. Cutaneous microvascular perfusion responses to insulin iontophoresis are differentially affected by insulin resistance after spinal cord injury.

    Science.gov (United States)

    La Fountaine, Michael F; Cirnigliaro, Christopher M; Azarelo, Frank; Hobson, Joshua C; Tascione, Oriana; Swonger, Kirsten N; Dyson-Hudson, Trevor; Bauman, William A

    2017-09-01

    What is the central question of this study? What impact does insulin resistance have on cutaneous perfusion responses to insulin iontophoresis in vascular beds with markedly reduced or functionally ablated sympathetic nervous system vasomotor function resulting from spinal cord injury? What is the main finding and its importance? Persons with spinal cord injury have sublesional microvascular endothelial dysfunction, as indicated by a blunted cutaneous perfusion response to acetylcholine iontophoresis, and the presence of insulin resistance has a further confounding effect on endothelium-mediated changes to cutaneous perfusion in the lower extremities. Endothelium-mediated mechanisms that regulate skin blood flow might play an integral role in optimizing skin perfusion in vascular beds with sympathetic nervous system vasomotor impairment, such as in spinal cord injury (SCI). Insulin is a vasoactive hormone and second messenger of nitric oxide that facilitates endothelium-mediated dilatation. The effects of insulin resistance (IR) on sublesional cutaneous perfusion responses to insulin provocation have yet to be described in persons with SCI. Persons with SCI and an able-bodied (AB) cohort were divided into subgroups based upon fasting plasma insulin concentration cut-offs for IR (≥13.13 mIU ml -1 ) or insulin sensitivity (IS; insulin, acetylcholine or placebo iontophoresis in the lower extremities; BPU responses were log 10 transformed to facilitate comparisons, and the net insulin response (NetIns) BPU response was calculated (insulin minus placebo BPU response). The NetIns was significantly greater in both IS groups compared with their corresponding IR group. The acetylcholine-mediated BPU responses in the SCI subgroups were significantly lower than those in the ABIS group. The proportional BPU responses of NetIns to acetylcholine in the IS cohorts (i.e. ABIS and SCIS) were significantly greater (P < 0.05) than that of each IR subgroup. The presence of IR

  19. Insulin-like growth factors and pancreas beta cells.

    NARCIS (Netherlands)

    Haeften, T.W. van; Twickler, M.

    2004-01-01

    Abstract Insulin-like growth factors (IGFs) have been implicated in normal growth, and especially foetal pancreas beta-cell development. As low birth weight has been implicated in the development of obesity and type 2 diabetes, much research has evolved into the importance of IGF and their

  20. Insulin-like growth factors and pancreas beta cells

    NARCIS (Netherlands)

    van Haeften, T. W.; Twickler, TB

    Insulin-like growth factors (IGFs) have been implicated in normal growth, and especially foetal pancreas beta-cell development. As low birth weight has been implicated in the development of obesity and type 2 diabetes, much research has evolved into the importance of IGF and their signalling

  1. Insulin-like growth factors and pancreas beta cells

    NARCIS (Netherlands)

    van Haeften, T. W.; Twickler, Th B.

    2004-01-01

    Abstract Insulin-like growth factors (IGFs) have been implicated in normal growth, and especially foetal pancreas beta-cell development. As low birth weight has been implicated in the development of obesity and type 2 diabetes, much research has evolved into the importance of IGF and their

  2. Rational Design of Glucose-Responsive Insulin Using Pharmacokinetic Modeling.

    Science.gov (United States)

    Bakh, Naveed A; Bisker, Gili; Lee, Michael A; Gong, Xun; Strano, Michael S

    2017-11-01

    A glucose responsive insulin (GRI) is a therapeutic that modulates its potency, concentration, or dosing of insulin in relation to a patient's dynamic glucose concentration, thereby approximating aspects of a normally functioning pancreas. Current GRI design lacks a theoretical basis on which to base fundamental design parameters such as glucose reactivity, dissociation constant or potency, and in vivo efficacy. In this work, an approach to mathematically model the relevant parameter space for effective GRIs is induced, and design rules for linking GRI performance to therapeutic benefit are developed. Well-developed pharmacokinetic models of human glucose and insulin metabolism coupled to a kinetic model representation of a freely circulating GRI are used to determine the desired kinetic parameters and dosing for optimal glycemic control. The model examines a subcutaneous dose of GRI with kinetic parameters in an optimal range that results in successful glycemic control within prescribed constraints over a 24 h period. Additionally, it is demonstrated that the modeling approach can find GRI parameters that enable stable glucose levels that persist through a skipped meal. The results provide a framework for exploring the parameter space of GRIs, potentially without extensive, iterative in vivo animal testing. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  3. Aging and insulin signaling differentially control normal and tumorous germline stem cells.

    Science.gov (United States)

    Kao, Shih-Han; Tseng, Chen-Yuan; Wan, Chih-Ling; Su, Yu-Han; Hsieh, Chang-Che; Pi, Haiwei; Hsu, Hwei-Jan

    2015-02-01

    Aging influences stem cells, but the processes involved remain unclear. Insulin signaling, which controls cellular nutrient sensing and organismal aging, regulates the G2 phase of Drosophila female germ line stem cell (GSC) division cycle in response to diet; furthermore, this signaling pathway is attenuated with age. The role of insulin signaling in GSCs as organisms age, however, is also unclear. Here, we report that aging results in the accumulation of tumorous GSCs, accompanied by a decline in GSC number and proliferation rate. Intriguingly, GSC loss with age is hastened by either accelerating (through eliminating expression of Myt1, a cell cycle inhibitory regulator) or delaying (through mutation of insulin receptor (dinR) GSC division, implying that disrupted cell cycle progression and insulin signaling contribute to age-dependent GSC loss. As flies age, DNA damage accumulates in GSCs, and the S phase of the GSC cell cycle is prolonged. In addition, GSC tumors (which escape the normal stem cell regulatory microenvironment, known as the niche) still respond to aging in a similar manner to normal GSCs, suggesting that niche signals are not required for GSCs to sense or respond to aging. Finally, we show that GSCs from mated and unmated females behave similarly, indicating that female GSC-male communication does not affect GSCs with age. Our results indicate the differential effects of aging and diet mediated by insulin signaling on the stem cell division cycle, highlight the complexity of the regulation of stem cell aging, and describe a link between ovarian cancer and aging. © 2014 The Authors. Aging Cell published by the Anatomical Society and John Wiley & Sons Ltd.

  4. Insulin

    Science.gov (United States)

    ... For Consumers Home For Consumers Consumer Information by Audience For Women Women's Health Topics Insulin Share Tweet ... I start having side effects? What is my target blood sugar level? How often should I check ...

  5. Fucosterol activates the insulin signaling pathway in insulin resistant HepG2 cells via inhibiting PTP1B.

    Science.gov (United States)

    Jung, Hyun Ah; Bhakta, Himanshu Kumar; Min, Byung-Sun; Choi, Jae Sue

    2016-10-01

    Insulin resistance is a characteristic feature of type 2 diabetes mellitus (T2DM) and is characterized by defects in insulin signaling. This study investigated the modulatory effects of fucosterol on the insulin signaling pathway in insulin-resistant HepG2 cells by inhibiting protein tyrosine phosphatase 1B (PTP1B). In addition, molecular docking simulation studies were performed to predict binding energies, the specific binding site of fucosterol to PTP1B, and to identify interacting residues using Autodock 4.2 software. Glucose uptake was determined using a fluorescent D-glucose analogue and the glucose tracer 2-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl) amino]-2-deoxyglucose, and the signaling pathway was detected by Western blot analysis. We found that fucosterol enhanced insulin-provoked glucose uptake and conjointly decreased PTP1B expression level in insulin-resistant HepG2 cells. Moreover, fucosterol significantly reduced insulin-stimulated serine (Ser307) phosphorylation of insulin receptor substrate 1 (IRS1) and increased phosphorylation of Akt, phosphatidylinositol-3-kinase, and extracellular signal- regulated kinase 1 at concentrations of 12.5, 25, and 50 µM in insulin-resistant HepG2 cells. Fucosterol inhibited caspase-3 activation and nuclear factor kappa B in insulin-resistant hepatocytes. These results suggest that fucosterol stimulates glucose uptake and improves insulin resistance by downregulating expression of PTP1B and activating the insulin signaling pathway. Thus, fucosterol has potential for development as an anti-diabetic agent.

  6. Relation between the insulin receptor number in cells, autophosphorylation and insulin-stimulated Ras.GTP formation

    NARCIS (Netherlands)

    Osterop, A.P.R.M.; Medema, R.H.; Bos, J.L.; Zon, G.C.M. van der; Moller, D.E.; Flier, J.S.; Möller, W.; Maassen, J.A.

    1992-01-01

    We showed previously that upon insulin stimulation of an insulin receptor overexpressing cell linme,o st of the p2lras warsa pidly converted into the GTP bound state (Burgering, B. M. T., Medema, R. H., Maassen, J. A., Van de Wetering, M. L., Van der Eb, A. J., McCormick, F., and Bos, J. L.

  7. Acute insulin resistance stimulates and insulin sensitization attenuates vascular smooth muscle cell migration and proliferation.

    Science.gov (United States)

    Cersosimo, Eugenio; Xu, Xiaojing; Upala, Sikarin; Triplitt, Curtis; Musi, Nicolas

    2014-08-01

    Differential activation/deactivation of insulin signaling, PI-3K and MAP-K pathways by high glucose and palmitate, with/out the insulin sensitizer pioglitazone (PIO), have been previously shown in vascular smooth muscle cells (VSMCs). To determine the biological impact of these molecular changes, we examined VSMC migration and proliferation ("M"&"P") patterns in similar conditions. VSMCs from healthy human coronary arteries were incubated in growth medium and "M"&"P" were analyzed after exposure to high glucose (25 mmol/L) ± palmitate (200 μmol/L) and ± PIO (8 μmol/L) for 5 h. "M"&"P" were assessed by: (1) polycarbonate membrane barrier with chemo-attractants and extended cell protrusions quantified by optical density (OD595 nm); (2) % change in radius area (2D Assay) using inverted microscopy images; and (3) cell viability assay expressed as cell absorbance (ABS) in media. "M" in 25 mmol/L glucose media increased by ~25% from baseline and % change in radius area rose from ~20% to ~30%. The addition of PIO was accompanied by a significant decrease in "M" from 0.25 ± 0.02 to 0.19 ± 0.02; a comparable decline from 0.25 ± 0.02 to 0.18 ± 0.02 was also seen with 25 mmol/L of glucose +200 μmol/L of palmitate. When PIO was coincubated with high glucose plus palmitate there was a 50% reduction in % change in radius. A ~10% increase in ABS, reflecting augmented "P" in media with 25 mmol/L glucose versus control was documented. The addition of PIO reduced ABS from 0.208 ± 0.03 to 0.183 ± 0.06. Both high glucose and palmitate showed ABS of ~0.140 ± 0.02, which decreased with PIO to ~0.120 ± 0.02, indicating "P" was reduced. These results confirm that high glucose and palmitate stimulate VSMCs migration and proliferation in vitro, which is attenuated by coincubation with the insulin sensitizer PIO. Although, we cannot ascertain whether these functional changes are coincident with the activation/deactivation of signal molecules, our findings are consistent with the

  8. Alteration in insulin action

    DEFF Research Database (Denmark)

    Tanti, J F; Gual, P; Grémeaux, T

    2004-01-01

    Insulin resistance, when combined with impaired insulin secretion, contributes to the development of type 2 diabetes. Insulin resistance is characterised by a decrease in insulin effect on glucose transport in muscle and adipose tIssue. Tyrosine phosphorylation of insulin receptor substrate 1 (IRS......-1) and its binding to phosphatidylinositol 3-kinase (PI 3-kinase) are critical events in the insulin signalling cascade leading to insulin-stimulated glucose transport. Modification of IRS-1 by serine phosphorylation could be one of the mechanisms leading to a decrease in IRS-1 tyrosine...... to phosphorylate these serine residues have been identified. These exciting results suggest that serine phosphorylation of IRS-1 is a possible hallmark of insulin resistance in biologically insulin responsive cells or tIssues. Identifying the pathways by which "diabetogenic" factors activate IRS-1 kinases...

  9. Mechanical stretch augments insulin-induced vascular smooth muscle cell proliferation by insulin-like growth factor-1 receptor

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Gang [Department of Pharmacology, Faculty of Medicine, Kagawa University, Kagawa (Japan); Department of Anesthesiology, First Affiliated Hospital of China Medical University, Shenyang (China); Hitomi, Hirofumi, E-mail: hitomi@kms.ac.jp [Department of Pharmacology, Faculty of Medicine, Kagawa University, Kagawa (Japan); Hosomi, Naohisa [Department of Cardiorenal and Cerebrovascular Medicine, Faculty of Medicine, Kagawa University, Kagawa (Japan); Lei, Bai; Nakano, Daisuke [Department of Pharmacology, Faculty of Medicine, Kagawa University, Kagawa (Japan); Deguchi, Kazushi; Mori, Hirohito; Masaki, Tsutomu [Department of Gastroenterology and Neurology, Faculty of Medicine, Kagawa University, Kagawa (Japan); Ma, Hong [Department of Anesthesiology, First Affiliated Hospital of China Medical University, Shenyang (China); Griendling, Kathy K. [Department of Medicine, Division of Cardiology, Emory University School of Medicine, Atlanta, GA (United States); Nishiyama, Akira [Department of Pharmacology, Faculty of Medicine, Kagawa University, Kagawa (Japan)

    2011-10-15

    Insulin resistance and hypertension have been implicated in the pathogenesis of cardiovascular disease; however, little is known about the roles of insulin and mechanical force in vascular smooth muscle cell (VSMC) remodeling. We investigated the contribution of mechanical stretch to insulin-induced VSMC proliferation. Thymidine incorporation was stimulated by insulin in stretched VSMCs, but not in un-stretched VSMCs. Insulin increased 2-deoxy-glucose incorporation in both stretched and un-stretched VSMCs. Mechanical stretch augmented insulin-induced extracellular signal-regulated kinase (ERK) and Akt phosphorylation. Inhibitors of epidermal growth factor (EGF) receptor tyrosine kinase and Src attenuated insulin-induced ERK and Akt phosphorylation, as well as thymidine incorporation, whereas 2-deoxy-glucose incorporation was not affected by these inhibitors. Moreover, stretch augmented insulin-like growth factor (IGF)-1 receptor expression, although it did not alter the expression of insulin receptor and insulin receptor substrate-1. Insulin-induced ERK and Akt activation, and thymidine incorporation were inhibited by siRNA for the IGF-1 receptor. Mechanical stretch augments insulin-induced VSMC proliferation via upregulation of IGF-1 receptor, and downstream Src/EGF receptor-mediated ERK and Akt activation. Similar to in vitro experiment, IGF-1 receptor expression was also augmented in hypertensive rats. These results provide a basis for clarifying the molecular mechanisms of vascular remodeling in hypertensive patients with hyperinsulinemia. -- Highlights: {yields} Mechanical stretch augments insulin-induced VSMC proliferation via IGF-1 receptor. {yields} Src/EGFR-mediated ERK and Akt phosphorylation are augmented in stretched VSMCs. {yields} Similar to in vitro experiment, IGF-1 receptor is increased in hypertensive rats. {yields} Results provide possible mechanisms of vascular remodeling in hypertension with DM.

  10. Atherosclerosis is a vascular stem cell disease caused by insulin.

    Science.gov (United States)

    Traunmüller, Friederike

    2018-07-01

    The present article proposes the hypothesis that when multipotent vascular stem cells are exposed to excessive insulin in a rhythmic pattern of sharply rising and falling concentrations, their differentiation is misdirected toward adipogenic and osteogenic cell lineages. This results in plaque-like accumulation of adipocytes with fat and cholesterol deposition from adipocyte debris, and osteogenic (progenitor) cells with a calcified matrix in advanced lesions. The ingrowth of capillaries and infiltration with macrophages, which upon uptake of lipids turn into foam cells, are unspecific pro-resolving reactions. Epidemiological, histopathological, pharmacological, and experimental evidence in favour of this hypothesis is summarised. Copyright © 2018. Published by Elsevier Ltd.

  11. The zinc transporter ZNT3 co-localizes with insulin in INS-1E pancreatic beta cells and influences cell survival, insulin secretion capacity, and ZNT8 expression

    DEFF Research Database (Denmark)

    Smidt, Kamille; Larsen, Agnete; Brønden, Andreas

    2016-01-01

    Zinc trafficking in pancreatic beta cells is tightly regulated by zinc transporting (ZNTs) proteins. The role of different ZNTs in the beta cells is currently being clarified. ZNT8 transports zinc into insulin granules and is critical for a correct insulin crystallization and storage in the granu......Zinc trafficking in pancreatic beta cells is tightly regulated by zinc transporting (ZNTs) proteins. The role of different ZNTs in the beta cells is currently being clarified. ZNT8 transports zinc into insulin granules and is critical for a correct insulin crystallization and storage...

  12. Glucose-responsive insulin by molecular and physical design

    Science.gov (United States)

    Bakh, Naveed A.; Cortinas, Abel B.; Weiss, Michael A.; Langer, Robert S.; Anderson, Daniel G.; Gu, Zhen; Dutta, Sanjoy; Strano, Michael S.

    2017-10-01

    The concept of a glucose-responsive insulin (GRI) has been a recent objective of diabetes technology. The idea behind the GRI is to create a therapeutic that modulates its potency, concentration or dosing relative to a patient's dynamic glucose concentration, thereby approximating aspects of a normally functioning pancreas. From the perspective of the medicinal chemist, the GRI is also important as a generalized model of a potentially new generation of therapeutics that adjust potency in response to a critical therapeutic marker. The aim of this Perspective is to highlight emerging concepts, including mathematical modelling and the molecular engineering of insulin itself and its potency, towards a viable GRI. We briefly outline some of the most important recent progress toward this goal and also provide a forward-looking viewpoint, which asks if there are new approaches that could spur innovation in this area as well as to encourage synthetic chemists and chemical engineers to address the challenges and promises offered by this therapeutic approach.

  13. Glycogen Synthase Kinase-3 regulates IGFBP-1 gene transcription through the Thymine-rich Insulin Response Element

    Directory of Open Access Journals (Sweden)

    Marquez Rodolfo

    2004-09-01

    Full Text Available Abstract Background Hepatic expression of several gene products involved in glucose metabolism, including phosphoenolpyruvate carboxykinase (PEPCK, glucose-6-phosphatase (G6Pase and insulin-like growth factor binding protein-1 (IGFBP-1, is rapidly and completely inhibited by insulin. This inhibition is mediated through the regulation of a DNA element present in each of these gene promoters, that we call the Thymine-rich Insulin Response Element (TIRE. The insulin signalling pathway that results in the inhibition of these gene promoters requires the activation of phosphatidylinositol 3-kinase (PI 3-kinase. However, the molecules that connect PI 3-kinase to these gene promoters are not yet fully defined. Glycogen Synthase Kinase 3 (GSK-3 is inhibited following activation of PI 3-kinase. We have shown previously that inhibitors of GSK-3 reduce the activity of two TIRE-containing gene promoters (PEPCK and G6Pase, whose products are required for gluconeogenesis. Results In this report we demonstrate that in H4IIE-C3 cells, four distinct classes of GSK-3 inhibitor mimic the effect of insulin on a third TIRE-containing gene, IGFBP-1. We identify the TIRE as the minimum requirement for inhibition by these agents, and demonstrate that the target of GSK-3 is unlikely to be the postulated TIRE-binding protein FOXO-1. Importantly, overexpression of GSK-3 in cells reduces the insulin regulation of TIRE activity as well as endogenous IGFBP-1 expression. Conclusions These results implicate GSK-3 as an intermediate in the pathway from the insulin receptor to the TIRE. Indeed, this is the first demonstration of an absolute requirement for GSK-3 inhibition in insulin regulation of gene transcription. These data support the potential use of GSK-3 inhibitors in the treatment of insulin resistant states such as Type 2 diabetes mellitus, but suggest that it will be important to identify all TIRE-containing genes to assess potential side effects of these agents.

  14. Nutrient partitioning and response to insulin challenge at different ...

    African Journals Online (AJOL)

    Unknown

    concentrations were depressed to a smaller extent by insulin in the Saanen crossbred than ... of endogenous body reserves mediated by differences in insulin sensitivity. ..... Adaptations of glucose metabolism during pregnancy and lactation.

  15. Incretin hormone and insulin responses to oral versus intravenous lipid administration in humans

    DEFF Research Database (Denmark)

    Lindgren, Ola; Carr, Richard D; Deacon, Carolyn F

    2011-01-01

    Context: The incretin effect is responsible for the higher insulin response to oral glucose than to iv glucose at matching glucose levels. It is notknownwhetherthis effect is restricted to glucose only. Objective: The aim of the study was to examine whether insulin and incretin hormone responses ...

  16. A Study on the Glucose and Immunoreactive Insulin Response during Oral Glucose Tolerance Test in Patients with Chronic Liver Diseases

    International Nuclear Information System (INIS)

    Choe, Kang Won; Lee, Hong Kyu; Koh, Chang Soon; Lee, Mu Ho

    1973-01-01

    The blood glucose and plasma immunoreactive insulin (IRI) levels were measured during aral glucose tolerance test in 7 healthy subjects and 6 patients with chronic liver diseases. The glucose tolerance was impaired in 5 of the 6 patients and normal in I. Plasma IRI responses were markedly increased and delayed in all patients, suggesting endogenous insulin resistance. Patients with more glucose intolerance showed less increase in plasma IRI than the group with less intolerance. lt is suggested that some insulin antagonists may decrease the peripheral insulin sensitivity and stimulate compensatory hyperactivity of pancreatic islets. If the compensatory hyperactivity is inadequate due to gemetic predisposition to diabetes mellitus or exhaustion of β-cells of pancreatic islets, the glucose intolerance and overt diabetes mellitus may ensue.

  17. A Study on the Glucose and Immunoreactive Insulin Response during Oral Glucose Tolerance Test in Patients with Chronic Liver Diseases

    Energy Technology Data Exchange (ETDEWEB)

    Choe, Kang Won; Lee, Hong Kyu; Koh, Chang Soon; Lee, Mu Ho [Seoul National University College of Medicine, Seoul (Korea, Republic of)

    1973-03-15

    The blood glucose and plasma immunoreactive insulin (IRI) levels were measured during aral glucose tolerance test in 7 healthy subjects and 6 patients with chronic liver diseases. The glucose tolerance was impaired in 5 of the 6 patients and normal in I. Plasma IRI responses were markedly increased and delayed in all patients, suggesting endogenous insulin resistance. Patients with more glucose intolerance showed less increase in plasma IRI than the group with less intolerance. lt is suggested that some insulin antagonists may decrease the peripheral insulin sensitivity and stimulate compensatory hyperactivity of pancreatic islets. If the compensatory hyperactivity is inadequate due to gemetic predisposition to diabetes mellitus or exhaustion of beta-cells of pancreatic islets, the glucose intolerance and overt diabetes mellitus may ensue.

  18. Insulin and leptin induce Glut4 plasma membrane translocation and glucose uptake in a human neuronal cell line by a phosphatidylinositol 3-kinase- dependent mechanism.

    Science.gov (United States)

    Benomar, Yacir; Naour, Nadia; Aubourg, Alain; Bailleux, Virginie; Gertler, Arieh; Djiane, Jean; Guerre-Millo, Michèle; Taouis, Mohammed

    2006-05-01

    The insulin-sensitive glucose transporter Glut4 is expressed in brain areas that regulate energy homeostasis and body adiposity. In contrast with peripheral tissues, however, the impact of insulin on Glut4 plasma membrane (PM) translocation in neurons is not known. In this study, we examined the role of two anorexic hormones (leptin and insulin) on Glut4 translocation in a human neuronal cell line that express endogenous insulin and leptin receptors. We show that insulin and leptin both induce Glut4 translocation to the PM of neuronal cells and activate glucose uptake. Wortmannin, a specific inhibitor of phosphatidylinositol 3-kinase, totally abolished insulin- and leptin-dependent Glut4 translocation and stimulation of glucose uptake. Thus, Glut4 translocation is a phosphatidylinositol 3-kinase-dependent mechanism in neuronal cells. Next, we investigated the impact of chronic insulin and leptin treatments on Glut4 expression and translocation. Chronic exposure of neuronal cells to insulin or leptin down-regulates Glut4 proteins and mRNA levels and abolishes the acute stimulation of glucose uptake in response to acute insulin or leptin. In addition, chronic treatment with either insulin or leptin impaired Glut4 translocation. A cross-desensitization between insulin and leptin was apparent, where exposure to insulin affects leptin-dependent Glut4 translocation and vice versa. This cross-desensitization could be attributed to the increase in suppressor of cytokine signaling-3 expression, which was demonstrated in response to each hormone. These results provide evidence to suggest that Glut4 translocation to neuronal PM is regulated by both insulin and leptin signaling pathways. These pathways might contribute to an in vivo glucoregulatory reflex involving a neuronal network and to the anorectic effect of insulin and leptin.

  19. Insulin sensitizers prevent fine particulate matter-induced vascular insulin resistance and changes in endothelial progenitor cell homeostasis.

    Science.gov (United States)

    Haberzettl, Petra; McCracken, James P; Bhatnagar, Aruni; Conklin, Daniel J

    2016-06-01

    Exposure to fine particular matter (PM2.5) increases the risk of developing cardiovascular disease and Type 2 diabetes. Because blood vessels are sensitive targets of air pollutant exposure, we examined the effects of concentrated ambient PM2.5 (CAP) on vascular insulin sensitivity and circulating levels of endothelial progenitor cells (EPCs), which reflect cardiovascular health. We found that CAP exposure for 9 days decreased insulin-stimulated Akt phosphorylation in the aorta of mice maintained on control diet. This change was accompanied by the induction of IL-1β and increases in the abundance of cleaved IL-18 and p10 subunit of Casp-1, consistent with the activation of the inflammasome pathway. CAP exposure also suppressed circulating levels of EPCs (Flk-1(+)/Sca-1(+) cells), while enhancing the bone marrow abundance of these cells. Although similar changes in vascular insulin signaling and EPC levels were observed in mice fed high-fat diet, CAP exposure did not exacerbate diet-induced changes in vascular insulin resistance or EPC homeostasis. Treatment with an insulin sensitizer, metformin or rosiglitazone, prevented CAP-induced vascular insulin resistance and NF-κB and inflammasome activation and restored peripheral blood and bone marrow EPC levels. These findings suggest that PM2.5 exposure induces diet-independent vascular insulin resistance and inflammation and prevents EPC mobilization, and that this EPC mobilization defect could be mediated by vascular insulin resistance. Impaired vascular insulin sensitivity may be an important mechanism underlying PM2.5-induced vascular injury, and pharmacological sensitization to insulin action could potentially prevent deficits in vascular repair and mitigate vascular inflammation due to exposure to elevated levels of ambient air pollution. Copyright © 2016 the American Physiological Society.

  20. Insulin-Producing Cells Differentiated from Human Bone Marrow Mesenchymal Stem Cells In Vitro Ameliorate Streptozotocin-Induced Diabetic Hyperglycemia.

    Directory of Open Access Journals (Sweden)

    Ying Xin

    Full Text Available The two major obstacles in the successful transplantation of islets for diabetes treatment are inadequate supply of insulin-producing tissue and immune rejection. Induction of the differentiation of human bone marrow-derived mesenchymal stem cells (hMSCs into insulin-producing cells (IPCs for autologous transplantation may alleviate those limitations.hMSCs were isolated and induced to differentiate into IPCs through a three-stage differentiation protocol in a defined media with high glucose, nicotinamide, and exendin-4. The physiological characteristics and functions of IPCs were then evaluated. Next, about 3 × 10(6 differentiated cells were transplanted into the renal sub-capsular space of streptozotocin (STZ-induced diabetic nude mice. Graft survival and function were assessed by immunohistochemistry, TUNEL staining and measurements of blood glucose levels in the mice.The differentiated IPCs were characterized by Dithizone (DTZ positive staining, expression of pancreatic β-cell markers, and human insulin secretion in response to glucose stimulation. Moreover, 43% of the IPCs showed L-type Ca2+ channel activity and similar changes in intracellular Ca2+ in response to glucose stimulation as that seen in pancreatic β-cells in the process of glucose-stimulated insulin secretion. Transplantation of functional IPCs into the renal subcapsular space of STZ-induced diabetic nude mice ameliorated the hyperglycemia. Immunofluorescence staining revealed that transplanted IPCs sustainably expressed insulin, c-peptide, and PDX-1 without apparent apoptosis in vivo.IPCs derived from hMSCs in vitro can ameliorate STZ-induced diabetic hyperglycemia, which indicates that these hMSCs may be a promising approach to overcome the limitations of islet transplantation.

  1. A modified method of insulin producing cells' generation from bone marrow-derived mesenchymal stem cells.

    Science.gov (United States)

    Czubak, Paweł; Bojarska-Junak, Agnieszka; Tabarkiewicz, Jacek; Putowski, Lechosław

    2014-01-01

    Type 1 diabetes mellitus is a result of autoimmune destruction of pancreatic insulin producing β-cells and so far it can be cured only by insulin injection, by pancreas transplantation, or by pancreatic islet cells' transplantation. The methods are, however, imperfect and have a lot of disadvantages. Therefore new solutions are needed. The best one would be the use of differentiated mesenchymal stem cells (MSCs). In the present study, we investigated the potential of the bone marrow-derived MSCs line for in vitro differentiation into insulin producing cells (IPSs). We applied an 18-day protocol to differentiate MSCs. Differentiating cells formed cell clusters some of which resembled pancreatic islet-like cells. Using dithizone we confirmed the presence of insulin in the cells. What is more, the expression of proinsulin C-peptide in differentiated IPCs was analyzed by flow cytometry. For the first time, we investigated the influence of growth factors' concentration on IPCs differentiation efficiency. We have found that an increase in the concentration of growth factors up to 60 ng/mL of β-FGF/EGF and 30 ng/mL of activin A/β-cellulin increases the percentage of IPCs. Further increase of growth factors does not show any increase of the percentage of differentiated cells. Our findings suggest that the presented protocol can be adapted for differentiation of insulin producing cells from stem cells.

  2. Induced Pluripotent Stem Cell-Derived Endothelial Cells in Insulin Resistance and Metabolic Syndrome.

    Science.gov (United States)

    Carcamo-Orive, Ivan; Huang, Ngan F; Quertermous, Thomas; Knowles, Joshua W

    2017-11-01

    Insulin resistance leads to a number of metabolic and cellular abnormalities including endothelial dysfunction that increase the risk of vascular disease. Although it has been particularly challenging to study the genetic determinants that predispose to abnormal function of the endothelium in insulin-resistant states, the possibility of deriving endothelial cells from induced pluripotent stem cells generated from individuals with detailed clinical phenotyping, including accurate measurements of insulin resistance accompanied by multilevel omic data (eg, genetic and genomic characterization), has opened new avenues to study this relationship. Unfortunately, several technical barriers have hampered these efforts. In the present review, we summarize the current status of induced pluripotent stem cell-derived endothelial cells for modeling endothelial dysfunction associated with insulin resistance and discuss the challenges to overcoming these limitations. © 2017 American Heart Association, Inc.

  3. Generation of high-yield insulin producing cells from human bone marrow mesenchymal stem cells.

    Science.gov (United States)

    Jafarian, Arefeh; Taghikhani, Mohammad; Abroun, Saeid; Pourpak, Zahra; Allahverdi, Amir; Soleimani, Masoud

    2014-07-01

    Allogenic islet transplantation is a most efficient approach for treatment of diabetes mellitus. However, the scarcity of islets and long term need for an immunosuppressant limits its application. Recently, cell replacement therapies that generate of unlimited sources of β cells have been developed to overcome these limitations. In this study we have described a stage specific differentiation protocol for the generation of insulin producing islet-like clusters from human bone marrow mesenchymal stem cells (hBM-MSCs). This specific stepwise protocol induced differentiation of hMSCs into definitive endoderm, pancreatic endoderm and pancreatic endocrine cells that expressed of sox17, foxa2, pdx1, ngn3, nkx2.2, insulin, glucagon, somatostatin, pancreatic polypeptide, and glut2 transcripts respectively. In addition, immunocytochemical analysis confirmed protein expression of the above mentioned genes. Western blot analysis discriminated insulin from proinsulin in the final differentiated cells. In derived insulin producing cells (IPCs), secreted insulin and C-peptide was in a glucose dependent manner. We have developed a protocol that generates effective high-yield human IPCs from hBM-MSCs in vitro. These finding suggest that functional IPCs generated by this procedure can be used as a cell-based approach for insulin dependent diabetes mellitus.

  4. The establishment of insulin resistance model in FL83B and L6 cell

    Science.gov (United States)

    Liu, Lanlan; Han, Jizhong; Li, Haoran; Liu, Mengmeng; Zeng, Bin

    2017-10-01

    The insulin resistance models of mouse liver epithelial and rat myoblasts cells were induced by three kinds of inducers: dexamethasone, high insulin and high glucose. The purpose is to select the optimal insulin resistance model, to provide a simple and reliable TR cell model for the study of the pathogenesis of TR and the improvement of TR drugs and functional foods. The MTT method is used for toxicity screening of three compounds, selecting security and suitable concentration. We performed a Glucose oxidase peroxidase (GOD-POD) method involving FL83B and L6 cell with dexamethasone, high insulin and high glucose-induced insulin resistance. Results suggested that FL83B cells with dexamethasone-induced (0.25uM) were established insulin resistance and L6 cells with high-glucose (30mM) and dexamethasone-induced (0.25uM) were established insulin resistance.

  5. De Novo Formation of Insulin-Producing “Neo-β Cell Islets” from Intestinal Crypts

    Directory of Open Access Journals (Sweden)

    Yi-Ju Chen

    2014-03-01

    Full Text Available The ability to interconvert terminally differentiated cells could serve as a powerful tool for cell-based treatment of degenerative diseases, including diabetes mellitus. To determine which, if any, adult tissues are competent to activate an islet β cell program, we performed an in vivo screen by expressing three β cell “reprogramming factors” in a wide spectrum of tissues. We report that transient intestinal expression of these factors—Pdx1, MafA, and Ngn3 (PMN—promotes rapid conversion of intestinal crypt cells into endocrine cells, which coalesce into “neoislets” below the crypt base. Neoislet cells express insulin and show ultrastructural features of β cells. Importantly, intestinal neoislets are glucose-responsive and able to ameliorate hyperglycemia in diabetic mice. Moreover, PMN expression in human intestinal “organoids” stimulates the conversion of intestinal epithelial cells into β-like cells. Our results thus demonstrate that the intestine is an accessible and abundant source of functional insulin-producing cells.

  6. Studies of insulin secretory responses and of arachidonic acid incorporation into phospholipids of stably transfected insulinoma cells that overexpress group VIA phospholipase A2 (iPLA2beta ) indicate a signaling rather than a housekeeping role for iPLA2beta.

    Science.gov (United States)

    Ma, Z; Ramanadham, S; Wohltmann, M; Bohrer, A; Hsu, F F; Turk, J

    2001-04-20

    A cytosolic 84-kDa group VIA phospholipase A(2) (iPLA(2)beta) that does not require Ca(2+) for catalysis has been cloned from several sources, including rat and human pancreatic islet beta-cells and murine P388D1 cells. Many potential iPLA(2)beta functions have been proposed, including a signaling role in beta-cell insulin secretion and a role in generating lysophosphatidylcholine acceptors for arachidonic acid incorporation into P388D1 cell phosphatidylcholine (PC). Proposals for iPLA(2)beta function rest in part on effects of inhibiting iPLA(2)beta activity with a bromoenol lactone (BEL) suicide substrate, but BEL also inhibits phosphatidate phosphohydrolase-1 and a group VIB phospholipase A(2). Manipulation of iPLA(2)beta expression by molecular biologic means is an alternative approach to study iPLA(2)beta functions, and we have used a retroviral construct containing iPLA(2)beta cDNA to prepare two INS-1 insulinoma cell clonal lines that stably overexpress iPLA(2)beta. Compared with parental INS-1 cells or cells transfected with empty vector, both iPLA(2)beta-overexpressing lines exhibit amplified insulin secretory responses to glucose and cAMP-elevating agents, and BEL substantially attenuates stimulated secretion. Electrospray ionization mass spectrometric analyses of arachidonic acid incorporation into INS-1 cell PC indicate that neither overexpression nor inhibition of iPLA(2)beta affects the rate or extent of this process in INS-1 cells. Immunocytofluorescence studies with antibodies directed against iPLA(2)beta indicate that cAMP-elevating agents increase perinuclear fluorescence in INS-1 cells, suggesting that iPLA(2)beta associates with nuclei. These studies are more consistent with a signaling than with a housekeeping role for iPLA(2)beta in insulin-secreting beta-cells.

  7. The glucose-dependent insulinotropic polypeptide and glucose-stimulated insulin response to exercise training and diet in obesity.

    Science.gov (United States)

    Kelly, Karen R; Brooks, Latina M; Solomon, Thomas P J; Kashyap, Sangeeta R; O'Leary, Valerie B; Kirwan, John P

    2009-06-01

    Aging and obesity are characterized by decreased beta-cell sensitivity and defects in the potentiation of nutrient-stimulated insulin secretion by GIP. Exercise and diet are known to improve glucose metabolism and the pancreatic insulin response to glucose, and this effect may be mediated through the incretin effect of GIP. The purpose of this study was to assess the effects of a 12-wk exercise training intervention (5 days/wk, 60 min/day, 75% Vo(2 max)) combined with a eucaloric (EX, n = 10) or hypocaloric (EX-HYPO, pre: 1,945 +/- 190, post: 1,269 +/- 70, kcal/day; n = 9) diet on the GIP response to glucose in older (66.8 +/- 1.5 yr), obese (34.4 +/- 1.7 kg/m(2)) adults with impaired glucose tolerance. In addition to GIP, plasma PYY(3-36), insulin, and glucose responses were measured during a 3-h, 75-g oral glucose tolerance test. Both interventions led to a significant improvement in Vo(2 max) (P HYPO (-8.3 +/- 1.1 vs. -2.8 +/- 0.5, P = 0.002). The glucose-stimulated insulin response was reduced after EX-HYPO (P = 0.02), as was the glucose-stimulated GIP response (P caloric restriction and exercise reduces the GIP response to ingested glucose, 2) GIP may mediate the attenuated glucose-stimulated insulin response after exercise/diet interventions, and 3) the increased PYY(3-36) response represents an improved capacity to regulate satiety and potentially body weight in older, obese, insulin-resistant adults.

  8. Attenuation of insulin-evoked responses in brain networks controlling appetite and reward in insulin resistance: the cerebral basis for impaired control of food intake in metabolic syndrome?

    Science.gov (United States)

    Anthony, Karen; Reed, Laurence J; Dunn, Joel T; Bingham, Emma; Hopkins, David; Marsden, Paul K; Amiel, Stephanie A

    2006-11-01

    The rising prevalence of obesity and type 2 diabetes is a global challenge. A possible mechanism linking insulin resistance and weight gain would be attenuation of insulin-evoked responses in brain areas relevant to eating in systemic insulin resistance. We measured brain glucose metabolism, using [(18)F]fluorodeoxyglucose positron emission tomography, in seven insulin-sensitive (homeostasis model assessment of insulin resistance [HOMA-IR] = 1.3) and seven insulin-resistant (HOMA-IR = 6.3) men, during suppression of endogenous insulin by somatostatin, with and without an insulin infusion that elevated insulin to 24.6 +/- 5.2 and 23.2 +/- 5.8 mU/l (P = 0.76), concentrations similar to fasting levels of the resistant subjects and approximately threefold above those of the insulin-sensitive subjects. Insulin-evoked change in global cerebral metabolic rate for glucose was reduced in insulin resistance (+7 vs. +17.4%, P = 0.033). Insulin was associated with increased metabolism in ventral striatum and prefrontal cortex and with decreased metabolism in right amygdala/hippocampus and cerebellar vermis (P reward. Diminishing the link be-tween control of food intake and energy balance may contribute to development of obesity in insulin resistance.

  9. Dual role of proapoptotic BAD in insulin secretion and beta cell survival.

    Science.gov (United States)

    Danial, Nika N; Walensky, Loren D; Zhang, Chen-Yu; Choi, Cheol Soo; Fisher, Jill K; Molina, Anthony J A; Datta, Sandeep Robert; Pitter, Kenneth L; Bird, Gregory H; Wikstrom, Jakob D; Deeney, Jude T; Robertson, Kirsten; Morash, Joel; Kulkarni, Ameya; Neschen, Susanne; Kim, Sheene; Greenberg, Michael E; Corkey, Barbara E; Shirihai, Orian S; Shulman, Gerald I; Lowell, Bradford B; Korsmeyer, Stanley J

    2008-02-01

    The proapoptotic BCL-2 family member BAD resides in a glucokinase-containing complex that regulates glucose-driven mitochondrial respiration. Here, we present genetic evidence of a physiologic role for BAD in glucose-stimulated insulin secretion by beta cells. This novel function of BAD is specifically dependent upon the phosphorylation of its BH3 sequence, previously defined as an essential death domain. We highlight the pharmacologic relevance of phosphorylated BAD BH3 by using cell-permeable, hydrocarbon-stapled BAD BH3 helices that target glucokinase, restore glucose-driven mitochondrial respiration and correct the insulin secretory response in Bad-deficient islets. Our studies uncover an alternative target and function for the BAD BH3 domain and emphasize the therapeutic potential of phosphorylated BAD BH3 mimetics in selectively restoring beta cell function. Furthermore, we show that BAD regulates the physiologic adaptation of beta cell mass during high-fat feeding. Our findings provide genetic proof of the bifunctional activities of BAD in both beta cell survival and insulin secretion.

  10. Geniposide regulates glucose-stimulated insulin secretion possibly through controlling glucose metabolism in INS-1 cells.

    Directory of Open Access Journals (Sweden)

    Jianhui Liu

    Full Text Available Glucose-stimulated insulin secretion (GSIS is essential to the control of metabolic fuel homeostasis. The impairment of GSIS is a key element of β-cell failure and one of causes of type 2 diabetes mellitus (T2DM. Although the KATP channel-dependent mechanism of GSIS has been broadly accepted for several decades, it does not fully describe the effects of glucose on insulin secretion. Emerging evidence has suggested that other mechanisms are involved. The present study demonstrated that geniposide enhanced GSIS in response to the stimulation of low or moderately high concentrations of glucose, and promoted glucose uptake and intracellular ATP levels in INS-1 cells. However, in the presence of a high concentration of glucose, geniposide exerted a contrary role on both GSIS and glucose uptake and metabolism. Furthermore, geniposide improved the impairment of GSIS in INS-1 cells challenged with a high concentration of glucose. Further experiments showed that geniposide modulated pyruvate carboxylase expression and the production of intermediates of glucose metabolism. The data collectively suggest that geniposide has potential to prevent or improve the impairment of insulin secretion in β-cells challenged with high concentrations of glucose, likely through pyruvate carboxylase mediated glucose metabolism in β-cells.

  11. Fibronectin and laminin promote differentiation of human mesenchymal stem cells into insulin producing cells through activating Akt and ERK

    Directory of Open Access Journals (Sweden)

    Chiou Shih-Hwa

    2010-07-01

    Full Text Available Abstract Background Islet transplantation provides a promising cure for Type 1 diabetes; however it is limited by a shortage of pancreas donors. Bone marrow-derived multipotent mesenchymal stem cells (MSCs offer renewable cells for generating insulin-producing cells (IPCs. Methods We used a four-stage differentiation protocol, containing neuronal differentiation and IPC-conversion stages, and combined with pellet suspension culture to induce IPC differentiation. Results Here, we report adding extracellular matrix proteins (ECM such as fibronectin (FN or laminin (LAM enhances pancreatic differentiation with increases in insulin and Glut2 gene expressions, proinsulin and insulin protein levels, and insulin release in response to elevated glucose concentration. Adding FN or LAM induced activation of Akt and ERK. Blocking Akt or ERK by adding LY294002 (PI3K specific inhibitor, PD98059 (MEK specific inhibitor or knocking down Akt or ERK failed to abrogate FN or LAM-induced enhancement of IPC differentiation. Only blocking both of Akt and ERK or knocking down Akt and ERK inhibited the enhancement of IPC differentiation by adding ECM. Conclusions These data prove IPC differentiation by MSCs can be modulated by adding ECM, and these stimulatory effects were mediated through activation of Akt and ERK pathways.

  12. Differences in beta-cell function and insulin secretion in Black vs. White obese adolescents: Do incretin hormones play a role?

    Science.gov (United States)

    Black youth are at higher risk for type 2 diabetes (T2D) than their White peers. Previously we demonstrated that for the same degree of insulin sensitivity, Black youth have an upregulated beta-cell function and insulin hypersecretion, in response to intravenous (IV) glucose, compared with Whites. T...

  13. Bimodal effect on pancreatic β-cells of secretory products from normal or insulin-resistant human skeletal muscle

    DEFF Research Database (Denmark)

    Bouzakri, Karim; Plomgaard, Peter; Berney, Thierry

    2011-01-01

    Type 2 diabetes is characterized by insulin resistance with a relative deficiency in insulin secretion. This study explored the potential communication between insulin-resistant human skeletal muscle and primary (human and rat) β-cells.......Type 2 diabetes is characterized by insulin resistance with a relative deficiency in insulin secretion. This study explored the potential communication between insulin-resistant human skeletal muscle and primary (human and rat) β-cells....

  14. Curcuma longa polyphenols improve insulin-mediated lipid accumulation and attenuate proinflammatory response of 3T3-L1 adipose cells during oxidative stress through regulation of key adipokines and antioxidant enzymes.

    Science.gov (United States)

    Septembre-Malaterre, Axelle; Le Sage, Fanny; Hatia, Sarah; Catan, Aurélie; Janci, Laurent; Gonthier, Marie-Paule

    2016-07-08

    Plant polyphenols may exert beneficial action against obesity-related oxidative stress and inflammation which promote insulin resistance. This study evaluated the effect of polyphenols extracted from French Curcuma longa on 3T3-L1 adipose cells exposed to H2 O2 -mediated oxidative stress. We found that Curcuma longa extract exhibited high amounts of curcuminoids identified as curcumin, demethoxycurcumin, and bisdemethoxycurcumin, which exerted free radical-scavenging activities. Curcuma longa polyphenols improved insulin-mediated lipid accumulation and upregulated peroxisome proliferator-activated receptor-gamma gene expression and adiponectin secretion which decreased in H2 O2 -treated cells. Curcuminoids attenuated H2 O2 -enhanced production of pro-inflammatory molecules such as interleukin-6, tumor necrosis factor-alpha, monocyte chemoattractant protein-1, and nuclear factor κappa B. Moreover, they reduced intracellular levels of reactive oxygen species elevated by H2 O2 and modulated the expression of genes encoding superoxide dismutase and catalase antioxidant enzymes. Collectively, these findings highlight that Curcuma longa polyphenols protect adipose cells against oxidative stress and may improve obesity-related metabolic disorders. © 2016 BioFactors, 42(4):418-430, 2016. © 2016 International Union of Biochemistry and Molecular Biology.

  15. Repression of COUP-TFI Improves Bone Marrow-Derived Mesenchymal Stem Cell Differentiation into Insulin-Producing Cells

    Directory of Open Access Journals (Sweden)

    Tao Zhang

    2017-09-01

    Full Text Available Identifying molecular mechanisms that regulate insulin expression in bone marrow-derived mesenchymal stem cells (bmMSCs can provide clues on how to stimulate the differentiation of bmMSCs into insulin-producing cells (IPCs, which can be used as a therapeutic approach against type 1 diabetes (T1D. As repression factors may inhibit differentiation, the efficiency of this process is insufficient for cell transplantation. In this study, we used the mouse insulin 2 (Ins2 promoter sequence and performed a DNA affinity precipitation assay combined with liquid chromatography-mass spectrometry to identify the transcription factor, chicken ovalbumin upstream promoter transcriptional factor I (COUP-TFI. Functionally, bmMSCs were reprogrammed into IPCs via COUP-TFI suppression and MafA overexpression. The differentiated cells expressed higher levels of genes specific for islet endocrine cells, and they released C-peptide and insulin in response to glucose stimulation. Transplantation of IPCs into streptozotocin-induced diabetic mice caused a reduction in hyperglycemia. Mechanistically, COUP-TFI bound to the DR1 (direct repeats with 1 spacer element in the Ins2 promoter, thereby negatively regulating promoter activity. Taken together, the data provide a novel mechanism by which COUP-TFI acts as a negative regulator in the Ins2 promoter. The differentiation of bmMSCs into IPCs could be improved by knockdown of COUP-TFI, which may provide a novel stem cell-based therapy for T1D. Keywords: siRNAs, differentiation, stem cell transplantation, diabetes, mesenchymal stem cells

  16. Altered insulin response to an acute bout of exercise in pediatric obesity.

    Science.gov (United States)

    Tran, Brian D; Leu, Szu-Yun; Oliver, Stacy; Graf, Scott; Vigil, Diana; Galassetti, Pietro

    2014-11-01

    Pediatric obesity typically induces insulin resistance, often later evolving into type 2 diabetes. While exercise, enhancing insulin sensitivity, is broadly used to prevent this transition, it is unknown whether alterations in the exercise insulin response pattern occur in obese children. Therefore, we measured exercise insulin responses in 57 healthy weight (NW), 20 overweight (OW), and 56 obese (Ob) children. Blood samples were drawn before and after 30 min of intermittent (2 min on, 1 min off) cycling at ~80% VO2max. In a smaller group (14 NW, 6 OW, 15 Ob), a high-fat meal was ingested 45 min preexercise. Baseline glycemia was similar and increased slightly and similarly in all groups during exercise. Basal insulin (pmol/L) was significantly higher in Ob vs. other groups; postexercise, insulin increased in NW (+7± 3) and OW (+5 ± 8), but decreased in Ob (-15±5, p feeding caused a rapid rise in insulin, promptly corrected by exercise. In Ob, however, insulin rose again 30 min postexercise. Our data indicates a distinct pattern of exercise-induced insulin modulation in pediatric obesity, possibly modulated by basal insulin concentrations.

  17. Determining pancreatic β-cell compensation for changing insulin sensitivity using an oral glucose tolerance test

    DEFF Research Database (Denmark)

    Solomon, Thomas; Malin, Steven K; Karstoft, Kristian

    2014-01-01

    Plasma glucose, insulin, and C-peptide responses during an OGTT are informative for both research and clinical practice in type 2 diabetes. The aim of this study was to use such information to determine insulin sensitivity and insulin secretion so as to calculate an oral glucose disposition index...

  18. Cell surface alteration in Epstein-Barr virus-transformed cells from patients with extreme insulin resistance

    International Nuclear Information System (INIS)

    Gorden, D.L.; Robert, A.; Moncada, V.Y.; Taylor, S.I.; Muehlhauser, J.C.; Carpentier, J.L.

    1990-01-01

    An abnormality was detected in the morphology of the cell surface of Epstein-Barr virus-transformed lymphocytes of patients with genetic forms of insulin resistance. In cells from two patients with leprechaunism and two patients with type A extreme insulin resistance, scanning electron microscopy demonstrated a decrease in the percentage of the cell surface occupied by microvilli in cells from the patients with leprechaunism and type A insulin resistance compared with control cells. When cells from a healthy control subject and one of the patients with leprechaunism (Lep/Ark-1) were incubated with 125 I-labeled insulin, there was a decrease in the percentage of 125 I-insulin associated with microvilli on the cell surface. Thus, the decreased localization of insulin receptors with the microvillous region of the cell surface was in proportion to the decrease in microvilli

  19. Effects of Rosiglitazone, Glyburide, and Metformin on β-Cell Function and Insulin Sensitivity in ADOPT

    Science.gov (United States)

    Kahn, Steven E.; Lachin, John M.; Zinman, Bernard; Haffner, Steven M.; Aftring, R. Paul; Paul, Gitanjali; Kravitz, Barbara G.; Herman, William H.; Viberti, Giancarlo; Holman, Rury R.

    2011-01-01

    OBJECTIVE ADOPT (A Diabetes Outcome Progression Trial) demonstrated that initial monotherapy with rosiglitazone provided superior durability of glycemic control compared with metformin and glyburide in patients with recently diagnosed type 2 diabetes. Herein, we examine measures of β-cell function and insulin sensitivity from an oral glucose tolerance test (OGTT) over a 4-year period among the three treatments. RESEARCH DESIGN AND METHODS Recently diagnosed, drug-naïve patients with type 2 diabetes (4,360 total) were treated for a median of 4.0 years with rosiglitazone, metformin, or glyburide and were examined with periodic metabolic testing using an OGTT. RESULTS Measures of β-cell function and insulin sensitivity from an OGTT showed more favorable changes over time with rosiglitazone versus metformin or glyburide. Persistent improvements were seen in those who completed 4 years of monotherapy and marked deterioration of β-cell function in those who failed to maintain adequate glucose control with initial monotherapy. CONCLUSIONS The favorable combined changes in β-cell function and insulin sensitivity over time with rosiglitazone appear to be responsible for its superior glycemic durability over metformin and glyburide as initial monotherapy in type 2 diabetes. PMID:21415383

  20. Mitochondrial Respiration in Insulin-Producing β-Cells: General Characteristics and Adaptive Effects of Hypoxia.

    Science.gov (United States)

    Hals, Ingrid K; Bruerberg, Simon Gustafson; Ma, Zuheng; Scholz, Hanne; Björklund, Anneli; Grill, Valdemar

    2015-01-01

    To provide novel insights on mitochondrial respiration in β-cells and the adaptive effects of hypoxia. Insulin-producing INS-1 832/13 cells were exposed to 18 hours of hypoxia followed by 20-22 hours re-oxygenation. Mitochondrial respiration was measured by high-resolution respirometry in both intact and permeabilized cells, in the latter after establishing three functional substrate-uncoupler-inhibitor titration (SUIT) protocols. Concomitant measurements included proteins of mitochondrial complexes (Western blotting), ATP and insulin secretion. Intact cells exhibited a high degree of intrinsic uncoupling, comprising about 50% of oxygen consumption in the basal respiratory state. Hypoxia followed by re-oxygenation increased maximal overall respiration. Exploratory experiments in peremabilized cells could not show induction of respiration by malate or pyruvate as reducing substrates, thus glutamate and succinate were used as mitochondrial substrates in SUIT protocols. Permeabilized cells displayed a high capacity for oxidative phosphorylation for both complex I- and II-linked substrates in relation to maximum capacity of electron transfer. Previous hypoxia decreased phosphorylation control of complex I-linked respiration, but not in complex II-linked respiration. Coupling control ratios showed increased coupling efficiency for both complex I- and II-linked substrates in hypoxia-exposed cells. Respiratory rates overall were increased. Also previous hypoxia increased proteins of mitochondrial complexes I and II (Western blotting) in INS-1 cells as well as in rat and human islets. Mitochondrial effects were accompanied by unchanged levels of ATP, increased basal and preserved glucose-induced insulin secretion. Exposure of INS-1 832/13 cells to hypoxia, followed by a re-oxygenation period increases substrate-stimulated respiratory capacity and coupling efficiency. Such effects are accompanied by up-regulation of mitochondrial complexes also in pancreatic islets

  1. Hypothesis: Musculin is a hormone secreted by skeletal muscle, the body's largest endocrine organ. Evidence for actions on the endocrine pancreas to restrain the beta-cell mass and to inhibit insulin secretion and on the hypothalamus to co-ordinate the neuroendocrine and appetite responses to exercise.

    Science.gov (United States)

    Engler, Dennis

    2007-01-01

    Recent studies indicate that skeletal muscle may act as an endocrine organ by secreting interleukin-6 (IL-6) into the systemic circulation. From an analysis of the actions of IL-6 and of additional literature, we postulate that skeletal muscle also secretes an unidentified hormone, which we have named Musculin (Latin: musculus = muscle), which acts on the pancreatic beta-cell to restrain the size of the (beta-cell mass and to tonically inhibit insulin secretion and biosynthesis. It is suggested that the amount of Musculin secreted is determined by, and is positively correlated with, the prevailing insulin sensitivity of skeletal muscle, thereby accounting for the hyperinsulinemia that occurs in insulin resistant disorders such as type 2 diabetes mellitus, obesity, and the polycystic ovary syndrome. In addition, it is postulated that Musculin acts on the hypothalamus (arcuate nucleus, dorsomedial hypothalamic nucleus) to co-ordinate the neuroendocrine and appetite responses to exercise. However, the possibilities that Musculin may act on additional central nervous system sites and that an additional hormone(s) may be responsible for these actions are not excluded. It is suggested that a search be made for Musculin, since analogues of such a substance may be of therapeutic benefit in the treatment of the current global diabetes and obesity epidemic.

  2. The glucose-dependent insulinotropic polypeptide and glucose-stimulated insulin response to exercise training and diet in obesity

    OpenAIRE

    Kelly, Karen R.; Brooks, Latina M.; Solomon, Thomas P. J.; Kashyap, Sangeeta R.; O'Leary, Valerie B.; Kirwan, John P.

    2009-01-01

    Aging and obesity are characterized by decreased β-cell sensitivity and defects in the potentiation of nutrient-stimulated insulin secretion by GIP. Exercise and diet are known to improve glucose metabolism and the pancreatic insulin response to glucose, and this effect may be mediated through the incretin effect of GIP. The purpose of this study was to assess the effects of a 12-wk exercise training intervention (5 days/wk, 60 min/day, 75% V̇o2 max) combined with a eucaloric (EX, n = 10) or ...

  3. Leptin responses to bovine interferon- α and insulin in cattle

    African Journals Online (AJOL)

    Egyptian Journal of Biochemistry and Molecular Biology ... IFN- α injection produced a rapid increase in glucose and insulin levels but leptin levels did not show any alteration after the injection. ... Insulin levels rapidly increased in the blood and consequently a significant decrease in blood glucose level was recorded.

  4. Immunohistochemical expression of insulin, glucagon, and somatostatin in pancreatic islets of horses with and without insulin resistance.

    Science.gov (United States)

    Newkirk, Kim M; Ehrensing, Gordon; Odoi, Agricola; Boston, Raymond C; Frank, Nicholas

    2018-02-01

    OBJECTIVE To assess insulin, glucagon, and somatostatin expression within pancreatic islets of horses with and without insulin resistance. ANIMALS 10 insulin-resistant horses and 13 insulin-sensitive horses. PROCEDURES For each horse, food was withheld for at least 10 hours before a blood sample was collected for determination of serum insulin concentration. Horses with a serum insulin concentration horses with a serum insulin concentration > 20 μU/mL underwent a frequently sampled IV glucose tolerance test to determine sensitivity to insulin by minimal model analysis. Horses with a sensitivity to insulin horses were euthanized with a barbiturate overdose, and pancreatic specimens were harvested and immunohistochemically stained for determination of insulin, glucagon, and somatostatin expression in pancreatic islets. Islet hormone expression was compared between insulin-resistant and insulin-sensitive horses. RESULTS Cells expressing insulin, glucagon, and somatostatin made up approximately 62%, 12%, and 7%, respectively, of pancreatic islet cells in insulin-resistant horses and 64%, 18%, and 9%, respectively, of pancreatic islet cells in insulin-sensitive horses. Expression of insulin and somatostatin did not differ between insulin-resistant and insulin-sensitive horses, but the median percentage of glucagon-expressing cells in the islets of insulin-resistant horses was significantly less than that in insulin-sensitive horses. CONCLUSIONS AND CLINICAL RELEVANCE Results suggested that, in insulin-resistant horses, insulin secretion was not increased but glucagon production might be downregulated as a compensatory response to hyperinsulinemia.

  5. Autocrine growth induced by the insulin-related factor in the insulin-independent teratoma cell line 1246-3A

    International Nuclear Information System (INIS)

    Yamada, Yukio; Serrero, G.

    1988-01-01

    An insulin-independent teratoma-derived cell line, called 1246-3A, has been isolated from the adipogenic cell line 1246, which stringently requires insulin for proliferation. The 1246-3A cell line, which can proliferate in the absence of exogenous insulin, produces in its conditioned medium a growth factor similar to pancreatic insulin by its biological and immunological properties. This factor, called insulin-related factor (IRF), was purified and iodinated to study its binding to cell surface receptors. 125 I-labeled IRF binding to intact 1246-3A cells is lower than to 1246 cells. Cell surface binding can be restored by culturing the 1246-3A cells in the presence of an anti-porcine insulin monoclonal antibody of by acid prewash of the cells prior to performing the binding. Scatchard analysis of binding indicates that IRF secreted by the 1246-3A cells partially occupies high-affinity binding sites on the producer cells. Moreover, insulin monoclonal antibody inhibits the proliferation of the IRF-producing 1246-3A cells, suggesting that these cells are dependent on the secreted IRF for growth in culture. The authors conclude that the insulin-related factor secreted by the insulin-independent 1246-3A cells stimulates their proliferation in an autocrine fashion

  6. Extrinsic Factors Involved in the Differentiation of Stem Cells into Insulin-Producing Cells: An Overview

    Directory of Open Access Journals (Sweden)

    Rebecca S. Y. Wong

    2011-01-01

    Full Text Available Diabetes mellitus is a chronic disease with many debilitating complications. Treatment of diabetes mellitus mainly revolves around conventional oral hypoglycaemic agents and insulin replacement therapy. Recently, scientists have turned their attention to the generation of insulin-producing cells (IPCs from stem cells of various sources. To date, many types of stem cells of human and animal origins have been successfully turned into IPCs in vitro and have been shown to exert glucose-lowering effect in vivo. However, scientists are still faced with the challenge of producing a sufficient number of IPCs that can in turn produce sufficient insulin for clinical use. A careful choice of stem cells, methods, and extrinsic factors for induction may all be contributing factors to successful production of functional beta-islet like IPCs. It is also important that the mechanism of differentiation and mechanism by which IPCs correct hyperglycaemia are carefully studied before they are used in human subjects.

  7. Insulin sensitivity affects corticolimbic brain responses to visual food cues in polycystic ovary syndrome patients.

    Science.gov (United States)

    Alsaadi, Hanin M; Van Vugt, Dean A

    2015-11-01

    This study examined the effect of insulin sensitivity on the responsiveness of appetite regulatory brain regions to visual food cues. Nineteen participants diagnosed with polycystic ovary syndrome (PCOS) were divided into insulin-sensitive (n=8) and insulin-resistant (n=11) groups based on the homeostatic model assessment of insulin resistance (HOMA2-IR). Subjects underwent functional magnetic resonance imaging (fMRI) while viewing food pictures following water or dextrose consumption. The corticolimbic blood oxygen level dependent (BOLD) responses to high-calorie (HC) or low-calorie (LC) food pictures were compared within and between groups. BOLD responses to food pictures were reduced during a glucose challenge in numerous corticolimbic brain regions in insulin-sensitive but not insulin-resistant subjects. Furthermore, the degree of insulin resistance positively correlated with the corticolimbic BOLD response in the medial prefrontal cortex (mPFC), orbitofrontal cortex (OFC), anterior cingulate and ventral tegmental area (VTA) in response to HC pictures, and in the dorsolateral prefrontal cortex (DLPFC), mPFC, anterior cingulate, and insula in response to LC pictures following a glucose challenge. BOLD signal in the OFC, midbrain, hippocampus, and amygdala following a glucose challenge correlated with HOMA2-IR in response to HC-LC pictures. We conclude that the normal inhibition of corticolimbic brain responses to food pictures during a glucose challenge is compromised in insulin-resistant subjects. The increase in brain responsiveness to food pictures during postprandial hyperinsulinemia may lead to greater non-homeostatic eating and perpetuate obesity in insulin-resistant subjects.

  8. Protein kinase C is differentially regulated by thrombin, insulin, and epidermal growth factor in human mammary tumor cells

    Energy Technology Data Exchange (ETDEWEB)

    Gomez, M.L.; Tellez-Inon, M.T. (Instituto de Ingenieria Genetica y Biologia Molecular, Buenos Aires (Argentina)); Medrano, E.E.; Cafferatta, E.G.A. (Instituto de Investigaciones Bioquimicas Fundacion Campomar, Buenos Aires (Argentina))

    1988-03-01

    The exposure of serum-deprived mammary tumor cells MCF-7 and T-47D to insulin, thrombin, and epidermal growth factor (EGF) resulted in dramatic modifications in the activity and in the translocation capacity of protein kinase C from cytosol to membrane fractions. Insulin induces a 600% activation of the enzyme after 5 h of exposure to the hormone in MCF-7 cells; thrombin either activates (200% in MCF-7) or down-regulates (in T-47D), and EGF exerts only a moderate effect. Thus, the growth factors studied modulate differentially the protein kinase C activity in human mammary tumor cells. The physiological significance of the results obtained are discussed in terms of the growth response elicited by insulin, thrombin, and EGF.

  9. Egr2 enhances insulin resistance via JAK2/STAT3/SOCS-1 pathway in HepG2 cells treated with palmitate.

    Science.gov (United States)

    Lu, Lin; Ye, Xinhua; Yao, Qing; Lu, Aijiao; Zhao, Zhen; Ding, Yang; Meng, Chuchen; Yu, Wenlong; Du, Yunfeng; Cheng, JinLuo

    2018-05-01

    Insulin resistance is generally responsible for the pathogenesis of type 2 diabetes mellitus (T2DM). Early growth response proteins-2 (Egr2) has been reported to be able to increase the expression of the suppressors of cytokine signaling-1 (SOCS-1), and impair insulin signaling pathway through suppression of insulin receptor substrates (IRS), including IRS-1 and IRS-2. However, whether Egr2 is directly involved in the development of insulin resistance, and how its potential contributions to insulin resistance still remain unknown. Here, our present investigation found that the expression levels of Egr2 were up-regulated when insulin resistance occurs, and knockdown of Egr2 abolished the effect of insulin resistance in HepG2 cells induced with palmitate (PA). Importantly, inhibition of Egr2 decreased the expression of SOCS-1 as well as reduced phosphorylation of JAK2 and STAT3. And, our data indicated that silencing of Egr2 accelerated hepatic glucose uptake and reversed the impaired lipid metabolism upon insulin resistance. In summary, the present study confirms that Egr2 could deteriorate insulin resistance via the pathway of JAK2/STAT3/SOCS-1 and may shed light on resolving insulin resistance and further the pathogenesis of T2DM. Copyright © 2017 Elsevier Inc. All rights reserved.

  10. Generation of insulin-producing cells from gnotobiotic porcine skin-derived stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Yang, Ji Hoon; Lee, Sung Ho; Heo, Young Tae [Department of Bioscience and Biotechnology, Bio-Organ Research Center, Konkuk University, Seoul 143-701 (Korea, Republic of); Uhm, Sang Jun [Department of Animal Biotechnology, Bio-Organ Research Center, Konkuk University, Seoul 143-701 (Korea, Republic of); Lee, Hoon Taek, E-mail: htl3675@konkuk.ac.kr [Department of Animal Biotechnology, Bio-Organ Research Center, Konkuk University, Seoul 143-701 (Korea, Republic of)

    2010-07-09

    A major problem in the treatment of type 1 diabetes mellitus is the limited availability of alternative sources of insulin-producing cells for islet transplantation. In this study, we investigated the effect of bone morphogenetic protein 4 (BMP-4) treatments of gnotobiotic porcine skin-derived stem cells (gSDSCs) on their reprogramming and subsequent differentiation into insulin-producing cells (IPCs). We isolated SDSCs from the ear skin of a gnotobiotic pig. During the proliferation period, the cells expressed stem-cell markers Oct-4, Sox-2, and CD90; nestin expression also increased significantly. The cells could differentiate into IPCs after treatments with activin-A, glucagon-like peptide-1 (GLP-1), and nicotinamide. After 15 days in the differentiation medium, controlled gSDSCs began expressing endocrine progenitor genes and proteins (Ngn3, Neuro-D, PDX-1, NKX2.2, NKX6.1, and insulin). The IPCs showed increased insulin synthesis after glucose stimulation. The results indicate that stem cells derived from the skin of gnotobiotic pigs can differentiate into IPCs under the appropriate conditions in vitro. Our three-stage induction protocol could be applied without genetic modification to source IPCs from stem cells in the skin of patients with diabetes for autologous transplantation.

  11. Generation of insulin-producing cells from gnotobiotic porcine skin-derived stem cells

    International Nuclear Information System (INIS)

    Yang, Ji Hoon; Lee, Sung Ho; Heo, Young Tae; Uhm, Sang Jun; Lee, Hoon Taek

    2010-01-01

    A major problem in the treatment of type 1 diabetes mellitus is the limited availability of alternative sources of insulin-producing cells for islet transplantation. In this study, we investigated the effect of bone morphogenetic protein 4 (BMP-4) treatments of gnotobiotic porcine skin-derived stem cells (gSDSCs) on their reprogramming and subsequent differentiation into insulin-producing cells (IPCs). We isolated SDSCs from the ear skin of a gnotobiotic pig. During the proliferation period, the cells expressed stem-cell markers Oct-4, Sox-2, and CD90; nestin expression also increased significantly. The cells could differentiate into IPCs after treatments with activin-A, glucagon-like peptide-1 (GLP-1), and nicotinamide. After 15 days in the differentiation medium, controlled gSDSCs began expressing endocrine progenitor genes and proteins (Ngn3, Neuro-D, PDX-1, NKX2.2, NKX6.1, and insulin). The IPCs showed increased insulin synthesis after glucose stimulation. The results indicate that stem cells derived from the skin of gnotobiotic pigs can differentiate into IPCs under the appropriate conditions in vitro. Our three-stage induction protocol could be applied without genetic modification to source IPCs from stem cells in the skin of patients with diabetes for autologous transplantation.

  12. Role of aryl hydrocarbon receptor nuclear translocator in KATP channel-mediated insulin secretion in INS-1 insulinoma cells

    International Nuclear Information System (INIS)

    Kim, Ji-Seon; Zheng Haifeng; Kim, Sung Joon; Park, Jong-Wan; Park, Kyong Soo; Ho, Won-Kyung; Chun, Yang-Sook

    2009-01-01

    Aryl hydrocarbon receptor nuclear translocator (ARNT) has been known to participate in cellular responses to xenobiotic and hypoxic stresses, as a common partner of aryl hydrocarbon receptor and hypoxia inducible factor-1/2α. Recently, it was reported that ARNT is essential for adequate insulin secretion in response to glucose input and that its expression is downregulated in the pancreatic islets of diabetic patients. In the present study, the authors addressed the mechanism by which ARNT regulates insulin secretion in the INS-1 insulinoma cell line. In ARNT knock-down cells, basal insulin release was elevated, but insulin secretion was not further stimulated by a high-glucose challenge. Electrophysiological analyses revealed that glucose-dependent membrane depolarization was impaired in these cells. Furthermore, K ATP channel activity and expression were reduced. Of two K ATP channel subunits, Kir6.2 was found to be positively regulated by ARNT at the mRNA and protein levels. Based on these results, the authors suggest that ARNT expresses K ATP channel and by so doing regulates glucose-dependent insulin secretion.

  13. Suppression of Adaptive Immune Cell Activation Does Not Alter Innate Immune Adipose Inflammation or Insulin Resistance in Obesity.

    Directory of Open Access Journals (Sweden)

    Manikandan Subramanian

    Full Text Available Obesity-induced inflammation in visceral adipose tissue (VAT is a major contributor to insulin resistance and type 2 diabetes. Whereas innate immune cells, notably macrophages, contribute to visceral adipose tissue (VAT inflammation and insulin resistance, the role of adaptive immunity is less well defined. To address this critical gap, we used a model in which endogenous activation of T cells was suppressed in obese mice by blocking MyD88-mediated maturation of CD11c+ antigen-presenting cells. VAT CD11c+ cells from Cd11cCre+Myd88fl/fl vs. control Myd88fl/fl mice were defective in activating T cells in vitro, and VAT T and B cell activation was markedly reduced in Cd11cCre+Myd88fl/fl obese mice. However, neither macrophage-mediated VAT inflammation nor systemic inflammation were altered in Cd11cCre+Myd88fl/fl mice, thereby enabling a focused analysis on adaptive immunity. Unexpectedly, fasting blood glucose, plasma insulin, and the glucose response to glucose and insulin were completely unaltered in Cd11cCre+Myd88fl/fl vs. control obese mice. Thus, CD11c+ cells activate VAT T and B cells in obese mice, but suppression of this process does not have a discernible effect on macrophage-mediated VAT inflammation or systemic glucose homeostasis.

  14. Reduced levels of SCD1 accentuate palmitate-induced stress in insulin-producing β-cells

    Directory of Open Access Journals (Sweden)

    Hovsepyan Meri

    2010-09-01

    Full Text Available Abstract Background Stearoyl-CoA desaturase 1 (SCD1 is an ER resident enzyme introducing a double-bond in saturated fatty acids. Global knockout of SCD1 in mouse increases fatty acid oxidation and insulin sensitivity which makes the animal resistant to diet-induced obesity. Inhibition of SCD1 has therefore been proposed as a potential therapy of the metabolic syndrome. Much of the work has focused on insulin target tissue and very little is known about how reduced levels of SCD1 would affect the insulin-producing β-cell, however. The aim of the present study was therefore to investigate how reduced levels of SCD1 affect the β-cell. Results Insulin-secreting MIN6 cells with reduced levels of SCD1 were established by siRNA mediated knockdown. When fatty acid oxidation was measured, no difference between cells with reduced levels of SCD1 and mock-transfected cells were found. Also, reducing levels of SCD1 did not affect insulin secretion in response to glucose. To investigate how SCD1 knockdown affected cellular mechanisms, differentially regulated proteins were identified by a proteomic approach. Cells with reduced levels of SCD1 had higher levels of ER chaperones and components of the proteasome. The higher amounts did not protect the β-cell from palmitate-induced ER stress and apoptosis. Instead, rise in levels of p-eIF2α and CHOP after palmitate exposure was 2-fold higher in cells with reduced levels of SCD1 compared to mock-transfected cells. Accordingly, apoptosis rose to higher levels after exposure to palmitate in cells with reduced levels of SCD1 compared to mock-transfected cells. Conclusions In conclusion, reduced levels of SCD1 augment palmitate-induced ER stress and apoptosis in the β-cell, which is an important caveat when considering targeting this enzyme as a treatment of the metabolic syndrome.

  15. Metabolic and fibrinolytic response to changed insulin sensitivity in users of oral contraceptives

    DEFF Research Database (Denmark)

    Petersen, Kresten R.; Christiansen, Erik; Madsbad, Sten

    1999-01-01

    systems, are relevant in the evaluation of the risk of developing vascular disorders or diabetes in OC users. We studied insulin sensitivity index (S(I)), glucose effectiveness (S(g)), and insulin response in young, healthy women by frequently sampled intravenous glucose tolerance tests before and after...... randomization to 6 months of treatment with ethinyl estradiol in triphasic combination with norgestimate (n = 17) or gestodene (n = 20). Measurements of fasting triglycerides and antigen concentrations of tissue-type plasminogen activator (t-PA) and plasminogen activator inhibitor type 1 (PAI-1) were also...... included. Both compounds increased fasting plasma insulin and reduced S(i) but did not affect S(g). The relationships between S(i) and insulin response were unchanged. No consistent correlation between insulin sensitivity and triglycerides, t-PA, or PAI-1 were demonstrated before or during treatment. We...

  16. Human adipose tissue-derived mesenchymal stem cells differentiate into insulin, somatostatin, and glucagon expressing cells

    International Nuclear Information System (INIS)

    Timper, Katharina; Seboek, Dalma; Eberhardt, Michael; Linscheid, Philippe; Christ-Crain, Mirjam; Keller, Ulrich; Mueller, Beat; Zulewski, Henryk

    2006-01-01

    Mesenchymal stem cells (MSC) from mouse bone marrow were shown to adopt a pancreatic endocrine phenotype in vitro and to reverse diabetes in an animal model. MSC from human bone marrow and adipose tissue represent very similar cell populations with comparable phenotypes. Adipose tissue is abundant and easily accessible and could thus also harbor cells with the potential to differentiate in insulin producing cells. We isolated human adipose tissue-derived MSC from four healthy donors. During the proliferation period, the cells expressed the stem cell markers nestin, ABCG2, SCF, Thy-1 as well as the pancreatic endocrine transcription factor Isl-1. The cells were induced to differentiate into a pancreatic endocrine phenotype by defined culture conditions within 3 days. Using quantitative PCR a down-regulation of ABCG2 and up-regulation of pancreatic developmental transcription factors Isl-1, Ipf-1, and Ngn3 were observed together with induction of the islet hormones insulin, glucagon, and somatostatin

  17. Studies on interaction of insulin and insulin receptor in rat liver cell membranes

    Energy Technology Data Exchange (ETDEWEB)

    Sakai, Y; Hara, H; Kawate, R; Kawasaki, T [Hiroshima Univ. (Japan). School of Medicine

    1975-07-01

    Rat liver was homogenized with a Polytron PT 20 ST and fractionated by differential centrifugation. Prepared plasma membranes (100 ..mu..g protein) were incubated with enzymatically iodinated /sup 125/I-insulin (0.3 ng, specific activity 107 ..mu..Ci/..mu..g) in 25 mM Tris-HCl buffer, pH 7.5, containing 0.9% NaCl and 1% bovine serum albumin. The 12,000xg- and 17,000xg-sediments obtained after subfractionation of liver homogenates showed almost equally high specific binding activity with /sup 125/I-insulin and less activity was detected in the 600 g-, 5,000 g- and 40,000 g- sediments and the 40,000 g- supernatant. Specific binding of insulin with the membrane fraction was time-, temperature- and ionic strength-dependent. The highest binding was obtained under conditions in which the membrane fraction was incubated with insulin for 24 hours at 4/sup 0/C in the buffer containing 1 M NaCl. Under these conditions, specific binding of /sup 125/I-insulin was 26.8% of the total radioactivity. The effect of native insulin on the binding of /sup 125/I-insulin with the membrane fraction was studied in the range of 0--6.4 x 10/sup 5/ ..mu..U/ml of unlabeled insulin and a distinct competitive displacement of /sup 125/I-insulin with native insulin was observed between 10 and 10/sup 4/ ..mu..U/ml. Kinetic studies by Scatchard plot analysis of the above results revealed heterogeneity in insulin receptors or receptor sites, one with a high affinity of 10/sup 9/ M/sup -1/ order and the other with a low affinity of 10/sup 8/ M/sup -1/ order. Both affinities were also affected by temperature and ionic strength.

  18. Insulin resistance and chronic inflammation

    Directory of Open Access Journals (Sweden)

    Natalia Matulewicz

    2016-12-01

    Full Text Available Insulin resistance is a condition of reduced biological response to insulin. Growing evidence indicates the role of the chronic low-grade inflammatory response in the pathogenesis of insulin resistance. Adipose tissue in obesity is characterized by increased lipolysis with the excessive release of free fatty acids, and is also a source of proinflammatory cytokines. Both these factors may inhibit insulin action. Proinflammatory cytokines exert their effect by stimulating major inflammatory NFκB and JNK pathways within the cells. Inflammatory processes in other insulin responsive tissues may also play a role in inducing insulin resistance. This paper is an overview of the chronic low-grade inflammation in adipose tissue, skeletal muscle, liver and endothelial cells during the development of insulin resistance.

  19. FoxO1 Plays an Important Role in Regulating ?-Cell Compensation for Insulin Resistance in Male Mice

    OpenAIRE

    Zhang, Ting; Kim, Dae Hyun; Xiao, Xiangwei; Lee, Sojin; Gong, Zhenwei; Muzumdar, Radhika; Calabuig-Navarro, Virtu; Yamauchi, Jun; Harashima, Hideyoshi; Wang, Rennian; Bottino, Rita; Alvarez-Perez, Juan Carlos; Garcia-Oca?a, Adolfo; Gittes, George; Dong, H. Henry

    2016-01-01

    ?-Cell compensation is an essential mechanism by which ?-cells increase insulin secretion for overcoming insulin resistance to maintain euglycemia in obesity. Failure of ?-cells to compensate for insulin resistance contributes to insulin insufficiency and overt diabetes. To understand the mechanism of ?-cell compensation, we characterized the role of forkhead box O1 (FoxO1) in ?-cell compensation in mice under physiological and pathological conditions. FoxO1 is a key transcription factor that...

  20. Glucose-Responsive Implantable Polymeric Microdevices for "Smart" Insulin Therapy of Diabetes

    Science.gov (United States)

    Chu, Michael Kok Loon

    Diabetes mellitus is a chronic illness manifested by improper blood glucose management, affecting over 350 million worldwide. As a result, all type 1 patients and roughly 20% of type 2 patients require exogenous insulin therapy to survive. Typically, daily multiple injections are taken to maintain normal glucose levels in response glucose spikes from meals. However, patient compliance and dosing accuracy can fluctuate with variation in meals, exercise, glucose metabolism or stress, leading to poor clinical outcomes. A 'smart', closed-loop insulin delivery system providing on-demand release kinetics responding to circulating glucose levels would be a boon for diabetes patients, replacing constant self monitoring and insulin. This thesis focuses on the development of a novel, 'smart' insulin microdevice that can provide on-demand insulin release in response to blood glucose levels. In the early stage, the feasibility of integrating a composite membrane with pH-responsive nanoparticles embedded in ethylcellulose membrane to provide pH-responsive in vitro release was examined and confirmed using a model drug, vitamin B12. In the second microdevice, glucose oxidase for generating pH signals from glucose oxidation, catalase and manganese dioxide nanoparticles, as peroxide scavengers, were used in a bioinorganic, albumin-based membrane cross-linked with a polydimethylsiloxane (PDMS) grid-microdevice system. This prototype device demonstrated insulin release in response to glucose levels in vitro and regulating plasma glucose in type 1 diabetic rats when implanted intraperitoneally. Advancement allowing for subcutaneous implantation and improved biocompatibility was achieved with surface modification of PDMS microdevices grafted with activated 20 kDa polyethylene glycol (PEG) chains, dramatically reducing immune response and local inflammation. When implanted subcutaneously in diabetic rats, glucose-responsive insulin delivery microdevices showed short and long

  1. Melatonin Modulates Neuronal Cell Death Induced by Endoplasmic Reticulum Stress under Insulin Resistance Condition.

    Science.gov (United States)

    Song, Juhyun; Kim, Oh Yoen

    2017-06-10

    Insulin resistance (IR) is an important stress factor in the central nervous system, thereby aggravating neuropathogenesis and triggering cognitive decline. Melatonin, which is an antioxidant phytochemical and synthesized by the pineal gland, has multiple functions in cellular responses such as apoptosis and survival against stress. This study investigated whether melatonin modulates the signaling of neuronal cell death induced by endoplasmic reticulum (ER) stress under IR condition using SH-SY5Y neuroblastoma cells. Apoptosis cell death signaling markers (cleaved Poly [ADP-ribose] polymerase 1 (PARP), p53, and Bax) and ER stress markers (phosphorylated eIF2α (p-eIF2α), ATF4, CHOP, p-IRE1 , and spliced XBP1 (sXBP1)) were measured using reverse transcription-PCR, quantitative PCR, and western blottings. Immunofluorescence staining was also performed for p-ASK1 and p-IRE1 . The mRNA or protein expressions of cell death signaling markers and ER stress markers were increased under IR condition, but significantly attenuated by melatonin treatment. Insulin-induced activation of ASK1 ( p-ASK1 ) was also dose dependently attenuated by melatonin treatment. The regulatory effect of melatonin on neuronal cells under IR condition was associated with ASK1 signaling. In conclusion, the result suggested that melatonin may alleviate ER stress under IR condition, thereby regulating neuronal cell death signaling.

  2. Rising Intracellular Zinc by Membrane Depolarization and Glucose in Insulin-Secreting Clonal HIT-T15 Beta Cells

    Directory of Open Access Journals (Sweden)

    Kira G. Slepchenko

    2012-01-01

    Full Text Available Zinc (Zn2+ appears to be intimately involved in insulin metabolism since insulin secretion is correlated with zinc secretion in response to glucose stimulation, but little is known about the regulation of zinc homeostasis in pancreatic beta-cells. This study set out to identify the intracellular zinc transient by imaging free cytosolic zinc in HIT-T15 beta-cells with fluorescent zinc indicators. We observed that membrane depolarization by KCl (30–60 mM was able to induce a rapid increase in cytosolic concentration of zinc. Multiple zinc transients of similar magnitude were elicited during repeated stimulations. The amplitude of zinc responses was not affected by the removal of extracellular calcium or zinc. However, the half-time of the rising slope was significantly slower after removing extracellular zinc with zinc chelator CaEDTA, suggesting that extracellular zinc affect the initial rising phase of zinc response. Glucose (10 mM induced substantial and progressive increases in intracellular zinc concentration in a similar way as KCl, with variation in the onset and the duration of zinc mobilization. It is known that the depolarization of beta-cell membrane is coupled with the secretion of insulin. Rising intracellular zinc concentration may act as a critical signaling factor in insulin metabolism of pancreatic beta-cells.

  3. Rising intracellular zinc by membrane depolarization and glucose in insulin-secreting clonal HIT-T15 beta cells.

    Science.gov (United States)

    Slepchenko, Kira G; Li, Yang V

    2012-01-01

    Zinc (Zn(2+)) appears to be intimately involved in insulin metabolism since insulin secretion is correlated with zinc secretion in response to glucose stimulation, but little is known about the regulation of zinc homeostasis in pancreatic beta-cells. This study set out to identify the intracellular zinc transient by imaging free cytosolic zinc in HIT-T15 beta-cells with fluorescent zinc indicators. We observed that membrane depolarization by KCl (30-60 mM) was able to induce a rapid increase in cytosolic concentration of zinc. Multiple zinc transients of similar magnitude were elicited during repeated stimulations. The amplitude of zinc responses was not affected by the removal of extracellular calcium or zinc. However, the half-time of the rising slope was significantly slower after removing extracellular zinc with zinc chelator CaEDTA, suggesting that extracellular zinc affect the initial rising phase of zinc response. Glucose (10 mM) induced substantial and progressive increases in intracellular zinc concentration in a similar way as KCl, with variation in the onset and the duration of zinc mobilization. It is known that the depolarization of beta-cell membrane is coupled with the secretion of insulin. Rising intracellular zinc concentration may act as a critical signaling factor in insulin metabolism of pancreatic beta-cells.

  4. Acrolein decreases endothelial cell migration and insulin sensitivity through induction of let-7a.

    Science.gov (United States)

    O'Toole, Timothy E; Abplanalp, Wesley; Li, Xiaohong; Cooper, Nigel; Conklin, Daniel J; Haberzettl, Petra; Bhatnagar, Aruni

    2014-08-01

    Acrolein is a major reactive component of vehicle exhaust, and cigarette and wood smoke. It is also present in several food substances and is generated endogenously during inflammation and lipid peroxidation. Although previous studies have shown that dietary or inhalation exposure to acrolein results in endothelial activation, platelet activation, and accelerated atherogenesis, the basis for these effects is unknown. Moreover, the effects of acrolein on microRNA (miRNA) have not been studied. Using AGILENT miRNA microarray high-throughput technology, we found that treatment of cultured human umbilical vein endothelial cells with acrolein led to a significant (>1.5-fold) upregulation of 12, and downregulation of 15, miRNAs. Among the miRNAs upregulated were members of the let-7 family and this upregulation was associated with decreased expression of their protein targets, β3 integrin, Cdc34, and K-Ras. Exposure to acrolein attenuated β3 integrin-dependent migration and reduced Akt phosphorylation in response to insulin. These effects of acrolein on endothelial cell migration and insulin signaling were reversed by expression of a let-7a inhibitor. Also, inhalation exposure of mice to acrolein (1 ppm x 6 h/day x 4 days) upregulated let-7a and led to a decrease in insulin-stimulated Akt phosphorylation in the aorta. These results suggest that acrolein exposure has broad effects on endothelial miRNA repertoire and that attenuation of endothelial cell migration and insulin signaling by acrolein is mediated in part by the upregulation of let-7a. This mechanism may be a significant feature of vascular injury caused by inflammation, oxidized lipids, and exposure to environmental pollutants. © The Author 2014. Published by Oxford University Press on behalf of the Society of Toxicology. All rights reserved. For permissions, please email: journals.permissions@oup.com.

  5. Differential insulin and steroidogenic signaling in insulin resistant and non-insulin resistant human luteinized granulosa cells-A study in PCOS patients.

    Science.gov (United States)

    Belani, Muskaan; Deo, Abhilash; Shah, Preeti; Banker, Manish; Singal, Pawan; Gupta, Sarita

    2018-04-01

    Insulin resistance (IR) is one of the significant aberrations in polycystic ovarian syndrome (PCOS), however is only observed in 70%-80% of obese PCOS and 20%-25% of lean PCOS. Hyperinsulinemia accompanies PCOS-IR along with hyperandrogenemia against normal insulin and androgen levels in PCOS-non insulin resistance (NIR). This could possibly be due to defects in the downstream signaling pathways. The study thus aims to unravel insulin and steroidogenic signaling pathways in luteinized granulosa cells isolated from PCOS-IR and NIR vs matched controls. Luteinized granulosa cells from 30 controls and 39 PCOS were classified for IR based on a novel method of down regulation of protein expression of insulin receptor-β (INSR- β) as shown in our previous paper. We evaluated expression of molecules involved in insulin, steroidogenic signaling and lipid metabolism in luteinized granulosa cells followed by analysis of estradiol, progesterone and testosterone in follicular fluid. Protein expression of INSR- β, pIRS (ser 307), PI(3)K, PKC-ζ, pAkt, ERK1/2, pP38MAPK and gene expression of IGF showed differential expression in the two groups. Increased protein expression of PPAR-γ was accompanied by up regulation in SREBP1c, FAS, CPT-1 and ACC-1 genes in PCOS-IR group. Expression of StAR, CYP19A1, 17 β- HSD and 3 β- HSD demonstrated significant decrease along with increase in CYP11A1, FSH-R and LH-R in both the groups. Follicular fluid testosterone increased and progesterone decreased in PCOS-IR group. This study shows how candidate molecules that were differentially expressed, aid in designing targeted therapy against the two phenotypes of PCOS. Copyright © 2018 Elsevier Ltd. All rights reserved.

  6. Expression of the growth hormone receptor gene in insulin producing cells

    DEFF Research Database (Denmark)

    Møldrup, Annette; Billestrup, N; Nielsen, Jens Høiriis

    1990-01-01

    Growth hormone (GH) plays a dual role in glucose homeostasis. On the one hand, it exerts an insulin antagonistic effect on the peripheral tissue, on the other hand, it stimulates insulin biosynthesis and beta-cell proliferation. The expression of GH-receptors on the rat insulinoma cell line RIN-5...

  7. Insulin redirects differentiation from cardiogenic mesoderm and endoderm to neuroectoderm in differentiating human embryonic stem cells.

    NARCIS (Netherlands)

    Freund, C.M.A.H.; Ward-van Oostwaard, D.; Monshouwer-Kloots, J.; van den Brink, S.; van Rooijen, M.A.; Xu, X.; Zweigerdt, R.; Mummery, C.L.; Passier, R.

    2008-01-01

    Human embryonic stem cells (hESC) can proliferate indefinitely while retaining the capacity to form derivatives of all three germ layers. We have reported previously that hESC differentiate into cardiomyocytes when cocultured with a visceral endoderm-like cell line (END-2). Insulin/insulin-like

  8. Insulin-producing Cells from Adult Human Bone Marrow Mesenchymal Stromal Cells Could Control Chemically Induced Diabetes in Dogs: A Preliminary Study.

    Science.gov (United States)

    Gabr, Mahmoud M; Zakaria, Mahmoud M; Refaie, Ayman F; Ismail, Amani M; Khater, Sherry M; Ashamallah, Sylvia A; Azzam, Maha M; Ghoneim, Mohamed A

    2018-01-01

    Ten mongrel dogs were used in this study. Diabetes was chemically induced in 7 dogs, and 3 dogs served as normal controls. For each diabetic dog, 5 million human bone marrow-derived mesenchymal stem cells/kg were differentiated to form insulin-producing cells using a trichostatin-based protocol. Cells were then loaded in 2 TheraCyte capsules which were transplanted under the rectus sheath. One dog died 4 d postoperatively from pneumonia. Six dogs were followed up with for 6 to 18 mo. Euglycemia was achieved in 4 dogs. Their glucose tolerance curves exhibited a normal pattern demonstrating that the encapsulated cells were glucose sensitive and insulin responsive. In the remaining 2 dogs, the fasting blood sugar levels were reduced but did not reach normal values. The sera of all transplanted dogs contained human insulin and C-peptide with a negligible amount of canine insulin. Removal of the transplanted capsules was followed by prompt return of diabetes. Intracytoplasmic insulin granules were seen by immunofluorescence in cells from the harvested capsules. Furthermore, all pancreatic endocrine genes were expressed. This study demonstrated that the TheraCyte capsule or a similar device can provide adequate immunoisolation, an important issue when stem cells are considered for the treatment of type 1 diabetes mellitus.

  9. A New Method for Generating Insulin-Secreting Cells from Human Pancreatic Epithelial Cells After Islet Isolation Transformed by NeuroD1

    Science.gov (United States)

    Shimoda, Masayuki; Chen, Shuyuan; Noguchi, Hirofumi; Takita, Morihito; Sugimoto, Koji; Itoh, Takeshi; Chujo, Daisuke; Iwahashi, Shuichi; Naziruddin, Bashoo; Levy, Marlon F.

    2014-01-01

    Abstract The generation of insulin-secreting cells from nonendocrine pancreatic epithelial cells (NEPEC) has been demonstrated for potential clinical use in the treatment of diabetes. However, previous methods either had limited efficacy or required viral vectors, which hinder clinical application. In this study, we aimed to establish an efficient method of insulin-secreting cell generation from NEPEC without viral vectors. We used nonislet fractions from both research-grade human pancreata from brain-dead donors and clinical pancreata after total pancreatectomy with autologous islet transplantation to treat chronic pancreatitis. It is of note that a few islets could be mingled in the nonislet fractions, but their influence could be limited. The NeuroD1 gene was induced into NEPEC using an effective triple lipofection method without viral vectors to generate insulin-secreting cells. The differentiation was promoted by adding a growth factor cocktail into the culture medium. Using the research-grade human pancreata, the effective method showed high efficacy in the differentiation of NEPEC into insulin-positive cells that secreted insulin in response to a glucose challenge and improved diabetes after being transplanted into diabetic athymic mice. Using the clinical pancreata, similar efficacy was obtained, even though those pancreata suffered chronic pancreatitis. In conclusion, our effective differentiation protocol with triple lipofection method enabled us to achieve very efficient insulin-secreting cell generation from human NEPEC without viral vectors. This method offers the potential for supplemental insulin-secreting cell transplantation for both allogeneic and autologous islet transplantation. PMID:24845703

  10. Protein kinase N2 regulates AMP kinase signaling and insulin responsiveness of glucose metabolism in skeletal muscle.

    Science.gov (United States)

    Ruby, Maxwell A; Riedl, Isabelle; Massart, Julie; Åhlin, Marcus; Zierath, Juleen R

    2017-10-01

    Insulin resistance is central to the development of type 2 diabetes and related metabolic disorders. Because skeletal muscle is responsible for the majority of whole body insulin-stimulated glucose uptake, regulation of glucose metabolism in this tissue is of particular importance. Although Rho GTPases and many of their affecters influence skeletal muscle metabolism, there is a paucity of information on the protein kinase N (PKN) family of serine/threonine protein kinases. We investigated the impact of PKN2 on insulin signaling and glucose metabolism in primary human skeletal muscle cells in vitro and mouse tibialis anterior muscle in vivo. PKN2 knockdown in vitro decreased insulin-stimulated glucose uptake, incorporation into glycogen, and oxidation. PKN2 siRNA increased 5'-adenosine monophosphate-activated protein kinase (AMPK) signaling while stimulating fatty acid oxidation and incorporation into triglycerides and decreasing protein synthesis. At the transcriptional level, PKN2 knockdown increased expression of PGC-1α and SREBP-1c and their target genes. In mature skeletal muscle, in vivo PKN2 knockdown decreased glucose uptake and increased AMPK phosphorylation. Thus, PKN2 alters key signaling pathways and transcriptional networks to regulate glucose and lipid metabolism. Identification of PKN2 as a novel regulator of insulin and AMPK signaling may provide an avenue for manipulation of skeletal muscle metabolism. Copyright © 2017 the American Physiological Society.

  11. Insulin receptor substrates 1 and 2 but not Shc can activate the insulin receptor independent of insulin and induce proliferation in CHO-IR cells

    International Nuclear Information System (INIS)

    Niessen, Markus; Jaschinski, Frank; Item, Flurin; McNamara, Morgan P.; Spinas, Giatgen A.; Trueb, Thomas

    2007-01-01

    Ligand-activated insulin receptor (IR) attracts and phosphorylates various substrates such as insulin receptor substrates 1-4 (IRS) and Shc. To investigate how binding affinity for substrate affects signalling we generated chimeric receptors with the β-chain of the insulin receptor containing NPXY motives with different affinities for receptor substrates. We found that the extent of receptor tyrosine phosphorylation positively correlates with binding affinity towards IRS1/2 but not towards Shc. Moreover, overexpression of IRS1 or IRS2 but not of Shc increased IR tyrosine phosphorylation in a dose-dependent manner, also independent of insulin. Molecular truncations of IRS1 revealed that neither the isolated PH and PTB domains nor the C-terminus with the tyrosine phosphorylation sites alone are sufficient for substrate-dependent receptor activation. Overexpression of IRS1 and IRS2 impaired insulin-induced internalization of the IR in a dose-dependent manner suggesting that IRS proteins prevent endosome-associated receptor dephosphorylation/inactivation. IRS1 and IRS2 could therefore target the activated IR to different cellular compartments. Overexpression of IRS1 and IRS2 inhibited insulin-stimulated activation of the MAP kinases Erk1/2 while it increased/induced activation of Akt/PKB. Finally, overexpression of IRS1 and IRS2 but not of Shc induced DNA synthesis in starved CHO-IR cells independent of exogenous growth factors. Our results demonstrate that variations in cellular IRS1 and IRS2 concentration affect insulin signalling both upstream and downstream and that IRS proteins could play instructive rather than just permissive roles in signal transmission

  12. Insulin-like growth factor 1 enhances the migratory capacity of mesenchymal stem cells

    International Nuclear Information System (INIS)

    Li, Yangxin; Yu, XiYong; Lin, ShuGuang; Li, XiaoHong; Zhang, Saidan; Song, Yao-Hua

    2007-01-01

    Mesenchymal stem cells (MSCs) are attractive candidates for cell based therapies. However, the mechanisms responsible for stem cell migration and homing after transplantation remain unknown. It has been shown that insulin-like growth factor-1 (IGF-1) induces proliferation and migration of some cell types, but its effects on stem cells have not been investigated. We isolated and cultured MSC from rat bone marrow, and found that IGF-1 increased the expression levels of the chemokine receptor CXCR4 (receptor for stromal cell-derived factor-1, SDF-1). Moreover, IGF-1 markedly increased the migratory response of MSC to SDF-1. The IGF-1-induced increase in MSC migration in response to SDF-1 was attenuated by PI3 kinase inhibitor (LY294002 and wortmannin) but not by mitogen-activated protein/ERK kinase inhibitor PD98059. Our data indicate that IGF-1 increases MSC migratory responses via CXCR4 chemokine receptor signaling which is PI3/Akt dependent. These findings provide a new paradigm for biological effects of IGF-1 on MSC and have implications for the development of novel stem cell therapeutic strategies

  13. The Role of lysophosphatidic acide (LPA) in the insulin resistence of the pancreatic β-cells

    OpenAIRE

    Mourad Agha, Zein

    2016-01-01

    The pathogenesis of the type-2-diabetes mellitus underlying is characterized by a combination of peripheral insulin resistance, β-cell dysfunction and reduction in the β cell mass. The increasing of FFA level or their metabolites lead to inhibition of insulin signaling. Consequent, the ability of insulin is reduced and therefore lead to insulin resistance. LPA is a lipid mediator that is associated with a progression of T2D. It has been suggested that LPA and the development of obesity are st...

  14. Atorvastatin inhibits insulin synthesis by inhibiting the Ras/Raf/ERK/CREB pathway in INS-1 cells

    Science.gov (United States)

    Sun, Hongxi; Li, Yu; Sun, Bei; Hou, Ningning; Yang, Juhong; Zheng, Miaoyan; Xu, Jie; Wang, Jingyu; Zhang, Yi; Zeng, Xianwei; Shan, Chunyan; Chang, Bai; Chen, Liming; Chang, Baocheng

    2016-01-01

    Abstract Backround: Type 2 diabetes has become a global epidemic disease. Atorvastatin has become a cornerstone in the prevention and treatment of atherosclerosis. However, increasing evidence showed that statins can dose-dependently increase the risk of diabetes mellitus. The mechanism is not clear. Objective: The Ras complex pathway (Ras/Raf/extracellular signal-regulated kinase [ERK]/cAMP response element-binding protein [CREB]) is the major pathway that regulates the gene transcription. Except for the inhibition of cholesterol synthesis by inhibiting the 3-hydroxy-3-methyl glutaryl coenzyme A (HMG-COA) reductase, statins can also downregulate the phosphorylation of a series of downstream substrates including the key proteins of the Ras complex pathway, therefore may inhibit the insulin syntheses in pancreatic beta cells. In our study, we investigated the inhibitory effect and the underlying mechanism of atorvastatin on insulin synthesis in rat islets. Methods: Islets were isolated from Wistar rats and cultured in Roswell Park Memorial Institute (RPMI)-1640 medium. The insulin content in the medium was measured by radioimmunoassay before and after the treatment of 50 μM atorvastatin. Effect of atorvastatin on the expression of insulin message Ribonucleic acid (mRNA) in pancreatic islet beta cells was also detected using quantitative real-time polymerase chain reaction. Western blotting was used to explore the possible role of the Ras complex pathway (Ras/Raf/ERK/CREB) in atorvastatin-inhibited insulin synthesis. The effects of atorvastatin on the binding of nuclear transcription factor p-CREB with CRE in INS-1 cells were examined via chromatin immunoprecipitation assay. Results: Compared with the control group, the insulin level decreased by 27.1% at 24 hours after atorvastatin treatment. Atorvastatin inhibited insulin synthesis by decreasing insulin mRNA expression of pancreatic islet beta cells. The activities of Ras, Raf-1, and p-CREB in the Ras complex

  15. Glucagon-insulin interaction on fat cell metabolism using c14 glucose

    International Nuclear Information System (INIS)

    Zewail, M.A.; Nielsen, J.H.

    1984-01-01

    Glucagon is known to stimulate the lipolysis in isolated fat cells from young rats, but not in fat cells from old heavy rate (Manganiello 1972). Insulin is known to counteract the lipolytic effect and to stimulate the synthesis of fatty acids from glucose. However, little is known about the interaction between the two hormones on the glucose metabolism. Experiments based on the use of various inhibitors of lipolysis have however, clearly shown that glucagon can also stimulate the entry and overall oxidation of glucose by mechanism which is distinct from its lipolysis stimulating mechanism (M. Blecher et al. 1969). Fat cells from old heavy rats are known to be less responsive to both the lipogenic action of insulin and the lipolytic action of glucagon than fat cells from young lean rats (E.G. Hansen, Nielsen and Gliemann, 1974). The aim of the present study was to see how glucagon affects glucose metabolism in fat cells, and whether this effect was dependent on the lipolytic action of glucagon

  16. Efficient Differentiation of Mouse Embryonic Stem Cells into Insulin-Producing Cells

    Directory of Open Access Journals (Sweden)

    Szu-Hsiu Liu

    2012-01-01

    Full Text Available Embryonic stem (ES cells are a potential source of a variety of differentiated cells for cell therapy, drug discovery, and toxicology screening. Here, we present an efficacy strategy for the differentiation of mouse ES cells into insulin-producing cells (IPCs by a two-step differentiation protocol comprising of (i the formation of definitive endoderm in monolayer culture by activin A, and (ii this monolayer endoderm being induced to differentiate into IPCs by nicotinamide, insulin, and laminin. Differentiated cells can be obtained within approximately 7 days. The differentiation IPCs combined application of RT-PCR, ELISA, and immunofluorescence to characterize phenotypic and functional properties. In our study, we demonstrated that IPCs produced pancreatic transcription factors, endocrine progenitor marker, definitive endoderm, pancreatic β-cell markers, and Langerhans α and δ cells. The IPCs released insulin in a manner that was dose dependent upon the amount of glucose added. These techniques may be able to be applied to human ES cells, which would have very important ramifications for treating human disease.

  17. Gender discrimination in the influence of hyperglycemia and hyperosmolarity on rat aortic tissue responses to insulin.

    Science.gov (United States)

    Wong, Nikki L; Achike, Francis I

    2010-08-09

    Hyperglycaemia initiates endothelial dysfunction causing diabetic macro- and micro-vasculopathy, the main causes of morbidity and mortality in diabetes mellitus. The vasculopathy exhibits gender peculiarities. We therefore explored gender differences in comparing the effects of hyperglycaemia (50 mM) per se with its hyperosmolar (50 mM) effects on vascular tissue responses to insulin. Endothelium-intact or denuded thoracic aortic rings from age-matched male and female Sprague-Dawley rats were incubated for 10 min or 6 h (acute versus chronic exposure) in normal, hyperglycaemic or hyperosmolar Krebs solution. Relaxant responses to insulin (6.9x10(-7)-6.9x10(-5) M) of the phenylephrine-contracted tissues were recorded. Endothelium denudation in both genders inhibited relaxation to insulin in all conditions, more significantly in female than in male tissues, suggesting the female response to insulin is more endothelium-dependent than the male. Acutely and chronically exposed normoglycemic endothelium-intact or -denuded tissues responded similarly to insulin. Chronic hyperglycemic or hyperosmolar exposure did not alter the endothelium-denuded tissue responses to insulin, whereas the responses of the endothelium-intact male and female hyperosmolar, and male hyperglycemic tissues were enhanced. The results show that insulin exerts an endothelium-dependent and independent relaxation with the female tissue responses more endothelium-dependent than the male. The data also suggest that hyperosmolarity per se enhances aortic tissue relaxant responses to insulin whereas hyperglycemia per se inhibits the same and more so in female than male tissues. These effects are endothelium-dependent. Copyright (c) 2010 Elsevier B.V. All rights reserved.

  18. Effects of a hypercaloric diet on β-cell responsivity in lean healthy men

    NARCIS (Netherlands)

    Brands, Myrte; Swat, Maciej; Lammers, Nicolette M.; Sauerwein, Hans P.; Endert, Erik; Ackermans, Mariëtte T.; Verhoeven, Arthur J.; Serlie, Mireille J.

    2013-01-01

    Insulin resistance and hyperinsulinaemia precede the onset of obesity-induced DM2. The early adaptation of the β-cell during the initial phase of overfeeding and weight gain has only been partly elucidated. We studied the early changes in insulin clearance and β-cell responsivity during a positive

  19. [In vitro generation of insulin-producing cells from the neonatal rat bone marrow mesenchymal stem cells].

    Science.gov (United States)

    Li, Xiaohu; Huang, Haiyan; Liu, Xirong; Xia, Hongxia; Li, Mincai

    2015-03-01

    To observe the differentiation of the neonatal rat bone marrow mesenchymal stem cells (MSCs) into insulin-producing cells and detect the expressions of insulin, pancreatic duodenal homebox-1 (PDX-1) and nestin. MSCs were isolated from the neonatal rats and cultured in the modified medium composed of 10 μg/L human epidermal growth factor (EGF), 10 μg/L basic fibroblast growth factor (bFGF), 10 μg/L hepatocyte growth factor (HGF), 10 μg/L human B cell regulin, 20 mmol/L nicotinamide and 20 g/L B27. After the induction, the mRNA expressions of insulin, PDX-1 and nestin were examined by reverse transcription-PCR, and the insulin, PDX-1 and nestin protein levels were detected by immunocytochemistry. The insulin and PDX-1 mRNA expressions increased and the nestin mRNA expression decreased in the differentiation of the neonatal rat MSCs into insulin-producing cells. The nestin, PDX-1 and insulin proteins were co-expressed in insulin-producing cells. MSCs can be induced to differentiate into insulin-producing cells.

  20. Effect of test meals of varying dietary fiber content on plasma insulin and glucose response.

    Science.gov (United States)

    Potter, J G; Coffman, K P; Reid, R L; Krall, J M; Albrink, M J

    1981-03-01

    To assess the effect of dietary fiber on glucose tolerance four different meals of varying fiber content but identical protein fat and carbohydrate content were fed to eight healthy men aged 22 to 45. Each meal provided 75 g of carbohydrate as liquid glucose formula, as brown rice, pinto beans, or All Bran. The mean plasma glucose and insulin responses were highest following the formula, and least for All Bran and pinto beans. Rice produced nearly as great a rise in insulin and glucose as did the formula. The rank of each meal by content of neutral detergent fiber was nearly the inverse of the rank by magnitude of the insulin response evoked, fiber content being greatest in All Bran (18 g) and pinto beans (16.2 g), low in rice (2.8 g) and absent from the formula. It was concluded that dietary fiber dampened the insulin response to a high carbohydrate meal.

  1. Brain Insulin Administration Triggers Distinct Cognitive and Neurotrophic Responses in Young and Aged Rats.

    Science.gov (United States)

    Haas, Clarissa B; Kalinine, Eduardo; Zimmer, Eduardo R; Hansel, Gisele; Brochier, Andressa W; Oses, Jean P; Portela, Luis V; Muller, Alexandre P

    2016-11-01

    Aging is a major risk factor for cognitive deficits and neurodegenerative disorders, and impaired brain insulin receptor (IR) signaling is mechanistically linked to these abnormalities. The main goal of this study was to investigate whether brain insulin infusions improve spatial memory in aged and young rats. Aged (24 months) and young (4 months) male Wistar rats were intracerebroventricularly injected with insulin (20 mU) or vehicle for five consecutive days. The animals were then assessed for spatial memory using a Morris water maze. Insulin increased memory performance in young rats, but not in aged rats. Thus, we searched for cellular and molecular mechanisms that might account for this distinct memory response. In contrast with our expectation, insulin treatment increased the proliferative activity in aged rats, but not in young rats, implying that neurogenesis-related effects do not explain the lack of insulin effects on memory in aged rats. Furthermore, the expression levels of the IR and downstream signaling proteins such as GSK3-β, mTOR, and presynaptic protein synaptophysin were increased in aged rats in response to insulin. Interestingly, insulin treatment increased the expression of the brain-derived neurotrophic factor (BDNF) and tropomyosin receptor kinase B (TrkB) receptors in the hippocampus of young rats, but not of aged rats. Our data therefore indicate that aged rats can have normal IR downstream protein expression but failed to mount a BDNF response after challenge in a spatial memory test. In contrast, young rats showed insulin-mediated TrkB/BDNF response, which paralleled with improved memory performance.

  2. Patients with neuroglycopenia after gastric bypass surgery have exaggerated incretin and insulin secretory responses to a mixed meal.

    Science.gov (United States)

    Goldfine, A B; Mun, E C; Devine, E; Bernier, R; Baz-Hecht, M; Jones, D B; Schneider, B E; Holst, J J; Patti, M E

    2007-12-01

    Hyperinsulinemic hypoglycemia is newly recognized as a rare but important complication after Roux-en-Y gastric bypass (GB). The etiology of the syndrome and metabolic characteristics remain incompletely understood. Recent studies suggest that levels of incretin hormones are increased after GB and may promote excessive beta-cell function and/or growth. We performed a cross-sectional analysis of metabolic variables, in both the fasting state and after a liquid mixed-meal challenge, in four subject groups: 1) with clinically significant hypoglycemia [neuroglycopenia (NG)] after GB surgery, 2) with no symptoms of hypoglycemia at similar duration after GB surgery, 3) without GB similar to preoperative body mass index of the surgical cohorts, and 4) without GB similar to current body mass index of the surgical cohorts. Insulin and C-peptide after the liquid mixed meal were both higher relative to the glucose level achieved in persons after GB with NG compared with asymptomatic individuals. Glucagon, glucagon-like peptide 1, and glucose-dependent insulinotropic peptide levels were higher in both post-GB surgical groups compared with both overweight and morbidly obese persons, and glucagon-like peptide 1 was markedly higher in the group with NG. Insulin resistance, assessed by homeostasis model assessment of insulin resistance, the composite insulin sensitivity index, or adiponectin, was similar in both post-GB groups. Dumping score was also higher in both GB groups but did not discriminate between asymptomatic and symptomatic patients. Notably, the frequency of asymptomatic hypoglycemia after a liquid mixed meal was high in post-GB patients. A robust insulin secretory response was associated with postprandial hypoglycemia in patients after GB presenting with NG. Increased incretin levels may contribute to the increased insulin secretory response.

  3. Detection of Transketolase in Bone Marrow—Derived Insulin-Producing Cells: Benfotiamine Enhances Insulin Synthesis and Glucose Metabolism

    OpenAIRE

    Oh, Seh-Hoon; Witek, Rafal P.; Bae, Si-Hyun; Darwiche, Houda; Jung, Youngmi; Pi, Liya; Brown, Alicia; Petersen, Bryon E.

    2009-01-01

    Adult bone marrow (BM)-derived insulin-producing cells (IPCs) are capable of regulating blood glucose levels in chemically induced hyperglycemic mice. Using cell transplantation therapy, fully functional BM-derived IPCs help to mediate treatment of diabetes mellitus. Here, we demonstrate the detection of the pentose phosphate pathway enzyme, transketolase (TK), in BM-derived IPCs cultured under high-glucose conditions. Benfotiamine, a known activator of TK, was not shown to affect the prolife...

  4. Pancreatic β-Cell Electrical Activity and Insulin Secretion: of Mice and Men

    Science.gov (United States)

    Rorsman, Patrik; Ashcroft, Frances M

    2018-01-01

    The pancreatic β-cell plays a key role in glucose homeostasis by secreting insulin, the only hormone capable of lowering the blood glucose concentration. Impaired insulin secretion results in the chronic hyperglycaemia that characterizes type 2 diabetes (T2DM), which currently afflicts >450 million people worldwide. The healthy β-cell acts as a glucose sensor matching its output to the circulating glucose concentration. It does so via metabolically induced changes in electrical activity, which culminate in an increase in the cytoplasmic Ca2+ concentration and initiation of Ca2+-dependent exocytosis of insulin-containing secretory granules. Here, we review recent advances in our understanding of the β-cell transcriptome, electrical activity and insulin exocytosis. We highlight salient differences between mouse and human β-cells, provide models of how the different ion channels contribute to their electrical activity and insulin secretion, and conclude by discussing how these processes become perturbed in T2DM. PMID:29212789

  5. Mitochondrial Dysfunction Contributes to Impaired Insulin Secretion in INS-1 Cells with Dominant-negative Mutations of HNF-1α and in HNF-1α-deficient Islets*

    OpenAIRE

    Pongratz, Rebecca L.; Kibbey, Richard G.; Kirkpatrick, Clare L.; Zhao, Xiaojian; Pontoglio, Marco; Yaniv, Moshe; Wollheim, Claes B.; Shulman, Gerald I.; Cline, Gary W.

    2009-01-01

    Maturity Onset Diabetes of the Young-type 3 (MODY-3) has been linked to mutations in the transcription factor hepatic nuclear factor (HNF)-1α, resulting in deficiency in glucose-stimulated insulin secretion. In INS-1 cells overexpressing doxycycline-inducible HNF-1α dominant-negative (DN-) gene mutations, and islets from Hnf-1α knock-out mice, insulin secretion was impaired in response to glucose (15 mm) and other nutrient secretagogues. Decreased rates of insulin secretion in response to glu...

  6. Characterization of Insulin-Immunoreactive Cells and Endocrine Cells Within the Duct System of the Adult Human Pancreas.

    Science.gov (United States)

    Li, Rong; Zhang, Xiaoxi; Yu, Lan; Zou, Xia; Zhao, Hailu

    2016-01-01

    The adult pancreatic duct system accommodates endocrine cells that have the potential to produce insulin. Here we report the characterization and distribution of insulin-immunoreactive cells and endocrine cells within the ductal units of adult human pancreas. Sequential pancreas sections from 12 nondiabetic adults were stained with biomarkers of ductal epithelial cells (cytokeratin 19), acinar cells (amylase), endocrine cells (chromogranin A; neuron-specific enolase), islet hormones (insulin, glucagon, somatostatin, pancreatic polypeptide), cell proliferation (Ki-67), and neogenesis (CD29). The number of islet hormone-immunoreactive cells increased from large ducts to the terminal branches. The insulin-producing cells outnumbered endocrine cells reactive for glucagon, somatostatin, or pancreatic polypeptide. The proportions of insulin-immunoreactive count compared with local islets (100% as a baseline) were 1.5% for the main ducts, 7.2% for interlobular ducts, 24.8% for intralobular ducts, 67.9% for intercalated ducts, and 348.9% for centroacinar cells. Both Ki-67- and CD29-labeled cells were predominantly localized in the terminal branches around the islets. The terminal branches also showed cells coexpressing islet hormones and cytokeratin 19. The adult human pancreatic ducts showed islet hormone-producing cells. The insulin-reactive cells predominantly localized in terminal branches where they may retain potential capability for β-cell neogenesis.

  7. Postprandial Glucose and Insulin Responses to Grain Products in ...

    African Journals Online (AJOL)

    Prof. Ogunji

    diabetes was low in women who consumed more cereal and vegetable fiber. Under normal ... Body Mass Index (BMI) was calculated, and obesity was defined as a ... much they could eat and be satisfied. There was no .... advantageous with respect to the insulin resistance syndrome since from this study the low glycaemic.

  8. Insulin response to oral glucose in healthy, lean young women and patients with polycystic ovary syndrome.

    Science.gov (United States)

    Kulshreshtha, Bindu; Ganie, Mohammed Ashraf; Praveen, Edavan Pulikkanath; Gupta, Nandita; Lal Khurana, Madan; Seith, Ashu; Dwivedi, Sadanand N; Kumar, Guresh; Ammini, Ariachery C

    2008-11-01

    Insulin resistance and consequent hyperinsulinemia are common among patients with polycystic ovary syndrome (PCOS). Ethnicity and dietary habits affect insulin levels. There is little published information from India on insulin levels in PCOS patients. Thus the present study aimed to determine the insulin response to oral glucose in women with PCOS and healthy women. In a case-control study design, women with PCOS and lean healthy women without a family history of diabetes mellitus underwent oral glucose tolerance testing. Samples were collected at 0, 1 and 2 h after glucose ingestion. Two hundred and eighty-five women with PCOS and 27 lean healthy young women were enrolled into the study. The mean age of controls was 22.8 +/- 4.5 years (range 15-32 years) and their mean body mass index (BMI) was 19.7 +/- 2.6 kg/m(2). Mean blood glucose at 0, 1 and 2 h was 88.2 +/- 7.2, 115.5 +/- 25.5 and 91.8 +/- 20.5 mg/dl, respectively. Corresponding plasma insulin levels were 5.8 +/- 1.1, 32.7 +/- 26.5 and 14.6 +/- 9.6 mIU/l. Peak insulin levels were seen at 1 h and these came down to less than 40% of the peak value by 2 h. Glucose/insulin ratio at 0, 1 and 2 h was 15.6 +/- 3.1, 7.0 +/- 3.1 and 11.4 +/- 7.0. Homeostasis model assessment of insulin resistance (HOMA-IR) was 1.2 +/- 0.2. The age of the PCOS women ranged from 15 to 40 years (mean 23.4 +/- 6.2 years) and their BMI ranged from 16.4 to 50.4 kg/m(2) (mean 27.7 +/- 6.3 kg/m(2)). One hundred and seventy-six (62%) PCOS patients had normal glucose tolerance (NGT), 39 (14%) had impaired fasting glucose (IFG), 49 (17%) had impaired glucose tolerance (IGT) and 21 (7%) had type 2 diabetes mellitus (T2DM). Insulin response was higher in women with PCOS. Peak insulin was observed at 1 h. The difference between 1-h and 2-h post-glucose insulin decreased with worsening glucose tolerance. Both plasma insulin and BMI showed a rising trend from NGT to IFG to IGT. There was no further increase in either insulin or BMI from IGT to T2DM

  9. Novel remodeling of the mouse heart mitochondrial proteome in response to acute insulin stimulation

    Science.gov (United States)

    Pedersen, Brian A; Yazdi, Puya G; Taylor, Jared F; Khattab, Omar S; Chen, Yu-Han; Chen, Yumay; Wang, Ping H

    2015-01-01

    Mitochondrial dysfunction contributes to the pathophysiology of diabetic cardiomyopathy. The aim of this study was to investigate the acute changes in the mitochondrial proteome in response to insulin stimulation. Cardiac mitochondria from C57BL/6 mice after insulin stimulation were analyzed using two-dimensional fluorescence difference gel electrophoresis. MALDI-TOF MS/MS was utilized to identify differences. Two enzymes involved in metabolism and four structural proteins were identified. Succinyl-CoA ligase [ADP forming] subunit beta was identified as one of the differentially regulated proteins. Upon insulin stimulation, a relatively more acidic isoform of this protein was increased by 53% and its functional activity was decreased by ∼32%. This proteomic remodeling in response to insulin stimulation may play an important role in the normal and diabetic heart. PMID:26610654

  10. Suppression of the Nuclear Factor Eny2 Increases Insulin Secretion in Poorly Functioning INS-1E Insulinoma Cells

    Directory of Open Access Journals (Sweden)

    P. Dames

    2012-01-01

    Full Text Available Eny2, the mammalian ortholog of yeast Sus1 and drosophila E(y2, is a nuclear factor that participates in several steps of gene transcription and in mRNA export. We had previously found that Eny2 expression changes in mouse pancreatic islets during the metabolic adaptation to pregnancy. We therefore hypothesized that the protein contributes to the regulation of islet endocrine cell function and tested this hypothesis in rat INS-1E insulinoma cells. Overexpression of Eny2 had no effect but siRNA-mediated knockdown of Eny2 resulted in markedly increased glucose and exendin-4-induced insulin secretion from otherwise poorly glucose-responsive INS-1E cells. Insulin content, cellular viability, and the expression levels of several key components of glucose sensing remained unchanged; however glucose-dependent cellular metabolism was higher after Eny2 knockdown. Suppression of Eny2 enhanced the intracellular incretin signal downstream of cAMP. The use of specific cAMP analogues and pathway inhibitors primarily implicated the PKA and to a lesser extent the EPAC pathway. In summary, we identified a potential link between the nuclear protein Eny2 and insulin secretion. Suppression of Eny2 resulted in increased glucose and incretin-induced insulin release from a poorly glucose-responsive INS-1E subline. Whether these findings extend to other experimental conditions or to in vivo physiology needs to be determined in further studies.

  11. Effect of insulin on the mitotic activity of bone marrow cells after irradiation. [Gamma radiation, rats

    Energy Technology Data Exchange (ETDEWEB)

    Barkalaya, A I

    1976-02-01

    A total of 236 white rats were given a whole-body gamma dose of 750 R. Part of the rats were given a subcutaneous insulin injection of 0.2 units/kg. After 10, 20, 30 min, 1, 2, 3, 5, 8, 10 and 12 hours the mitotic index was determined in both groups of rats in the bone marrow of the femur. The content of glucose and insulin in the blood was determined. The mitotic index was found to be higher on administering insulin. The use of insulin in radiation sickness intensifies the mitotic activity of bone marrow cells and stimulates the recovery of bone marrow hematopoiesis. 5 references.

  12. Food fried in extra-virgin olive oil improves postprandial insulin response in obese, insulin-resistant women.

    Science.gov (United States)

    Farnetti, Sara; Malandrino, Noemi; Luciani, Davide; Gasbarrini, Giovanni; Capristo, Esmeralda

    2011-03-01

    The benefits of low glycemic load (GL) diets on clinical outcome in several metabolic and cardiovascular diseases have extensively been demonstrated. The GL of a meal can be affected by modulating the bioavailability of carbohydrates or by changing food preparation. We investigated the effect on plasma glucose and insulin response in lean and obese women of adding raw or fried extra-virgin olive oil to a carbohydrate-containing meal. After an overnight fast, 12 obese insulin-resistant women (body mass index [BMI], 32.8 ± 2.2 kg/m(2)) and five lean subjects (BMI, 22.2 ± 1.2 kg/m(2)) were randomly assigned to receive two different meals (designated A and B). Meal A was composed of 60 g of pasta made from wheat flour and 150 g of grilled courgettes with 25 g of uncooked oil. Meal B included 15 g of oil in the 150 g of deep-fried courgettes and 10 g of oil in the 60 g of stir-fried pasta. Both meals included 150 g of apple. Blood samples were collected at baseline and every 30 minutes over a 3-hour post-meal period and were tested for levels of glucose, insulin, C-peptide, and triglycerides. The area under the curve (AUC) values were calculated. In obese women the AUCs for C-peptide were significantly higher after meal A than after meal B at 120 minutes (W [Wilcoxon sign rank test] = 27.5, P = .0020), 150 minutes (W = 26.5, P = .0039), and 180 minutes (W = 26.5, P = .0039). No differences were found in lean subjects. This study demonstrated that in obese, insulin-resistant women, food fried in extra-virgin olive oil significantly reduced both insulin and C-peptide responses after a meal.

  13. A clinical study on insulin receptors of mononuclear cells in diabetes

    International Nuclear Information System (INIS)

    Dalimunthe, D.

    1980-01-01

    125 I-insulin binding activity to mononuclear cells was studied in 75 noninsulin-dependent diabetic subjects and 31 normal subjects and the following results were obtained. 1. 125 I-insulin binding is directly proportional to the mononuclear cell concentrations. There is a linear increase of specific 125 I-insulin binding. 2. The binding of 125 I-insulin to mononuclear cells is displaced by the increasing concentration of native insulin. 3. The 125 I-insulin degradation in the incubation medium after incubation of mononuclear cells for 24 hours at 4 0 C was almost 5% in this study. 4. The insulin binding activity in diabetic subjects was lower than that in normal subjects (P < 0.001) without any significant difference in affinity constant. 5. The relationship of binding activity to age of diabetics (r = 0.06, N.S), relative body weitht (r = 0.06, N.S) and duration of diabetes from onset was not significant. 6. In untreated noninsulin-dependent diabetics the insulin binding activity was inversely correlated to fasting blood glucose level (r = 0.78, P < 0.001) and slightly inversely correlated to serum insulin level (r = 0.47, P < 0.01). A slight inverse correlation was also observed in serum triglyceride level (r = 0.53, P < 0.01) and in total cholesterol level (r = 0.29, P < 0.05). 7. No significant difference between the binding activity was observed by grade of diabetic retinopathy. 8. After treatment with diet and/or sulfonylurea, the diabetics exhibited a significant increase in insulin binding activity (P < 0.005) but no significant difference in plasma insulin level, body weight and plasma lipid levels was observed. (author)

  14. Dual effect of insulin resistance and cadmium on human granulosa cells - In vitro study

    Energy Technology Data Exchange (ETDEWEB)

    Belani, Muskaan, E-mail: muskaanbelani@gmail.com [Department of Biochemistry, Faculty of Science, The Maharaja Sayajirao University of Baroda, Vadodara 390 002, Gujarat, India. (India); Shah, Preeti, E-mail: preeti.shah@novaivifertility.com [Nova IVI Fertility, Behind Xavier' s Ladies Hostel, 108, Swastik Society Rd., Navrangpura, Ahmedabad 390009, Gujarat, India. (India); Banker, Manish, E-mail: manish.banker@novaivifertility.com [Nova IVI Fertility, Behind Xavier' s Ladies Hostel, 108, Swastik Society Rd., Navrangpura, Ahmedabad 390009, Gujarat, India. (India); Gupta, Sarita, E-mail: saritagupta9@gmail.com [Department of Biochemistry, Faculty of Science, The Maharaja Sayajirao University of Baroda, Vadodara 390 002, Gujarat, India. (India)

    2016-12-15

    Combined exposure of cadmium (Cd) and insulin resistance (IR) might be responsible for subfertility. In the present study, we investigated the effects of Cd in vitro in IR human granulosa cells. Isolated human granulosa cells from control and polycystic ovary syndrome (PCOS) follicular fluid samples were confirmed for IR by decrease in protein expression of insulin receptor-β. Control and IR human granulosa cells were then incubated with or without 32 μM Cd. The combined effect of IR with 32 μM Cd in granulosa cells demonstrated significant decrease in expression of StAR, CYP11A1, CYP19A1, 17β-HSD, 3β-HSD, FSH-R and LH-R. Decrease was also observed in progesterone and estradiol concentrations as compared to control. Additionally, increase in protein expression of cleaved PARP-F2, active caspase-3 and a positive staining for Annexin V and PI indicated apoptosis as the mode of increased cell death ultimately leading to decreased steroidogenesis, as observed through the combined exposure. Taken together the results suggest decrease in steroidogenesis ultimately leading to abnormal development of the follicle thus compromising fertility at the level of preconception. - Highlights: • Protein expression of INSR-β in granulosa cells to differentiate PCOS-IR and NIR • Cd and IR together decrease steroidogenesis in human granulosa cells in vitro. • Cd and IR increase human granulosa cell death by increase in apoptosis. • Environment and life style are set to hamper pregnancies at preconception level.

  15. Dual effect of insulin resistance and cadmium on human granulosa cells - In vitro study

    International Nuclear Information System (INIS)

    Belani, Muskaan; Shah, Preeti; Banker, Manish; Gupta, Sarita

    2016-01-01

    Combined exposure of cadmium (Cd) and insulin resistance (IR) might be responsible for subfertility. In the present study, we investigated the effects of Cd in vitro in IR human granulosa cells. Isolated human granulosa cells from control and polycystic ovary syndrome (PCOS) follicular fluid samples were confirmed for IR by decrease in protein expression of insulin receptor-β. Control and IR human granulosa cells were then incubated with or without 32 μM Cd. The combined effect of IR with 32 μM Cd in granulosa cells demonstrated significant decrease in expression of StAR, CYP11A1, CYP19A1, 17β-HSD, 3β-HSD, FSH-R and LH-R. Decrease was also observed in progesterone and estradiol concentrations as compared to control. Additionally, increase in protein expression of cleaved PARP-F2, active caspase-3 and a positive staining for Annexin V and PI indicated apoptosis as the mode of increased cell death ultimately leading to decreased steroidogenesis, as observed through the combined exposure. Taken together the results suggest decrease in steroidogenesis ultimately leading to abnormal development of the follicle thus compromising fertility at the level of preconception. - Highlights: • Protein expression of INSR-β in granulosa cells to differentiate PCOS-IR and NIR • Cd and IR together decrease steroidogenesis in human granulosa cells in vitro. • Cd and IR increase human granulosa cell death by increase in apoptosis. • Environment and life style are set to hamper pregnancies at preconception level.

  16. Pathways for insulin access to the brain: the role of the microvascular endothelial cell.

    Science.gov (United States)

    Meijer, Rick I; Gray, Sarah M; Aylor, Kevin W; Barrett, Eugene J

    2016-11-01

    Insulin affects multiple important central nervous system (CNS) functions including memory and appetite, yet the pathway(s) by which insulin reaches brain interstitial fluid (bISF) has not been clarified. Recent studies demonstrate that to reach bISF, subarachnoid cerebrospinal fluid (CSF) courses through the Virchow-Robin space (VRS) which sheaths penetrating pial vessels down to the capillary level. Whether insulin predominantly enters the VRS and bISF by local transport through the blood-brain barrier, or by being secreted into the CSF by the choroid plexus, is unknown. We injected 125 I-TyrA14-insulin or regular insulin intravenously and compared the rates of insulin reaching subarachnoid CSF with its plasma clearance by brain tissue samples (an index of microvascular endothelial cell binding/uptake/transport). The latter process was more than 40-fold more rapid. We then showed that selective insulin receptor blockade or 4 wk of high-fat feeding each inhibited microvascular brain 125 I-TyrA14-insulin clearance. We further confirmed that 125 I-TyrA14-insulin was internalized by brain microvascular endothelial cells, indicating that the in vivo tissue association reflected cellular transport, not simply microvascular tracer binding. Copyright © 2016 the American Physiological Society.

  17. High performance liquid chromatographic analysis of insulin degradation products from a cultured kidney cell line

    International Nuclear Information System (INIS)

    Duckworth, W.C.; Hamel, F.G.; Liepnieks, J.; Frank, B.H.; Yagil, C.; Rabkin, R.

    1988-01-01

    The kidney is a major site for insulin removal and degradation, but the subcellular processes and enzymes involved have not been established. We have examined this process by analyzing insulin degradation products by HPLC. Monoiodoinsulin specifically labeled on either the A14 or B26 tyrosine residue was incubated with a cultured kidney epithelial cell line, and both intracellular and extracellular products were examined on HPLC. The products were then compared with products of known structure generated by hepatocytes and the enzyme insulin protease. Intracellular and extracellular products were different, suggesting two different degradative pathways, as previously shown in liver. The extracellular degradation products eluted from HPLC both before and after sulfitolysis similarly with hepatocyte products and products generated by insulin protease. The intracellular products also eluted identically with hepatocyte products. Based on comparisons with identified products, the kidney cell generates two fragments from the A chain of intact insulin, one with a cleavage at A13-A14 and the other at A14-A15. The B chain of intact insulin is cleaved in a number of different sites, resulting in peptides that elute identically with B chain peptides cleaved at B9-B10, B13-B14, B16-B17, B24-B25, and B25-B26. These similarities with hepatocytes and insulin protease suggest that liver and kidney have similar mechanisms for insulin degradation and that insulin protease or a very similar enzyme is involved in both tissues

  18. Differentiation of human-induced pluripotent stem cells into insulin-producing clusters.

    Science.gov (United States)

    Shaer, Anahita; Azarpira, Negar; Vahdati, Akbar; Karimi, Mohammad Hosein; Shariati, Mehrdad

    2015-02-01

    In diabetes mellitus type 1, beta cells are mostly destroyed; while in diabetes mellitus type 2, beta cells are reduced by 40% to 60%. We hope that soon, stem cells can be used in diabetes therapy via pancreatic beta cell replacement. Induced pluripotent stem cells are a kind of stem cell taken from an adult somatic cell by "stimulating" certain genes. These induced pluripotent stem cells may be a promising source of cell therapy. This study sought to produce isletlike clusters of insulin-producing cells taken from induced pluripotent stem cells. A human-induced pluripotent stem cell line was induced into isletlike clusters via a 4-step protocol, by adding insulin, transferrin, and selenium (ITS), N2, B27, fibroblast growth factor, and nicotinamide. During differentiation, expression of pancreatic β-cell genes was evaluated by reverse transcriptase-polymerase chain reaction; the morphologic changes of induced pluripotent stem cells toward isletlike clusters were observed by a light microscope. Dithizone staining was used to stain these isletlike clusters. Insulin produced by these clusters was evaluated by radio immunosorbent assay, and the secretion capacity was analyzed with a glucose challenge test. Differentiation was evaluated by analyzing the morphology, dithizone staining, real-time quantitative polymerase chain reaction, and immunocytochemistry. Gene expression of insulin, glucagon, PDX1, NGN3, PAX4, PAX6, NKX6.1, KIR6.2, and GLUT2 were documented by analyzing real-time quantitative polymerase chain reaction. Dithizone-stained cellular clusters were observed after 23 days. The isletlike clusters significantly produced insulin. The isletlike clusters could increase insulin secretion after a glucose challenge test. This work provides a model for studying the differentiation of human-induced pluripotent stem cells to insulin-producing cells.

  19. Insulin Resistance Induced by Hyperinsulinemia Coincides with a Persistent Alteration at the Insulin Receptor Tyrosine Kinase Domain

    Science.gov (United States)

    Catalano, Karyn J.; Maddux, Betty A.; Szary, Jaroslaw; Youngren, Jack F.; Goldfine, Ira D.; Schaufele, Fred

    2014-01-01

    Insulin resistance, the diminished response of target tissues to insulin, is associated with the metabolic syndrome and a predisposition towards diabetes in a growing proportion of the worldwide population. Under insulin resistant states, the cellular response of the insulin signaling pathway is diminished and the body typically responds by increasing serum insulin concentrations to maintain insulin signaling. Some evidence indicates that the increased insulin concentration may itself further dampen insulin response. If so, insulin resistance would worsen as the level of circulating insulin increases during compensation, which could contribute to the transition of insulin resistance to more severe disease. Here, we investigated the consequences of excess insulin exposure to insulin receptor (IR) activity. Cells chronically exposed to insulin show a diminished the level of IR tyrosine and serine autophosphorylation below that observed after short-term insulin exposure. The diminished IR response did not originate with IR internalization since IR amounts at the cell membrane were similar after short- and long-term insulin incubation. Förster resonance energy transfer between fluorophores attached to the IR tyrosine kinase (TK) domain showed that a change in the TK domain occurred upon prolonged, but not short-term, insulin exposure. Even though the altered ‘insulin refractory’ IR TK FRET and IR autophosphorylation levels returned to baseline (non-stimulated) levels after wash-out of the original insulin stimulus, subsequent short-term exposure to insulin caused immediate re-establishment of the insulin-refractory levels. This suggests that some cell-based ‘memory’ of chronic hyperinsulinemic exposure acts directly at the IR. An improved understanding of that memory may help define interventions to reset the IR to full insulin responsiveness and impede the progression of insulin resistance to more severe disease states. PMID:25259572

  20. Insulin resistance induced by hyperinsulinemia coincides with a persistent alteration at the insulin receptor tyrosine kinase domain.

    Directory of Open Access Journals (Sweden)

    Karyn J Catalano

    Full Text Available Insulin resistance, the diminished response of target tissues to insulin, is associated with the metabolic syndrome and a predisposition towards diabetes in a growing proportion of the worldwide population. Under insulin resistant states, the cellular response of the insulin signaling pathway is diminished and the body typically responds by increasing serum insulin concentrations to maintain insulin signaling. Some evidence indicates that the increased insulin concentration may itself further dampen insulin response. If so, insulin resistance would worsen as the level of circulating insulin increases during compensation, which could contribute to the transition of insulin resistance to more severe disease. Here, we investigated the consequences of excess insulin exposure to insulin receptor (IR activity. Cells chronically exposed to insulin show a diminished the level of IR tyrosine and serine autophosphorylation below that observed after short-term insulin exposure. The diminished IR response did not originate with IR internalization since IR amounts at the cell membrane were similar after short- and long-term insulin incubation. Förster resonance energy transfer between fluorophores attached to the IR tyrosine kinase (TK domain showed that a change in the TK domain occurred upon prolonged, but not short-term, insulin exposure. Even though the altered 'insulin refractory' IR TK FRET and IR autophosphorylation levels returned to baseline (non-stimulated levels after wash-out of the original insulin stimulus, subsequent short-term exposure to insulin caused immediate re-establishment of the insulin-refractory levels. This suggests that some cell-based 'memory' of chronic hyperinsulinemic exposure acts directly at the IR. An improved understanding of that memory may help define interventions to reset the IR to full insulin responsiveness and impede the progression of insulin resistance to more severe disease states.

  1. Multiple growth hormone-binding proteins are expressed on insulin-producing cells

    DEFF Research Database (Denmark)

    Møldrup, A; Billestrup, N; Thorn, N A

    1989-01-01

    The insulin-producing rat islet tumor cell line, RIN-5AH, expresses somatogen binding sites and responds to GH by increased proliferation and insulin production. Affinity cross-linking shows that RIN-5AH cells contain two major GH-binding subunits of Mr 100-130K (110K), which appear to exist as d....... It is concluded that the RIN-5AH cells have multiple GH-binding proteins which may mediate signals for either proliferation and/or insulin production....

  2. Effects of insulin on the survival of irradiated chinese hamster lung cells

    Energy Technology Data Exchange (ETDEWEB)

    Lin, P S; Kwock, L; Hefter, K; Wallach, D F.H.; Brotman, R [Tufts-New England Medical Center, Boston, Mass. (USA)

    1977-01-01

    Insulin treatment (10/sup -7/-10/sup -9/ M) before ..gamma.. irradiation (50 to 500 rads) increases the long term survival of Chinese hamster lung cells (DON). Our data indicates that the radioprotective effect of insulin is not due to a modulation of cyclic-adenosine-3',5'-monophosphate levels within these cells. The results suggest that the radiosensitive plasma membrane component postulated to be involved in the interphase death of thymocytes and protected by insulin may have a counterpart in DON cells.

  3. Intracellular and extracellular adenosine triphosphate in regulation of insulin secretion from pancreatic β cells (β).

    Science.gov (United States)

    Wang, Chunjiong; Geng, Bin; Cui, Qinghua; Guan, Youfei; Yang, Jichun

    2014-03-01

    Adenosine triphosphate (ATP) synthesis and release in mitochondria play critical roles in regulating insulin secretion in pancreatic β cells. Mitochondrial dysfunction is mainly characterized by a decrease in ATP production, which is a central event in the progression of pancreatic β cell dysfunction and diabetes. ATP has been demonstrated to regulate insulin secretion via several pathways: (i) Intracellular ATP directly closes ATP-sensitive potassium channel to open L-type calcium channel, leading to an increase in free cytosolic calcium levels and exocytosis of insulin granules; (ii) A decrease in ATP production is always associated with an increase in production of reactive oxygen species, which exerts deleterious effects on pancreatic β cell survival and insulin secretion; and (iii) ATP can be co-secreted with insulin from pancreatic β cells, and the released ATP functions as an autocrine signal to modulate insulin secretory process via P2 receptors on the cell membrane. In this review, the recent findings regarding the role and mechanism of ATP synthesis and release in regulation of insulin secretion from pancreatic β cells will be summarized and discussed. © 2013 Ruijin Hospital, Shanghai Jiaotong University School of Medicine and Wiley Publishing Asia Pty Ltd.

  4. Chitosan-modified porous silicon microparticles for enhanced permeability of insulin across intestinal cell monolayers.

    Science.gov (United States)

    Shrestha, Neha; Shahbazi, Mohammad-Ali; Araújo, Francisca; Zhang, Hongbo; Mäkilä, Ermei M; Kauppila, Jussi; Sarmento, Bruno; Salonen, Jarno J; Hirvonen, Jouni T; Santos, Hélder A

    2014-08-01

    Porous silicon (PSi) based particulate systems are emerging as an important drug delivery system due to its advantageous properties such as biocompatibility, biodegradability and ability to tailor the particles' physicochemical properties. Here, annealed thermally hydrocarbonized PSi (AnnTHCPSi) and undecylenic acid modified AnnTHCPSi (AnnUnTHCPSi) microparticles were developed as a PSi-based platform for oral delivery of insulin. Chitosan (CS) was used to modify the AnnUnTHCPSi microparticles to enhance the intestinal permeation of insulin. Surface modification with CS led to significant increase in the interaction of PSi microparticles with Caco-2/HT-29 cell co-culture monolayers. Compared to pure insulin, the CS-conjugated microparticles significantly improved the permeation of insulin across the Caco-2/HT-29 cell monolayers, with ca. 20-fold increase in the amount of insulin permeated and ca. 7-fold increase in the apparent permeability (P(app)) value. Moreover, among all the investigated particles, the CS-conjugated microparticles also showed the highest amount of insulin associated with the mucus layer and the intestinal Caco-2 cells and mucus secreting HT-29 cells. Our results demonstrate that CS-conjugated AnnUnTHCPSi microparticles can efficiently enhance the insulin absorption across intestinal cells, and thus, they are promising microsystems for the oral delivery of proteins and peptides across the intestinal cell membrane. Copyright © 2014 Elsevier Ltd. All rights reserved.

  5. The effect of dietary fiber and other factors on insulin response: role in obesity.

    Science.gov (United States)

    Ullrich, I H; Albrink, M J

    1985-07-01

    Epidemiologic evidence favors the hypothesis that obesity may result from the fiber-depleted diet of industrialized societies. Since hyperinsulinemia is a universal characteristic and perhaps causal of obesity, the possibility is considered that dietary factors causing excess insulin secretion might lead to obesity. Dietary glucose causes a slightly greater insulin rise than cooked starch containing an equal amount of carbohydrate, and high fiber starchy foods cause a much lesser insulin response than does glucose in solution. Doubling the dose of carbohydrate in a meal causes only a small increase in glucose response but a large increase in insulin response. Dietary fiber could act by displacing some of the carbohydrate that would normally be absorbable in the small intestine, or could translocate the carbohydrate to a point lower in the intestinal tract where less effect on insulin secretion would be observed. Evidence is presented that a higher fiber diet is associated with a higher concentration of fasting circulating free fatty acids, a lesser post-cibal decrease in circulating free fatty acids and triglycerides and less chronic increase in fasting triglycerides than a low fiber diet. These differences are associated with a lesser insulin response to high fiber meals. The extreme fluctuations between the fed and fasted states seen with low fiber diets are thus dampened by high fiber diets. The less complete inhibition of lipolysis during the fed state, and more intense lipolysis during fasting, suggested by the above data, might tend to prevent obesity. The mechanisms of the lesser insulin response to high rather than low fiber meals are not known, but the possibility that dietary fiber decreases the GIP response is considered.

  6. Insulin secretion and signaling in response to dietary restriction and subsequent re-alimentation in cattle.

    Science.gov (United States)

    Keogh, Kate; Kenny, David A; Kelly, Alan K; Waters, Sinéad M

    2015-08-01

    The objectives of this study were to examine systemic insulin response to a glucose tolerance test (GTT) and transcript abundance of genes of the insulin signaling pathway in skeletal muscle, during both dietary restriction and re-alimentation-induced compensatory growth. Holstein Friesian bulls were blocked to one of two groups: 1) restricted feed allowance for 125 days (period 1) (RES, n = 15) followed by ad libitum feeding for 55 days (period 2) or 2) ad libitum access to feed throughout (periods 1 and 2) (ADLIB, n = 15). On days 90 and 36 of periods 1 and 2, respectively, a GTT was performed. M. longissimus dorsi biopsies were harvested from all bulls on days 120 and 15 of periods 1 and 2, respectively, and RNA-Seq analysis was performed. RES displayed a lower growth rate during period 1 (RES: 0.6 kg/day, ADLIB: 1.9 kg/day; P alimentation (RES: 2.5 kg/day, ADLIB: 1.4 kg/day; P alimentation (P > 0.05). Genes differentially expressed in the insulin signaling pathway suggested a greater sensitivity to insulin in skeletal muscle, with pleiotropic effects of insulin signaling interrupted during dietary restriction. Collectively, these results indicate increased sensitivity to glucose clearance and skeletal muscle insulin signaling during dietary restriction; however, no overall role for insulin was apparent in expressing compensatory growth. Copyright © 2015 the American Physiological Society.

  7. Zinc stimulates glucose oxidation and glycemic control by modulating the insulin signaling pathway in human and mouse skeletal muscle cell lines.

    Science.gov (United States)

    Norouzi, Shaghayegh; Adulcikas, John; Sohal, Sukhwinder Singh; Myers, Stephen

    2018-01-01

    Zinc is a metal ion that is an essential cell signaling molecule. Highlighting this, zinc is an insulin mimetic, activating cellular pathways that regulate cellular homeostasis and physiological responses. Previous studies have linked dysfunctional zinc signaling with several disease states including cancer, obesity, cardiovascular disease and type 2 diabetes. The present study evaluated the insulin-like effects of zinc on cell signaling molecules including tyrosine, PRSA40, Akt, ERK1/2, SHP-2, GSK-3β and p38, and glucose oxidation in human and mouse skeletal muscle cells. Insulin and zinc independently led to the phosphorylation of these proteins over a 60-minute time course in both mouse and human skeletal muscle cells. Similarly, utilizing a protein array we identified that zinc could active the phosphorylation of p38, ERK1/2 and GSK-3B in human and ERK1/2 and GSK-3B in mouse skeletal muscle cells. Glucose oxidation assays were performed on skeletal muscle cells treated with insulin, zinc, or a combination of both and resulted in a significant induction of glucose consumption in mouse (pzinc alone. Insulin, as expected, increased glucose oxidation in mouse (pzinc and insulin did not augment glucose consumption in these cells. Zinc acts as an insulin mimetic, activating key molecules implicated in cell signaling to maintain glucose homeostasis in mouse and human skeletal muscle cells. Zinc is an important metal ion implicated in several biological processes. The role of zinc as an insulin memetic in activating key signaling molecules involved in glucose homeostasis could provide opportunities to utilize this ion therapeutically in treating disorders associated with dysfunctional zinc signaling.

  8. The potential role of SOCS-3 in the interleukin-1beta-induced desensitization of insulin signaling in pancreatic beta-cells

    DEFF Research Database (Denmark)

    Emanuelli, Brice; Glondu, Murielle; Filloux, Chantal

    2004-01-01

    insulin signaling is required for the optimal beta-cell function, we assessed the effect of IL-1beta on the insulin pathway in a rat pancreatic beta-cell line. We show that IL-1beta decreases insulin-induced tyrosine phosphorylation of the insulin receptor (IR) and insulin receptor substrate (IRS...

  9. Inter-domain tagging implicates caveolin-1 in insulin receptor trafficking and Erk signaling bias in pancreatic beta-cells

    Directory of Open Access Journals (Sweden)

    Tobias Boothe

    2016-05-01

    Full Text Available Objective: The role and mechanisms of insulin receptor internalization remain incompletely understood. Previous trafficking studies of insulin receptors involved fluorescent protein tagging at their termini, manipulations that may be expected to result in dysfunctional receptors. Our objective was to determine the trafficking route and molecular mechanisms of functional tagged insulin receptors and endogenous insulin receptors in pancreatic beta-cells. Methods: We generated functional insulin receptors tagged with pH-resistant fluorescent proteins between domains. Confocal, TIRF and STED imaging revealed a trafficking pattern of inter-domain tagged insulin receptors and endogenous insulin receptors detected with antibodies. Results: Surprisingly, interdomain-tagged and endogenous insulin receptors in beta-cells bypassed classical Rab5a- or Rab7-mediated endocytic routes. Instead, we found that removal of insulin receptors from the plasma membrane involved tyrosine-phosphorylated caveolin-1, prior to trafficking within flotillin-1-positive structures to lysosomes. Multiple methods of inhibiting caveolin-1 significantly reduced Erk activation in vitro or in vivo, while leaving Akt signaling mostly intact. Conclusions: We conclude that phosphorylated caveolin-1 plays a role in insulin receptor internalization towards lysosomes through flotillin-1-positive structures and that caveolin-1 helps bias physiological beta-cell insulin signaling towards Erk activation. Author Video: Author Video Watch what authors say about their articles Keywords: Insulin receptor internalization, Insulin resistance, Pancreatic islet beta-cells, Autocrine insulin signaling

  10. Redifferentiation of insulin-secreting cells after in vitro expansion of adult human pancreatic islet tissue

    International Nuclear Information System (INIS)

    Lechner, Andreas; Nolan, Anna L.; Blacken, Robyn A.; Habener, Joel F.

    2005-01-01

    Cellular replacement therapy holds promise for the treatment of diabetes mellitus but donor tissue is severely limited. Therefore, we investigated whether insulin-secreting cells could be differentiated in vitro from a monolayer of cells expanded from human donor pancreatic islets. We describe a three-step culture protocol that allows for the efficient generation of insulin-producing cell clusters from in vitro expanded, hormone-negative cells. These clusters express insulin at levels of up to 34% that of average freshly isolated human islets and secrete C-peptide upon membrane depolarization. They also contain cells expressing the other major islet hormones (glucagon, somatostatin, and pancreatic polypeptide). The source of the newly differentiated endocrine cells could either be indigenous stem/progenitor cells or the proliferation-associated dedifferentiation and subsequent redifferentiation of mature endocrine cells. The in vitro generated cell clusters may be efficacious in providing islet-like tissue for transplantation into diabetic recipients

  11. Insulin induces a transcriptional activation of epiregulin, HB-EGF and amphiregulin, by a PI3K-dependent mechanism: Identification of a specific insulin-responsive promoter element

    International Nuclear Information System (INIS)

    Ornskov, Dorthe; Nexo, Ebba; Sorensen, Boe S.

    2007-01-01

    Previously we have shown that insulin-stimulation of RT4 bladder cancer cells leads to increased proliferation, which require HER1 activation, and is accompanied by increased mRNA expression of the EGF-ligands heparin-binding EGF-like growth factor (HB-EGF), amphiregulin (AR), and epiregulin (EPI) [D. Ornskov, E. Nexo, B.S. Sorensen, Insulin-induced proliferation of bladder cancer cells is mediated through activation of the epidermal growth factor system, FEBS J. 273 (2006) 5479-5489]. In the present paper, we have investigated the molecular mechanism leading to this insulin-induced expression. We monitored the decay of mRNA after inhibiting transcription with Actinomycin D and demonstrated that the insulin-mediated increase was not caused by enhanced mRNA stability. In untreated cells, HB-EGF mRNA was the least stable, whereas AR and EPI mRNA decayed with slower kinetics. However, promoter analysis of HB-EGF and EPI demonstrated that insulin stimulated transcription. Studies on the EPI promoter identified the insulin-responsive element to be located in the region -564 to -365 bp. This region contains potential binding sites for the transcription factors SP1, AP1, and NF-κB. Interestingly, all three transcription factors can be activated by PI3K. We demonstrate that the insulin-induced expression of HB-EGF, AR, and EPI mRNA is completely prevented by the specific PI3K inhibitor Wortmannin, suggesting an involvement of the PI3K

  12. Estradiol represses Insulin-like 3 expression and promoter activity in MA-10 Leydig cells

    International Nuclear Information System (INIS)

    Lague, Eric; Tremblay, Jacques J.

    2009-01-01

    There are increasing evidence in the literature reporting the detrimental effects of endocrine disruptors on the development and function of the male reproductive system. One example is cryptorchidism, or undescended testis, caused by exposure to excessive estrogens. Estrogens, acting through the estrogen receptor α (ERα), have been shown to repress expression of the gene encoding insulin-like 3 (INSL3), a small peptide produced by testicular Leydig cells that is essential for normal testis descent. The molecular mechanism of estrogen/ER action on Insl3 expression, however, remains poorly understood. Here we report estradiol (E 2 ) represses Insl3 mRNA levels in MA-10 cells, a Leydig cell line model. We also found that E 2 represses the activity of the human and mouse Insl3 promoter in these cells. The E 2 -responsive region of the human INSL3 promoter was located to the proximal INSL3 promoter. This region does not contain a consensus estrogen response element indicating an indirect mechanism of action. In agreement with this, we found that E 2 -responsiveness was lost when two previously characterized binding sites for the nuclear receptors NUR77 and SF1 were mutated. Finally we show that the E 2 repressive effect could be overcome by cotreatment with testosterone, a positive regulator of Insl3 transcription. Collectively our data provide important new insights into the molecular mechanism of estrogen action in Insl3 transcription in Leydig cells

  13. Sirt1 regulates insulin secretion by repressing UCP2 in pancreatic beta cells.

    Directory of Open Access Journals (Sweden)

    Laura Bordone

    2006-02-01

    Full Text Available Sir2 and insulin/IGF-1 are the major pathways that impinge upon aging in lower organisms. In Caenorhabditis elegans a possible genetic link between Sir2 and the insulin/IGF-1 pathway has been reported. Here we investigate such a link in mammals. We show that Sirt1 positively regulates insulin secretion in pancreatic beta cells. Sirt1 represses the uncoupling protein (UCP gene UCP2 by binding directly to the UCP2 promoter. In beta cell lines in which Sirt1 is reduced by SiRNA, UCP2 levels are elevated and insulin secretion is blunted. The up-regulation of UCP2 is associated with a failure of cells to increase ATP levels after glucose stimulation. Knockdown of UCP2 restores the ability to secrete insulin in cells with reduced Sirt1, showing that UCP2 causes the defect in glucose-stimulated insulin secretion. Food deprivation induces UCP2 in mouse pancreas, which may occur via a reduction in NAD (a derivative of niacin levels in the pancreas and down-regulation of Sirt1. Sirt1 knockout mice display constitutively high UCP2 expression. Our findings show that Sirt1 regulates UCP2 in beta cells to affect insulin secretion.

  14. Insulin resistance enhances the mitogen-activated protein kinase signaling pathway in ovarian granulosa cells.

    Directory of Open Access Journals (Sweden)

    Linghui Kong

    Full Text Available The ovary is the main regulator of female fertility. Granulosa cell dysfunction may be involved in various reproductive endocrine disorders. Here we investigated the effect of insulin resistance on the metabolism and function of ovarian granulosa cells, and dissected the functional status of the mitogen-activated protein kinase signaling pathway in these cells. Our data showed that dexamethasone-induced insulin resistance in mouse granulosa cells reduced insulin sensitivity, accompanied with an increase in phosphorylation of p44/42 mitogen-activated protein kinase. Furthermore, up-regulation of cytochrome P450 subfamily 17 and testosterone and down-regulation of progesterone were observed in insulin-resistant mouse granulosa cells. Inhibition of p44/42 mitogen-activated protein kinase after induction of insulin resistance in mouse granulosa cells decreased phosphorylation of p44/42 mitogen-activated protein kinase, downregulated cytochrome P450 subfamily 17 and lowered progesterone production. This insulin resistance cell model can successfully demonstrate certain mechanisms such as hyperandrogenism, which may inspire a new strategy for treating reproductive endocrine disorders by regulating cell signaling pathways.

  15. Response of the adrenal medulla to exogenous insulin in head X-irradiated dogs

    Energy Technology Data Exchange (ETDEWEB)

    Yamashita, K; Mieno, M; Shimizu, T [Nagasaki Univ. (Japan). School of Medicine

    1976-12-01

    The observed results on the secretory response of the adrenal medulla to exogenous insulin in dogs whose heads had been irradiated with 200 rad X-irradiation at a rate of 60.6 rad/min are reported. Approximately 20 h after irradiation 2 i.u. insulin/kg body weight was administered to both irradiated and control (non-irradiated) dogs. The secretion of both adrenaline and noradrenaline in five control dogs which received insulin had increased markedly 30 min after the injection and it was sustained over the first 120 min. Five irradiated dogs also responded to the injection of insulin by secreting appreciable amounts of both adrenaline and noradrenaline, but the responsiveness was considerably lower; adrenaline and noradrenaline secretion 30 and 60 min after the injection was 59 and 33% less than that caused by insulin in control animals respectively, and these differences were significant (P < 0.01). Since insulin induces hypoglycaemia and thereby leads to an increase in the adrenal medullary secretion through the central mechanism, the present results observed 1 day after X-irradiation of the head indicate that the susceptibility of the adrenomedullary activating mechanism in the central nervous system may be diminished considerably after X-irradiation, even at doses as low as 200 rad.

  16. Response of the adrenal medulla to exogenous insulin in head X-irradiated dogs

    International Nuclear Information System (INIS)

    Yamashita, K.; Mieno, M.; Shimizu, T.

    1976-01-01

    The observed results on the secretory response of the adrenal medulla to exogenous insulin in dogs whose heads had been irradiated with 200 rad X-irradiation at a rate of 60.6 rad/min are reported. Approximately 20 h after irradiation 2 i.u. insulin/kg body weight was administered to both irradiated and control (non-irradiated) dogs. The secretion of both adrenaline and noradrenaline in five control dogs which received insulin had increased markedly 30 min after the injection and it was sustained over the first 120 min. Five irradiated dogs also responded to the injection of insulin by secreting appreciable amounts of both adrenaline and noradrenaline, but the responsiveness was considerably lower; adrenaline and noradrenaline secretion 30 and 60 min after the injection was 59 and 33% less than that caused by insulin in control animals respectively, and these differences were significant (P < 0.01). Since insulin induces hypoglycaemia and thereby leads to an increase in the adrenal medullary secretion through the central mechanism, the present results observed 1 day after X-irradiation of the head indicate that the susceptibility of the adrenomedullary activating mechanism in the central nervous system may be diminished considerably after X-irradiation, even at doses as low as 200 rad. (U.K.)

  17. Generation of insulin-producing human mesenchymal stem cells using recombinant adeno-associated virus.

    Science.gov (United States)

    Kim, Jeong Hwan; Park, Si-Nae; Suh, Hwal

    2007-02-28

    The purpose of current experiment is the generation of insulin-producing human mesenchymal stem cells as therapeutic source for the cure of type 1 diabetes. Type 1 diabetes is generally caused by insulin deficiency accompanied by the destruction of islet beta-cells. In various trials for the treatment of type 1 diabetes, cell-based gene therapy using stem cells is considered as one of the most useful candidate for the treatment. In this experiment, human mesenchymal stem cells were transduced with AAV which is containing furin-cleavable human preproinsulin gene to generate insulin-producing cells as surrogate beta-cells for the type 1 diabetes therapy. In the rAAV production procedure, rAAV was generated by transfection of AD293 cells. Human mesenchymal stems cells were transduced using rAAV with a various multiplicity of infection. Transduction of recombinant AAV was also tested using beta-galactosidse expression. Cell viability was determined by using MTT assay to evaluate the toxicity of the transduction procedure. Expression and production of Insulin were tested using reverse transcriptase-polymerase chain reaction and immunocytochemistry. Secretion of human insulin and C-peptide from the cells was assayed using enzyme-linked immunosorbent assay. Production of insulin and C-peptide from the test group represented a higher increase compared to the control group. In this study, we examined generation of insulin-producing cells from mesenchymal stem cells by genetic engineering for diabetes therapy. This work might be valuable to the field of tissue engineering for diabetes treatment.

  18. Structural and functional study of the GlnB22-insulin mutant responsible for maturity-onset diabetes of the young.

    Directory of Open Access Journals (Sweden)

    Květoslava Křížková

    Full Text Available The insulin gene mutation c.137G>A (R46Q, which changes an arginine at the B22 position of the mature hormone to glutamine, causes the monogenic diabetes variant maturity-onset diabetes of the young (MODY. In MODY patients, this mutation is heterozygous, and both mutant and wild-type (WT human insulin are produced simultaneously. However, the patients often depend on administration of exogenous insulin. In this study, we chemically synthesized the MODY mutant [GlnB22]-insulin and characterized its biological and structural properties. The chemical synthesis of this insulin analogue revealed that its folding ability is severely impaired. In vitro and in vivo tests showed that its binding affinity and biological activity are reduced (both approximately 20% that of human insulin. Comparison of the solution structure of [GlnB22]-insulin with the solution structure of native human insulin revealed that the most significant structural effect of the mutation is distortion of the B20-B23 β-turn, leading to liberation of the B chain C-terminus from the protein core. The distortion of the B20-B23 β-turn is caused by the extended conformational freedom of the GlnB22 side chain, which is no longer anchored in a hydrogen bonding network like the native ArgB22. The partially disordered [GlnB22]-insulin structure appears to be one reason for the reduced binding potency of this mutant and may also be responsible for its low folding efficiency in vivo. The altered orientation and flexibility of the B20-B23 β-turn may interfere with the formation of disulfide bonds in proinsulin bearing the R46Q (GlnB22 mutation. This may also have a negative effect on the WT proinsulin simultaneously biosynthesized in β-cells and therefore play a major role in the development of MODY in patients producing [GlnB22]-insulin.

  19. The Investigation of ADAMTS16 in Insulin-Induced Human Chondrosarcoma Cells.

    Science.gov (United States)

    Cakmak, Ozlem; Comertoglu, Ismail; Firat, Ridvan; Erdemli, Haci Kemal; Kursunlu, S Fatih; Akyol, Sumeyya; Ugurcu, Veli; Altuntas, Aynur; Adam, Bahattin; Demircan, Kadir

    2015-08-01

    A disintegrin-like metalloproteinase with thrombospondin motifs (ADAMTS) is a group of proteins that have enzymatic activity secreted by cells to the outside extracellular matrix. Insulin induces proteoglycan biosynthesis in chondrosarcoma chondrocytes. The purpose of the present in vitro study is to assess the time course effects of insulin on ADAMTS16 expression in OUMS-27 (human chondrosarcoma) cell line to examine whether insulin regulates ADAMTS16 expression as well as proteoglycan biosynthesis with multifaceted properties or not. Chondrosarcoma cells were cultured in Dulbecco's modified Eagle's medium having either 10 μg/mL insulin or not. While the experiment was going on, the medium containing insulin had been changed every other day. Cells were harvested at 1st, 3rd, 7th, and 11th days; subsequently, RNA and proteins were isolated in every experimental group according to their time interval. RNA expression of ADAMTS was estimated by quantitative real-time polymerase chain reaction (qRT-PCR) by using primers. Immunoreactive protein levels were encountered by the western blot protein detection technique by using proper anti-ADAMTS16 antibodies. ADAMTS16 mRNA expression level of chondrosarcoma cells was found to be insignificantly decreased in chondrosarcoma cells induced by insulin detected by the qRT-PCR instrument. On the other hand, there was a gradual decrease in immune-reactant ADAMTS16 protein amount by the time course in insulin-treated cell groups when compared with control cells. It has been suggested that insulin might possibly regulate ADAMTS16 levels/activities in OUMS-27 chondrosarcoma cells taking a role in extracellular matrix turnover.

  20. Effect of starvation on human muscle protein metabolism and its response to insulin

    International Nuclear Information System (INIS)

    Fryburg, D.A.; Barrett, E.J.; Louard, R.J.; Gelfand, R.A.

    1990-01-01

    To assess the effect of fasting on muscle protein turnover in the basal state and in response to insulin, we measured forearm amino acid kinetics, using [3H]phenylalanine (Phe) and [14C]leucine (Leu) infused systemically, in eight healthy subjects after 12 (postabsorptive) and 60 h of fasting. After a 150-min basal period, forearm local insulin concentration was selectively raised by approximately 25 muU/ml for 150 min by intra-arterial insulin infusion (0.02 mU.kg-1. min-1). The 60-h fast increased urine nitrogen loss and whole body Leu flux and oxidation (by 50-75%, all P less than 0.02). Post-absorptively, forearm muscle exhibited a net release of Phe and Leu, which increased two- to threefold after the 60-h fast (P less than 0.05); this effect was mediated exclusively by accelerated local rates of amino acid appearance (Ra), with no reduction in rates of disposal (Rd). Local hyperinsulinemia in the postabsorptive condition caused a twofold increase in forearm glucose uptake (P less than 0.01) and completely suppressed the net forearm output of Phe and Leu (P less than 0.02). After the 60-h fast, forearm glucose disposal was depressed basally and showed no response to insulin; in contrast, insulin totally abolished the accelerated net forearm release of Phe and Leu. The action of insulin to reverse the augmented net release of Phe and Leu was mediated exclusively by approximately 40% suppression of Ra (P less than 0.02) rather than a stimulation of Rd. We conclude that in short-term fasted humans (1) muscle amino acid output accelerates due to increased proteolysis rather than reduced protein synthesis, and (2) despite its catabolic state and a marked impairment in insulin-mediated glucose disposal, muscle remains sensitive to insulin's antiproteolytic action

  1. Effect of starvation on human muscle protein metabolism and its response to insulin

    Energy Technology Data Exchange (ETDEWEB)

    Fryburg, D.A.; Barrett, E.J.; Louard, R.J.; Gelfand, R.A. (Yale Univ. School of Medicine, New Haven, CT (USA))

    1990-10-01

    To assess the effect of fasting on muscle protein turnover in the basal state and in response to insulin, we measured forearm amino acid kinetics, using (3H)phenylalanine (Phe) and (14C)leucine (Leu) infused systemically, in eight healthy subjects after 12 (postabsorptive) and 60 h of fasting. After a 150-min basal period, forearm local insulin concentration was selectively raised by approximately 25 muU/ml for 150 min by intra-arterial insulin infusion (0.02 mU.kg-1. min-1). The 60-h fast increased urine nitrogen loss and whole body Leu flux and oxidation (by 50-75%, all P less than 0.02). Post-absorptively, forearm muscle exhibited a net release of Phe and Leu, which increased two- to threefold after the 60-h fast (P less than 0.05); this effect was mediated exclusively by accelerated local rates of amino acid appearance (Ra), with no reduction in rates of disposal (Rd). Local hyperinsulinemia in the postabsorptive condition caused a twofold increase in forearm glucose uptake (P less than 0.01) and completely suppressed the net forearm output of Phe and Leu (P less than 0.02). After the 60-h fast, forearm glucose disposal was depressed basally and showed no response to insulin; in contrast, insulin totally abolished the accelerated net forearm release of Phe and Leu. The action of insulin to reverse the augmented net release of Phe and Leu was mediated exclusively by approximately 40% suppression of Ra (P less than 0.02) rather than a stimulation of Rd. We conclude that in short-term fasted humans (1) muscle amino acid output accelerates due to increased proteolysis rather than reduced protein synthesis, and (2) despite its catabolic state and a marked impairment in insulin-mediated glucose disposal, muscle remains sensitive to insulin's antiproteolytic action.

  2. Decreased insulin response in dairy cows following a four-day fast to induce hepatic lipidosis.

    Science.gov (United States)

    Oikawa, S; Oetzel, G R

    2006-08-01

    Negative energy balance has been implicated in the development of fatty liver, insulin resistance, and impaired health in dairy cows. A 4-d fasting model previously was reported to increase liver triglycerides more than 2.5-fold. The purpose of the present study was to evaluate insulin response in this fasting model. Nonlactating, nonpregnant Holstein cows were fasted for 4 d (6 cows) or fed continuously as control cows (4 cows). Samples were collected 5 d before fasting, during fasting, and immediately after the 4-d fast, 8 d after the fast, and 16 d after the fast. Fasted cows had greater liver triglyceride content (49.4 vs. 16.2 mg/g, wet-weight basis) at the end of the fasting period compared with control cows. Fasted cows also had increased plasma nonesterified fatty acid (NEFA) concentrations (1.24 vs. 0.21 mmol/L) and increased plasma beta-hydroxybutyrate (BHBA) concentrations at the end of the fasting period. Liver triglyceride, plasma NEFA, and plasma BHBA in fasted cows returned to prefasting concentrations by the end of the experiment. Plasma glucose concentrations were not affected by fasting. Plasma insulin concentrations were decreased (6.3 vs. 14.1 microU/mL) and insulin-stimulated blood glucose reduction was decreased (24.9 vs. 48.6%) in the fasted cows compared with control cows at the end of the fast, indicating reduced insulin response. Insulin response was negatively correlated with plasma NEFA and liver triglycerides. Decreased insulin response may be an important complication of negative energy balance and hepatic lipidosis.

  3. Histone deacetylase regulates insulin signaling via two pathways in pancreatic β cells.

    Directory of Open Access Journals (Sweden)

    Yukina Kawada

    Full Text Available Recent studies demonstrated that insulin signaling plays important roles in the regulation of pancreatic β cell mass, the reduction of which is known to be involved in the development of diabetes. However, the mechanism underlying the alteration of insulin signaling in pancreatic β cells remains unclear. The involvement of epigenetic control in the onset of diabetes has also been reported. Thus, we analyzed the epigenetic control of insulin receptor substrate 2 (IRS2 expression in the MIN6 mouse insulinoma cell line. We found concomitant IRS2 up-regulation and enhanced insulin signaling in MIN6 cells, which resulted in an increase in cell proliferation. The H3K9 acetylation status of the Irs2 promoter was positively associated with IRS2 expression. Treatment of MIN6 cells with histone deacetylase inhibitors led to increased IRS2 expression, but this occurred in concert with low insulin signaling. We observed increased IRS2 lysine acetylation as a consequence of histone deacetylase inhibition, a modification that was coupled with a decrease in IRS2 tyrosine phosphorylation. These results suggest that insulin signaling in pancreatic β cells is regulated by histone deacetylases through two novel pathways affecting IRS2: the epigenetic control of IRS2 expression by H3K9 promoter acetylation, and the regulation of IRS2 activity through protein modification. The identification of the histone deacetylase isoform(s involved in these mechanisms would be a valuable approach for the treatment of type 2 diabetes.

  4. Chinese hamster ovary (CHO-K1) cells expressed native insulin-like ...

    African Journals Online (AJOL)

    GREGORY

    2011-12-16

    Dec 16, 2011 ... ... University Malaysia (IIUM), P.O. Box 10, 50728, Kuala Lumpur, Malaysia. Accepted 7 November, 2011. Insulin-like growth factor-1 (IGF-1) has been shown to promote cell proliferation and inhibit apoptosis of cells. These are two characteristics of mammalian cell culture which may lead to high density cell.

  5. Prolonged inorganic arsenite exposure suppresses insulin-stimulated AKT S473 phosphorylation and glucose uptake in 3T3-L1 adipocytes: involvement of the adaptive antioxidant response.

    Science.gov (United States)

    Xue, Peng; Hou, Yongyong; Zhang, Qiang; Woods, Courtney G; Yarborough, Kathy; Liu, Huiyu; Sun, Guifan; Andersen, Melvin E; Pi, Jingbo

    2011-04-08

    There is growing evidence that chronic exposure of humans to inorganic arsenic, a potent environmental oxidative stressor, is associated with the incidence of type 2 diabetes (T2D). One critical feature of T2D is insulin resistance in peripheral tissues, especially in mature adipocytes, the hallmark of which is decreased insulin-stimulated glucose uptake (ISGU). Despite the deleterious effects of reactive oxygen species (ROS), they have been recognized as a second messenger serving an intracellular signaling role for insulin action. Nuclear factor erythroid 2-related factor 2 (NRF2) is a central transcription factor regulating cellular adaptive response to oxidative stress. This study proposes that in response to arsenic exposure, the NRF2-mediated adaptive induction of endogenous antioxidant enzymes blunts insulin-stimulated ROS signaling and thus impairs ISGU. Exposure of differentiated 3T3-L1 cells to low-level (up to 2 μM) inorganic arsenite (iAs³(+)) led to decreased ISGU in a dose- and time-dependent manner. Concomitant to the impairment of ISGU, iAs³(+) exposure significantly attenuated insulin-stimulated intracellular ROS accumulation and AKT S473 phosphorylation, which could be attributed to the activation of NRF2 and induction of a battery of endogenous antioxidant enzymes. In addition, prolonged iAs³(+) exposure of 3T3-L1 adipocytes resulted in significant induction of inflammatory response genes and decreased expression of adipogenic genes and glucose transporter type 4 (GLUT4), suggesting chronic inflammation and reduction in GLUT4 expression may also be involved in arsenic-induced insulin resistance in adipocytes. Taken together our studies suggest that prolonged low-level iAs³(+) exposure activates the cellular adaptive oxidative stress response, which impairs insulin-stimulated ROS signaling that is involved in ISGU, and thus causes insulin resistance in adipocytes. Copyright © 2011 Elsevier Inc. All rights reserved.

  6. The effect of feeding frequency on insulin and ghrelin responses in human subjects

    DEFF Research Database (Denmark)

    Solomon, Thomas; Chambers, Edward S; Jeukendrup, Asker E

    2008-01-01

    Recent work shows that increased meal frequency reduces ghrelin responses in sheep. Human research suggests there is an interaction between insulin and ghrelin. The effect of meal frequency on this interaction is unknown. Therefore, we investigated the effect of feeding frequency on insulin...... and ghrelin responses in human subjects. Five healthy male volunteers were recruited from the general population: age 24 (SEM 2)years, body mass 75.7 (SEM 3.2) kg and BMI 23.8 (SEM 0.8) kg/m(2). Volunteers underwent three 8-h feeding regimens: fasting (FAST); low-frequency(two) meal ingestion (LOFREQ......(MEAL)); high-frequency (twelve) meal ingestion (HIFREQ(MEAL)). Meals were equi-energetic within trials,consisting of 64% carbohydrate, 23% fat and 13% protein. Total energy intake was equal between feeding trials. Total area under the curve for serum insulin and plasma ghrelin responses did not differ between...

  7. Human Liver Cells Expressing Albumin and Mesenchymal Characteristics Give Rise to Insulin-Producing Cells

    Directory of Open Access Journals (Sweden)

    Irit Meivar-Levy

    2011-01-01

    Full Text Available Activation of the pancreatic lineage in the liver has been suggested as a potential autologous cell replacement therapy for diabetic patients. Transcription factors-induced liver-to-pancreas reprogramming has been demonstrated in numerous species both in vivo and in vitro. However, human-derived liver cells capable of acquiring the alternate pancreatic repertoire have never been characterized. It is yet unknown whether hepatic-like stem cells or rather adult liver cells give rise to insulin-producing cells. Using an in vitro experimental system, we demonstrate that proliferating adherent human liver cells acquire mesenchymal-like characteristics and a considerable level of cellular plasticity. However, using a lineage-tracing approach, we demonstrate that insulin-producing cells are primarily generated in cells enriched for adult hepatic markers that coexpress both albumin and mesenchymal markers. Taken together, our data suggest that adult human hepatic tissue retains a substantial level of developmental plasticity, which could be exploited in regenerative medicine approaches.

  8. Mathematical modeling and statistical analysis of calcium-regulated insulin granule exocytosis in ß-cells from mice and humans

    DEFF Research Database (Denmark)

    Pedersen, Morten Gram; Cortese, Giuliana; Eliasson, Lena

    2011-01-01

    Insulin is released from pancreatic ß-cells as a result of Ca2+-evoked exocytosis of dense-core granules. Secretion is biphasic, which has been suggested to correspond to the release of different granule pools. Here we review and carefully reanalyze previously published patch-clamp data on depola......Insulin is released from pancreatic ß-cells as a result of Ca2+-evoked exocytosis of dense-core granules. Secretion is biphasic, which has been suggested to correspond to the release of different granule pools. Here we review and carefully reanalyze previously published patch-clamp data...... on depolarization-evoked Ca2+-currents and corresponding capacitance measurements. Using a statistical mixed-effects model, we show that the data indicate that pool depletion is negligible in response to short depolarizations in mouse ß-cells. We then review mathematical models of granule dynamics and exocytosis...

  9. Evaluation of Insulin Medium or Chondrogenic Medium on Proliferation and Chondrogenesis of ATDC5 Cells

    OpenAIRE

    Yao, Yongchang; Zhai, Zhichen; Wang, Yingjun

    2014-01-01

    Background. The ATDC5 cell line is regarded as an excellent cell model for chondrogenesis. In most studies with ATDC5 cells, insulin medium (IM) was used to induce chondrogenesis while chondrogenic medium (CM), which was usually applied in chondrogenesis of mesenchymal stem cells (MSCs), was rarely used for ATDC5 cells. This study was mainly designed to investigate the effect of IM, CM, and growth medium (GM) on chondrogenesis of ATDC5 cells. Methods. ATDC5 cells were, respectively, cultured ...

  10. Effects of 7 days of exercise training on insulin sensitivity and responsiveness in type 2 diabetes mellitus

    DEFF Research Database (Denmark)

    Kirwan, John P; Solomon, Thomas; Wojta, Daniel M

    2009-01-01

    sensitivity and responsiveness and 2) short-term exercise training results in improved suppression of hepatic glucose production by insulin. Fourteen obese patients with type 2 diabetes, age 64 +/- 2 yr, underwent a two-stage hyperinsulinemic euglycemic clamp procedure, first stage 40 mU.m(-2).min(-1) insulin......The objectives of this study were to determine whether 1) the improvement in insulin action induced by short-term exercise training in patients with type 2 diabetes is due to an improvement in insulin sensitivity, an improvement in insulin responsiveness, or a combination of improved insulin...... infusion, second stage 1,000 mU.m(-2).min(-1) insulin infusion, together with a [3-(3)H]glucose infusion, before and after 7 days of exercise. The training consisted of 30 min of cycling and 30 min of treadmill walking at approximately 70% of maximal aerobic capacity daily for 7 days. The exercise program...

  11. Insulin sensitivity and beta-cell function after carbohydrate oral loading in hip replacement surgery: a double-blind, randomised controlled clinical trial.

    Science.gov (United States)

    Ljunggren, Stefan; Hahn, Robert G; Nyström, Thomas

    2014-06-01

    Surgery initiates a series of physiological stress processes in the body, inducing transient insulin resistance. Preoperative carbohydrate treatment can reduce the latter phenomenon. We investigated the effects of carbohydrate loading on insulin sensitivity and beta-cell function after elective hip replacement. Twenty-three nondiabetic patients (mean age of 68 years) who underwent elective hip replacement surgery participated in this double-blind controlled study. The patients were randomised to a nutrition group, which ingested a carbohydrate-rich fluid (50 kcal/100 ml) (Preop(®)), or a control group (tap water flavoured with lemon) 800 ml + 400 ml before the surgery. The insulin response (beta-cell function) and the insulin sensitivity were measured with an intravenous glucose tolerance test (IVGTT) and a hyperinsulinaemic euglycaemic glucose clamp, respectively, one day before and two days after the surgery. Insulin sensitivity decreased by 51% (median; 25-75th percentiles 35-61) after ingesting Preop(®) and by 39% (21-51) after ingesting in the control group (n.s.). The postoperative IVGTT in the nutrition group was followed by a significantly larger area under the curve (AUC) for plasma insulin (+54% versus the preoperative IVGTT) compared to the control group (+7%). This difference was already apparent during the first phase (0-10 min) of insulin secretion (+20 and -21%, respectively; P water prior to the surgery demonstrated a significant but similar decrease in insulin sensitivity. The carbohydrates increased the beta-cell function as a compensatory response to the disposition index, resulting in a smaller reduction in surgery-induced insulin resistance compared to the tap water. The study was registered at http://www.clinicaltrials.gov (NCT01774084). Copyright © 2013 Elsevier Ltd and European Society for Clinical Nutrition and Metabolism. All rights reserved.

  12. Increased Incretin But Not Insulin Response after Oral versus Intravenous Branched Chain Amino Acids.

    Science.gov (United States)

    Gojda, Jan; Straková, Radka; Plíhalová, Andrea; Tůma, Petr; Potočková, Jana; Polák, Jan; Anděl, Michal

    2017-01-01

    Branched chain amino acids (BCAAs) are known to exert an insulinotropic effect. Whether this effect is mediated by incretins (glucagon like peptide 1 [GLP-1] or glucose-dependent insulinotropic peptide [GIP]) is not known. The aim of this study was to show whether an equivalent dose of BCAA elicits a greater insulin and incretin response when administered orally than intravenously (IV). Eighteen healthy, male subjects participated in 3 tests: IV application of BCAA solution, oral ingestion of BCAA and placebo in an equivalent dose (30.7 ± 1.1 g). Glucose, insulin, C-peptide, glucagon, GLP-1, GIP, valine, leucine and isoleucine concentrations were measured. Rise in serum BCAA was achieved in both BCAA tests, with incremental areas under the curve (iAUC) being 2.1 times greater for IV BCAA compared with those of the oral BCAA test (p BCAA induced comparable insulin response greater than placebo (240 min insulin iAUC: oral 3,411 ± 577 vs. IV 2,361 ± 384 vs. placebo 961.2 ± 175 pmol/L, p = 0.0006). Oral BCAA induced higher GLP-1 (p BCAA tests with no change in the placebo group. An equivalent dose of BCAA elicited a comparable insulin and greater incretin response when administered orally and not when administered through IV. We conclude that insulinotropic effects of BCAA are partially incretin dependent. © 2017 S. Karger AG, Basel.

  13. Generation of insulin-producing cells from rat mesenchymal stem cells using an aminopyrrole derivative XW4.4.

    Science.gov (United States)

    Ouyang, Jingfeng; Huang, Wei; Yu, Wanwan; Xiong, Wei; Mula, Ramanjaneya V R; Zou, Hongbin; Yu, Yongping

    2014-02-05

    Type 1 diabetes mellitus (T1DM), a multisystem disease with both biochemical and anatomical/structural consequences, is a major health concern worldwide. Pancreatic islet transplantation provides a promising treatment for T1DM. However, the limited availability of islet tissue or new sources of insulin producing cells (IPCs) that are responsive to glucose hinder this promising approach. Though slow, the development of pancreatic beta-cell lines from rodent or human origin has been steadily progressing. Bone marrow-derived mesenchymal stem cells (MSCs) are multipotent, culture-expanded, non-hematopoietic cells that are currently being investigated as a novel cellular therapy. The in vitro differentiation potential of IPCs has raised hopes for a treatment of clinical diseases associated with autoimmunity. We screened for small molecules that induce pancreatic differentiation of IPCs. There are some compounds which showed positive effects on the DTZ staining. The aminopyrrole derivative compound XW4.4 which shows the best activity among them was found to induce pancreatic differentiation of rat MSCs (rMSCs). The in vitro studies indicated that treatment of rMSCs with compound XW4.4 resulted in differentiated cells with characteristics of IPCs including islet-like clusters, spherical, grape-like morphology, insulin secretion, positive for dithizone, glucose stimulation and expression of pancreatic endocrine cell marker genes. The data has also suggested that hepatocyte nuclear factor 3β (HNF 3β) may be involved in pancreatic differentiation of rMSCs when treated with XW4.4. Results indicate that XW4.4 induced rMSCs support the efforts to derive functional IPCs and serve as a means to alleviate limitations surrounding islet cell transplantation in the treatment of T1DM. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  14. Prolonged inorganic arsenite exposure suppresses insulin-stimulated AKT S473 phosphorylation and glucose uptake in 3T3-L1 adipocytes: Involvement of the adaptive antioxidant response

    International Nuclear Information System (INIS)

    Xue, Peng; Hou, Yongyong; Zhang, Qiang; Woods, Courtney G.; Yarborough, Kathy; Liu, Huiyu; Sun, Guifan; Andersen, Melvin E.; Pi, Jingbo

    2011-01-01

    Highlights: → In 3T3-L1 adipocytes iAs 3+ decreases insulin-stimulated glucose uptake. → iAs 3+ attenuates insulin-induced phosphorylation of AKT S473. → iAs 3+ activates the cellular adaptive oxidative stress response. → iAs 3+ impairs insulin-stimulated ROS signaling. → iAs 3+ decreases expression of adipogenic genes and GLUT4. -- Abstract: There is growing evidence that chronic exposure of humans to inorganic arsenic, a potent environmental oxidative stressor, is associated with the incidence of type 2 diabetes (T2D). One critical feature of T2D is insulin resistance in peripheral tissues, especially in mature adipocytes, the hallmark of which is decreased insulin-stimulated glucose uptake (ISGU). Despite the deleterious effects of reactive oxygen species (ROS), they have been recognized as a second messenger serving an intracellular signaling role for insulin action. Nuclear factor erythroid 2-related factor 2 (NRF2) is a central transcription factor regulating cellular adaptive response to oxidative stress. This study proposes that in response to arsenic exposure, the NRF2-mediated adaptive induction of endogenous antioxidant enzymes blunts insulin-stimulated ROS signaling and thus impairs ISGU. Exposure of differentiated 3T3-L1 cells to low-level (up to 2 μM) inorganic arsenite (iAs 3+ ) led to decreased ISGU in a dose- and time-dependent manner. Concomitant to the impairment of ISGU, iAs 3+ exposure significantly attenuated insulin-stimulated intracellular ROS accumulation and AKT S473 phosphorylation, which could be attributed to the activation of NRF2 and induction of a battery of endogenous antioxidant enzymes. In addition, prolonged iAs 3+ exposure of 3T3-L1 adipocytes resulted in significant induction of inflammatory response genes and decreased expression of adipogenic genes and glucose transporter type 4 (GLUT4), suggesting chronic inflammation and reduction in GLUT4 expression may also be involved in arsenic-induced insulin resistance in

  15. Prolonged inorganic arsenite exposure suppresses insulin-stimulated AKT S473 phosphorylation and glucose uptake in 3T3-L1 adipocytes: Involvement of the adaptive antioxidant response

    Energy Technology Data Exchange (ETDEWEB)

    Xue, Peng [The Hamner Institutes for Health Sciences, Research Triangle Park, NC 27709 (United States); School of Public Health, China Medical University, Shenyang 110001 (China); Hou, Yongyong; Zhang, Qiang; Woods, Courtney G.; Yarborough, Kathy; Liu, Huiyu [The Hamner Institutes for Health Sciences, Research Triangle Park, NC 27709 (United States); Sun, Guifan [School of Public Health, China Medical University, Shenyang 110001 (China); Andersen, Melvin E. [The Hamner Institutes for Health Sciences, Research Triangle Park, NC 27709 (United States); Pi, Jingbo, E-mail: jpi@thehamner.org [The Hamner Institutes for Health Sciences, Research Triangle Park, NC 27709 (United States)

    2011-04-08

    Highlights: {yields} In 3T3-L1 adipocytes iAs{sup 3+} decreases insulin-stimulated glucose uptake. {yields} iAs{sup 3+} attenuates insulin-induced phosphorylation of AKT S473. {yields} iAs{sup 3+} activates the cellular adaptive oxidative stress response. {yields} iAs{sup 3+} impairs insulin-stimulated ROS signaling. {yields} iAs{sup 3+} decreases expression of adipogenic genes and GLUT4. -- Abstract: There is growing evidence that chronic exposure of humans to inorganic arsenic, a potent environmental oxidative stressor, is associated with the incidence of type 2 diabetes (T2D). One critical feature of T2D is insulin resistance in peripheral tissues, especially in mature adipocytes, the hallmark of which is decreased insulin-stimulated glucose uptake (ISGU). Despite the deleterious effects of reactive oxygen species (ROS), they have been recognized as a second messenger serving an intracellular signaling role for insulin action. Nuclear factor erythroid 2-related factor 2 (NRF2) is a central transcription factor regulating cellular adaptive response to oxidative stress. This study proposes that in response to arsenic exposure, the NRF2-mediated adaptive induction of endogenous antioxidant enzymes blunts insulin-stimulated ROS signaling and thus impairs ISGU. Exposure of differentiated 3T3-L1 cells to low-level (up to 2 {mu}M) inorganic arsenite (iAs{sup 3+}) led to decreased ISGU in a dose- and time-dependent manner. Concomitant to the impairment of ISGU, iAs{sup 3+} exposure significantly attenuated insulin-stimulated intracellular ROS accumulation and AKT S473 phosphorylation, which could be attributed to the activation of NRF2 and induction of a battery of endogenous antioxidant enzymes. In addition, prolonged iAs{sup 3+} exposure of 3T3-L1 adipocytes resulted in significant induction of inflammatory response genes and decreased expression of adipogenic genes and glucose transporter type 4 (GLUT4), suggesting chronic inflammation and reduction in GLUT4

  16. Synaptotagmin-7 phosphorylation mediates GLP-1-dependent potentiation of insulin secretion from β-cells

    DEFF Research Database (Denmark)

    Wu, Bingbing; Wei, Shunhui; Petersen, Natalia

    2015-01-01

    Glucose stimulates insulin secretion from β-cells by increasing intracellular Ca(2+). Ca(2+) then binds to synaptotagmin-7 as a major Ca(2+) sensor for exocytosis, triggering secretory granule fusion and insulin secretion. In type-2 diabetes, insulin secretion is impaired; this impairment...... is ameliorated by glucagon-like peptide-1 (GLP-1) or by GLP-1 receptor agonists, which improve glucose homeostasis. However, the mechanism by which GLP-1 receptor agonists boost insulin secretion remains unclear. Here, we report that GLP-1 stimulates protein kinase A (PKA)-dependent phosphorylation...... of synaptotagmin-7 at serine-103, which enhances glucose- and Ca(2+)-stimulated insulin secretion and accounts for the improvement of glucose homeostasis by GLP-1. A phospho-mimetic synaptotagmin-7 mutant enhances Ca(2+)-triggered exocytosis, whereas a phospho-inactive synaptotagmin-7 mutant disrupts GLP-1...

  17. Effects of aging and insulin resistant states on protein anabolic responses in older adults.

    Science.gov (United States)

    Morais, Jose A; Jacob, Kathryn Wright; Chevalier, Stéphanie

    2018-07-15

    Insulin is the principal postprandial anabolic hormone and resistance to its action could contribute to sarcopenia. We developed different types of hyperinsulinemic clamp protocols to measure simultaneously glucose and protein metabolism in insulin resistant states (older adults, obesity, diabetes, etc.). To define effects of healthy aging in response to insulin, we employed the hyperinsulinemic, euglycemic and isoaminoacidemic (HYPER-1) clamp. The net whole-body anabolic (protein balance) response to hyperinsulinemia was lower in the elderly vs young (p = 0.007) and was highly correlated with the clamp glucose rate of disposal (r = 0.671, p anabolism compared with young ones. As most of the anabolism occurs during feeding, we studied the fed-state metabolic responses with aging using the hyperinsulinemic, hyperglycemic and hyperaminoacidemic clamp, including muscle biopsies. Older women showed comparable whole-body protein anabolic responses and stimulation of mixed-muscle protein synthesis by feeding to the young. The responses of skeletal muscle insulin signaling through the Akt-mTORC1 pathway were also unaltered, and therefore consistent with muscle protein synthesis results. Given that type 2 diabetes infers insulin resistance of protein metabolism with aging, we studied 10 healthy, 8 obese, and 8 obese type 2 diabetic elderly women using the HYPER-1 clamp. When compared to the group of young lean women to define the effects of obesity and diabetes with aging, whole-body change in net protein balance with hyperinsulinemia was similarly blunted in obese and diabetic older women. However, only elderly obese women with diabetes had lower net balance than lean older women. We conclude that with usual aging, the blunted whole-body anabolic response in elderly subjects is mediated by the failure of insulin to stimulate protein synthesis to the same extent as in the young, especially in men. This blunted response can be overcome at the whole-body and muscle

  18. TET1-GPER-PI3K/AKT pathway is involved in insulin-driven endometrial cancer cell proliferation

    International Nuclear Information System (INIS)

    Xie, Bing-ying; Lv, Qiao-ying; Ning, Cheng-cheng; Yang, Bing-yi; Shan, Wei-wei; Cheng, Ya-li; Gu, Chao; Luo, Xue-zhen; Zhang, Zhen-bo; Chen, Xiao-jun; Xi, Xiao-wei; Feng, You-ji

    2017-01-01

    Large amount of clinical evidence has demonstrated that insulin resistance is closely related to oncogenesis of endometrial cancer (EC). Despite recent studies showed the up-regulatory role of insulin in G protein-coupled estrogen receptor (GPER/GPR30) expression, GPER expression was not decreased compared to control when insulin receptor was blocked even in insulin treatment. The purpose of this study was to explore the possible mechanism by which insulin up-regulates GPER that drives EC cell proliferation. For this purpose, we first investigated the GPER expression in tissues of endometrial lesions, further explored the effect of GPER on EC cell proliferation in insulin resistance context. Then we analyzed the role of Ten-Eleven Translocation 1 (TET1) in insulin-induced GEPR expression and EC cell proliferation. The results showed that GPER was highly expressed in endometrial atypical hyperplasia and EC tissues. Mechanistically, insulin up-regulated TET1 expression and the latter played an important role in up-regulating GPER expression and activating PI3K/AKT signaling pathway. TET1 mediated GPER up-regulation was another mechanism that insulin promotes EC cell proliferation. - Highlights: • GPER acts as an oncogene to drive EC cell growth in insulin resistance context. • TET1 is associated with insulin-induced GPER expression. • Insulin resistance contributed to EC through TET1-GPER-PI3K/AKT pathway.

  19. Enrichment of human embryonic stem cell-derived NKX6.1-expressing pancreatic progenitor cells accelerates the maturation of insulin-secreting cells in vivo.

    Science.gov (United States)

    Rezania, Alireza; Bruin, Jennifer E; Xu, Jean; Narayan, Kavitha; Fox, Jessica K; O'Neil, John J; Kieffer, Timothy J

    2013-11-01

    Human embryonic stem cells (hESCs) are considered a potential alternative to cadaveric islets as a source of transplantable cells for treating patients with diabetes. We previously described a differentiation protocol to generate pancreatic progenitor cells from hESCs, composed of mainly pancreatic endoderm (PDX1/NKX6.1-positive), endocrine precursors (NKX2.2/synaptophysin-positive, hormone/NKX6.1-negative), and polyhormonal cells (insulin/glucagon-positive, NKX6.1-negative). However, the relative contributions of NKX6.1-negative versus NKX6.1-positive cell fractions to the maturation of functional β-cells remained unclear. To address this question, we generated two distinct pancreatic progenitor cell populations using modified differentiation protocols. Prior to transplant, both populations contained a high proportion of PDX1-expressing cells (~85%-90%) but were distinguished by their relatively high (~80%) or low (~25%) expression of NKX6.1. NKX6.1-high and NKX6.1-low progenitor populations were transplanted subcutaneously within macroencapsulation devices into diabetic mice. Mice transplanted with NKX6.1-low cells remained hyperglycemic throughout the 5-month post-transplant period whereas diabetes was reversed in NKX6.1-high recipients within 3 months. Fasting human C-peptide levels were similar between groups throughout the study, but only NKX6.1-high grafts displayed robust meal-, glucose- and arginine-responsive insulin secretion as early as 3 months post-transplant. NKX6.1-low recipients displayed elevated fasting glucagon levels. Theracyte devices from both groups contained almost exclusively pancreatic endocrine tissue, but NKX6.1-high grafts contained a greater proportion of insulin-positive and somatostatin-positive cells, whereas NKX6.1-low grafts contained mainly glucagon-expressing cells. Insulin-positive cells in NKX6.1-high, but not NKX6.1-low grafts expressed nuclear MAFA. Collectively, this study demonstrates that a pancreatic endoderm

  20. Effect of High Fat and High Sugar Diet on Glucose Tolerance, Insulin Response to Glucose Load and Insulin Sensitivity in Rats

    OpenAIRE

    岡﨑, 悟

    1987-01-01

    To investigate the precipitating effects of the westernized diet on diabetes mellitus, glucose tolerance and insulin response to oral glucose load (1.5g/kg body weight) and insulin sensitivity to exogenous insulin (0.2U/kg) were studied in rats fed an experimental diet for 8 weeks. Four experimental diets were used : low fat-no sugar diet (energy ratio of 10% fat, 70% starch, a model of the traditional Japanese diet), high fat-high sugar diet (40% fat, 20% starch, 20% sugar, a model of the we...

  1. Maternal Rat Diabetes Mellitus Deleteriously Affects Insulin Sensitivity and Beta-Cell Function in the Offspring

    Directory of Open Access Journals (Sweden)

    Abdel-Baset M. Aref

    2013-01-01

    Full Text Available This study was designed to assess the effect of maternal diabetes in rats on serum glucose and insulin concentrations, insulin resistance, histological architecture of pancreas and glycogen content in liver of offspring. The pregnant rat females were allocated into two main groups: normal control group and streptozotocin-induced diabetic group. After birth, the surviving offspring were subjected to biochemical and histological examination immediately after delivery and at the end of the 1st and 2nd postnatal weeks. In comparison with the offspring of normal control dams, the fasting serum glucose level of offspring of diabetic mothers was significantly increased at the end of the 1st and 2nd postnatal weeks. Serum insulin level of offspring of diabetic dams was significantly higher at birth and decreased significantly during the following 2 postnatal weeks, while in normal rat offspring, it was significantly increased with progress of time. HOMA Insulin Resistance (HOMA-IR was significantly increased in the offspring of diabetic dams at birth and after 1 week than in normal rat offspring, while HOMA insulin sensitivity (HOMA-IS was significantly decreased. HOMA beta-cell function was significantly decreased at all-time intervals in offspring of diabetic dams. At birth, islets of Langerhans as well as beta cells in offspring of diabetic dams were hypertrophied. The cells constituting islets seemed to have a high division rate. However, beta-cells were degenerated during the following 2 post-natal weeks and smaller insulin secreting cells predominated. Vacuolation and necrosis of the islets of Langerhans were also observed throughout the experimental period. The carbohydrate content in liver of offspring of diabetic dams was at all-time intervals lower than that in control. The granule distribution was more random. Overall, the preexisting maternal diabetes leads to glucose intolerance, insulin resistance, and impaired insulin sensitivity and β-cell

  2. Autophagy is essential for the differentiation of porcine PSCs into insulin-producing cells.

    Science.gov (United States)

    Ren, Lipeng; Yang, Hong; Cui, Yanhua; Xu, Shuanshuan; Sun, Fen; Tian, Na; Hua, Jinlian; Peng, Sha

    2017-07-01

    Porcine pancreatic stem cells (PSCs) are seed cells with potential use for diabetes treatment. Stem cell differentiation requires strict control of protein turnover and lysosomal digestion of organelles. Autophagy is a highly conserved process that controls the turnover of organelles and proteins within cells and contributes to the balance of cellular components. However, whether autophagy plays roles in PSC differentiation remains unknown. In this study, we successfully induced porcine PSCs into insulin-producing cells and found that autophagy was activated during the second induction stage. Inhibition of autophagy in the second stage resulted in reduced differentiational efficiency and impaired glucose-stimulated insulin secretion. Moreover, the expression of active β-catenin increased while autophagy was activated but was suppressed when autophagy was inhibited. Therefore, autophagy is essential to the formation of insulin-producing cells, and the effects of autophagy on differentiation may be regulated by canonical Wnt signalling pathway. Copyright © 2017 Elsevier Inc. All rights reserved.

  3. Fibroblast growth factor 21 improves insulin sensitivity and synergizes with insulin in human adipose stem cell-derived (hASC adipocytes.

    Directory of Open Access Journals (Sweden)

    Darwin V Lee

    Full Text Available Fibroblast growth factor 21 (FGF21 has evolved as a major metabolic regulator, the pharmacological administration of which causes weight loss, insulin sensitivity and glucose control in rodents and humans. To understand the molecular mechanisms by which FGF21 exerts its metabolic effects, we developed a human in vitro model of adipocytes to examine crosstalk between FGF21 and insulin signaling. Human adipose stem cell-derived (hASC adipocytes were acutely treated with FGF21 alone, insulin alone, or in combination. Insulin signaling under these conditions was assessed by measuring tyrosine phosphorylation of insulin receptor (InsR, insulin receptor substrate-1 (IRS-1, and serine 473 phosphorylation of Akt, followed by a functional assay using 14C-2-deoxyglucose [14C]-2DG to measure glucose uptake in these cells. FGF21 alone caused a modest increase of glucose uptake, but treatment with FGF21 in combination with insulin had a synergistic effect on glucose uptake in these cells. The presence of FGF21 also effectively lowered the insulin concentration required to achieve the same level of glucose uptake compared to the absence of FGF21 by 10-fold. This acute effect of FGF21 on insulin signaling was not due to IR, IGF-1R, or IRS-1 activation. Moreover, we observed a substantial increase in basal S473-Akt phosphorylation by FGF21 alone, in contrast to the minimal shift in basal glucose uptake. Taken together, our data demonstrate that acute co-treatment of hASC-adipocytes with FGF21 and insulin can result in a synergistic improvement in glucose uptake. These effects were shown to occur at or downstream of Akt, or separate from the canonical insulin signaling pathway.

  4. Genetically Engineered Islets and Alternative Sources of Insulin-Producing Cells for Treating Autoimmune Diabetes: Quo Vadis?

    Directory of Open Access Journals (Sweden)

    Feng-Cheng Chou

    2012-01-01

    Full Text Available Islet transplantation is a promising therapy for patients with type 1 diabetes that can provide moment-to-moment metabolic control of glucose and allow them to achieve insulin independence. However, two major problems need to be overcome: (1 detrimental immune responses, including inflammation induced by the islet isolation/transplantation procedure, recurrence autoimmunity, and allorejection, can cause graft loss and (2 inadequate numbers of organ donors. Several gene therapy approaches and pharmaceutical treatments have been demonstrated to prolong the survival of pancreatic islet grafts in animal models; however, the clinical applications need to be investigated further. In addition, for an alternative source of pancreatic β-cell replacement therapy, the ex vivo generation of insulin-secreting cells from diverse origins of stem/progenitor cells has become an attractive option in regenerative medicine. This paper focuses on the genetic manipulation of islets during transplantation therapy and summarizes current strategies to obtain functional insulin-secreting cells from stem/progenitor cells.

  5. High fat feeding results in a decrease in insulin responsiveness of isolated solei

    International Nuclear Information System (INIS)

    Grundleger, M.L.; Preves, D.M.

    1986-01-01

    The relationship between diet and insulin responsiveness was examined in isolated solei from 6 week old female Sprague-Dawley rats. Weanling rats were fed either a high fat (HF) (67%kcal) or a high carbohydrate diet (HC) (67% kcal) for 21 days. A significant decrease in plasma insulin (I) but not glucose was observed in the HF fed rats. Insulin stimulated (IS) glucose (G) metabolism was examined using a maximal concentration of I (20 mU/m1). G uptake was estimated using 14 C-2 deoxyglucose (2DG). Basal and IS 2DG uptake decreased in HF rats. However, I sensitivity but not responsiveness remained intact in the HF rats. Total G utilization (GU) was estimated by the sum of the rate of formation of: 3 H 2 O from 5- 3 H-glucose [glycolysis- (GL)] and 3 H-glycogen (GLY). IS GU decreased in HF versus HC fed rats. I failed to stimulate GL while GLY remained sensitive. Glucose oxidation (GO) was measured by 14 CO 2 . I failed to stimulated GO. Intracellular metabolite concentrations (IC) were measured in solei from HF and HC fed rats. IS IC-G6P decreased in HF compared to HC fed rats. Basal IC-F6P but not IC-F 1.6 BP increased in HF compared to HC fed rats. I failed to stimulate an increase in IC-F 1,6BP concentrations. Glycolytic activators were determined. HF produced a significant decrease in F2, 6BP concentration when compared to HC rats. Prostaglandins (PG) have been implicated in mediating insulin action. HF produced a significant decrease in basal and insulin stimulated PGE 2 . These data demonstrate that postreceptor - postmembrane alterations are in part responsible for the decreased insulin responsiveness observed after HF feeding

  6. Transformation of Nonfunctioning Pancreatic Neuroendocrine Carcinoma Cells into Insulin Producing Cells after Treatment with Sunitinib

    Directory of Open Access Journals (Sweden)

    Jung Hun Ohn

    2013-06-01

    Full Text Available We report a rare case of severe hypoglycemia after sunitinib treatment for pancreatic neuroendocrine carcinoma. We describe the initial clinical presentation, laboratory results, pathologic findings, and managment in a patient with a nonfunctioning pancreatic neuroendocrine carcinoma with liver metastases who developed life threatening hypoglycemia after 2 months of sunitinib therapy. A 46-year-old woman presented to the emergency department with loss of consciousness from hypoglycemia. Serum C-peptide and insulin levels at fasting state revealed that the hypoglycemia resulted from endogenous hyperinsulinemia. She had been diagnosed with nonfunctioning pancreatic neuroendocrine carcinoma based on a biopsy of metastatic cervical lymph node and was being treated with sunitinib, a small molecule tyrosine kinase inhibitor. Immunohistochemical stain of the metastatic liver mass demonstrated that the initially nonfunctioning neuroendocrine carcinoma cells had changed into insulin-producing cells after sunitinib therapy. Transarterial chemoembolization of the liver masses and systemic chemotherapy with streptozotocin/adriamycin relieved the hypoglycemia. A nonfunctioning pancreatic neuroendocrine carcinoma was transformed into an insulin-producing tumor after treatment with sunitinib, causing endogenous hyperinsulinemia and severe hypoglycemia.

  7. Olive Component Oleuropein Promotes β-Cell Insulin Secretion and Protects β-Cells from Amylin Amyloid-Induced Cytotoxicity.

    Science.gov (United States)

    Wu, Ling; Velander, Paul; Liu, Dongmin; Xu, Bin

    2017-09-26

    Oleuropein, a natural product derived from olive leaves, has reported anti-diabetic functions. However, detailed molecular mechanisms for how it affects β-cell functions remain poorly understood. Here, we present evidence that oleuropein promotes glucose-stimulated insulin secretion (GSIS) in β-cells. The effect is dose-dependent and stimulates the ERK/MAPK signaling pathway. We further demonstrated that oleuropein inhibits the cytotoxicity induced by amylin amyloids, a hallmark feature of type 2 diabetes. We demonstrated that these dual functions are structure-specific: we identified the 3-hydroxytyrosol moiety of oleuropein as the main functional entity responsible for amyloid inhibition, but the novel GSIS function requires the entire structure scaffold of the molecule.

  8. Cell response to surgery.

    LENUS (Irish Health Repository)

    Ni Choileain, Niamh

    2012-02-03

    OBJECTIVES: To describe the profound alterations in host immunity that are produced by major surgery as demonstrated by experimental and clinical studies, and to evaluate the benefits of therapeutic strategies aimed at attenuating perioperative immune dysfunction. DATA SOURCES: A review of the English-language literature was conducted, incorporating searches of the MEDLINE, EMBASE, and Cochrane collaboration databases to identify laboratory and clinical studies investigating the cellular response to surgery. STUDY SELECTION: Original articles and case reports describing immune dysfunction secondary to surgical trauma were included. DATA EXTRACTION: The results were compiled to show outcomes of different studies and were compared. DATA SYNTHESIS: Current evidence indicates that the early systemic inflammatory response syndrome observed after major surgery that is characterized by proinflammatory cytokine release, microcirculatory disturbance, and cell-mediated immune dysfunction is followed by a compensatory anti-inflammatory response syndrome, which predisposes the patient to opportunistic infection, multiple organ dysfunction syndrome, and death. Because there are currently no effective treatment options for multiple organ dysfunction syndrome, measures to prevent its onset should be initiated at an early stage. Accumulating experimental evidence suggests that targeted therapeutic strategies involving immunomodulatory agents such as interferon gamma, granulocyte colony-stimulating factor, the prostaglandin E(2) antagonist, indomethacin, and pentoxifylline may be used for the treatment of systemic inflammatory response syndrome to prevent the onset of multiple organ dysfunction syndrome. CONCLUSIONS: Surgical trauma produces profound immunological dysfunction. Therapeutic strategies directed at restoring immune homeostasis should aim to redress the physiological proinflammatory-anti-inflammatory cell imbalance associated with major surgery.

  9. Role of Insulin-Like Growth Factor-1 Signaling Pathway in Cisplatin-Resistant Lung Cancer Cells

    International Nuclear Information System (INIS)

    Sun Yunguang; Zheng Siyuan; Torossian, Artour; Speirs, Christina K.; Schleicher, Stephen; Giacalone, Nicholas J.; Carbone, David P.; Zhao Zhongming; Lu Bo

    2012-01-01

    Purpose: The development of drug-resistant phenotypes has been a major obstacle to cisplatin use in non–small-cell lung cancer. We aimed to identify some of the molecular mechanisms that underlie cisplatin resistance using microarray expression analysis. Methods and Materials: H460 cells were treated with cisplatin. The differences between cisplatin-resistant lung cancer cells and parental H460 cells were studied using Western blot, MTS, and clonogenic assays, in vivo tumor implantation, and microarray analysis. The cisplatin-R cells were treated with human recombinant insulin-like growth factor (IGF) binding protein-3 and siRNA targeting IGF-1 receptor. Results: Cisplatin-R cells illustrated greater expression of the markers CD133 and aldehyde dehydrogenase, more rapid in vivo tumor growth, more resistance to cisplatin- and etoposide-induced apoptosis, and greater survival after treatment with cisplatin or radiation than the parental H460 cells. Also, cisplatin-R demonstrated decreased expression of insulin-like growth factor binding protein-3 and increased activation of IGF-1 receptor signaling compared with parental H460 cells in the presence of IGF-1. Human recombinant IGF binding protein-3 reversed cisplatin resistance in cisplatin-R cells and targeting of IGF-1 receptor using siRNA resulted in sensitization of cisplatin-R-cells to cisplatin and radiation. Conclusions: The IGF-1 signaling pathway contributes to cisplatin-R to cisplatin and radiation. Thus, this pathway represents a potential target for improved lung cancer response to treatment.

  10. Role of Insulin-Like Growth Factor-1 Signaling Pathway in Cisplatin-Resistant Lung Cancer Cells

    Energy Technology Data Exchange (ETDEWEB)

    Sun Yunguang [Department of Radiation Oncology, Vanderbilt University Medical Center, Nashville, TN (United States); Zheng Siyuan [Department of Biomedical Informatics, Vanderbilt University Medical Center, Nashville, TN (United States); Torossian, Artour; Speirs, Christina K.; Schleicher, Stephen; Giacalone, Nicholas J. [Department of Radiation Oncology, Vanderbilt University Medical Center, Nashville, TN (United States); Carbone, David P. [Department of Hematology and Oncology, Vanderbilt University Medical Center, Nashville, TN (United States); Zhao Zhongming, E-mail: zhongming.zhao@vanderbilt.edu [Department of Biomedical Informatics, Vanderbilt University Medical Center, Nashville, TN (United States); Lu Bo, E-mail: bo.lu@vanderbilt.edu [Department of Radiation Oncology, Vanderbilt University Medical Center, Nashville, TN (United States)

    2012-03-01

    Purpose: The development of drug-resistant phenotypes has been a major obstacle to cisplatin use in non-small-cell lung cancer. We aimed to identify some of the molecular mechanisms that underlie cisplatin resistance using microarray expression analysis. Methods and Materials: H460 cells were treated with cisplatin. The differences between cisplatin-resistant lung cancer cells and parental H460 cells were studied using Western blot, MTS, and clonogenic assays, in vivo tumor implantation, and microarray analysis. The cisplatin-R cells were treated with human recombinant insulin-like growth factor (IGF) binding protein-3 and siRNA targeting IGF-1 receptor. Results: Cisplatin-R cells illustrated greater expression of the markers CD133 and aldehyde dehydrogenase, more rapid in vivo tumor growth, more resistance to cisplatin- and etoposide-induced apoptosis, and greater survival after treatment with cisplatin or radiation than the parental H460 cells. Also, cisplatin-R demonstrated decreased expression of insulin-like growth factor binding protein-3 and increased activation of IGF-1 receptor signaling compared with parental H460 cells in the presence of IGF-1. Human recombinant IGF binding protein-3 reversed cisplatin resistance in cisplatin-R cells and targeting of IGF-1 receptor using siRNA resulted in sensitization of cisplatin-R-cells to cisplatin and radiation. Conclusions: The IGF-1 signaling pathway contributes to cisplatin-R to cisplatin and radiation. Thus, this pathway represents a potential target for improved lung cancer response to treatment.

  11. COPII-Dependent ER Export: A Critical Component of Insulin Biogenesis and β-Cell ER Homeostasis.

    Science.gov (United States)

    Fang, Jingye; Liu, Ming; Zhang, Xuebao; Sakamoto, Takeshi; Taatjes, Douglas J; Jena, Bhanu P; Sun, Fei; Woods, James; Bryson, Tim; Kowluru, Anjaneyulu; Zhang, Kezhong; Chen, Xuequn

    2015-08-01

    Pancreatic β-cells possess a highly active protein synthetic and export machinery in the endoplasmic reticulum (ER) to accommodate the massive production of proinsulin. ER homeostasis is vital for β-cell functions and is maintained by the delicate balance between protein synthesis, folding, export, and degradation. Disruption of ER homeostasis by diabetes-causing factors leads to β-cell death. Among the 4 components to maintain ER homeostasis in β-cells, the role of ER export in insulin biogenesis is the least understood. To address this knowledge gap, the present study investigated the molecular mechanism of proinsulin ER export in MIN6 cells and primary islets. Two inhibitory mutants of the secretion-associated RAS-related protein (Sar)1 small GTPase, known to specifically block coat protein complex II (COPII)-dependent ER export, were overexpressed in β-cells using recombinant adenoviruses. Results from this approach, as well as small interfering RNA-mediated Sar1 knockdown, demonstrated that defective Sar1 function blocked proinsulin ER export and abolished its conversion to mature insulin in MIN6 cells, isolated mouse, and human islets. It is further revealed, using an in vitro vesicle formation assay, that proinsulin was packaged into COPII vesicles in a GTP- and Sar1-dependent manner. Blockage of COPII-dependent ER exit by Sar1 mutants strongly induced ER morphology change, ER stress response, and β-cell apoptosis. These responses were mediated by the PKR (double-stranded RNA-dependent kinase)-like ER kinase (PERK)/eukaryotic translation initiation factor 2α (p-eIF2α) and inositol-requiring protein 1 (IRE1)/x-box binding protein 1 (Xbp1) pathways but not via activating transcription factor 6 (ATF6). Collectively, results from the study demonstrate that COPII-dependent ER export plays a vital role in insulin biogenesis, ER homeostasis, and β-cell survival.

  12. Insulin and C peptide response, and antibody levels in hepatitis C related chronic liver disease

    International Nuclear Information System (INIS)

    Abbas, Z.; Tariq, N.; Iqbal, M.; Shah, M.A.

    2002-01-01

    Objective: Patients with cirrhosis due to hepatitis C (HC) have an increased prevalence of diabetes mellitus. The pathogenic mechanism by which HC predisposes to DM is not clear. The objective of this study was to determine the insulin and C-peptide response to 75 gram oral glucose load and measure anti phospholipid antibody levels in patients with chronic liver disease due to HC. Design: a prospective study. Place and duration of study: This study was conducted at the department of medicine, Jinnah postgraduate medical centre over period of three months. Subjects and methods: An analytical case control study was carried out on 37 patients (m-18,f=19); none of these patients had received interferon. They were divided into four groups: (a) HC cirrhosis with DM (n=9 ), (b) HC cirrhosis without DM (n=11), (c) hepatitis B (HB) cirrhosis without DM (n=7), (d) chronic hepatitis C without DM (n=10). Group C and D were taken as controls. Fasting blood samples were taken and repeated after 2 hours of 75 gram oral glucose load (2 h PG). Result: mean ages of group A,B,C and D were (yr +- SD) 51.3 +- 7.6,48.9 +- 2.4, 33.7 +-10.8 and 31.7 +- 8.8 respectively. There was no statistically significant difference in the age, Pugh score and body mass index of HC cirrhotic patients with and without DM. Patients of group A had higher fasting and 2 h PG glucose levels (P=0.003 and 0.000) and higher fasting insulin level (p=0.045). However, increments in insulin and c peptide levels 2 h PG were much less (p=0.048 and 0.003). HB cirrhotics without diabetes (group C behaved just like HC cirrhotic without diabetes (group B). Patients of group D had normal glucose tolerance and insulin and C peptide levels. All four groups had normal anti phospholipid antibody levels. Conclusion: Patients with cirrhosis due to HC nd HB show evidence of glucose intolerance in spite of hyperinsulinaemia probably due to insulin resistance. HC cirrhotics with diabetes have fasting hyperglycemia in spite of

  13. Celastrol Protects against Antimycin A-Induced Insulin Resistance in Human Skeletal Muscle Cells

    Directory of Open Access Journals (Sweden)

    Mohamad Hafizi Abu Bakar

    2015-05-01

    Full Text Available Mitochondrial dysfunction and inflammation are widely accepted as key hallmarks of obesity-induced skeletal muscle insulin resistance. The aim of the present study was to evaluate the functional roles of an anti-inflammatory compound, celastrol, in mitochondrial dysfunction and insulin resistance induced by antimycin A (AMA in human skeletal muscle cells. We found that celastrol treatment improved insulin-stimulated glucose uptake activity of AMA-treated cells, apparently via PI3K/Akt pathways, with significant enhancement of mitochondrial activities. Furthermore, celastrol prevented increased levels of cellular oxidative damage where the production of several pro-inflammatory cytokines in cultures cells was greatly reduced. Celastrol significantly increased protein phosphorylation of insulin signaling cascades with amplified expression of AMPK protein and attenuated NF-κB and PKC θ activation in human skeletal muscle treated with AMA. The improvement of insulin signaling pathways by celastrol was also accompanied by augmented GLUT4 protein expression. Taken together, these results suggest that celastrol may be advocated for use as a potential therapeutic molecule to protect against mitochondrial dysfunction-induced insulin resistance in human skeletal muscle cells.

  14. Selective Insulin Resistance in Adipocytes*

    Science.gov (United States)

    Tan, Shi-Xiong; Fisher-Wellman, Kelsey H.; Fazakerley, Daniel J.; Ng, Yvonne; Pant, Himani; Li, Jia; Meoli, Christopher C.; Coster, Adelle C. F.; Stöckli, Jacqueline; James, David E.

    2015-01-01

    Aside from glucose metabolism, insulin regulates a variety of pathways in peripheral tissues. Under insulin-resistant conditions, it is well known that insulin-stimulated glucose uptake is impaired, and many studies attribute this to a defect in Akt signaling. Here we make use of several insulin resistance models, including insulin-resistant 3T3-L1 adipocytes and fat explants prepared from high fat-fed C57BL/6J and ob/ob mice, to comprehensively distinguish defective from unaffected aspects of insulin signaling and its downstream consequences in adipocytes. Defective regulation of glucose uptake was observed in all models of insulin resistance, whereas other major actions of insulin such as protein synthesis and anti-lipolysis were normal. This defect corresponded to a reduction in the maximum response to insulin. The pattern of change observed for phosphorylation in the Akt pathway was inconsistent with a simple defect at the level of Akt. The only Akt substrate that showed consistently reduced phosphorylation was the RabGAP AS160 that regulates GLUT4 translocation. We conclude that insulin resistance in adipose tissue is highly selective for glucose metabolism and likely involves a defect in one of the components regulating GLUT4 translocation to the cell surface in response to insulin. PMID:25720492

  15. From the Cover: Cell-replacement therapy for diabetes: Generating functional insulin-producing tissue from adult human liver cells

    Science.gov (United States)

    Sapir, Tamar; Shternhall, Keren; Meivar-Levy, Irit; Blumenfeld, Tamar; Cohen, Hamutal; Skutelsky, Ehud; Eventov-Friedman, Smadar; Barshack, Iris; Goldberg, Iris; Pri-Chen, Sarah; Ben-Dor, Lya; Polak-Charcon, Sylvie; Karasik, Avraham; Shimon, Ilan; Mor, Eytan; Ferber, Sarah

    2005-05-01

    Shortage in tissue availability from cadaver donors and the need for life-long immunosuppression severely restrict the large-scale application of cell-replacement therapy for diabetic patients. This study suggests the potential use of adult human liver as alternate tissue for autologous beta-cell-replacement therapy. By using pancreatic and duodenal homeobox gene 1 (PDX-1) and soluble factors, we induced a comprehensive developmental shift of adult human liver cells into functional insulin-producing cells. PDX-1-treated human liver cells express insulin, store it in defined granules, and secrete the hormone in a glucose-regulated manner. When transplanted under the renal capsule of diabetic, immunodeficient mice, the cells ameliorated hyperglycemia for prolonged periods of time. Inducing developmental redirection of adult liver offers the potential of a cell-replacement therapy for diabetics by allowing the patient to be the donor of his own insulin-producing tissue. pancreas | transdifferentiation

  16. Glucagon and insulin response to meals in non-obese and obese Dutch women

    NARCIS (Netherlands)

    Hill, P.; Garbaczewski, L.; Koppeschaar, H.; Thijssen, J.H.H.; Waard, F. de

    1987-01-01

    Many digestive complaints are associated with abnormalities in gastrointestinal peptide hormone function. To investigate the effect of obesity on the release of pancreatic peptide hormones, we have compared the release of insulin and glucagon in non-obese-obese Dutch women in response to isocaloric

  17. Insulin Downregulates the Transcriptional Coregulator CITED2, an Inhibitor of Proangiogenic Function in Endothelial Cells

    DEFF Research Database (Denmark)

    Wang, Xuanchun; Lockhart, Samuel M; Rathjen, Thomas

    2016-01-01

    In patients with atherosclerotic complications of diabetes, impaired neovascularization of ischemic tissue in the myocardium and lower limb limits the ability of these tissues to compensate for poor perfusion. We identified 10 novel insulin-regulated genes, among them Adm, Cited2 and Ctgf, which...... were downregulated in endothelial cells by insulin through FoxO1. CITED2, which was downregulated by insulin by up to 54%, is an important negative regulator of hypoxia-inducible factor (HIF) and impaired HIF signaling is a key mechanism underlying the impairment of angiogenesis in diabetes. Consistent...... with impairment of vascular insulin action, CITED2 was increased in cardiac endothelial cells from mice with diet-induced obesity and from db/db mice and was 3.8-fold higher in arterial tissue from patients with type 2 diabetes than non-diabetic controls. CITED2 knockdown promoted endothelial tube formation...

  18. Third Exposure to a Reduced Carbohydrate Meal Lowers Evening Postprandial Insulin and GIP Responses and HOMA-IR Estimate of Insulin Resistance.

    Science.gov (United States)

    Lin, Po-Ju; Borer, Katarina T

    2016-01-01

    Postprandial hyperinsulinemia, hyperglycemia, and insulin resistance increase the risk of type 2 diabetes (T2D) and cardiovascular disease mortality. Postprandial hyperinsulinemia and hyperglycemia also occur in metabolically healthy subjects consuming high-carbohydrate diets particularly after evening meals and when carbohydrate loads follow acute exercise. We hypothesized the involvement of dietary carbohydrate load, especially when timed after exercise, and mediation by the glucose-dependent insulinotropic peptide (GIP) in this phenomenon, as this incretin promotes insulin secretion after carbohydrate intake in insulin-sensitive, but not in insulin-resistant states. Four groups of eight metabolically healthy weight-matched postmenopausal women were provided with three isocaloric meals (a pre-trial meal and two meals during the trial day) containing either 30% or 60% carbohydrate, with and without two-hours of moderate-intensity exercise before the last two meals. Plasma glucose, insulin, glucagon, GIP, glucagon-like peptide 1 (GLP-1), free fatty acids (FFAs), and D-3-hydroxybutyrate concentrations were measured during 4-h postprandial periods and 3-h exercise periods, and their areas under the curve (AUCs) were analyzed by mixed-model ANOVA, and insulin resistance during fasting and meal tolerance tests within each diet was estimated using homeostasis-model assessment (HOMA-IR). The third low-carbohydrate meal, but not the high-carbohydrate meal, reduced: (1) evening insulin AUC by 39% without exercise and by 31% after exercise; (2) GIP AUC by 48% without exercise and by 45% after exercise, and (3) evening insulin resistance by 37% without exercise and by 24% after exercise. Pre-meal exercise did not alter insulin-, GIP- and HOMA-IR- lowering effects of low-carbohydrate diet, but exacerbated evening hyperglycemia. Evening postprandial insulin and GIP responses and insulin resistance declined by over 30% after three meals that limited daily carbohydrate intake to

  19. Effects of dietary energy allowance and decline in dry matter intake during the dry period on responses to glucose and insulin in transition dairy cows.

    Science.gov (United States)

    Salin, S; Vanhatalo, A; Elo, K; Taponen, J; Boston, R C; Kokkonen, T

    2017-07-01

    We assessed whether high energy intake during the early dry period [144% of metabolizable energy (ME) requirements/d] followed by a gradual restriction of energy intake in the close-up dry period (119% of ME/d; HEI) impaired whole-body insulin sensitivity compared with a controlled energy intake (100% of ME/d; CEI) throughout the 6-wk dry period. Multiparous Ayrshire dairy cows (n = 16) were blocked by body weight, body condition score, and expected date of parturition and were used in a randomized complete block design until 10 d after parturition. Cows were fed either HEI or CEI diets based on grass silage during the first 3 wk of the dry period and grass silage supplemented with a commercial concentrate (30% of ME intake) during the final 3 wk of gestation. After calving, all cows were fed grass silage ad libitum and an increasing amount of commercial concentrate (maximum 9 kg at d 10 postpartum). Intravenous glucose tolerance tests (IVGTT) and intravenous insulin challenges were performed -10 ± 5 d (n = 15) and +10 ± 1 d (n = 14) relative to parturition. Following glucose injection, we did not find any treatment effects on glucose and insulin responses. The prepartal nonesterified fatty acid (NEFA) response of the HEI group was blunted, basal NEFA and the decrement of NEFA were smaller, and the area under the response curve (AUC) of NEFA was less negative in HEI cows than in CEI cows. The NEFA response reversed after parturition; the NEFA AUC of the HEI group was more negative than that of the CEI group. We did not find similar responses after insulin injection. Across the treatments, NEFA AUC correlated strongly with the basal NEFA concentration during the IVGTT pre- and postpartum. Calculated and model-based indices characterizing the overall glucose tolerance and β-cell function and the insulin sensitivity were higher after parturition than during the dry period. Consistent with the lower basal insulin, the acute insulin release after the glucose infusion

  20. Matrix stiffness and oxigen tension modulate epigenetic conversion of mouse dermal fibroblasts into insulin producing cells.

    Directory of Open Access Journals (Sweden)

    Alessandro Zenobi

    2017-05-01

    Full Text Available In vivo, cells are surrounded by a three-dimensional (3-D organization of supporting matrix, neighboring cells and a gradient of chemical and mechanical signals (Antoni, et al., 2015. However, the present understanding of many biological processes is mainly based on two-dimensional (2-D systems that typically provides a static environment. In the present study, we tested two different 3-D culture systems and apply them to the epigenetic conversion of mouse dermal fibroblasts into insulin producing-cells (Pennarossa, et al., 2013; Brevini, et al., 2015, combining also the use of two oxygen tensions. In particular, cells were differentiated using the Polytetrafluoroethylene micro-bioreactor (PTFE and the Polyacrylamide (PAA gels with different stiffness (1 kPa; 4 kPa, maintained either in the standard 20% or in the more physiological 5% oxygen tensions. Standard differentiation performed on plastic substrates was assessed as a control. Cell morphology (Fig.1A, insulin expression and release were analyzed to evaluate the role of both stiffness and oxygen tension in the process. The results obtained showed that 1 kPa PAA gel and PTFE system induced a significantly higher insulin expression and release than plastic and 4 kPa PAA gel, especially in low oxygen condition (Fig.1B. Furthermore, comparing the efficiency of the two systems tested, 1 kPa PAA gel ensured a higher insulin transcription than PTFE (Fig.1C. Recent studies show the direct influence of substrates on lineage commitment and cell differentiation (Engler, et al., 2006; Evans, et al., 2009. The evidence here presented confirm that the use of an appropriate stiffness (similar to the pancreatic tissue, combined with a physiological oxygen tension, promote β-cell differentiation, with beneficial effects on cell functional activity and insulin release. The present results highlight the importance of 3-D cell rearrangement and oxigen tension to promote in vitro epigenetic conversion of

  1. Intracellular serotonin modulates insulin secretion from pancreatic beta-cells by protein serotonylation.

    Directory of Open Access Journals (Sweden)

    Nils Paulmann

    2009-10-01

    Full Text Available While serotonin (5-HT co-localization with insulin in granules of pancreatic beta-cells was demonstrated more than three decades ago, its physiological role in the etiology of diabetes is still unclear. We combined biochemical and electrophysiological analyses of mice selectively deficient in peripheral tryptophan hydroxylase (Tph1-/- and 5-HT to show that intracellular 5-HT regulates insulin secretion. We found that these mice are diabetic and have an impaired insulin secretion due to the lack of 5-HT in the pancreas. The pharmacological restoration of peripheral 5-HT levels rescued the impaired insulin secretion in vivo. These findings were further evidenced by patch clamp experiments with isolated Tph1-/- beta-cells, which clearly showed that the secretory defect is downstream of Ca(2+-signaling and can be rescued by direct intracellular application of 5-HT via the clamp pipette. In elucidating the underlying mechanism further, we demonstrate the covalent coupling of 5-HT by transglutaminases during insulin exocytosis to two key players in insulin secretion, the small GTPases Rab3a and Rab27a. This renders them constitutively active in a receptor-independent signaling mechanism we have recently termed serotonylation. Concordantly, an inhibition of such activating serotonylation in beta-cells abates insulin secretion. We also observed inactivation of serotonylated Rab3a by enhanced proteasomal degradation, which is in line with the inactivation of other serotonylated GTPases. Our results demonstrate that 5-HT regulates insulin secretion by serotonylation of GTPases within pancreatic beta-cells and suggest that intracellular 5-HT functions in various microenvironments via this mechanism in concert with the known receptor-mediated signaling.

  2. Inconsistent formation and nonfunction of insulin-positive cells from pancreatic endoderm derived from human embryonic stem cells in athymic nude rats.

    Science.gov (United States)

    Matveyenko, Aleksey V; Georgia, Senta; Bhushan, Anil; Butler, Peter C

    2010-11-01

    Embryonic stem cell therapy has been proposed as a therapeutic strategy to restore β-cell mass and function in T1DM. Recently, a group from Novocell (now ViaCyte) reported successful development of glucose-responsive islet-like structures after implantation of pancreatic endoderm (PE) derived from human embryonic stem cells (hESC) into immune-deficient mice. Our objective was to determine whether implantation of hESC-derived pancreatic endoderm from Novocell into athymic nude rats results in development of viable glucose-responsive pancreatic endocrine tissue. Athymic nude rats were implanted with PE derived from hESC either via implantation into the epididymal fat pads or by subcutaneous implantation into TheraCyte encapsulation devices for 20 wk. Blood glucose, weight, and human insulin/C-peptide secretion were monitored by weekly blood draws. Graft β-cell function was assessed by a glucose tolerance test, and graft morphology was assessed by immunohistochemistry and immunofluorescence. At 20 wk postimplantation, epididymal fat-implanted PE progressed to develop islet-like structures in 50% of implants, with a mean β-cell fractional area of 0.8 ± 0.3%. Human C-peptide and insulin were detectable, but at very low levels (C-peptide = 50 ± 26 pmol/l and insulin = 15 ± 7 pmol/l); however, there was no increase in human C-peptide/insulin levels after glucose challenge. There was no development of viable pancreatic tissue or meaningful secretory function when human PE was implanted in the TheraCyte encapsulation devices. These data confirm that islet-like structures develop from hESC differentiated to PE by the protocol developed by NovoCell. However, the extent of endocrine cell formation and secretory function is not yet sufficient to be clinically relevant.

  3. C peptide and insulin releasing RIA test for the investigation of β cell function in diabetic patients

    International Nuclear Information System (INIS)

    Shi Ailan; Zhu Chengmo; Wang Qiyu; Wang Ping

    1993-01-01

    Results of C-peptide releasing RIA test in 15 normals, and 100 diabetes were summarized and compared with glucose tolerance test and serum insulin for investigating the characteristics in different types of diabetes and evaluating the functional state of islet β cell. In 36 cases of IDDM the fasting blood sugar was significantly increased, and further elevated after eating of bread, but its peak time delay in 2 hours (normalin 1 hour). The level of basal C-peptide is very low, but shows slightly weak on no response after bread stimulating test, all of this denotes that β cell function of islets severely injured. The increasing of fasting blood sugar in 64 cases of NIDDM was lower than those of IDDM. Fasting C-peptide and insulin was normal or increased, their peak value increased after bread stimulation with peak time delayed also at 2 hours. Above results demonstrated that the function of islets B cell decreased but not fully deprived. It is concluded that C-peptide and insulin stimulating test, together with OGTT can accurately assess the islets β cell function, and also have important significance in the pathogenesis, classification and staging, prognostic evaluation and monitoring of therapeutic effects in diabetes

  4. Rat liver responsiveness to gluconeogenic substrates during insulin-induced hypoglycemia

    Directory of Open Access Journals (Sweden)

    H.M. de Souza

    2001-06-01

    Full Text Available Hepatic responsiveness to gluconeogenic substrates during insulin-induced hypoglycemia was investigated. For this purpose, livers were perfused with a saturating concentration of 2 mM glycerol, 5 mM L-alanine or 5 mM L-glutamine as gluconeogenic substrates. All experiments were performed 1 h after an ip injection of saline (CN group or 1 IU/kg of insulin (IN group. The IN group showed higher (P<0.05 hepatic glucose production from glycerol, L-alanine and L-glutamine and higher (P<0.05 production of L-lactate, pyruvate and urea from L-alanine and L-glutamine. In addition, ip injection of 100 mg/kg glycerol, L-alanine and L-glutamine promoted glucose recovery. The results indicate that the hepatic capacity to produce glucose from gluconeogenic precursors was increased during insulin-induced hypoglycemia.

  5. Glucose-responsive insulin delivery for type 1 diabetes: The artificial pancreas story.

    Science.gov (United States)

    Bally, Lia; Thabit, Hood; Hovorka, Roman

    2018-06-15

    Insulin replacement therapy is integral to the management of type 1 diabetes, which is characterised by absolute insulin deficiency. Optimal glycaemic control, as assessed by glycated haemoglobin, and avoidance of hyper- and hypoglycaemic excursions have been shown to prevent diabetes-related complications. Insulin pump use has increased considerably over the past decade with beneficial effects on glycaemic control, quality of life and treatment satisfaction. The advent and progress of ambulatory glucose sensor technology has enabled continuous glucose monitoring based on real-time glucose levels to be integrated with insulin therapy. Low glucose and predictive low glucose suspend systems are currently used in clinical practice to mitigate against hypoglycaemia, and provide the first step towards feedback glucose control. The more advanced technology approach, an artificial pancreas or a closed-loop system, gradually increases and decreases insulin delivery in a glucose-responsive fashion to mitigate against hyper- and hypoglycaemia. Randomised outpatient clinical trials over the past 5 years have demonstrated the feasibility, safety and efficacy of the approach, and the recent FDA approval of the first single hormone closed-loop system establishes a new standard of care for people with type 1 diabetes. Copyright © 2017 Elsevier B.V. All rights reserved.

  6. Determination of Insulin Resistance and Beta Cell Function in Healthy Obese and Non-obese Individuals

    International Nuclear Information System (INIS)

    Kazmi, A.; Sattar, A.; Tariq, K. M.; Najamussahar; Hashim, R.; Almani, M. I.

    2013-01-01

    Objective: To determine insulin resistance and beta cell function in healthy obese and nonobese individuals of the local population. Study Design: Case control study. Place and Duration of Study: AFIP Rawalpindi in collaboration with department of medicine military hospital(MH) Rawalpindi, from Aug 2008 to Mar 2009. Methods: Eighty obese(n=40) and non-obese(n=40) subjects were selected by non-probability convenience sampling. Plasma insulin, glucose, and serum total cholestrol were estimated in fasting state. Insulin resistance was calculated by HOMA-IR and beta cell function by HOMA- equation. Results: Significant differences were observed between obese and non-obese individuals regarding insulin resistance, beta cell function, and BMI and serum total cholesterol. Mean insulin resistance in obese group was found to be 11.1 +- 5.1(range 7.0-16.2) and in non-obese group it was 0.9+-0.4 (range 0.5-1.3). This difference was highly significant (p=0.001). There was a highly significant difference between the two groups in term of beta cell function with mean rank 60.1 for obese group and 20.9 non obese groups (Asym sig. 2 tailed 0.000). Also the correlation (r = 0.064) between insulin resistance and beta cell function in obese group is highly significant (p = 0.000). Mean serum leptin levels were lower (6.3 ng/ml) in non-obese, and high (57.2 ng/ml) in the obese group. Conclusions: Insulin resistance is found higher in obese individuals. Beta cell function is significantly different between obese and non-obese groups. (author)

  7. Insulin activates single amiloride-blockable Na channels in a distal nephron cell line (A6).

    Science.gov (United States)

    Marunaka, Y; Hagiwara, N; Tohda, H

    1992-09-01

    Using the patch-clamp technique, we studied the effect of insulin on an amiloride-blockable Na channel in the apical membrane of a distal nephron cell line (A6) cultured on permeable collagen films for 10-14 days. NPo (N, number of channels per patch membrane; Po, average value of open probability of individual channels in the patch) under baseline conditions was 0.88 +/- 0.12 (SE)(n = 17). After making cell-attached patches on the apical membrane which contained Na channels, insulin (1 mU/ml) was applied to the serosal bath. While maintaining the cell-attached patch, NPo significantly increased to 1.48 +/- 0.19 (n = 17; P less than 0.001) after 5-10 min of insulin application. The open probability of Na channels was 0.39 +/- 0.01 (n = 38) under baseline condition, and increased to 0.66 +/- 0.03 (n = 38, P less than 0.001) after addition of insulin. The baseline single-channel conductance was 4pS, and neither the single-channel conductance nor the current-voltage relationship was significantly changed by insulin. These results indicate that insulin increases Na absorption in the distal nephron by increasing the open probability of the amiloride-blockable Na channel.

  8. Osteoinductive activity of insulin-functionalized cell culture surfaces obtained using diazonium chemistry

    Science.gov (United States)

    Mikulska, Anna; Filipowska, Joanna; Osyczka, Anna; Nowakowska, Maria; Szczubiałka, Krzysztof

    2014-12-01

    Polymeric surfaces suitable for cell culture (DR/Pec) were constructed from diazoresin (DR) and pectin (Pec) in a form of ultrathin films using the layer-by-layer (LbL) technique. The surfaces were functionalized with insulin using diazonium chemistry. Such functionalized surfaces were used to culture human mesenchymal stem cells (hMSCs) to assess their suitability for bone tissue engineering and regeneration. The activity of insulin immobilized on the surfaces (DR/Pec/Ins) was compared to that of insulin dissolved in the culture medium. Human MSC grown on insulin-immobilized DR/Pec surfaces displayed increased proliferation and higher osteogenic activity. The latter was determined by means of alkaline phosphatase (ALP) activity, which increases at early stages of osteoblasts differentiation. Insulin dissolved in the culture medium did not stimulate cell proliferation and its osteogenic activity was significantly lower. Addition of recombinant human bone morphogenetic protein 2 (rhBMP-2) to the culture medium further increased ALP activity in hMSCs indicating additive osteogenic action of immobilized insulin and rhBMP-2

  9. Osteoinductive activity of insulin-functionalized cell culture surfaces obtained using diazonium chemistry

    Directory of Open Access Journals (Sweden)

    Anna eMikulska

    2015-01-01

    Full Text Available Polymeric surfaces suitable for cell culture (DR/Pec were constructed from diazoresin (DR and pectin (Pec in a form of ultrathin films using the layer-by-layer (LbL technique. The surfaces were functionalized with insulin using diazonium chemistry. Such functionalized surfaces were used to culture human mesenchymal stem cells (hMSCs to assess their suitability for bone tissue engineering and regeneration. The activity of insulin immobilized on the surfaces (DR/Pec/Ins was compared to that of insulin dissolved in the culture medium. Human MSC grown on insulin-immobilized DR/Pec surfaces displayed increased proliferation and higher osteogenic activity. The latter was determined by means of alkaline phosphatase (ALP activity, which increases at early stages of osteoblasts differentiation. Insulin dissolved in the culture medium did not stimulate cell proliferation and its osteogenic activity was significantly lower. Addition of recombinant human bone morphogenetic protein 2 (rhBMP-2 to the culture medium further increased ALP activity in hMSCs indicating additive osteogenic action of immobilized insulin and rhBMP-2

  10. Insulin like growth factor 2 regulation of aryl hydrocarbon receptor in MCF-7 breast cancer cells

    International Nuclear Information System (INIS)

    Tomblin, Justin K.; Salisbury, Travis B.

    2014-01-01

    Highlights: •IGF-2 stimulates concurrent increases in AHR and CCND1 expression. •IGF-2 promotes the binding of AHR to the endogenous cyclin D1 promoter. •AHR knockdown inhibits IGF-2 stimulated increases in CCND1 mRNA and protein. •AHR knockdown inhibits IGF-2 stimulated increases in MCF-7 proliferation. -- Abstract: Insulin like growth factor (IGF)-1 and IGF-2 stimulate normal growth, development and breast cancer cell proliferation. Cyclin D1 (CCND1) promotes cell cycle by inhibiting retinoblastoma protein (RB1). The aryl hydrocarbon receptor (AHR) is a major xenobiotic receptor that also regulates cell cycle. The purpose of this study was to investigate whether IGF-2 promotes MCF-7 breast cancer proliferation by inducing AHR. Western blot and quantitative real time PCR (Q-PCR) analysis revealed that IGF-2 induced an approximately 2-fold increase (P < .001) in the expression of AHR and CCND1. Chromatin immunoprecipitation (ChIP), followed by Q-PCR indicated that IGF-2 promoted (P < .001) a 7-fold increase in AHR binding on the CCND1 promoter. AHR knockdown significantly (P < .001) inhibited IGF-2 stimulated increases in CCND1 mRNA and protein. AHR knockdown cells were less (P < .001) responsive to the proliferative effects of IGF-2 than control cells. Collectively, our findings have revealed a new regulatory mechanism by which IGF-2 induction of AHR promotes the expression of CCND1 and the proliferation of MCF-7 cells. This previously uncharacterized pathway could be important for the proliferation of IGF responsive cancer cells that also express AHR

  11. Insulin like growth factor 2 regulation of aryl hydrocarbon receptor in MCF-7 breast cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Tomblin, Justin K.; Salisbury, Travis B., E-mail: salisburyt@marshall.edu

    2014-01-17

    Highlights: •IGF-2 stimulates concurrent increases in AHR and CCND1 expression. •IGF-2 promotes the binding of AHR to the endogenous cyclin D1 promoter. •AHR knockdown inhibits IGF-2 stimulated increases in CCND1 mRNA and protein. •AHR knockdown inhibits IGF-2 stimulated increases in MCF-7 proliferation. -- Abstract: Insulin like growth factor (IGF)-1 and IGF-2 stimulate normal growth, development and breast cancer cell proliferation. Cyclin D1 (CCND1) promotes cell cycle by inhibiting retinoblastoma protein (RB1). The aryl hydrocarbon receptor (AHR) is a major xenobiotic receptor that also regulates cell cycle. The purpose of this study was to investigate whether IGF-2 promotes MCF-7 breast cancer proliferation by inducing AHR. Western blot and quantitative real time PCR (Q-PCR) analysis revealed that IGF-2 induced an approximately 2-fold increase (P < .001) in the expression of AHR and CCND1. Chromatin immunoprecipitation (ChIP), followed by Q-PCR indicated that IGF-2 promoted (P < .001) a 7-fold increase in AHR binding on the CCND1 promoter. AHR knockdown significantly (P < .001) inhibited IGF-2 stimulated increases in CCND1 mRNA and protein. AHR knockdown cells were less (P < .001) responsive to the proliferative effects of IGF-2 than control cells. Collectively, our findings have revealed a new regulatory mechanism by which IGF-2 induction of AHR promotes the expression of CCND1 and the proliferation of MCF-7 cells. This previously uncharacterized pathway could be important for the proliferation of IGF responsive cancer cells that also express AHR.

  12. Increased response to insulin of glucose metabolism in the 6-day unloaded rat soleus muscle

    Science.gov (United States)

    Henriksen, Erik J.; Tischler, Marc E.; Johnson, David G.

    1986-01-01

    Hind leg muscles of female rats were unloaded by tail cast suspension for 6 days. In the fresh-frozen unloaded soleus, the significantly greater concentration of glycogen correlated with a lower activity ratio of glycogen phosphorylase (p less than 0.02). The activity ratio of glycogen synthase also was lower (p less than 0.001), possibly due to the higher concentration of glycogen. In isolated unloaded soleus, insulin (0.1 milliunit/ml) increased the oxidation of D(U-C-14) glucose, release of lactate and pyruvate, incorporation of D-(U-C-14) glucose into glycogen, and the concentration of glucose 6-phosphate more (p less than 0.05) than in the weight-bearing soleus. At physiological doses of insulin, the percent of maximal uptake of 2-deoxy-D-(1,2-H-3) glucose/muscle also was greater in the unloaded soleus. Unloading of the soleus increased, by 50 percent the concentration of insuling receptors, due to no decrease in total receptor number during muscle atrophy. This increase may account for the greater response of glucose metabolism to insulin in this muscle. The extensor digitorum longus, which generally shows little response to unloading, displayed no differential response of glucose metabolism to insulin.

  13. Tissue-specific methylation of human insulin gene and PCR assay for monitoring beta cell death.

    Directory of Open Access Journals (Sweden)

    Mohamed I Husseiny

    Full Text Available The onset of metabolic dysregulation in type 1 diabetes (T1D occurs after autoimmune destruction of the majority of pancreatic insulin-producing beta cells. We previously demonstrated that the DNA encoding the insulin gene is uniquely unmethylated in these cells and then developed a methylation-specific PCR (MSP assay to identify circulating beta cell DNA in streptozotocin-treated mice prior to the rise in blood glucose. The current study extends to autoimmune non-obese diabetic (NOD mice and humans, showing in NOD mice that beta cell death occurs six weeks before the rise in blood sugar and coincides with the onset of islet infiltration by immune cells, demonstrating the utility of MSP for monitoring T1D. We previously reported unique patterns of methylation of the human insulin gene, and now extend this to other human tissues. The methylation patterns of the human insulin promoter, intron 1, exon 2, and intron 2 were determined in several normal human tissues. Similar to our previous report, the human insulin promoter was unmethylated in beta cells, but methylated in all other tissues tested. In contrast, intron 1, exon 2 and intron 2 did not exhibit any tissue-specific DNA methylation pattern. Subsequently, a human MSP assay was developed based on the methylation pattern of the insulin promoter and human islet DNA was successfully detected in circulation of T1D patients after islet transplantation therapy. Signal levels of normal controls and pre-transplant samples were shown to be similar, but increased dramatically after islet transplantation. In plasma the signal declines with time but in whole blood remains elevated for at least two weeks, indicating that association of beta cell DNA with blood cells prolongs the signal. This assay provides an effective method to monitor beta cell destruction in early T1D and in islet transplantation therapy.

  14. β cell membrane remodelling and procoagulant events occur in inflammation-driven insulin impairment: a GLP-1 receptor dependent and independent control.

    Science.gov (United States)

    Gleizes, Céline; Kreutter, Guillaume; Abbas, Malak; Kassem, Mohamad; Constantinescu, Andrei Alexandru; Boisramé-Helms, Julie; Yver, Blandine; Toti, Florence; Kessler, Laurence

    2016-02-01

    Inflammation and hyperglycaemia are associated with a prothrombotic state. Cell-derived microparticles (MPs) are the conveyors of active procoagulant tissue factor (TF) and circulate at high concentration in diabetic patients. Liraglutide, a glucagon-like peptide (GLP)-1 analogue, is known to promote insulin secretion and β-cell preservation. In this in vitro study, we examined the link between insulin impairment, procoagulant activity and plasma membrane remodelling, under inflammatory conditions. Rin-m5f β-cell function, TF activity mediated by MPs and their modulation by 1 μM liraglutide were examined in a cell cross-talk model. Methyl-β-cyclodextrine (MCD), a cholesterol depletor, was used to evaluate the involvement of raft on TF activity, MP shedding and insulin secretion as well as Soluble N-éthylmaleimide-sensitive-factor Attachment protein Receptor (SNARE)-dependent exocytosis. Cytokines induced a two-fold increase in TF activity at MP surface that was counteracted by liraglutide. Microparticles prompted TF activity on the target cells and a two-fold decrease in insulin secretion via protein kinase A (PKA) and p38 signalling, that was also abolished by liraglutide. Large lipid raft clusters were formed in response to cytokines and liraglutide or MCD-treated cells showed similar patterns. Cells pre-treated by saturating concentration of the GLP-1r antagonist exendin (9-39), showed a partial abolishment of the liraglutide-driven insulin secretion and liraglutide-decreased TF activity. Measurement of caspase 3 cleavage and MP shedding confirmed the contribution of GLP-1r-dependent and -independent pathways. Our results confirm an integrative β-cell response to GLP-1 that targets receptor-mediated signalling and membrane remodelling pointing at the coupling of insulin secretion and inflammation-driven procoagulant events. © 2015 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and

  15. The Prohormone VGF Regulates β Cell Function via Insulin Secretory Granule Biogenesis

    Directory of Open Access Journals (Sweden)

    Samuel B. Stephens

    2017-09-01

    Full Text Available The prohormone VGF is expressed in neuroendocrine and endocrine tissues and regulates nutrient and energy status both centrally and peripherally. We and others have shown that VGF-derived peptides have direct action on the islet β cell as secretagogues and cytoprotective agents; however, the endogenous function of VGF in the β cell has not been described. Here, we demonstrate that VGF regulates secretory granule formation. VGF loss-of-function studies in both isolated islets and conditional knockout mice reveal a profound decrease in stimulus-coupled insulin secretion. Moreover, VGF is necessary to facilitate efficient exit of granule cargo from the trans-Golgi network and proinsulin processing. It also functions to replenish insulin granule stores following nutrient stimulation. Our data support a model in which VGF operates at a critical node of granule biogenesis in the islet β cell to coordinate insulin biosynthesis with β cell secretory capacity.

  16. The islet beta-cell: fuel responsive and vulnerable.

    Science.gov (United States)

    Nolan, Christopher J; Prentki, Marc

    2008-10-01

    The pancreatic beta-cell senses blood nutrient levels and is modulated by neurohormonal signals so that it secretes insulin according to the need of the organism. Nutrient sensing involves marked metabolic activation, resulting in the production of coupling signals that promote insulin biosynthesis and secretion. The beta-cell's high capacity for nutrient sensing, however, necessitates reduced protection to nutrient toxicity. This potentially explains why in susceptible individuals, chronic fuel surfeit results in beta-cell failure and type 2 diabetes. Here we discuss recent insights into first, the biochemical basis of beta-cell signaling in response to glucose, amino acids and fatty acids, and second, beta-cell nutrient detoxification. We emphasize the emerging role of glycerolipid/fatty acid cycling in these processes.

  17. Urea impairs β cell glycolysis and insulin secretion in chronic kidney disease

    Science.gov (United States)

    Koppe, Laetitia; Nyam, Elsa; Vivot, Kevin; Manning Fox, Jocelyn E.; Dai, Xiao-Qing; Nguyen, Bich N.; Attané, Camille; Moullé, Valentine S.; MacDonald, Patrick E.; Ghislain, Julien

    2016-01-01

    Disorders of glucose homeostasis are common in chronic kidney disease (CKD) and are associated with increased mortality, but the mechanisms of impaired insulin secretion in this disease remain unclear. Here, we tested the hypothesis that defective insulin secretion in CKD is caused by a direct effect of urea on pancreatic β cells. In a murine model in which CKD is induced by 5/6 nephrectomy (CKD mice), we observed defects in glucose-stimulated insulin secretion in vivo and in isolated islets. Similarly, insulin secretion was impaired in normal mouse and human islets that were cultured with disease-relevant concentrations of urea and in islets from normal mice treated orally with urea for 3 weeks. In CKD mouse islets as well as urea-exposed normal islets, we observed an increase in oxidative stress and protein O-GlcNAcylation. Protein O-GlcNAcylation was also observed in pancreatic sections from CKD patients. Impairment of insulin secretion in both CKD mouse and urea-exposed islets was associated with reduced glucose utilization and activity of phosphofructokinase 1 (PFK-1), which could be reversed by inhibiting O-GlcNAcylation. Inhibition of O-GlcNAcylation also restored insulin secretion in both mouse models. These results suggest that insulin secretory defects associated with CKD arise from elevated circulating levels of urea that increase islet protein O-GlcNAcylation and impair glycolysis. PMID:27525435

  18. Phenolic excipients of insulin formulations induce cell death, pro-inflammatory signaling and MCP-1 release

    Directory of Open Access Journals (Sweden)

    Claudia Weber

    2015-01-01

    Insulin solutions displayed cytotoxic and pro-inflammatory potential caused by phenol or m-cresol. We speculate that during insulin pump therapy phenol and m-cresol might induce cell death and inflammatory reactions at the infusion site in vivo. Inflammation is perpetuated by release of MCP-1 by activated monocytic cells leading to enhanced recruitment of inflammatory cells. To minimize acute skin complications caused by phenol/m-cresol accumulation, a frequent change of infusion sets and rotation of the infusion site is recommended.

  19. Differentiation of Human Mesenchymal Stem Cells into Insulin Producing Cells by Using A Lentiviral Vector Carrying PDX1.

    Science.gov (United States)

    Allahverdi, Amir; Abroun, Saied; Jafarian, Arefeh; Soleimani, Masoud; Taghikhani, Mohammad; Eskandari, Fatemeh

    2015-01-01

    Type I diabetes is an immunologically-mediated devastation of insulin producing cells (IPCs) in the pancreatic islet. Stem cells that produce β-cells are a new promising tool. Adult stem cells such as mesenchymal stem cells (MSCs) are self renewing multi potent cells showing capabilities to differentiate into ectodermal, mesodermal and endodermal tissues. Pancreatic and duodenal homeobox factor 1 (PDX1) is a master regulator gene required for embryonic development of the pancreas and is crucial for normal pancreatic islets activities in adults. We induced the over-expression of the PDX1 gene in human bone marrow MSCs (BM-MSCs) by Lenti-PDX1 in order to generate IPCs. Next, we examine the ability of the cells by measuring insulin/c-peptide production and INSULIN and PDX1 gene expressions. After transduction, MSCs changed their morphology at day 5 and gradually differentiated into IPCs. INSULIN and PDX1 expressions were confirmed by real time polymerase chain reaction (RT-PCR) and immunostaining. IPC secreted insulin and C-peptide in the media that contained different glucose concentrations. MSCs differentiated into IPCs by genetic manipulation. Our result showed that lentiviral vectors could deliver PDX1 gene to MSCs and induce pancreatic differentiation.

  20. Identification of a growth hormone-responsive STAT5-binding element in the rat insulin 1 gene

    DEFF Research Database (Denmark)

    Galsgaard, E D; Gouilleux, F; Groner, B

    1996-01-01

    promoter activity 2-fold, and this stimulation was abolished by introduction of a block mutation in a gamma-interferon-activated sequence (GAS)-like element (GLE) with the sequence 5'-TTCTGGGAA-3' located in the rat insulin 1 enhancer at position -330 to -322. This element, termed Ins-GLE, was able...... transfected with STAT5 and GH receptor cDNAs, it was found that expression of STAT5 was necessary for GH induction of these two DNA-binding complexes. These results suggest that GH stimulates insulin 1 promoter activity by inducing the binding of STAT5 to Ins-GLE.......GH and PRL stimulate both proliferation and insulin production in pancreatic beta-cells as well as in the rat insulinoma cell line RIN-5AH, We report here that human GH increases insulin mRNA levels in RIN-5AH cells via both somatogenic and lactogenic receptors. GH stimulated the rat insulin 1...

  1. DEFECTS IN INSULIN-SECRETION IN NIDDM - B-CELL GLUCOSE INSENSITIVITY OR GLUCOSE TOXICITY

    NARCIS (Netherlands)

    VANHAEFTEN, TW

    In NIDDM, first-phase insulin release to glucose is (almost) absent. However, in contrast to older studies which suggested that in NIDDM the B-cell is ''blind'' for glucose, recent evidence indicates that the B-cell is not insensitive for glucose as far as second phase release is concerned. This

  2. Efficient and simple production of insulin-producing cells from embryonal carcinoma stem cells using mouse neonate pancreas extract, as a natural inducer.

    Directory of Open Access Journals (Sweden)

    Marzieh Ebrahimie

    Full Text Available An attractive approach to replace the destroyed insulin-producing cells (IPCs is the generation of functional β cells from stem cells. Embryonal carcinoma (EC stem cells are pluripotent cells which can differentiate into all cell types. The present study was carried out to establish a simple nonselective inductive culture system for generation of IPCs from P19 EC cells by 1-2 weeks old mouse pancreas extract (MPE. Since, mouse pancreatic islets undergo further remodeling and maturation for 2-3 weeks after birth, we hypothesized that the mouse neonatal MPE contains essential factors to induce in vitro differentiation of pancreatic lineages. Pluripotency of P19 cells were first confirmed by expression analysis of stem cell markers, Oct3/4, Sox-2 and Nanog. In order to induce differentiation, the cells were cultured in a medium supplemented by different concentrations of MPE (50, 100, 200 and 300 µg/ml. The results showed that P19 cells could differentiate into IPCs and form dithizone-positive cell clusters. The generated P19-derived IPCs were immunoreactive to proinsulin, insulin and insulin receptor beta. The expression of pancreatic β cell genes including, PDX-1, INS1 and INS2 were also confirmed. The peak response at the 100 µg/ml MPE used for investigation of EP300 and CREB1 gene expression. When stimulated with glucose, these cells synthesized and secreted insulin. Network analysis of the key transcription factors (PDX-1, EP300, CREB1 during the generation of IPCs resulted in introduction of novel regulatory candidates such as MIR17, and VEZF1 transcription factors, as well as MORN1, DKFZp761P0212, and WAC proteins. Altogether, we demonstrated the possibility of generating IPCs from undifferentiated EC cells, with the characteristics of pancreatic β cells. The derivation of pancreatic cells from EC cells which are ES cell siblings would provide a valuable experimental tool in study of pancreatic development and function as well as rapid

  3. Relationship between red cell membrane fatty acids and adipokines in individuals with varying insulin sensitivity.

    Science.gov (United States)

    Min, Y; Lowy, C; Islam, S; Khan, F S; Swaminathan, R

    2011-06-01

    Plasma leptin and adiponectin, and membrane phospholipid fatty acid composition are implicated into the mechanism of insulin resistance but no clear pattern has emerged. Hence, this study examined these variables in subjects presenting to the diabetic clinic for a diagnostic glucose tolerance test. Body composition, glucose, glycated hemoglobin, insulin, leptin, adiponectin, and red cell and plasma phospholipid fatty acids were assessed from 42 normal and 28 impaired glucose tolerant subjects. Insulin sensitivity was determined by homeostatic model assessment. The plasma phosphatidylcholine fatty acid composition of the impaired glucose tolerant subjects was similar to that of normal subjects. However, the impaired glucose tolerant subjects had significantly lower linoleic (Pphosphatidylcholine and phosphatidylethanolamine compared with the normal subjects. Moreover, red cell phosphatidylcholine docosahexaenoic acid correlated positively with adiponectin (r=0.290, Pinsulin (r=-0.335, Pinsulin resistance (r=-0.322, Pinsulin level whereas insulin was the only component that predicted the membrane fatty acids. We postulate that membrane phospholipids fatty acids have an indirect role in determining insulin concentration but insulin has a major role in determining membrane fatty acid composition.

  4. Insulin is essential for in vitro chondrogenesis of mesenchymal progenitor cells and influences chondrogenesis in a dose-dependent manner.

    Science.gov (United States)

    Mueller, Michael B; Blunk, Torsten; Appel, Bernhard; Maschke, Angelika; Goepferich, Achim; Zellner, Johannes; Englert, Carsten; Prantl, Lukas; Kujat, Richard; Nerlich, Michael; Angele, Peter

    2013-01-01

    Insulin is a commonly used additive in chondrogenic media for differentiating mesenchymal stem cells (MSCs). The indispensability of other bioactive factors like TGF-β or dexamethasone in these medium formulations has been shown, but the role of insulin is unclear. The purpose of this study was to investigate whether insulin is essential for MSC chondrogenesis and if there is a dose-dependent effect of insulin on MSC chondrogenesis. We cultivated human MSCs in pellet culture in serum-free chondrogenic medium with insulin concentrations between 0 and 50 μg/ml and assessed the grade of chondrogenic differentiation by histological evaluation and determination of glycosaminoglycan (GAG), total collagen and DNA content. We further tested whether insulin can be delivered in an amount sufficient for MSC chondrogenesis via a drug delivery system in insulin-free medium. Chondrogenesis was not induced by standard chondrogenic medium without insulin and the expression of cartilage differentiation markers was dose-dependent at insulin concentrations between 0 and 10 μg/ml. An insulin concentration of 50 μg/ml had no additional effect compared with 10 μg/ml. Insulin was delivered by a release system into the cell culture under insulin-free conditions in an amount sufficient to induce chondrogenesis. Insulin is essential for MSC chondrogenesis in this system and chondrogenic differentiation is influenced by insulin in a dose-dependent manner. Insulin can be provided in a sufficient amount by a drug delivery system. Therefore, insulin is a suitable and inexpensive indicator substance for testing drug release systems in vitro.

  5. Fibroblast growth factor regulates insulin-like growth factor-binding protein production by vascular smooth muscle cells.

    Science.gov (United States)

    Ververis, J; Ku, L; Delafontaine, P

    1994-02-01

    Insulin-like growth factor I is an important mitogen for vascular smooth muscle cells, and its effects are regulated by several binding proteins. Western ligand blotting of conditioned medium from rat aortic smooth muscle cells detected a 24 kDa binding protein and a 28 kDa glycosylated variant of this protein, consistent with insulin-like growth factor binding protein-4 by size. Low amounts of a glycosylated 38 to 42 kDa doublet (consistent with binding protein-3) and a 31 kDa non-glycosylated protein also were present. Basic fibroblast growth factor markedly increased secretion of the 24 kDa binding protein and its 28 kDa glycosylated variant. This effect was dose- and time-dependent and was inhibited by co-incubation with cycloheximide. Crosslinking of [125I]-insulin-like growth factor I to cell monolayers revealed no surface-associated binding proteins, either basally or after agonist treatment. Induction of binding protein production by fibroblast growth factor at sites of vascular injury may be important in vascular proliferative responses in vivo.

  6. Mechanisms of estradiol-induced insulin secretion by the G protein-coupled estrogen receptor GPR30/GPER in pancreatic beta-cells.

    Science.gov (United States)

    Sharma, Geetanjali; Prossnitz, Eric R

    2011-08-01

    Sexual dimorphism and supplementation studies suggest an important role for estrogens in the amelioration of glucose intolerance and diabetes. Because little is known regarding the signaling mechanisms involved in estradiol-mediated insulin secretion, we investigated the role of the G protein-coupled receptor 30, now designated G protein-coupled estrogen receptor (GPER), in activating signal transduction cascades in β-cells, leading to secretion of insulin. GPER function in estradiol-induced signaling in the pancreatic β-cell line MIN6 was assessed using small interfering RNA and GPER-selective ligands (G-1 and G15) and in islets isolated from wild-type and GPER knockout mice. GPER is expressed in MIN6 cells, where estradiol and the GPER-selective agonist G-1 mediate calcium mobilization and activation of ERK and phosphatidylinositol 3-kinase. Both estradiol and G-1 induced insulin secretion under low- and high-glucose conditions, which was inhibited by pretreatment with GPER antagonist G15 as well as depletion of GPER by small interfering RNA. Insulin secretion in response to estradiol and G-1 was dependent on epidermal growth factor receptor and ERK activation and further modulated by phosphatidylinositol 3-kinase activity. In islets isolated from wild-type mice, the GPER antagonist G15 inhibited insulin secretion induced by estradiol and G-1, both of which failed to induce insulin secretion in islets obtained from GPER knockout mice. Our results indicate that GPER activation of the epidermal growth factor receptor and ERK in response to estradiol treatment plays a critical role in the secretion of insulin from β-cells. The results of this study suggest that the activation of downstream signaling pathways by the GPER-selective ligand G-1 could represent a novel therapeutic strategy in the treatment of diabetes.

  7. Mechanisms of Estradiol-Induced Insulin Secretion by the G Protein-Coupled Estrogen Receptor GPR30/GPER in Pancreatic β-Cells

    Science.gov (United States)

    Sharma, Geetanjali

    2011-01-01

    Sexual dimorphism and supplementation studies suggest an important role for estrogens in the amelioration of glucose intolerance and diabetes. Because little is known regarding the signaling mechanisms involved in estradiol-mediated insulin secretion, we investigated the role of the G protein-coupled receptor 30, now designated G protein-coupled estrogen receptor (GPER), in activating signal transduction cascades in β-cells, leading to secretion of insulin. GPER function in estradiol-induced signaling in the pancreatic β-cell line MIN6 was assessed using small interfering RNA and GPER-selective ligands (G-1 and G15) and in islets isolated from wild-type and GPER knockout mice. GPER is expressed in MIN6 cells, where estradiol and the GPER-selective agonist G-1 mediate calcium mobilization and activation of ERK and phosphatidylinositol 3-kinase. Both estradiol and G-1 induced insulin secretion under low- and high-glucose conditions, which was inhibited by pretreatment with GPER antagonist G15 as well as depletion of GPER by small interfering RNA. Insulin secretion in response to estradiol and G-1 was dependent on epidermal growth factor receptor and ERK activation and further modulated by phosphatidylinositol 3-kinase activity. In islets isolated from wild-type mice, the GPER antagonist G15 inhibited insulin secretion induced by estradiol and G-1, both of which failed to induce insulin secretion in islets obtained from GPER knockout mice. Our results indicate that GPER activation of the epidermal growth factor receptor and ERK in response to estradiol treatment plays a critical role in the secretion of insulin from β-cells. The results of this study suggest that the activation of downstream signaling pathways by the GPER-selective ligand G-1 could represent a novel therapeutic strategy in the treatment of diabetes. PMID:21673097

  8. Glucose Induces Mouse β-Cell Proliferation via IRS2, MTOR, and Cyclin D2 but Not the Insulin Receptor

    Science.gov (United States)

    Stamateris, Rachel E.; Sharma, Rohit B.; Kong, Yahui; Ebrahimpour, Pantea; Panday, Deepika; Ranganath, Pavana; Zou, Baobo; Levitt, Helena; Parambil, Nisha Abraham; O’Donnell, Christopher P.; García-Ocaña, Adolfo

    2016-01-01

    An important goal in diabetes research is to understand the processes that trigger endogenous β-cell proliferation. Hyperglycemia induces β-cell replication, but the mechanism remains debated. A prime candidate is insulin, which acts locally through the insulin receptor. Having previously developed an in vivo mouse hyperglycemia model, we tested whether glucose induces β-cell proliferation through insulin signaling. By using mice lacking insulin signaling intermediate insulin receptor substrate 2 (IRS2), we confirmed that hyperglycemia-induced β-cell proliferation requires IRS2 both in vivo and ex vivo. Of note, insulin receptor activation was not required for glucose-induced proliferation, and insulin itself was not sufficient to drive replication. Glucose and insulin caused similar acute signaling in mouse islets, but chronic signaling differed markedly, with mammalian target of rapamycin (MTOR) and extracellular signal–related kinase (ERK) activation by glucose and AKT activation by insulin. MTOR but not ERK activation was required for glucose-induced proliferation. Cyclin D2 was necessary for glucose-induced β-cell proliferation. Cyclin D2 expression was reduced when either IRS2 or MTOR signaling was lost, and restoring cyclin D2 expression rescued the proliferation defect. Human islets shared many of these regulatory pathways. Taken together, these results support a model in which IRS2, MTOR, and cyclin D2, but not the insulin receptor, mediate glucose-induced proliferation. PMID:26740601

  9. Leucine metabolism in regulation of insulin secretion from pancreatic beta cells

    OpenAIRE

    Yang, Jichun; Chi, Yujing; Burkhardt, Brant R.; Guan, Youfei; Wolf, Bryan A

    2010-01-01

    Leucine, a the branched-chain amino acids that must be supplied in daily diet, plays an important role in controlling protein synthesis and regulating cell metabolism in various cell types. In pancreatic β cells, leucine acutely stimulates insulin secretion by serving as both metabolic fuel and allosteric activator of glutamate dehydrogenase to enhance glutaminolysis. Leucine has also been shown to regulate gene transcription and protein synthesis in pancreatic islet β cells via both mTOR-dep...

  10. Beta-cell lines derived from transgenic mice expressing a hybrid insulin gene-oncogene

    DEFF Research Database (Denmark)

    Efrat, S; Linde, S; Kofod, Hans

    1988-01-01

    Three pancreatic beta-cell lines have been established from insulinomas derived from transgenic mice carrying a hybrid insulin-promoted simian virus 40 tumor antigen gene. The beta tumor cell (beta TC) lines maintain the features of differentiated beta cells for about 50 passages in culture. The ...... both to immortalize a rare cell type and to provide a selection for the maintenance of its differentiated phenotype....

  11. Nuclear SREBP-1a causes loss of pancreatic β-cells and impaired insulin secretion

    International Nuclear Information System (INIS)

    Iwasaki, Yuko; Iwasaki, Hitoshi; Yatoh, Shigeru; Ishikawa, Mayumi; Kato, Toyonori; Matsuzaka, Takashi; Nakagawa, Yoshimi; Yahagi, Naoya; Kobayashi, Kazuto; Takahashi, Akimitsu; Suzuki, Hiroaki; Yamada, Nobuhiro; Shimano, Hitoshi

    2009-01-01

    Transgenic mice expressing nuclear sterol regulatory element-binding protein-1a under the control of the insulin promoter were generated to determine the role of SREBP-1a in pancreatic β-cells. Only low expressors could be established, which exhibited mild hyperglycemia, impaired glucose tolerance, and reduced plasma insulin levels compared to C57BL/6 controls. The islets isolated from the transgenic mice were fewer and smaller, and had decreased insulin content and unaltered glucagon staining. Both glucose- and potassium-stimulated insulin secretions were decreased. The transgenic islets consistently expressed genes for fatty acids and cholesterol synthesis, resulting in accumulation of triglycerides but not cholesterol. PDX-1, ΒΕΤΑ2, MafA, and IRS-2 were suppressed, partially explaining the loss and dysfunction of β-cell mass. The transgenic mice on a high fat/high sucrose diet still exhibited impaired insulin secretion and continuous β-cell growth defect. Therefore, nuclear SREBP-1a, even at a low level, strongly disrupts β-cell mass and function.

  12. Valsartan Improves β-Cell Function and Insulin Sensitivity in Subjects With Impaired Glucose Metabolism

    Science.gov (United States)

    van der Zijl, Nynke J.; Moors, Chantalle C.M.; Goossens, Gijs H.; Hermans, Marc M.H.; Blaak, Ellen E.; Diamant, Michaela

    2011-01-01

    OBJECTIVE Recently, the Nateglinide and Valsartan in Impaired Glucose Tolerance Outcomes Research Trial demonstrated that treatment with the angiotensin receptor blocker (ARB) valsartan for 5 years resulted in a relative reduction of 14% in the incidence of type 2 diabetes in subjects with impaired glucose metabolism (IGM). We investigated whether improvements in β-cell function and/or insulin sensitivity underlie these preventive effects of the ARB valsartan in the onset of type 2 diabetes. RESEARCH DESIGN AND METHODS In this randomized controlled, double-blind, two-center study, the effects of 26 weeks of valsartan (320 mg daily; n = 40) or placebo (n = 39) on β-cell function and insulin sensitivity were assessed in subjects with impaired fasting glucose and/or impaired glucose tolerance, using a combined hyperinsulinemic-euglycemic and hyperglycemic clamp with subsequent arginine stimulation and a 2-h 75-g oral glucose tolerance test (OGTT). Treatment effects were analyzed using ANCOVA, adjusting for center, glucometabolic status, and sex. RESULTS Valsartan increased first-phase (P = 0.028) and second-phase (P = 0.002) glucose-stimulated insulin secretion compared with placebo, whereas the enhanced arginine-stimulated insulin secretion was comparable between groups (P = 0.25). In addition, valsartan increased the OGTT-derived insulinogenic index (representing first-phase insulin secretion after an oral glucose load; P = 0.027). Clamp-derived insulin sensitivity was significantly increased with valsartan compared with placebo (P = 0.049). Valsartan treatment significantly decreased systolic and diastolic blood pressure compared with placebo (P valsartan treatment increased glucose-stimulated insulin release and insulin sensitivity in normotensive subjects with IGM. These findings may partly explain the beneficial effects of valsartan in the reduced incidence of type 2 diabetes. PMID:21330640

  13. Derivation of Insulin Producing Cells From Human Endometrial Stromal Stem Cells and Use in the Treatment of Murine Diabetes

    OpenAIRE

    Santamaria, Xavier; Massasa, Efi E; Feng, Yuzhe; Wolff, Erin; Taylor, Hugh S

    2011-01-01

    Pancreatic islet cell transplantation is an effective approach to treat type 1 diabetes, however the shortage of cadaveric donors and limitations due to rejection require alternative solutions. Multipotent cells derived from the uterine endometrium have the ability to differentiate into mesodermal and ectodermal cellular lineages, suggesting the existence of mesenchymal stem cells in this tissue. We differentiated human endometrial stromal stem cells (ESSC) into insulin secreting cells using ...

  14. Role of sialic acid in insulin action and the insulin resistance of diabetes mellitus

    International Nuclear Information System (INIS)

    Salhanick, A.I.; Amatruda, J.M.

    1988-01-01

    Adipocytes treated with neuraminidase show markedly reduced responsiveness to insulin without any alteration in insulin binding. In addition, several studies have separately demonstrated both insulin resistance and decreases in membrane sialic acid content and associated biosynthetic enzymes in diabetes mellitus. In the present study, the authors investigated the role that sialic acid residues may play in insulin action and in the hepatic insulin resistance associated with nonketotic diabetes. Primary cultures of hepatocytes from normal rats treated with neuraminidase demonstrated a dose-dependent decrease in insulin-stimulated lipogenesis. At a concentration of neuraminidase that decreases insulin action by 50%, 23% of total cellular sialic acid content was released. Neuraminidase-releasable sialic acid was significantly decreased in hepatocytes from diabetic rats and this was associated with significant insulin resistance. Treatment of hepatocytes from diabetic rats with cytidine 5'-monophospho-N-acetylneuraminic acid (CMP-NANA) enhanced insulin responsiveness 39%. The enhanced insulin responsiveness induced by CMP-NANA was blocked by cytidine 5'-monophosphate (CMP) suggesting that the CMP-NANA effect was catalyzed by a cell surface sialyl-transferase. CMP reduced neuraminidase-releasable [ 14 C]sialic acid incorporation into hepatocytes by 43%. The data demonstrate a role for cell surface sialic acid residues in hepatic insulin action and support a role for decreased cell surface sialic acid residues in the insulin resistance of diabetes mellitus

  15. Novel nuclear localization and potential function of insulin-like growth factor-1 receptor/insulin receptor hybrid in corneal epithelial cells.

    Directory of Open Access Journals (Sweden)

    Yu-Chieh Wu

    Full Text Available BACKGROUND: Type I insulin-like growth factor receptor (IGF-1R and insulin receptor (INSR are highly homologous molecules, which can heterodimerize to form an IGF-1R/INSR hybrid (Hybrid-R. The presence and biological significance of the Hybrid-R in human corneal epithelium has not yet been established. In addition, while nuclear localization of IGF-1R was recently reported in cancer cells and human corneal epithelial cells, the function and profile of nuclear IGF-1R is unknown. In this study, we characterized the nuclear localization and function of the Hybrid-R and the role of IGF-1/IGF-1R and Hybrid-R signaling in the human corneal epithelium. METHODOLOGY/PRINCIPLE FINDINGS: IGF-1-mediated signaling and cell growth were examined in a human telomerized corneal epithelial (hTCEpi cell line using co-immunoprecipitation, immunoblotting and cell proliferation assays. The presence of Hybrid-R in hTCEpi and primary cultured human corneal epithelial cells was confirmed by immunofluorescence and reciprocal immunoprecipitation of whole cell lysates. We found that IGF-1 stimulated Akt and promoted cell growth through IGF-1R activation, which was independent of the Hybrid-R. The presence of Hybrid-R, but not IGF-1R/IGF-1R, was detected in nuclear extracts. Knockdown of INSR by small interfering RNA resulted in depletion of the INSR/INSR and preferential formation of Hybrid-R. Chromatin-immunoprecipitation sequencing assay with anti-IGF-1R or anti-INSR was subsequently performed to identify potential genomic targets responsible for critical homeostatic regulatory pathways. CONCLUSION/SIGNIFICANCE: In contrast to previous reports on nuclear localized IGF-1R, this is the first report identifying the nuclear localization of Hybrid-R in an epithelial cell line. The identification of a nuclear Hybrid-R and novel genomic targets suggests that IGF-1R traffics to the nucleus as an IGF-1R/INSR heterotetrameric complex to regulate corneal epithelial homeostatic

  16. Targeting non-small cell lung cancer cells by dual inhibition of the insulin receptor and the insulin-like growth factor-1 receptor.

    Directory of Open Access Journals (Sweden)

    Emma E Vincent

    Full Text Available Phase III trials of the anti-insulin-like growth factor-1 receptor (IGF1R antibody figitumumab in non-small cell lung cancer (NSCLC patients have been discontinued owing to lack of survival benefit. We investigated whether inhibition of the highly homologous insulin receptor (IR in addition to the IGF1R would be more effective than inhibition of the IGF1R alone at preventing the proliferation of NSCLC cells. Signalling through IGF1R and IR in the NSCLC cell lines A549 and Hcc193 was stimulated by a combination of IGF1, IGF2 and insulin. It was inhibited by antibodies that block ligand binding, αIR3 (IGF1R and IR47-9 (IR, and by the ATP-competitive small molecule tyrosine kinase inhibitors AZ12253801 and NVPAWD742 which inhibit both IGF1R and IR tyrosine kinases. The effect of inhibitors was determined by an anchorage-independent proliferation assay and by analysis of Akt phosphorylation. In Hcc193 cells the reduction in cell proliferation and Akt phosphorylation due to anti-IGF1R antibody was enhanced by antibody-mediated inhibition of the IR whereas in A549 cells, with a relatively low IR:IGF1R expression ratio, it was not. In each cell line proliferation and Akt phosphorylation were more effectively inhibited by AZ12253801 and NVPAWD742 than by combined αIR3 and IR47-9. When the IGF1R alone is inhibited, unencumbered signalling through the IR can contribute to continued NSCLC cell proliferation. We conclude that small molecule inhibitors targeting both the IR and IGF1R more effectively reduce NSCLC cell proliferation in a manner independent of the IR:IGF1R expression ratio, providing a therapeutic rationale for the treatment of this disease.

  17. Effects of the antitumor drug OSI-906, a dual inhibitor of IGF-1 receptor and insulin receptor, on the glycemic control, β-cell functions, and β-cell proliferation in male mice.

    Science.gov (United States)

    Shirakawa, Jun; Okuyama, Tomoko; Yoshida, Eiko; Shimizu, Mari; Horigome, Yuka; Tuno, Takayuki; Hayasaka, Moe; Abe, Shiori; Fuse, Masahiro; Togashi, Yu; Terauchi, Yasuo

    2014-06-01

    The IGF-1 receptor has become a therapeutic target for the treatment of cancer. The efficacy of OSI-906 (linstinib), a dual inhibitor of IGF-1 receptor and insulin receptor, for solid cancers has been examined in clinical trials. The effects of OSI-906, however, on the blood glucose levels and pancreatic β-cell functions have not yet been reported. We investigated the impact of OSI-906 on glycemic control, insulin secretion, β-cell mass, and β-cell proliferation in male mice. Oral administration of OSI-906 worsened glucose tolerance in a dose-dependent manner in the wild-type mice. OSI-906 at a dose equivalent to the clinical daily dose (7.5 mg/kg) transiently evoked glucose intolerance and hyperinsulinemia. Insulin receptor substrate (IRS)-2-deficient mice and mice with diet-induced obesity, both models of peripheral insulin resistance, exhibited more severe glucose intolerance after OSI-906 administration than glucokinase-haploinsufficient mice, a model of impaired insulin secretion. Phloridzin improved the hyperglycemia induced by OSI-906 in mice. In vitro, OSI-906 showed no effect on insulin secretion from isolated islets. After daily administration of OSI-906 for a week to mice, the β-cell mass and β-cell proliferation rate were significantly increased. The insulin signals in the β-cells were apparently unaffected in those mice. Taken together, the results suggest that OSI-906 could exacerbate diabetes, especially in patients with insulin resistance. On the other hand, the results suggest that the β-cell mass may expand in response to chemotherapy with this drug.

  18. A novel dual-color reporter for identifying insulin-producing beta-cells and classifying heterogeneity of insulinoma cell lines.

    Directory of Open Access Journals (Sweden)

    Nan Sook Lee

    Full Text Available Many research studies use immortalized cell lines as surrogates for primary beta- cells. We describe the production and use of a novel "indirect" dual-fluorescent reporter system that leads to mutually exclusive expression of EGFP in insulin-producing (INS(+ beta-cells or mCherry in non-beta-cells. Our system uses the human insulin promoter to initiate a Cre-mediated shift in reporter color within a single transgene construct and is useful for FACS selection of cells from single cultures for further analysis. Application of our reporter to presumably clonal HIT-T15 insulinoma cells, as well as other presumably clonal lines, indicates that these cultures are in fact heterogeneous with respect to INS(+ phenotype. Our strategy could be easily applied to other cell- or tissue-specific promoters. We anticipate its utility for FACS purification of INS(+ and glucose-responsive beta-like-cells from primary human islet cell isolates or in vitro differentiated pluripotent stem cells.

  19. Relationship between serum adiponectin concentration, body condition score, and peripheral tissue insulin response of dairy cows during the dry period.

    Science.gov (United States)

    De Koster, J; Urh, C; Hostens, M; Van den Broeck, W; Sauerwein, H; Opsomer, G

    2017-04-01

    The aim of the present study was to describe the relationship between serum adiponectin concentration and peripheral tissue insulin response in dairy cows with a variable body condition score (BCS) during the dry period. Cows were selected at the beginning of the dry period based on BCS (BCS 3.75, n = 5). Animals were followed from the beginning of the dry period by weekly blood sampling and assessment of BCS and backfat thickness. Weekly blood samples were analyzed for adiponectin concentration using a bovine specific ELISA. Hyperinsulinemic euglycemic clamp tests were performed at the end of the dry period to measure peripheral tissue insulin response. Insulin dose response curves were established for both glucose and fatty acid metabolism. Regression analysis revealed that the serum concentrations of adiponectin dropped at the end of the dry period (P insulin responsiveness (reflecting the maximal effect of insulin; r = 0.76, P insulin sensitivity (reflecting the insulin concentration needed to achieve halfmaximal effect; r = -0.54, P = 0.13). At the level of the fatty acid metabolism, greater adiponectin concentrations were negatively correlated with lower NEFA levels during the HEC test reflecting the insulin responsiveness of the NEFA metabolism (r = -0.61, P = 0.08), whereas there was no association with the insulin sensitivity of the NEFA metabolism (r = -0.16, P = 0.67). In conclusion, serum concentrations of adiponectin were negatively associated with the BCS of dairy cows during the dry period and positively associated with insulin responsiveness of the glucose and fatty acid metabolism. Copyright © 2016 Elsevier Inc. All rights reserved.

  20. The level of menadione redox-cycling in pancreatic β-cells is proportional to the glucose concentration: role of NADH and consequences for insulin secretion.

    Science.gov (United States)

    Heart, Emma; Palo, Meridith; Womack, Trayce; Smith, Peter J S; Gray, Joshua P

    2012-01-15

    Pancreatic β-cells release insulin in response to elevation of glucose from basal (4-7mM) to stimulatory (8-16mM) levels. Metabolism of glucose by the β-cell results in the production of low levels of reactive oxygen intermediates (ROI), such as hydrogen peroxide (H(2)O(2)), a newly recognized coupling factor linking glucose metabolism to insulin secretion. However, high and toxic levels of H(2)O(2) inhibit insulin secretion. Menadione, which produces H(2)O(2) via redox cycling mechanism in a dose-dependent manner, was investigated for its effect on β-cell metabolism and insulin secretion in INS-1 832/13, a rat β-cell insulinoma cell line, and primary rodent islets. Menadione-dependent redox cycling and resulting H(2)O(2) production under stimulatory glucose exceeded several-fold those reached at basal glucose. This was paralleled by a differential effect of menadione (0.1-10μM) on insulin secretion, which was enhanced at basal, but inhibited at stimulatory glucose. Redox cycling of menadione and H(2)O(2) formation was dependent on glycolytically-derived NADH, as inhibition of glycolysis and application of non-glycogenic insulin secretagogues did not support redox cycling. In addition, activity of plasma membrane electron transport, a system dependent in part on glycolytically-derived NADH, was also inhibited by menadione. Menadione-dependent redox cycling was sensitive to the NQO1 inhibitor dicoumarol and the flavoprotein inhibitor diphenylene iodonium, suggesting a role for NQO1 and other oxidoreductases in this process. These data may explain the apparent dichotomy between the stimulatory and inhibitory effects of H(2)O(2) and menadione on insulin secretion. Published by Elsevier Inc.

  1. NKT Cell Responses to B Cell Lymphoma.

    Science.gov (United States)

    Li, Junxin; Sun, Wenji; Subrahmanyam, Priyanka B; Page, Carly; Younger, Kenisha M; Tiper, Irina V; Frieman, Matthew; Kimball, Amy S; Webb, Tonya J

    2014-06-01

    Natural killer T (NKT) cells are a unique subset of CD1d-restricted T lymphocytes that express characteristics of both T cells and natural killer cells. NKT cells mediate tumor immune-surveillance; however, NKT cells are numerically reduced and functionally impaired in lymphoma patients. Many hematologic malignancies express CD1d molecules and co-stimulatory proteins needed to induce anti-tumor immunity by NKT cells, yet most tumors are poorly immunogenic. In this study, we sought to investigate NKT cell responses to B cell lymphoma. In the presence of exogenous antigen, both mouse and human NKT cell lines produce cytokines following stimulation by B cell lymphoma lines. NKT cell populations were examined ex vivo in mouse models of spontaneous B cell lymphoma, and it was found that during early stages, NKT cell responses were enhanced in lymphoma-bearing animals compared to disease-free animals. In contrast, in lymphoma-bearing animals with splenomegaly and lymphadenopathy, NKT cells were functionally impaired. In a mouse model of blastoid variant mantle cell lymphoma, treatment of tumor-bearing mice with a potent NKT cell agonist, α-galactosylceramide (α-GalCer), resulted in a significant decrease in disease pathology. Ex vivo studies demonstrated that NKT cells from α-GalCer treated mice produced IFN-γ following α-GalCer restimulation, unlike NKT cells from vehicle-control treated mice. These data demonstrate an important role for NKT cells in the immune response to an aggressive hematologic malignancy like mantle cell lymphoma.

  2. Insulin-induced inhibition of gluconeogenesis genes, including glutamic pyruvic transaminase 2, is associated with reduced histone acetylation in a human liver cell line.

    Science.gov (United States)

    Honma, Kazue; Kamikubo, Michiko; Mochizuki, Kazuki; Goda, Toshinao

    2017-06-01

    Hepatic glutamic pyruvic transaminase (GPT; also known as alanine aminotransferase) is a gluconeogenesis enzyme that catalyzes conversions between alanine and pyruvic acid. It is also used as a blood biomarker for hepatic damage. In this study, we investigated whether insulin regulates GPT expression, as it does for other gluconeogenesis genes, and if this involves the epigenetic modification of histone acetylation. Human liver-derived HepG2 cells were cultured with 0.5-100nM insulin for 8h, and the mRNA expression of GPT, glutamic-oxaloacetic transaminase (GOT), γ-glutamyltransferase (GGT), PCK1, G6PC and FBP1 was measured. We also investigated the extent of histone acetylation around these genes. Insulin suppressed the mRNA expression of gluconeogenesis genes (GPT2, GOT1, GOT2, GGT1, GGT2, G6PC, and PCK1) in HepG2 cells in a dose-dependent manner. mRNA levels of GPT2, but not GPT1, were decreased by insulin. Histone acetylation was also reduced around GPT2, G6PC, and PCK1 in response to insulin. The expression of GPT2 and other gluconeogenesis genes such as G6PC and PCK1 was suppressed by insulin, in association with decreases in histone H3 and H4 acetylation surrounding these genes. Copyright © 2017 Elsevier Inc. All rights reserved.

  3. Mesenchymal Stem Cell Differentiation into Adipocytes Is Equally Induced by Insulin and Proinsulin In Vitro.

    Science.gov (United States)

    Pfützner, Andreas; Schipper, Dorothee; Pansky, Andreas; Kleinfeld, Claudia; Roitzheim, Barbara; Tobiasch, Edda

    2017-11-30

    In advanced β -cell dysfunction, proinsulin is increasingly replacing insulin as major component of the secretion product. It has been speculated that proinsulin has at least the same adipogenic potency than insulin, leading to an increased tendency of lipid tissue formation in patients with late stage β -cell dysfunction. Mesenchymal stem cells obtained from liposuction material were grown in differentiation media containing insulin (0.01 μmol), proinsulin (0.01 μmol) or insulin+proinsulin (each 0.005 μmol). Cell culture supernatants were taken from these experiments and an untreated control at weeks 1, 2, and 3, and were stored at -80°C until analysis. Cell differentiation was microscopically supervised and adiponectin concentrations were measured as marker for differentiation into mature lipid cells. This experiment was repeated three times. No growth of lipid cells and no change in adiponectin values was observed in the negative control group (after 7/14/12 days: 3.2±0.5/3.3±0.1/4.4±0.5 ng/ml/12 h). A continuous differentiation into mature adipocytes (also confirmed by Red-Oil-staining) and a corresponding increase in adiponectin values was observed in the experiments with insulin (3.6±1.9/5.1±1.4/13.3±1.5 ng/ml/12 h; p<0.05 week 1 vs. week 3) and proinsulin (3.3±1.2/3.5±0.3/12.2±1.2 ng/ml/12 h; p<0.05). Comparable effects were seen with the insulin/proinsulin combination. Proinsulin has the same adipogenic potential than insulin in vitro. Proinsulin has only 10∼20% of the glucose-lowering effect of insulin. It can be speculated that the adipogenic potential of proinsulin may be a large contributor to the increased body weight problems in patients with type 2 diabetes and advanced β -cell dysfunction.

  4. Radiation block of bone marrow cell mitoses and the effect of insulin

    Energy Technology Data Exchange (ETDEWEB)

    Barkalaya, A I

    1976-01-01

    Insulin (0.15 - 0.2 units/kg) has been administered to white rats immediately after the exposure to 750 R, at the background of hypercorticoidism. This resulted in the inhibition of the development of the post-irradiation-stress-hyperglycemia; and the mitotic index of the bone marrow cells at the time of the mitosis block was higher than in the control irradiated rats. Insulin administration at the peak of radiation sickness during hypercorticoidism levelled hyperglycemia, stimulated the mitotic activity of cells of the bone marrow and the regeneration of the latter.

  5. Superior Glycemic Control with a Glucose-Responsive Insulin Analog: Hepatic and Nonhepatic Impacts.

    Science.gov (United States)

    Moore, Mary Courtney; Kelley, David E; Camacho, Raul C; Zafian, Peter; Ye, Tian; Lin, Songnian; Kaarsholm, Niels C; Nargund, Ravi; Kelly, Terri M; Van Heek, Margaret; Previs, Stephen F; Moyes, Christopher; Smith, Marta S; Farmer, Ben; Williams, Phil; Cherrington, Alan D

    2018-03-14

    We evaluated the hepatic and nonhepatic responses to glucose-responsive insulin (GRI). Eight dogs received GRI or regular human insulin (HI) in random order. A primed, continuous intravenous infusion of [3- 3 H]glucose began at -120 min. Basal sampling (-30 to 0 min) was followed by 2 study periods (150 min each), P1 and P2. At 0 min, somatostatin and GRI (36±3 pmol/kg/min) or HI (1.8 pmol/kg/min) were infused IV; basal glucagon was replaced intraportally. Glucose was infused intravenously to clamp plasma glucose at 80 mg/dL (P1) and 240 mg/dL (P2). Whole body insulin clearance (WBIC) and insulin concentrations were not different in P1 vs P2 with HI, but WBIC was 23% higher and arterial insulin 16% lower in P1 vs P2 with GRI. Net hepatic glucose output was similar between treatments in P1. In P2, both treatments induced net hepatic glucose uptake (2.1±0.5 [HI] vs 3.3±0.4 [GRI] mg/kg/min). Nonhepatic glucose uptake (nonHGU, mg/kg/min) in P1 and P2, respectively, differed between treatments (2.6±0.3 and 7.4±0.6 with HI; 2.0±0.2 and 8.1±0.8 with GRI). Thus, glycemia impacted GRI but not HI clearance, with resultant differential effects on HGU and nonHGU. GRI holds promise for decreasing hypoglycemia risk while enhancing glucose uptake under hyperglycemic conditions. © 2018 by the American Diabetes Association.

  6. DPP-4 inhibitor des-F-sitagliptin treatment increased insulin exocytosis from db/db mice {beta} cells

    Energy Technology Data Exchange (ETDEWEB)

    Nagamatsu, Shinya, E-mail: shinya@ks.kyorin-u.ac.jp [Department of Biochemistry, Kyorin University School of Medicine, Mitaka, Tokyo 181-8611 (Japan); Ohara-Imaizumi, Mica; Nakamichi, Yoko; Aoyagi, Kyota; Nishiwaki, Chiyono [Department of Biochemistry, Kyorin University School of Medicine, Mitaka, Tokyo 181-8611 (Japan)

    2011-09-09

    Highlights: {yields} Anti-diabetic new drug, DPP-4 inhibitor, can affect the insulin exocytosis. {yields} DPP-4 inhibitor treatment altered syntaxin 1 expression. {yields} Treatment of db/db mice with DPP-4 inhibitor increased insulin release. -- Abstract: Incretin promotes insulin secretion acutely. Recently, orally-administered DPP-4 inhibitors represent a new class of anti-hyperglycemic agents. Indeed, inhibitors of dipeptidyl peptidase-IV (DPP-4), sitagliptin, has just begun to be widely used as therapeutics for type 2 diabetes. However, the effects of sitagliptin-treatment on insulin exocytosis from single {beta}-cells are yet unknown. We therefore investigated how sitagliptin-treatment in db/db mice affects insulin exocytosis by treating db/db mice with des-F-sitagliptin for 2 weeks. Perfusion studies showed that 2 weeks-sitagliptin treatment potentiated insulin secretion. We then analyzed insulin granule motion and SNARE protein, syntaxin 1, by TIRF imaging system. TIRF imaging of insulin exocytosis showed the increased number of docked insulin granules and increased fusion events from them during first-phase release. In accord with insulin exocytosis data, des-F-sitagliptin-treatment increased the number of syntaxin 1 clusters on the plasma membrane. Thus, our data demonstrated that 2-weeks des-F-sitagliptin-treatment increased the fusion events of insulin granules, probably via increased number of docked insulin granules and that of syntaxin 1 clusters.

  7. Differentiation of human induced pluripotent stem cells into insulin-like cell clusters with miR-186 and miR-375 by using chemical transfection.

    Science.gov (United States)

    Shaer, Anahita; Azarpira, Negar; Karimi, Mohammad Hosein

    2014-09-01

    Diabetes mellitus is characterized by either the inability to produce insulin or insensitivity to insulin secreted by the body. Islet cell replacement is an effective approach for diabetes treatment; however, it is not sufficient for all the diabetic patients. MicroRNAs (miRNAs) are a class of small noncoding RNAs that play an important role in mediating a broad and expanding range of biological activities, such as pancreas development. The present study aimed to develop a protocol to efficiently differentiate human induced pluripotent stem (iPS) cells into islet-like cell clusters (ILCs) in vitro by using miR-186 and miR-375. The human iPS colonies were transfected with hsa-miR-186 and hsa-miR-375 by using siPORT™ NeoFX™ Transfection Agent, and the differentiation was compared to controls. Total RNA was extracted 24 and 48 h after transfection. The gene expressions of insulin, NGN3, GLUT2, PAX4, PAX6, KIR6.2, NKX6.1, PDX1, Glucagon, and OCT4 were then evaluated through real-time qPCR. On the third day, the potency of the clusters was assessed in response to high glucose levels. Dithizone (DTZ) was used to identify the existence of the β-cells. Besides, the presence of insulin and NGN3 proteins was investigated by immunocytochemistry. Morphological changes were observed on the first day after the chemical transfection, and cell clusters were formed on the third day. The expression of pancreatic specific transcription factors was increased on the first day and significantly increased on the second day. The ILCs were positive for insulin and NGN3 proteins in the immunocytochemistry. Besides, the clusters were stained with DTZ and secreted insulin in glucose challenge test. Overexpression of miR-186 and miR-375 can be an alternative strategy for producing ILCs from the iPS cells in a short time. This work provides a new approach by using patient-specific iPSCs for β-cell replacement therapy in diabetic patients.

  8. Short-Term Exercise Training Improves Insulin Sensitivity but Does Not Inhibit Inflammatory Pathways in Immune Cells from Insulin-Resistant Subjects

    Directory of Open Access Journals (Sweden)

    Sara M. Reyna

    2013-01-01

    Full Text Available Background. Exercise has an anti-inflammatory effect against, and immune cells play critical roles in the development, of insulin resistance and atherosclerotic vascular disease (AVD. Thus, the goal of this study was to determine whether exercise improves insulin sensitivity in insulin-resistant subjects by downregulating proinflammatory signaling in immune cells. Methods. Seventeen lean, 8 obese nondiabetic, and 11 obese type 2 diabetic individuals underwent an aerobic exercise program for 15 days and an insulin clamp before and after exercise. Peripheral mononuclear cells (PMNC were obtained for determination of Toll-like receptor (TLR 2 and 4 protein content and mitogen-activated protein kinase phosphorylation. Results. Compared with that in lean individuals, TLR4 protein content was increased by 4.2-fold in diabetic subjects. This increase in TLR4 content was accompanied by a 3.0-fold increase in extracellular signal-regulated kinase (ERK phosphorylation. Exercise improved insulin sensitivity in the lean, obese, and type 2 diabetes groups. However, exercise did not affect TLR content or ERK phosphorylation. Conclusions. TLR4 content and ERK phosphorylation are increased in PMNC of type 2 diabetic individuals. While exercise improves insulin sensitivity, this effect is not related to changes in TLR2/TLR4 content or ERK phosphorylation in PMNC of type 2 diabetic individuals.

  9. Short-term exercise training improves insulin sensitivity but does not inhibit inflammatory pathways in immune cells from insulin-resistant subjects.

    Science.gov (United States)

    Reyna, Sara M; Tantiwong, Puntip; Cersosimo, Eugenio; Defronzo, Ralph A; Sriwijitkamol, Apiradee; Musi, Nicolas

    2013-01-01

    Background. Exercise has an anti-inflammatory effect against, and immune cells play critical roles in the development, of insulin resistance and atherosclerotic vascular disease (AVD). Thus, the goal of this study was to determine whether exercise improves insulin sensitivity in insulin-resistant subjects by downregulating proinflammatory signaling in immune cells. Methods. Seventeen lean, 8 obese nondiabetic, and 11 obese type 2 diabetic individuals underwent an aerobic exercise program for 15 days and an insulin clamp before and after exercise. Peripheral mononuclear cells (PMNC) were obtained for determination of Toll-like receptor (TLR) 2 and 4 protein content and mitogen-activated protein kinase phosphorylation. Results. Compared with that in lean individuals, TLR4 protein content was increased by 4.2-fold in diabetic subjects. This increase in TLR4 content was accompanied by a 3.0-fold increase in extracellular signal-regulated kinase (ERK) phosphorylation. Exercise improved insulin sensitivity in the lean, obese, and type 2 diabetes groups. However, exercise did not affect TLR content or ERK phosphorylation. Conclusions. TLR4 content and ERK phosphorylation are increased in PMNC of type 2 diabetic individuals. While exercise improves insulin sensitivity, this effect is not related to changes in TLR2/TLR4 content or ERK phosphorylation in PMNC of type 2 diabetic individuals.

  10. Depletion of Regulatory T Cells in Visceral Adipose Tissues Contributes to Insulin Resistance in Hashimoto's Thyroiditis

    Directory of Open Access Journals (Sweden)

    Min Yang

    2018-02-01

    Full Text Available Hashimoto's Thyroiditis (HT is a common organ-specific autoimmune disorder associated with a high incidence, and insulin resistance is highly related to autoimmune. Here, we examined the insulin sensitivity in HT patients and found decreased insulin sensitivity occurred in HT patients. To explore the relationship between impaired insulin sensitivity and immune status, we established HT model mice which showed similar pathological features and immune features to HT patients. In HT model mice, reinfusion of regulatory T cells (Tregs from peripheral blood of normal mice could improve insulin sensitivity and decrease the inflammation. Anti-CD25 antibodies blocked beneficial effects from reinfusion of Tregs, but delayed administration of anti-CD25 antibodies could not abolished the effect from Tregs. Delayed administration of anti-CD25 antibodies abolished exogenous Tregs in peripheral blood, but there were increased exogenous Tregs located to visceral adipose tissues (VATs which modulated the expression of cytokines in VATs. These findings suggest that insulin resistance exists in HT patients and it associates with the decreased Tregs and increased inflammation in the VATs.

  11. The role of polyunsaturated fatty acids (n-3 PUFAs) on the pancreatic β-cells and insulin action.

    Science.gov (United States)

    Baynes, Habtamu Wondifraw; Mideksa, Seifu; Ambachew, Sintayehu

    2018-03-14

    Polyunsaturated Fatty acids have multiple effects in peripheral tissues and pancreatic beta cell function. The n-3 Polyunsaturated Fatty acids prevent and reverse high-fat-diet induced adipose tissue inflammation and insulin resistance. Insulin secretion is stimulated by glucose, amino acids, and glucagon- like peptide-1 in tissue containing high levels of n-3 Polyunsaturated Fatty acids than lower level of n-3 Polyunsaturated Fatty acids. Also, n-3 Polyunsaturated Fatty acids led to decreased production of prostaglandin, which in turn contributed to the elevation of insulin secretion. N-3 polyunsaturated fatty acids prevent cytokine-induced cell death in pancreatic islets. Supplementation of n-3 Polyunsaturated Fatty acids for human subjects prevent beta cell destruction and insulin resistance. It also enhances insulin secretion, reduction in lipid profiles and glucose concentration particularly in type II diabetes patients. Therefore there should be a focus on the treatment mechanism of insulin related obesity and diabetes by n-3 polyunsaturated fatty acids.

  12. Interaction of insulin-like growth factor I with porcine thyroid cells cultured in monolayer

    International Nuclear Information System (INIS)

    Saji, M.; Tsushima, T.; Isozaki, O.; Murakami, H.; Ohba, Y.; Sato, K.; Arai, M.; Mariko, A.; Shizume, K.

    1987-01-01

    The interaction of insulin-like growth factor I (IGF-I) with porcine thyroid cells cultured in monolayer was studied. Specific binding of [ 125 I]iodo-IGF-I to thyroid cells was a reversible process dependent on the time and temperature of incubation. A steady state was achieved in 18 h at 4 C and averaged 14.2 +/- 2% (mean +/- SD)/10(6) cells. Binding of [ 125 I]iodo-IGF-I was inhibited by unlabeled IGF-I; half-maximal inhibition occurred at concentrations of 2-5 ng/ml. Multiplication-stimulating activity (rat IGF-II) and pork insulin had relative potencies of 1:20 and 1:300 compared with IGF-I. Scatchard analysis of binding data revealed a single class of IGF-I receptors with a Ka of 4.3 X 10(10) M-1, 49,000 binding sites were estimated per cell. Affinity cross-linking and autoradiography demonstrated the presence of type I IGF receptors. Thyroid cells also had specific receptors for insulin, but specific binding of [ 125 I]iodoinsulin was much lower than that of [ 125 I]iodo-IGF-I. Preincubation of thyroid cells with IGF-I or insulin caused a concentration-dependent decrease in [ 125 I]iodo-IGF-I binding due to an apparent loss of receptors. Preincubation with epidermal growth factor, fibroblast growth factor, platelet-derived growth factor, or TSH did not alter subsequent binding of [ 125 I]iodo-IGF-I. Low concentrations of IGF-I stimulated DNA synthesis and proliferation of thyroid cells and acted synergistically with epidermal growth factor. Multiplication-stimulating activity and insulin had relative potencies in stimulating DNA synthesis comparable to their abilities to inhibit the binding of [ 125 I]iodo-IGF-I to thyroid cells

  13. Insulin autoimmune syndrome: case report

    Directory of Open Access Journals (Sweden)

    Rodrigo Oliveira Moreira

    Full Text Available CONTEXT: Insulin autoimmune syndrome (IAS, Hirata disease is a rare cause of hypoglycemia in Western countries. It is characterized by hypoglycemic episodes, elevated insulin levels, and positive insulin antibodies. Our objective is to report a case of IAS identified in South America. CASE REPORT: A 56-year-old Caucasian male patient started presenting neuroglycopenic symptoms during hospitalization due to severe trauma. Biochemical evaluation confirmed hypoglycemia and abnormally high levels of insulin. Conventional imaging examinations were negative for pancreatic tumor. Insulin antibodies were above the normal range. Clinical remission of the episodes was not achieved with verapamil and steroids. Thus, a subtotal pancreatectomy was performed due to the lack of response to conservative treatment and because immunosuppressants were contraindicated due to bacteremia. Histopathological examination revealed diffuse hypertrophy of beta cells. The patient continues to have high insulin levels but is almost free of hypoglycemic episodes.

  14. Glucose-Responsive Insulin Delivery by Microneedle-Array Patches Loaded with Hypoxia-Sensitive Vesicles.

    Science.gov (United States)

    Yu, Jicheng; Zhang, Yuqi; Gu, Zhen

    2017-01-01

    In this chapter, we describe the preparation of glucose-responsive vesicles (GRVs) and the fabrication of GRV-loaded microneedle-array patches for insulin delivery. The GRVs were formed of hypoxia-sensitive hyaluronic acid (HS-HA), the synthesis of which is presented in detail. We also describe the procedure to evaluate the in vivo efficacy of this smart patch in a mouse model of chemically induced type 1 diabetes through transcutaneous administration.

  15. The glycolipid sulfatide protects insulin-producing cells against cytokine-induced apoptosis, a possible role in diabetes

    DEFF Research Database (Denmark)

    Roeske-Nielsen, A; Dalgaard, L T; Månsson, Sven-Erik

    2010-01-01

    these is NO production. The glycosphingolipid sulfatide is present in ß-cells in the secretory granules in varying amounts and is secreted together with insulin. We now investigate whether sulfatide is able to protect insulin-producing cells against the pro-apoptotic effect of interleukin-1ß, interferon-¿ and tumour...

  16. RFX6 Regulates Insulin Secretion by Modulating Ca2+ Homeostasis in Human β Cells

    Directory of Open Access Journals (Sweden)

    Vikash Chandra

    2014-12-01

    Full Text Available Development and function of pancreatic β cells involve the regulated activity of specific transcription factors. RFX6 is a transcription factor essential for mouse β cell differentiation that is mutated in monogenic forms of neonatal diabetes. However, the expression and functional roles of RFX6 in human β cells, especially in pathophysiological conditions, are poorly explored. We demonstrate the presence of RFX6 in adult human pancreatic endocrine cells. Using the recently developed human β cell line EndoC-βH2, we show that RFX6 regulates insulin gene transcription, insulin content, and secretion. Knockdown of RFX6 causes downregulation of Ca2+-channel genes resulting in the reduction in L-type Ca2+-channel activity that leads to suppression of depolarization-evoked insulin exocytosis. We also describe a previously unreported homozygous missense RFX6 mutation (p.V506G that is associated with neonatal diabetes, which lacks the capacity to activate the insulin promoter and to increase Ca2+-channel expression. Our data therefore provide insights for understanding certain forms of neonatal diabetes.

  17. Localization of glucagon and insulin cells and its variation with respect to physiological events in Eutropis carinata

    Directory of Open Access Journals (Sweden)

    Vidya. R. Chandavar

    2011-06-01

    Full Text Available The aim of the present investigation was to localize glucagon and insulin immunoreactive (IR cells of pancreas during annual seasonal cycle of reproduction and to find out whether they had any effect on the regulation of plasma glucose level in the skink Eutropis carinata. Immunolocalized pancreatic cells revealed significantly different mean numbers in different periods of reproduction. The numbers of glucagon-IR and insulin-IR cells were highest in recrudescent period which was corresponded with low plasma glucose level. Unlike other lizards the arrangement of insulin cells in the central core and glucagon cells at the periphery was absent instead glucagon-IR and insulin-IR cells were paracrine in arrangement. Among the two immunoreactive cells glucagon-IR cells were predominant. Morphological differences between two cell types were observed by electron microscopy after staining with uranyl acetate and lead citrate. Plasma glucose showed cyclic change being highest during reproductive period.

  18. Pancreatic Transdifferentiation and Glucose-Regulated Production of Human Insulin in the H4IIE Rat Liver Cell Line

    Directory of Open Access Journals (Sweden)

    Binhai Ren

    2016-04-01

    Full Text Available Due to the limitations of current treatment regimes, gene therapy is a promising strategy being explored to correct blood glucose concentrations in diabetic patients. In the current study, we used a retroviral vector to deliver either the human insulin gene alone, the rat NeuroD1 gene alone, or the human insulin gene and rat NeuroD1 genes together, to the rat liver cell line, H4IIE, to determine if storage of insulin and pancreatic transdifferentiation occurred. Stable clones were selected and expanded into cell lines: H4IIEins (insulin gene alone, H4IIE/ND (NeuroD1 gene alone, and H4IIEins/ND (insulin and NeuroD1 genes. The H4IIEins cells did not store insulin; however, H4IIE/ND and H4IIEins/ND cells stored 65.5 ± 5.6 and 1475.4 ± 171.8 pmol/insulin/5 × 106 cells, respectively. Additionally, several β cell transcription factors and pancreatic hormones were expressed in both H4IIE/ND and H4IIEins/ND cells. Electron microscopy revealed insulin storage vesicles in the H4IIE/ND and H4IIEins/ND cell lines. Regulated secretion of insulin to glucose (0–20 mmol/L was seen in the H4IIEins/ND cell line. The H4IIEins/ND cells were transplanted into diabetic immunoincompetent mice, resulting in normalization of blood glucose. This data shows that the expression of NeuroD1 and insulin in liver cells may be a useful strategy for inducing islet neogenesis and reversing diabetes.

  19. Modulation of the effect of acetylcholine on insulin release by the membrane potential of B cells

    International Nuclear Information System (INIS)

    Hermans, M.P.; Schmeer, W.; Henquin, J.C.

    1987-01-01

    Mouse islets were used to test the hypothesis that the B cell membrane must be depolarized for acetylcholine to increase insulin release. The resting membrane potential of B cells (at 3 mM glucose) was slightly decreased (5 mV) by acetylcholine, but no electrical activity appeared. This depolarization was accompanied by a Ca-independent acceleration of 86 Rb and 45 Ca efflux but no insulin release. When the B cell membrane was depolarized by a stimulatory concentration of glucose (10 mM), acetylcholine potentiated electrical activity, accelerated 86 Rb and 45 Ca efflux, and increased insulin release. This latter effect, but not the acceleration of 45 Ca efflux, was totally dependent on extracellular Ca. If glucose-induced depolarization of the B cell membrane was prevented by diazoxide, acetylcholine lost all effects but those produced at low glucose. In contrast, when the B cell membrane was depolarized by leucine or tolbutamide (at 3 mM glucose), acetylcholine triggered a further depolarization with appearance of electrical activity, accelerated 86 Rb and 45 Ca efflux, and stimulated insulin release. Acetylcholine produced similar effects (except for electrical activity) in the presence of high K or arginine which, unlike the above test agents, depolarize the B cell membrane by a mechanism other than a decrease in K+ permeability. Omission of extracellular Ca abolished the releasing effect of acetylcholine under all conditions but only partially decreased the stimulation of 45 Ca efflux. The results show thus that acetylcholine stimulation of insulin release does not result from mobilization of cellular Ca but requires that the B cell membrane be sufficiently depolarized to reach the threshold potential where Ca channels are activated. This may explain why acetylcholine alone does not initiate release but becomes active in the presence of a variety of agents

  20. Glucose-stimulated insulin response in pregnant sheep following acute suppression of plasma non-esterified fatty acid concentrations

    Directory of Open Access Journals (Sweden)

    Sriskandarajah Nadarajah

    2004-09-01

    Full Text Available Abstract Background Elevated non-esterified fatty acids (NEFA concentrations in non-pregnant animals have been reported to decrease pancreatic responsiveness. As ovine gestation advances, maternal insulin concentrations fall and NEFA concentrations increase. Experiments were designed to examine if the pregnancy-associated rise in NEFA concentration is associated with a reduced pancreatic sensitivity to glucose in vivo. We investigated the possible relationship of NEFA concentrations in regulating maternal insulin concentrations during ovine pregnancy at three physiological states, non-pregnant, non-lactating (NPNL, 105 and 135 days gestational age (dGA, term 147+/- 3 days. Methods The plasma concentrations of insulin, growth hormone (GH and ovine placental lactogen (oPL were determined by double antibody radioimmunoassay. Insulin responsiveness to glucose was measured using bolus injection and hyperglycaemic clamp techniques in 15 non-pregnant, non-lactating ewes and in nine pregnant ewes at 105 dGA and near term at 135 dGA. Plasma samples were also collected for hormone determination. In addition to bolus injection glucose and insulin Area Under Curve calculations, the Mean Plasma Glucose Increment, Glucose Infusion Rate and Mean Plasma Insulin Increment and Area Under Curve were determined for the hyperglycaemic clamp procedures. Statistical analysis of data was conducted with Students t-tests, repeated measures ANOVA and 2-way ANOVA. Results Maternal growth hormone, placental lactogen and NEFA concentrations increased, while basal glucose and insulin concentrations declined with advancing gestation. At 135 dGA following bolus glucose injections, peak insulin concentrations and insulin area under curve (AUC profiles were significantly reduced in pregnant ewes compared with NPNL control ewes (p Conclusions Results suggest that despite an acute suppression of circulating NEFA concentrations during pregnancy, the associated steroids and hormones

  1. Early growth response-1 negative feedback regulates skeletal muscle postprandial insulin sensitivity via activating Ptp1b transcription.

    Science.gov (United States)

    Wu, Jing; Tao, Wei-Wei; Chong, Dan-Yang; Lai, Shan-Shan; Wang, Chuang; Liu, Qi; Zhang, Tong-Yu; Xue, Bin; Li, Chao-Jun

    2018-03-15

    Postprandial insulin desensitization plays a critical role in maintaining whole-body glucose homeostasis by avoiding the excessive absorption of blood glucose; however, the detailed mechanisms that underlie how the major player, skeletal muscle, desensitizes insulin action remain to be elucidated. Herein, we report that early growth response gene-1 ( Egr-1) is activated by insulin in skeletal muscle and provides feedback inhibition that regulates insulin sensitivity after a meal. The inhibition of the transcriptional activity of Egr-1 enhanced the phosphorylation of the insulin receptor (InsR) and Akt, thus increasing glucose uptake in L6 myotubes after insulin stimulation, whereas overexpression of Egr-1 decreased insulin sensitivity. Furthermore, deletion of Egr-1 in the skeletal muscle improved systemic insulin sensitivity and glucose tolerance, which resulted in lower blood glucose levels after refeeding. Mechanistic analysis demonstrated that EGR-1 inhibited InsR phosphorylation and glucose uptake in skeletal muscle by binding to the proximal promoter region of protein tyrosine phosphatase-1B (PTP1B) and directly activating transcription. PTP1B knockdown largely restored insulin sensitivity and enhanced glucose uptake, even under conditions of EGR-1 overexpression. Our results indicate that EGR-1/PTP1B signaling negatively regulates postprandial insulin sensitivity and suggest a potential therapeutic target for the prevention and treatment of excessive glucose absorption.-Wu, J., Tao, W.-W., Chong, D.-Y., Lai, S.-S., Wang, C., Liu, Q., Zhang, T.-Y., Xue, B., Li, C.-J. Early growth response-1 negative feedback regulates skeletal muscle postprandial insulin sensitivity via activating Ptp1b transcription.

  2. Human beta-cell precursors mature into functional insulin-producing cells in an immunoisolation device: implications for diabetes cell therapies.

    Science.gov (United States)

    Lee, Seung-Hee; Hao, Ergeng; Savinov, Alexei Y; Geron, Ifat; Strongin, Alex Y; Itkin-Ansari, Pamela

    2009-04-15

    Islet transplantation is limited by the need for chronic immunosuppression and the paucity of donor tissue. As new sources of human beta-cells are developed (e.g., stem cell-derived tissue), transplanting them in a durable device could obviate the need for immunosuppression, while also protecting the patient from any risk of tumorigenicity. Here, we studied (1) the survival and function of encapsulated human beta-cells and their progenitors and (2) the engraftment of encapsulated murine beta-cells in allo- and autoimmune settings. Human islets and human fetal pancreatic islet-like cell clusters were encapsulated in polytetrafluorethylene devices (TheraCyte) and transplanted into immunodeficient mice. Graft survival and function was measured by immunohistochemistry, circulating human C-peptide levels, and blood glucose levels. Bioluminescent imaging was used to monitor encapsulated neonatal murine islets. Encapsulated human islet-like cell clusters survived, replicated, and acquired a level of glucose responsive insulin secretion sufficient to ameliorate hyperglycemia in diabetic mice. Bioluminescent imaging of encapsulated murine neonatal islets revealed a dynamic process of cell death followed by regrowth, resulting in robust long-term allograft survival. Further, in the non-obese diabetic (NOD) mouse model of type I diabetes, encapsulated primary beta-cells ameliorated diabetes without stimulating a detectable T-cell response. We demonstrate for the first time that human beta-cells function is compatible with encapsulation in a durable, immunoprotective device. Moreover, our study suggests that encapsulation of beta-cells before terminal differentiation will be a successful approach for new cell-based therapies for diabetes, such as those derived from stem cells.

  3. Human β-cell Precursors Mature Into Functional Insulin-producing Cells in an Immunoisolation Device: Implications for Diabetes Cell Therapies

    Science.gov (United States)

    Lee, Seung-Hee; Hao, Ergeng; Savinov, Alexei Y.; Geron, Ifat; Strongin, Alex Y.; Itkin-Ansari, Pamela

    2009-01-01

    Background Islet transplantation is limited by the need for chronic immunosuppression and the paucity of donor tissue. As new sources of human β-cells are developed (e.g., stem cell-derived tissue), transplanting them in a durable device could obviate the need for immunosuppression, while also protecting the patient from any risk of tumorigenicity. Here, we studied (1) the survival and function of encapsulated human β-cells and their progenitors and (2) the engraftment of encapsulated murine β-cells in allo- and autoimmune settings. Methods Human islets and human fetal pancreatic islet-like cell clusters were encapsulated in polytetrafluorethylene devices (TheraCyte) and transplanted into immunodeficient mice. Graft survival and function was measured by immunohistochemistry, circulating human C-peptide levels, and blood glucose levels. Bioluminescent imaging was used to monitor encapsulated neonatal murine islets. Results Encapsulated human islet-like cell clusters survived, replicated, and acquired a level of glucose responsive insulin secretion sufficient to ameliorate hyperglycemia in diabetic mice. Bioluminescent imaging of encapsulated murine neonatal islets revealed a dynamic process of cell death followed by regrowth, resulting in robust long-term allograft survival. Further, in the non-obese diabetic (NOD) mouse model of type I diabetes, encapsulated primary β-cells ameliorated diabetes without stimulating a detectable T-cell response. Conclusions We demonstrate for the first time that human β-cells function is compatible with encapsulation in a durable, immunoprotective device. Moreover, our study suggests that encapsulation of β-cells before terminal differentiation will be a successful approach for new cell-based therapies for diabetes, such as those derived from stem cells. PMID:19352116

  4. Leptin and insulin up-regulate miR-4443 to suppress NCOA1 and TRAF4, and decrease the invasiveness of human colon cancer cells

    International Nuclear Information System (INIS)

    Meerson, Ari; Yehuda, Hila

    2016-01-01

    Obesity is a risk factor for colorectal cancer (CRC). Normal and tumor cells respond to metabolic hormones, such as leptin and insulin. Thus, obesity-associated resistance to these hormones likely leads to changes in gene expression and behavior of tumor cells. However, the mechanisms affected by leptin and insulin signaling in CRC cells remain mostly unknown. We hypothesized that microRNAs (miRNAs) are involved in the regulation of tumorigenesis-related gene expression in CRC cells by leptin and insulin. To test this hypothesis, miRNA levels in the CRC-derived cell lines HCT-116, HT-29 and DLD-1 were profiled, following leptin and insulin treatment. Candidate miRNAs were validated by RT-qPCR. Predicted miRNA targets with known roles in cancer, were validated by immunoblots and reporter assays in HCT-116 cells. Transfection of HCT-116 cells with candidate miRNA mimic was used to test in vitro effects on proliferation and invasion. Of ~800 miRNAs profiled, miR-4443 was consistently up-regulated by leptin and insulin in HCT-116 and HT-29, but not in DLD-1, which lacked normal leptin receptor expression. Dose response experiments showed that leptin at 100 ng/ml consistently up-regulated miR-4443 in HCT-116 cells, concomitantly with a significant decrease in cell invasion ability. Transfection with miR-4443 mimic decreased invasion and proliferation of HCT-116 cells. Moreover, leptin and miR-4443 transfection significantly down-regulated endogenous NCOA1 and TRAF4, both predicted targets of miR-4443 with known roles in cancer metastasis. miR-4443 was found to directly regulate TRAF4 and NCOA1, as validated by a reporter assay. The up-regulation of miR-4443 by leptin or insulin was attenuated by the inhibition of MEK1/2. Our findings suggest that miR-4443 acts in a tumor-suppressive manner by down-regulating TRAF4 and NCOA1 downstream of MEK-C/EBP-mediated leptin and insulin signaling, and that insulin and/or leptin resistance (e.g. in obesity) may suppress this pathway

  5. Glucose, other secretagogues, and nerve growth factor stimulate mitogen-activated protein kinase in the insulin-secreting beta-cell line, INS-1

    DEFF Research Database (Denmark)

    Frödin, M; Sekine, N; Roche, E

    1995-01-01

    The signaling pathways whereby glucose and hormonal secretagogues regulate insulin-secretory function, gene transcription, and proliferation of pancreatic beta-cells are not well defined. We show that in the glucose-responsive beta-cell line INS-1, major secretagogue-stimulated signaling pathways...... converge to activate 44-kDa mitogen-activated protein (MAP) kinase. Thus, glucose-induced insulin secretion was found to be associated with a small stimulatory effect on 44-kDa MAP kinase, which was synergistically enhanced by increased levels of intracellular cAMP and by the hormonal secretagogues......-1. Phorbol ester, an activator of protein kinase C, stimulated 44-kDa MAP kinase by both Ca(2+)-dependent and -independent pathways. Nerve growth factor, independently of changes in cytosolic Ca2+, efficiently stimulated 44-kDa MAP kinase without causing insulin release, indicating that activation...

  6. Intraportal injection of insulin-producing cells generated from human bone marrow mesenchymal stem cells decreases blood glucose level in diabetic rats.

    Science.gov (United States)

    Tsai, Pei-Jiun; Wang, Hwai-Shi; Lin, Chi-Hung; Weng, Zen-Chung; Chen, Tien-Hua; Shyu, Jia-Fwu

    2014-01-01

    We studied the process of trans-differentiation of human bone marrow mesenchymal stem cells (hBM-MSCs) into insulin-producing cells. Streptozotocin (STZ)-induced diabetic rat model was used to study the effect of portal vein transplantation of these insulin-producing cells on blood sugar levels. The BM-MSCs were differentiated into insulin-producing cells under defined conditions. Real-time PCR, immunocytochemistry and glucose challenge were used to evaluate in vitro differentiation. Flow cytometry showed that hBM-MSCs were strongly positive for CD44, CD105 and CD73 and negative for hematopoietic markers CD34, CD38 and CD45. Differentiated cells expressed C-peptide as well as β-cells specific genes and hormones. Glucose stimulation increased C-peptide secretion in these cells. The insulin-producing, differentiated cells were transplanted into the portal vein of STZ-induced diabetic rats using a Port-A catheter. The insulin-producing cells were localized in the liver of the recipient rat and expressed human C-peptide. Blood glucose levels were reduced in diabetic rats transplanted with insulin-producing cells. We concluded that hBM-MSCs could be trans-differentiated into insulin-producing cells in vitro. Portal vein transplantation of insulin-producing cells alleviated hyperglycemia in diabetic rats.

  7. Rates of insulin secretion in INS-1 cells are enhanced by coupling to anaplerosis and Kreb’s cycle flux independent of ATP synthesis

    International Nuclear Information System (INIS)

    Cline, Gary W.; Pongratz, Rebecca L.; Zhao, Xiaojian; Papas, Klearchos K.

    2011-01-01

    Highlights: ► We studied media effects on mechanisms of insulin secretion of INS-1 cells. ► Insulin secretion was higher in DMEM than KRB despite identical ATP synthesis rates. ► Insulin secretion rates correlated with rates of anaplerosis and TCA cycle. ► Mitochondria metabolism and substrate cycles augment secretion signal of ATP. -- Abstract: Mechanistic models of glucose stimulated insulin secretion (GSIS) established in minimal media in vitro, may not accurately describe the complexity of coupling metabolism with insulin secretion that occurs in vivo. As a first approximation, we have evaluated metabolic pathways in a typical growth media, DMEM as a surrogate in vivo medium, for comparison to metabolic fluxes observed under the typical experimental conditions using the simple salt-buffer of KRB. Changes in metabolism in response to glucose and amino acids and coupling to insulin secretion were measured in INS-1 832/13 cells. Media effects on mitochondrial function and the coupling efficiency of oxidative phosphorylation were determined by fluorometrically measured oxygen consumption rates (OCRs) combined with 31 P NMR measured rates of ATP synthesis. Substrate preferences and pathways into the TCA cycle, and the synthesis of mitochondrial 2nd messengers by anaplerosis were determined by 13 C NMR isotopomer analysis of the fate of [U- 13 C] glucose metabolism. Despite similar incremental increases in insulin secretion, the changes of OCR in response to increasing glucose from 2.5 to 15 mM were blunted in DMEM relative to KRB. Basal and stimulated rates of insulin secretion rates were consistently higher in DMEM, while ATP synthesis rates were identical in both DMEM and KRB, suggesting greater mitochondrial uncoupling in DMEM. The relative rates of anaplerosis, and hence synthesis and export of 2nd messengers from the mitochondria were found to be similar in DMEM to those in KRB. And, the correlation of total PC flux with insulin secretion rates in DMEM

  8. Rates of insulin secretion in INS-1 cells are enhanced by coupling to anaplerosis and Kreb's cycle flux independent of ATP synthesis

    Energy Technology Data Exchange (ETDEWEB)

    Cline, Gary W., E-mail: gary.cline@yale.edu [The Department of Internal Medicine, Yale University School of Medicine, New Haven, CT 06520 (United States); Department of Surgery, University of Minnesota-Twin Cities, Minneapolis, MN 55455 (United States); Pongratz, Rebecca L.; Zhao, Xiaojian [The Department of Internal Medicine, Yale University School of Medicine, New Haven, CT 06520 (United States); Papas, Klearchos K. [Department of Surgery, University of Minnesota-Twin Cities, Minneapolis, MN 55455 (United States)

    2011-11-11

    Highlights: Black-Right-Pointing-Pointer We studied media effects on mechanisms of insulin secretion of INS-1 cells. Black-Right-Pointing-Pointer Insulin secretion was higher in DMEM than KRB despite identical ATP synthesis rates. Black-Right-Pointing-Pointer Insulin secretion rates correlated with rates of anaplerosis and TCA cycle. Black-Right-Pointing-Pointer Mitochondria metabolism and substrate cycles augment secretion signal of ATP. -- Abstract: Mechanistic models of glucose stimulated insulin secretion (GSIS) established in minimal media in vitro, may not accurately describe the complexity of coupling metabolism with insulin secretion that occurs in vivo. As a first approximation, we have evaluated metabolic pathways in a typical growth media, DMEM as a surrogate in vivo medium, for comparison to metabolic fluxes observed under the typical experimental conditions using the simple salt-buffer of KRB. Changes in metabolism in response to glucose and amino acids and coupling to insulin secretion were measured in INS-1 832/13 cells. Media effects on mitochondrial function and the coupling efficiency of oxidative phosphorylation were determined by fluorometrically measured oxygen consumption rates (OCRs) combined with {sup 31}P NMR measured rates of ATP synthesis. Substrate preferences and pathways into the TCA cycle, and the synthesis of mitochondrial 2nd messengers by anaplerosis were determined by {sup 13}C NMR isotopomer analysis of the fate of [U-{sup 13}C] glucose metabolism. Despite similar incremental increases in insulin secretion, the changes of OCR in response to increasing glucose from 2.5 to 15 mM were blunted in DMEM relative to KRB. Basal and stimulated rates of insulin secretion rates were consistently higher in DMEM, while ATP synthesis rates were identical in both DMEM and KRB, suggesting greater mitochondrial uncoupling in DMEM. The relative rates of anaplerosis, and hence synthesis and export of 2nd messengers from the mitochondria were found

  9. Optimal dose and timing of insulin Aspart to mimic first phase insulin response in patients with recently onset type 2 diabetes

    DEFF Research Database (Denmark)

    Gredal, C.; Rosenfalck, A.; Dejgaard, A.

    2008-01-01

    OBJECTIVE: To assess the optimal dose and timing of subcutaneous injection of insulin Aspart (IAsp) in relation to meal to mimic first phase insulin response in patients with recently diagnosed type 2 diabetes. DESIGN AND METHODS: Twenty patients were randomised in a double blind, double dummy...... design to four standard meal tests with pre-meal injection of insulin Aspart 0.08 IU/kg BW 30 min before the meal, insulin Aspart 0.04 IU/kg BW 30 or 15 min before the meal and placebo. RESULTS: All three insulin regimes significantly reduced postprandial glucose increment (area under the curve AUC(-30...... injection of IAsp 0.08 IU/kg BW. No difference in postprandial glucose profile was demonstrated whether IAsp 0.04 IU/kg BW was administrated 15 or 30 min before mealtime. CONCLUSIONS: IAsp 0.04IU/kg BW injected subcutaneously 15 or 30 min before meal reduced the postprandial blood glucose increment without...

  10. Developmental programming of polycystic ovary syndrome (PCOS): prenatal androgens establish pancreatic islet α/β cell ratio and subsequent insulin secretion.

    Science.gov (United States)

    Ramaswamy, S; Grace, C; Mattei, A A; Siemienowicz, K; Brownlee, W; MacCallum, J; McNeilly, A S; Duncan, W C; Rae, M T

    2016-06-06

    Exogenous androgenic steroids applied to pregnant sheep programmes a PCOS-like phenotype in female offspring. Via ultrasound guidance we applied steroids directly to ovine fetuses at d62 and d82 of gestation, and examined fetal (day 90 gestation) and postnatal (11 months old) pancreatic structure and function. Of three classes of steroid agonists applied (androgen - Testosterone propionate (TP), estrogen - Diethystilbesterol (DES) and glucocorticoid - Dexamethasone (DEX)), only androgens (TP) caused altered pancreatic development. Beta cell numbers were significantly elevated in prenatally androgenised female fetuses (P = 0.03) (to approximately the higher numbers found in male fetuses), whereas alpha cell counts were unaffected, precipitating decreased alpha:beta cell ratios in the developing fetal pancreas (P = 0.001), sustained into adolescence (P = 0.0004). In adolescence basal insulin secretion was significantly higher in female offspring from androgen-excess pregnancies (P = 0.045), and an exaggerated, hyperinsulinaemic response to glucose challenge (P = 0.0007) observed, whereas prenatal DES or DEX treatment had no effects upon insulin secretion. Postnatal insulin secretion correlated with beta cell numbers (P = 0.03). We conclude that the pancreas is a primary locus of androgenic stimulation during development, giving rise to postnatal offspring whose pancreas secreted excess insulin due to excess beta cells in the presence of a normal number of alpha cells.

  11. Insulin receptor in mouse neuroblastoma cell line N18TG2: binding properties and visualization with colloidal gold.

    Science.gov (United States)

    Sartori, C; Stefanini, S; Bernardo, A; Augusti-Tocco, G

    1992-08-01

    Insulin function in the nervous system is still poorly understood. Possible roles as a neuromodulator and as a growth factor have been proposed (Baskin et al., 1987, Ann. Rev. Physiol. 49, 335-347). Stable cell lines may provide an appropriate experimental system for the analysis of insulin action on the various cellular components of the central nervous system. We report here a study to investigate the presence and the properties of insulin specific binding sites in the murine neuroblastoma line, N18TG2, together with insulin action on cell growth and metabolism. Also, receptor internalization has been studied. Binding experiments, carried out in standard conditions at 20 degrees C, enabled us to demonstrate that these cells bind insulin in a specific manner, thus confirming previous findings on other cell lines. Saturation curves showed the presence of two binding sites with Kd 0.3 and 9.7 nM. Competition experiments with porcine and bovine insulin showed an IC50 of 1 and 10 nM, respectively. Competition did not occur in the presence of the unrelated hormones ACTH and FSH. Dissociation experiments indicated the existence of an internalization process of the ligand-receptor complex; this was confirmed by an ultrastructural study using gold conjugated insulin. As far as the insulin action in N18TG2 cells is concerned, physiological concentrations stimulate cell proliferation, whereas no stimulation of glucose uptake was observed, indicating that insulin action in these cells is not mediated by general metabolic effects. On the basis of these data, N18TG2 line appears to be a very suitable model for further studies of the neuronal type insulin receptors, and possibly insulin specific action on the nervous system.

  12. Evaluation of fasting state-/oral glucose tolerance test-derived measures of insulin release for the detection of genetically impaired β-cell function.

    Directory of Open Access Journals (Sweden)

    Silke A Herzberg-Schäfer

    Full Text Available BACKGROUND: To date, fasting state- and different oral glucose tolerance test (OGTT-derived measures are used to estimate insulin release with reasonable effort in large human cohorts required, e.g., for genetic studies. Here, we evaluated twelve common (or recently introduced fasting state-/OGTT-derived indices for their suitability to detect genetically determined β-cell dysfunction. METHODOLOGY/PRINCIPAL FINDINGS: A cohort of 1364 White European individuals at increased risk for type 2 diabetes was characterized by OGTT with glucose, insulin, and C-peptide measurements and genotyped for single nucleotide polymorphisms (SNPs known to affect glucose- and incretin-stimulated insulin secretion. One fasting state- and eleven OGTT-derived indices were calculated and statistically evaluated. After adjustment for confounding variables, all tested SNPs were significantly associated with at least two insulin secretion measures (p≤0.05. The indices were ranked according to their associations' statistical power, and the ranks an index obtained for its associations with all the tested SNPs (or a subset were summed up resulting in a final ranking. This approach revealed area under the curve (AUC(Insulin(0-30/AUC(Glucose(0-30 as the best-ranked index to detect SNP-dependent differences in insulin release. Moreover, AUC(Insulin(0-30/AUC(Glucose(0-30, corrected insulin response (CIR, AUC(C-Peptide(0-30/AUC(Glucose(0-30, AUC(C-Peptide(0-120/AUC(Glucose(0-120, two different formulas for the incremental insulin response from 0-30 min, i.e., the insulinogenic indices (IGI(2 and IGI(1, and insulin 30 min were significantly higher-ranked than homeostasis model assessment of β-cell function (HOMA-B; p<0.05. AUC(C-Peptide(0-120/AUC(Glucose(0-120 was best-ranked for the detection of SNPs involved in incretin-stimulated insulin secretion. In all analyses, HOMA-β displayed the highest rank sums and, thus, scored last. CONCLUSIONS/SIGNIFICANCE: With AUC(Insulin(0

  13. Activity of insulin growth factors and shrimp neurosecretory organ extracts on a lepidopteran cell line.

    Science.gov (United States)

    Hatt, P J; Liebon, C; Morinière, M; Oberlander, H; Porcheron, P

    1997-01-01

    Ecdysteroids, or molting hormones, have been proven to be key differentiation regulators for epidermal cells in the postembryonic development of arthropods. Regulators of cell proliferation, however, remain largely unknown. To date, no diffusible insect peptidic growth factors have been characterized. Molecules structurally related to insulin have been discovered in insects, as in other eucaryotes. We developed in vitro tests for the preliminary characterization of potential growth factors in arthropods by adapting the procedures designed to detect such factors in vertebrates to an insect cell line (IAL-PID2) established from imaginal discs of the Indian meal moth. We verified the ability of these tests to measure the proliferation of IAL-PID2 cells. We tested mammalian insulin and insulin-like growth factors (IGF-I, IGF-II). Following an arrest of cell proliferation by serum deprivation, IGF-I and IGF-II caused partial resumption of the cell cycle, evidenced by DNA synthesis. In contrast, the addition of 20-hydroxyecdysone arrested the proliferation of the IAL-PID2 cells. The cell line was then used in a test for functional characterization of potential growth factors originating from the penaeid shrimp, Penaeus vannamei. Crude extracts of neurosecretory and nervous tissues, eyestalks, and ventral neural chain compensated for serum deprivation and stimulated completion of mitosis. Arch.

  14. Effect of iron on pancreatic beta cell function and insulin resistance ...

    African Journals Online (AJOL)

    Background: Increase in total body iron store has been reported in the aetiology and development of diabetes mellitus. The effect of iron supplementation in female with respect to the incidence of diabetes mellitus was investigated on the pancreatic beta cell function and insulin resistance in normal female rats. Methods: ...

  15. Islet amyloid polypeptide and insulin expression are controlled differently in primary and transformed islet cells

    DEFF Research Database (Denmark)

    Madsen, O D; Michelsen, Bo Thomas; Westermark, P

    1991-01-01

    in unstable heterogeneous clones such as NHI-6F. This clone is composed of primarily glucagon-producing cells in vitro, but insulin gene expression becomes dominant after passage in vivo. Interestingly, IAPP was hyperexpressed with glucagon under in vitro conditions in this clone. We conclude that the tissue...

  16. Imaging of beta-Cell Mass and Insulitis in Insulin-Dependent (Type 1) Diabetes Mellitus

    NARCIS (Netherlands)

    Di Gialleonardo, Valentina; de Vries, Erik F. J.; Di Girolamo, Marco; Quintero, Ana M.; Dierckx, Rudi A. J. O.; Signore, Alberto

    2012-01-01

    Insulin-dependent (type 1) diabetes mellitus is a metabolic disease with a complex multifactorial etiology and a poorly understood pathogenesis. Genetic and environmental factors cause an autoimmune reaction against pancreatic beta-cells, called insulitis, confirmed in pancreatic samples obtained at

  17. Plasma membrane events associated with the meiotic divisions in the amphibian oocyte: insights into the evolution of insulin transduction systems and cell signaling

    Directory of Open Access Journals (Sweden)

    Morrill Gene A

    2013-01-01

    Full Text Available Abstract Background Insulin and its plasma membrane receptor constitute an ancient response system critical to cell growth and differentiation. Studies using intact Rana pipiens oocytes have shown that insulin can act at receptors on the oocyte surface to initiate resumption of the first meiotic division. We have reexamined the insulin-induced cascade of electrical and ion transport-related plasma membrane events using both oocytes and intact plasma membranes in order to characterize the insulin receptor-steroid response system associated with the meiotic divisions. Results [125I]Insulin binding (Kd = 54 ± 6 nM at the oocyte plasma membrane activates membrane serine protease(s, followed by the loss of low affinity ouabain binding sites, with a concomitant 3–4 fold increase in high affinity ouabain binding sites. The changes in protease activity and ouabain binding are associated with increased Na+/Ca2+ exchange, increased endocytosis, decreased Na+ conductance resulting in membrane hyperpolarization, increased 2-deoxy-D-glucose uptake and a sustained elevation of intracellular pH (pHi. Hyperpolarization is largely due to Na+-channel inactivation and is the main driving force for glucose uptake by the oocyte via Na+/glucose cotransport. The Na+ sym- and antiporter systems are driven by the Na+ free energy gradient generated by Na+/K+-ATPase. Shifts in α and/or β Na+-pump subunits to caveolar (lipid raft membrane regions may activate Na/K-ATPase and contribute to the Na+ free energy gradient and the increase in both Na+/glucose co-transport and pHi. Conclusions Under physiological conditions, resumption of meiosis results from the concerted action of insulin and progesterone at the cell membrane. Insulin inactivates Na+ channels and mobilizes fully functional Na+-pumps, generating a Na+ free energy gradient which serves as the energy source for several membrane anti- and symporter systems.

  18. p38 MAPK activation upregulates proinflammatory pathways in skeletal muscle cells from insulin-resistant type 2 diabetic patients

    DEFF Research Database (Denmark)

    Brown, Audrey E; Palsgaard, Jane; Borup, Rehannah

    2015-01-01

    Skeletal muscle is the key site of peripheral insulin resistance in type 2 diabetes. Insulin-stimulated glucose uptake is decreased in differentiated diabetic cultured myotubes, which is in keeping with a retained genetic/epigenetic defect of insulin action. We investigated differences in gene...... expression during differentiation between diabetic and control muscle cell cultures. Microarray analysis was performed using skeletal muscle cell cultures established from type 2 diabetic patients with a family history of type 2 diabetes and clinical evidence of marked insulin resistance and nondiabetic...... significantly, it did not improve insulin-stimulated glucose uptake. Increased cytokine expression driven by increased p38 MAPK activation is a key feature of cultured myotubes derived from insulin-resistant type 2 diabetic patients. p38 MAPK inhibition decreased cytokine expression but did not affect...

  19. The H+/K+ ATPase Inhibitor SCH-28080 Inhibits Insulin Secretion and Induces Cell Death in INS-1E Rat Insulinoma Cells

    Directory of Open Access Journals (Sweden)

    Martin Jakab

    2017-10-01

    Full Text Available Background/Aims: Glucose-stimulated insulin secretion (GSIS of pancreatic β-cells involves glucose uptake and metabolism, closure of KATP channels and depolarization of the cell membrane potential (Vmem, activation of voltage-activated Ca2+ currents (ICav and influx of Ca2+, which eventually triggers hormone exocytosis. Beside this classical pathway, KATP-independent mechanisms such as changes in intracellular pH (pHi or cell volume, which also affect β-cell viability, can elicit or modify insulin release. In β-cells the regulation of pHi is mainly accomplished by Na+/H+ exchangers (NHEs. To investigate if other proton extrusion mechanisms than NHEs are involved in pH regulation, we tested for the presence of the non-gastric H+/K+ ATPase in rat insulinoma cells and assessed effects of the H+/K+ ATPase inhibitor SCH-28080 on insulin secretion, cell viability and apoptosis. Methods: In INS-1E cell cultures, H+/K+ ATPase gene and protein expression was analyzed by reverse transcription PCR and Western blotting. Intracellular pH (pHi recovery after acute acidic load was measured by NH4Cl prepulsing using BCECF. Insulin secretion was determined by ELISA from the cell culture supernatant. Vmem, K+ and Ca2+ currents were recorded using patch clamp. Overall cell responses were determined using resazurin (viability and cytotoxicity assays. The mean cell volume (MCV, cell granularity (side-scatter; SSC, phosphatidylserine (PS exposure, cell membrane integrity, caspase activity and the mitochondrial membrane potential (ΔΨm were measured by flow cytometry. Results: We found that the α-subunit of the non-gastric H+/K+ ATPase (HKα2 is expressed on mRNA and protein level. However, compared to rat colon tissue, in INS-1E cells mRNA abundance was very low. In NH4Cl prepulsing experiments no K+-dependent pHi recovery was observed under Na+-free extracellular conditions. Nonetheless within 1 h, 20 µM SCH-28080 inhibited GSIS by ∼50%, while basal release

  20. Mitochondrial Dysfunction Contributes to Impaired Insulin Secretion in INS-1 Cells with Dominant-negative Mutations of HNF-1α and in HNF-1α-deficient Islets*

    Science.gov (United States)

    Pongratz, Rebecca L.; Kibbey, Richard G.; Kirkpatrick, Clare L.; Zhao, Xiaojian; Pontoglio, Marco; Yaniv, Moshe; Wollheim, Claes B.; Shulman, Gerald I.; Cline, Gary W.

    2009-01-01

    Maturity Onset Diabetes of the Young-type 3 (MODY-3) has been linked to mutations in the transcription factor hepatic nuclear factor (HNF)-1α, resulting in deficiency in glucose-stimulated insulin secretion. In INS-1 cells overexpressing doxycycline-inducible HNF-1α dominant-negative (DN-) gene mutations, and islets from Hnf-1α knock-out mice, insulin secretion was impaired in response to glucose (15 mm) and other nutrient secretagogues. Decreased rates of insulin secretion in response to glutamine plus leucine and to methyl pyruvate, but not potassium depolarization, indicate defects specific to mitochondrial metabolism. To identify the biochemical mechanisms responsible for impaired insulin secretion, we used 31P NMR measured mitochondrial ATP synthesis (distinct from glycolytic ATP synthesis) together with oxygen consumption measurements to determine the efficiency of mitochondrial oxidative phosphorylation. Mitochondrial uncoupling was significantly higher in DN-HNF-1α cells, such that rates of ATP synthesis were decreased by approximately one-half in response to the secretagogues glucose, glutamine plus leucine, or pyruvate. In addition to closure of the ATP-sensitive K+ channels with mitochondrial ATP synthesis, mitochondrial production of second messengers through increased anaplerotic flux has been shown to be critical for coupling metabolism to insulin secretion. 13C-Isotopomer analysis and tandem mass spectrometry measurement of Krebs cycle intermediates revealed a negative impact of DN-HNF-1α and Hnf-1α knock-out on mitochondrial second messenger production with glucose but not amino acids. Taken together, these results indicate that, in addition to reduced glycolytic flux, uncoupling of mitochondrial oxidative phosphorylation contributes to impaired nutrient-stimulated insulin secretion with either mutations or loss of HNF-1α. PMID:19376774

  1. Expression, purification, and functional characterization of the insulin-responsive facilitative glucose transporter GLUT4.

    Science.gov (United States)

    Kraft, Thomas E; Hresko, Richard C; Hruz, Paul W

    2015-12-01

    The insulin-responsive facilitative glucose transporter GLUT4 is of fundamental importance for maintenance of glucose homeostasis. Despite intensive effort, the ability to express and purify sufficient quantities of structurally and functionally intact protein for biophysical analysis has previously been exceedingly difficult. We report here the development of novel methods to express, purify, and functionally reconstitute GLUT4 into detergent micelles and proteoliposomes. Rat GLUT4 containing FLAG and His tags at the amino and carboxy termini, respectively, was engineered and stably transfected into HEK-293 cells. Overexpression in suspension culture yielded over 1.5 mg of protein per liter of culture. Systematic screening of detergent solubilized GLUT4-GFP fusion protein via fluorescent-detection size exclusion chromatography identified lauryl maltose neopentyl glycol (LMNG) as highly effective for isolating monomeric GLUT4 micelles. Preservation of structural integrity and ligand binding was demonstrated via quenching of tryptophan fluorescence and competition of ATB-BMPA photolabeling by cytochalasin B. GLUT4 was reconstituted into lipid nanodiscs and proper folding was confirmed. Reconstitution of purified GLUT4 with amphipol A8-35 stabilized the transporter at elevated temperatures for extended periods of time. Functional activity of purified GLUT4 was confirmed by reconstitution of LMNG-purified GLUT4 into proteoliposomes and measurement of saturable uptake of D-glucose over L-glucose. Taken together, these data validate the development of an efficient means to generate milligram quantities of stable and functionally intact GLUT4 that is suitable for a wide array of biochemical and biophysical analyses. © 2015 The Protein Society.

  2. Insulin-like peptide response to nutritional input in honey bee workers.

    Science.gov (United States)

    Ihle, Kate E; Baker, Nicholas A; Amdam, Gro V

    2014-10-01

    The rise in metabolic disorders in the past decades has heightened focus on achieving a healthy dietary balance in humans. This is also an increasingly important issue in the management of honey bees (Apis mellifera) where poor nutrition has negative effects on health and productivity in agriculture, and nutrition is suggested as a contributing factor in the recent global declines in honey bee populations. As in other organisms, the insulin/insulin-like signaling (IIS) pathway is likely involved in maintaining nutrient homeostasis in honey bees. Honey bees have two insulin-like peptides (Ilps) with differing spatial expression patterns in the fat body suggesting that AmIlp1 potentially functions in lipid metabolism while AmIlp2 is a more general indicator of nutritional status. We fed caged worker bees artificial diets high in carbohydrates, proteins or lipids and measured expression of AmIlp1, AmIlp2, and the insulin receptor substrate (IRS) to test their responses to dietary macronutrients. We also measured lifespan, worker weight and gustatory sensitivity to sugar as measures of individual physical condition. We found that expression of AmIlp1 was affected by diet composition and was highest on a diet high in protein. Expression of AmIlp2 and AmIRS were not affected by diet. Workers lived longest on a diet high in carbohydrates and low in protein and lipids. However, bees fed this diet weighed less than those that received a diet high in protein and low in carbohydrates and lipids. Bees fed the high carbohydrates diet were also more responsive to sugar, potentially indicating greater levels of hunger. These results support a role for AmIlp1 in nutritional homeostasis and provide new insight into how unbalanced diets impact individual honey bee health. Copyright © 2014 Elsevier Ltd. All rights reserved.

  3. Rates of insulin secretion in INS-1 cells are enhanced by coupling to anaplerosis and Kreb's cycle flux independent of ATP synthesis.

    Science.gov (United States)

    Cline, Gary W; Pongratz, Rebecca L; Zhao, Xiaojian; Papas, Klearchos K

    2011-11-11

    Mechanistic models of glucose stimulated insulin secretion (GSIS) established in minimal media in vitro, may not accurately describe the complexity of coupling metabolism with insulin secretion that occurs in vivo. As a first approximation, we have evaluated metabolic pathways in a typical growth media, DMEM as a surrogate in vivo medium, for comparison to metabolic fluxes observed under the typical experimental conditions using the simple salt-buffer of KRB. Changes in metabolism in response to glucose and amino acids and coupling to insulin secretion were measured in INS-1 832/13 cells. Media effects on mitochondrial function and the coupling efficiency of oxidative phosphorylation were determined by fluorometrically measured oxygen consumption rates (OCRs) combined with (31)P NMR measured rates of ATP synthesis. Substrate preferences and pathways into the TCA cycle, and the synthesis of mitochondrial 2nd messengers by anaplerosis were determined by (13)C NMR isotopomer analysis of the fate of [U-(13)C] glucose metabolism. Despite similar incremental increases in insulin secretion, the changes of OCR in response to increasing glucose from 2.5 to 15mM were blunted in DMEM relative to KRB. Basal and stimulated rates of insulin secretion rates were consistently higher in DMEM, while ATP synthesis rates were identical in both DMEM and KRB, suggesting greater mitochondrial uncoupling in DMEM. The relative rates of anaplerosis, and hence synthesis and export of 2nd messengers from the mitochondria were found to be similar in DMEM to those in KRB. And, the correlation of total PC flux with insulin secretion rates in DMEM was found to be congruous with the correlation in KRB. Together, these results suggest that signaling mechanisms associated with both TCA cycle flux and with anaplerotic flux, but not ATP production, may be responsible for the enhanced rates of insulin secretion in more complex, and physiologically-relevant media. Copyright © 2011 Elsevier Inc. All

  4. Activation of PPARd and RXRa stimulates fatty acid oxidatin and insulin secretion inpancreatic beta-cells

    DEFF Research Database (Denmark)

    Børgesen, Michael; Ravnskjær, Kim; Frigerio, Francesca

    as a central effector of unsaturated fatty acids in pancreatic ß-cells. Interestingly, activation of PPARd increases basal as well as glucose-stimulated insulin secretion of INS-1E cells. This increase is further potentiated by RXR agonists. This observation suggests that PPARd may mediate some of the positive......ACTIVATION OF PPARd AND RXRa STIMULATES FATTY ACID OXIDATION AND INSULIN SECRETION IN PANCREATIC b-CELLS Michael Boergesen1, Kim Ravnskjaer2, Francesca Frigerio3, Allan E. Karlsen4, Pierre Maechler3 and Susanne Mandrup1 1 Department of Biochemistry and Molecular Biology, University of Southern...... of genes as PPARd specific agonists and stimulates ß-oxidation. Importantly, oleate-induction of gene expression and ß-oxidation in INS-1E cells is abolished by knock-down of PPARd using adenoviral transfer of shRNA. Thus, PPARd appears to be a central regulator of fatty acid metabolism as well...

  5. Periodontitis induced by Porphyromonas gingivalis drives periodontal microbiota dysbiosis and insulin resistance via an impaired adaptive immune response.

    Science.gov (United States)

    Blasco-Baque, Vincent; Garidou, Lucile; Pomié, Céline; Escoula, Quentin; Loubieres, Pascale; Le Gall-David, Sandrine; Lemaitre, Mathieu; Nicolas, Simon; Klopp, Pascale; Waget, Aurélie; Azalbert, Vincent; Colom, André; Bonnaure-Mallet, Martine; Kemoun, Philippe; Serino, Matteo; Burcelin, Rémy

    2017-05-01

    To identify a causal mechanism responsible for the enhancement of insulin resistance and hyperglycaemia following periodontitis in mice fed a fat-enriched diet. We set-up a unique animal model of periodontitis in C57Bl/6 female mice by infecting the periodontal tissue with specific and alive pathogens like Porphyromonas gingivalis ( Pg ), Fusobacterium nucleatum and Prevotella intermedia . The mice were then fed with a diabetogenic/non-obesogenic fat-enriched diet for up to 3 months. Alveolar bone loss, periodontal microbiota dysbiosis and features of glucose metabolism were quantified. Eventually, adoptive transfer of cervical (regional) and systemic immune cells was performed to demonstrate the causal role of the cervical immune system. Periodontitis induced a periodontal microbiota dysbiosis without mainly affecting gut microbiota. The disease concomitantly impacted on the regional and systemic immune response impairing glucose metabolism. The transfer of cervical lymph-node cells from infected mice to naive recipients guarded against periodontitis-aggravated metabolic disease. A treatment with inactivated Pg prior to the periodontal infection induced specific antibodies against Pg and protected the mouse from periodontitis-induced dysmetabolism. Finally, a 1-month subcutaneous chronic infusion of low rates of lipopolysaccharides from Pg mimicked the impact of periodontitis on immune and metabolic parameters. We identified that insulin resistance in the high-fat fed mouse is enhanced by pathogen-induced periodontitis. This is caused by an adaptive immune response specifically directed against pathogens and associated with a periodontal dysbiosis. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/.

  6. Insulin regulates Glut4 confinement in plasma membrane clusters in adipose cells.

    Science.gov (United States)

    Lizunov, Vladimir A; Stenkula, Karin; Troy, Aaron; Cushman, Samuel W; Zimmerberg, Joshua

    2013-01-01

    Insulin-stimulated delivery of glucose transporter-4 (GLUT4) to the plasma membrane (PM) is the hallmark of glucose metabolism. In this study we examined insulin's effects on GLUT4 organization in PM of adipose cells by direct microscopic observation of single monomers tagged with photoswitchable fluorescent protein. In the basal state, after exocytotic delivery only a fraction of GLUT4 is dispersed into the PM as monomers, while most of the GLUT4 stays at the site of fusion and forms elongated clusters (60-240 nm). GLUT4 monomers outside clusters diffuse freely and do not aggregate with other monomers. In contrast, GLUT4 molecule collision with an existing cluster can lead to immediate confinement and association with that cluster. Insulin has three effects: it shifts the fraction of dispersed GLUT4 upon delivery, it augments the dissociation of GLUT4 monomers from clusters ∼3-fold and it decreases the rate of endocytic uptake. All together these three effects of insulin shift most of the PM GLUT4 from clustered to dispersed states. GLUT4 confinement in clusters represents a novel kinetic mechanism for insulin regulation of glucose homeostasis.

  7. Insulin regulates Glut4 confinement in plasma membrane clusters in adipose cells.

    Directory of Open Access Journals (Sweden)

    Vladimir A Lizunov

    Full Text Available Insulin-stimulated delivery of glucose transporter-4 (GLUT4 to the plasma membrane (PM is the hallmark of glucose metabolism. In this study we examined insulin's effects on GLUT4 organization in PM of adipose cells by direct microscopic observation of single monomers tagged with photoswitchable fluorescent protein. In the basal state, after exocytotic delivery only a fraction of GLUT4 is dispersed into the PM as monomers, while most of the GLUT4 stays at the site of fusion and forms elongated clusters (60-240 nm. GLUT4 monomers outside clusters diffuse freely and do not aggregate with other monomers. In contrast, GLUT4 molecule collision with an existing cluster can lead to immediate confinement and association with that cluster. Insulin has three effects: it shifts the fraction of dispersed GLUT4 upon delivery, it augments the dissociation of GLUT4 monomers from clusters ∼3-fold and it decreases the rate of endocytic uptake. All together these three effects of insulin shift most of the PM GLUT4 from clustered to dispersed states. GLUT4 confinement in clusters represents a novel kinetic mechanism for insulin regulation of glucose homeostasis.

  8. Dual role of proapoptotic BAD in insulin secretion and beta cell survival

    OpenAIRE

    Danial, Nika N.; Walensky, Loren D.; Zhang, Chen-Yu; Choi, Cheol Soo; Fisher, Jill K.; Molina, Anthony J. A.; Datta, Sandeep Robert; Pitter, Kenneth L.; Bird, Gregory H.; Wikstrom, Jakob D.; Deeney, Jude T.; Robertson, Kirsten; Morash, Joel; Kulkarni, Ameya; Neschen, Susanne

    2008-01-01

    The proapoptotic BCL-2 family member BAD resides in a glucokinase-containing complex that regulates glucose-driven mitochondrial respiration. Here, we present genetic evidence of a physiologic role for BAD in glucose-stimulated insulin secretion by beta cells. This novel function of BAD is specifically dependent upon the phosphorylation of its BH3 sequence, previously defined as an essential death domain. We highlight the pharmacologic relevance of phosphorylated BAD BH3 by using cell-permeab...

  9. Mitochondrial oxidative stress contributes differently to rat pancreatic islet cell apoptosis and insulin secretory defects after prolonged culture in a low non-stimulating glucose concentration.

    Science.gov (United States)

    Roma, L P; Pascal, S M; Duprez, J; Jonas, J-C

    2012-08-01

    Pancreatic beta cells chronically exposed to low glucose concentrations show signs of oxidative stress, loss of glucose-stimulated insulin secretion (GSIS) and increased apoptosis. Our aim was to confirm the role of mitochondrial oxidative stress in rat islet cell apoptosis under these culture conditions and to evaluate whether its reduction similarly improves survival and GSIS. Apoptosis, oxidative stress-response gene mRNA expression and glucose-induced stimulation of mitochondrial metabolism, intracellular Ca(2+) concentration and insulin secretion were measured in male Wistar rat islets cultured for 1 week in RPMI medium containing 5-10 mmol/l glucose with or without manganese(III)tetrakis(4-benzoic acid)porphyrin (MnTBAP) or N-acetyl-L-: cysteine (NAC). Oxidative stress was measured in islet cell clusters cultured under similar conditions using cytosolic and mitochondrial redox-sensitive green fluorescent protein (roGFP1/mt-roGFP1). Prolonged culture in 5 vs 10 mmol/l glucose increased mt-roGFP1 (but not roGFP1) oxidation followed by beta cell apoptosis and loss of GSIS resulting from reduced insulin content, mitochondrial metabolism, Ca(2+) influx and Ca(2+)-induced secretion. Tolbutamide-induced, but not high K(+)-induced, Ca(2+) influx was also suppressed. Under these conditions, MnTBAP, but not NAC, triggered parallel ~50-70% reductions in mt-roGFP1 oxidation and beta cell apoptosis, but failed to protect against the loss of GSIS despite significant improvement in glucose-induced and tolbutamide-induced Ca(2+) influx. Mitochondrial oxidative stress contributes differently to rat pancreatic islet cell apoptosis and insulin secretory defects during culture in a low glucose concentration. Thus, targeting beta cell survival may not be sufficient to restore insulin secretion when beta cells suffer from prolonged mitochondrial oxidative stress, e.g. in the context of reduced glucose metabolism.

  10. Interleukin-33-Activated Islet-Resident Innate Lymphoid Cells Promote Insulin Secretion through Myeloid Cell Retinoic Acid Production.

    Science.gov (United States)

    Dalmas, Elise; Lehmann, Frank M; Dror, Erez; Wueest, Stephan; Thienel, Constanze; Borsigova, Marcela; Stawiski, Marc; Traunecker, Emmanuel; Lucchini, Fabrizio C; Dapito, Dianne H; Kallert, Sandra M; Guigas, Bruno; Pattou, Francois; Kerr-Conte, Julie; Maechler, Pierre; Girard, Jean-Philippe; Konrad, Daniel; Wolfrum, Christian; Böni-Schnetzler, Marianne; Finke, Daniela; Donath, Marc Y

    2017-11-21

    Pancreatic-islet inflammation contributes to the failure of β cell insulin secretion during obesity and type 2 diabetes. However, little is known about the nature and function of resident immune cells in this context or in homeostasis. Here we show that interleukin (IL)-33 was produced by islet mesenchymal cells and enhanced by a diabetes milieu (glucose, IL-1β, and palmitate). IL-33 promoted β cell function through islet-resident group 2 innate lymphoid cells (ILC2s) that elicited retinoic acid (RA)-producing capacities in macrophages and dendritic cells via the secretion of IL-13 and colony-stimulating factor 2. In turn, local RA signaled to the β cells to increase insulin secretion. This IL-33-ILC2 axis was activated after acute β cell stress but was defective during chronic obesity. Accordingly, IL-33 injections rescued islet function in obese mice. Our findings provide evidence that an immunometabolic crosstalk between islet-derived IL-33, ILC2s, and myeloid cells fosters insulin secretion. Copyright © 2017 Elsevier Inc. All rights reserved.

  11. Restitution of defective glucose-stimulated insulin secretion in diabetic GK rat by acetylcholine uncovers paradoxical stimulatory effect of beta-cell muscarinic receptor activation on cAMP production.

    Science.gov (United States)

    Dolz, Manuel; Bailbé, Danielle; Giroix, Marie-Hélène; Calderari, Sophie; Gangnerau, Marie-Noelle; Serradas, Patricia; Rickenbach, Katharina; Irminger, Jean-Claude; Portha, Bernard

    2005-11-01

    Because acetylcholine (ACh) is a recognized potentiator of glucose-stimulated insulin release in the normal beta-cell, we have studied ACh's effect on islets of the Goto-Kakizaki (GK) rat, a spontaneous model of type 2 diabetes. We first verified that ACh was able to restore the insulin secretory glucose competence of the GK beta-cell. Then, we demonstrated that in GK islets 1) ACh elicited a first-phase insulin release at low glucose, whereas it had no effect in Wistar; 2) total phospholipase C activity, ACh-induced inositol phosphate production, and intracellular free calcium concentration ([Ca2+]i) elevation were normal; 3) ACh triggered insulin release, even in the presence of thapsigargin, which induced a reduction of the ACh-induced [Ca2+]i response (suggesting that ACh produces amplification signals that augment the efficacy of elevated [Ca2+]i on GK exocytosis); 4) inhibition of protein kinase C did not affect [Ca2+]i nor the insulin release responses to ACh; and 5) inhibition of cAMP-dependent protein kinases (PKAs), adenylyl cyclases, or cAMP generation, while not affecting the [Ca2+]i response, significantly lowered the insulinotropic response to ACh (at low and high glucose). In conclusion, ACh acts mainly through activation of the cAMP/PKA pathway to potently enhance Ca2+-stimulated insulin release in the GK beta-cell and, in doing so, normalizes its defective glucose responsiveness.

  12. Akt kinase C-terminal modifications control activation loop dephosphorylation and enhance insulin response.

    Science.gov (United States)

    Chan, Tung O; Zhang, Jin; Tiegs, Brian C; Blumhof, Brian; Yan, Linda; Keny, Nikhil; Penny, Morgan; Li, Xue; Pascal, John M; Armen, Roger S; Rodeck, Ulrich; Penn, Raymond B

    2015-10-01

    The Akt protein kinase, also known as protein kinase B, plays key roles in insulin receptor signalling and regulates cell growth, survival and metabolism. Recently, we described a mechanism to enhance Akt phosphorylation that restricts access of cellular phosphatases to the Akt activation loop (Thr(308) in Akt1 or protein kinase B isoform alpha) in an ATP-dependent manner. In the present paper, we describe a distinct mechanism to control Thr(308) dephosphorylation and thus Akt deactivation that depends on intramolecular interactions of Akt C-terminal sequences with its kinase domain. Modifications of amino acids surrounding the Akt1 C-terminal mTORC2 (mammalian target of rapamycin complex 2) phosphorylation site (Ser(473)) increased phosphatase resistance of the phosphorylated activation loop (pThr(308)) and amplified Akt phosphorylation. Furthermore, the phosphatase-resistant Akt was refractory to ceramide-dependent dephosphorylation and amplified insulin-dependent Thr(308) phosphorylation in a regulated fashion. Collectively, these results suggest that the Akt C-terminal hydrophobic groove is a target for the development of agents that enhance Akt phosphorylation by insulin. © 2015 Authors; published by Portland Press Limited.

  13. Differentiation of PDX1 gene-modified human umbilical cord mesenchymal stem cells into insulin-producing cells in vitro.

    Science.gov (United States)

    He, Dongmei; Wang, Juan; Gao, Yangjun; Zhang, Yuan

    2011-12-01

    Mesenchymal stem cells (MSCs) have significant advantages over other stem cell types, and greater potential for immediate clinical application. MSCs would be an interesting cellular source for treatment of type 1 diabetes. In this study, MSCs from human umbilical cord were differentiated into functional insulin-producing cells in vitro by introduction of the pancreatic and duodenal homeobox factor 1 (PDX1) and in the presence of induction factors. The expressions of cell surface antigens were detected by flow cytometry. After induction in an adipogenic medium or an osteogenic medium, the cells were observed by Oil Red O staining and alkaline phosphatase staining. Recombinant adenovirus carrying the PDX1 gene was constructed and MSCs were infected by the recombinant adenovirus, then treated with several inducing factors for differentiation into islet β-like cells. The expression of the genes and protein related to islet β-cells was detected by immunocytochemistry, RT-PCR and Western blot analysis. Insulin and C-peptide secretion were assayed. Our results show that the morphology and immunophenotype of MSCs from human umbilical cord were similar to those present in human bone marrow. The MSCs could be induced to differentiate into osteocytes and adipocytes. After induction by recombined adenovirus vector with induction factors, MSCs were aggregated and presented islet-like bodies. Dithizone staining of these cells was positive. The genes' expression related to islet β-cells was found. After induction, insulin and C-peptide secretion in the supernatant were significantly increased. In conclusion, our results demonstrated that PDX1 gene-modified human umbilical cord mesenchymal stem cells could be differentiated into insulin-producing cells in vitro.

  14. Role of pancreatic polypeptide as a market of transplanted insulin-producing fetal pig cells.

    Science.gov (United States)

    Tuch, B E; Tabiin, M T; Casamento, F M; Yao, M; Georges, P; Amaratunga, A; Pinto, A N

    2001-01-01

    Transplantation of insulin-producing fetal pancreatic tissue into diabetic recipients has been shown to normalize blood glucose levels after several months. This time period is required for the growth and maturation of the fetal tissue so insulin levels cannot be used as a marker of graft function while the beta-cell is immature. Therefore, we have examined the use of another pancreatic endocrine hormone, pancreatic polypeptide (PP), to monitor graft function. The cell that produces this hormone has been shown to be the first mature endocrine cell in the fetal pancreas. Fetal pig pancreatic tissue, both in the form of 1 mm3 explants and islet-like cell clusters (ICCs), was transplanted into immunodeficient SCID mice and the levels of PP and insulin were measured in plasma and in the graft for up to 12 weeks. PP was detected in the untransplanted explants (0.58 pmol/mg) and ICCs (0.06 pmol/ICC) and the PP to insulin ratio was 2.7% and 5.8%, respectively. PP (but not porcine C-peptide, a marker of insulin secretion) was detectable in the plasma of SCID mice from 4 days to 3 weeks after transplantation, but not thereafter. The highest values were obtained at 4 days to 1 week. In the grafted tissue PP and insulin were present at all time points and the ratio of PP to insulin was 59%, 87%, 75%, 56%, 7%, 8%, and 7% at 4 days, 1, 2, 3, 6, 9, and 12 weeks, respectively. The decline in PP levels 3 weeks after transplantation was associated with beta-cell development in the graft. PP was also secreted by fetal pig pancreatic explants transplanted into diabetic NOD/SCID mice, with plasma levels measurable in the first week after the tissue was grafted. In immunocompetent BALB/c mice transplanted with the tissue, PP was detectable in plasma for 2 days after transplantation but not at 4 days, when cellular rejection commenced, or thereafter. We conclude that plasma PP levels can be used as a marker of the viability of fetal porcine pancreatic tissue in the first 3 weeks after

  15. Effects of metformin hydrochloride on blood glucose and insulin responses to oral dextrose in horses.

    Science.gov (United States)

    Rendle, D I; Rutledge, F; Hughes, K J; Heller, J; Durham, A E

    2013-11-01

    Metformin is a potential therapeutic agent for the treatment of insulin resistance (IR). In laboratory animals, orally administered metformin reduces intestinal glucose absorption and may therefore affect insulinaemic responses to oral carbohydrate ingestion. To determine whether pretreatment with metformin reduces plasma glucose concentration and insulin responses following consumption of dextrose in horses. Therapeutic cross-over study. Seven healthy Standardbred and Thoroughbred geldings were subjected to an oral dextrose challenge test on 4 occasions: with and without metformin, before and after induction of IR with dexamethasone. Metformin was administered by nasogastric tube at 30 mg/kg bwt 1 h before administration of dextrose. Glucose and insulin concentrations in plasma/serum were measured at regular intervals during each test. Linear mixed models were specified for each predetermined outcome variable, and for each model the 'treatment' was included as a fixed effect with 4 categorical levels (none, metformin, dexamethasone and dexamethasone with metformin) and horse accounted for as a random effect. In healthy horses, the administration of metformin resulted in a statistically significant reduction in peak glucose concentration (P = 0.002), area under the glucose curve (Pdextrose administration (P = 0.011). Following the induction of IR, administration of metformin was associated with significant differences in peak glucose concentration (Pdextrose administration (P = 0.014). Metformin resulted in reduced glycaemic and insulinaemic responses both in healthy horses and in horses with experimentally induced IR. Metformin may benefit horses with naturally acquired IR by reducing glycaemic and insulinaemic responses to dietary nonstructural carbohydrates. Further investigations into the mechanisms of action of metformin in horses and controlled clinical trials are warranted. © 2013 EVJ Ltd.

  16. Insulin-Independent GABAA Receptor-Mediated Response in the Barrel Cortex of Mice with Impaired Met Activity.

    Science.gov (United States)

    Lo, Fu-Sun; Erzurumlu, Reha S; Powell, Elizabeth M

    2016-03-30

    Autism spectrum disorder (ASD) is a neurodevelopmental disorder caused by genetic variants, susceptibility alleles, and environmental perturbations. The autism associated geneMETtyrosine kinase has been implicated in many behavioral domains and endophenotypes of autism, including abnormal neural signaling in human sensory cortex. We investigated somatosensory thalamocortical synaptic communication in mice deficient in Met activity in cortical excitatory neurons to gain insights into aberrant somatosensation characteristic of ASD. The ratio of excitation to inhibition is dramatically increased due to decreased postsynaptic GABAAreceptor-mediated inhibition in the trigeminal thalamocortical pathway of mice lacking active Met in the cerebral cortex. Furthermore, in contrast to wild-type mice, insulin failed to increase GABAAreceptor-mediated response in the barrel cortex of mice with compromised Met signaling. Thus, lacking insulin effects may be a risk factor in ASD pathogenesis. A proposed common cause of neurodevelopmental disorders is an imbalance in excitatory neural transmission, provided by the glutamatergic neurons, and the inhibitory signals from the GABAergic interneurons. Many genes associated with autism spectrum disorders impair synaptic transmission in the expected cell type. Previously, inactivation of the autism-associated Met tyrosine kinase receptor in GABAergic interneurons led to decreased inhibition. In thus report, decreased Met signaling in glutamatergic neurons had no effect on excitation, but decimated inhibition. Further experiments indicate that loss of Met activity downregulates GABAAreceptors on glutamatergic neurons in an insulin independent manner. These data provide a new mechanism for the loss of inhibition and subsequent abnormal excitation/inhibition balance and potential molecular candidates for treatment or prevention. Copyright © 2016 the authors 0270-6474/16/363691-07$15.00/0.

  17. Effect of cooling heat-stressed dairy cows during the dry period on insulin response.

    Science.gov (United States)

    Tao, S; Thompson, I M; Monteiro, A P A; Hayen, M J; Young, L J; Dahl, G E

    2012-09-01

    affect the insulin responses to GTT and IC during the transition period and glucose responses to GTT and IC at -14 and 28 DRC were not affected by treatments. At 7 DRC, CL cows tended to have slower glucose clearance to GTT and weaker glucose response to IC relative to HT cows. Cows from the cooling treatment had stronger nonesterified fatty acid responses to IC postpartum but not prepartum compared with HT. In conclusion, cooling heat-stressed dairy cows in the dry period reduced insulin effects on peripheral tissues in early lactation but not in the dry period. Copyright © 2012 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

  18. Insulin-like growth factor I has independent effects on bone matrix formation and cell replication

    International Nuclear Information System (INIS)

    Hock, J.M.; Centrella, M.; Canalis, E.

    1988-01-01

    The effects of insulin-like growth factor-I (IGF-I) and insulin on bone matrix synthesis and bone cell replication were studied in cultured 21-day-old fetal rat calvariae. Histomorphometry techniques were developed to measure the incorporation of [2,3- 3 H]proline and [methyl- 3 H]thymidine into bone matrix and bone cell nuclei, respectively, using autoradiographs of sagittal sections of calvariae cultured with IGF-I, insulin, or vehicle for up to 96 h. To confirm an effect on bone formation, IGF-I was also studied for its effects on [ 3 H]proline incorporation into collagenase-digestible protein (CDP) and noncollagen protein and on [ 3 H]thymidine incorporation into acid-precipitable material (DNA). IGF-I at 10(-9)-10(-7) M significantly increased the rate of bone matrix apposition and CDP after 24 h by 45-50% and increased cell labeling by 8-fold in the osteoprogenitor cell zone, by 4-fold in the osteoblast cell zone, and by 2-fold in the periosteal fibroblast zone. Insulin at 10(-9)-10(-6) M also increased matrix apposition rate and CDP by 40-50%, but increased cell labeling by 2-fold only at a concentration of 10(-7) M or higher and then only in the osteoprogenitor cell zone. When hydroxyurea was added to IGF-I-treated bones, the effects of IGF-I on DNA synthesis were abolished, but the increase in bone matrix apposition induced by IGF-I was only partly diminished. In conclusion, IGF-I stimulates matrix synthesis in calvariae, an effect that is partly, although not completely, dependent on its stimulatory effect on DNA synthesis

  19. Interrelations between glucose-induced insulin response, metabolic indicators, and time of first ovulation in high-yielding dairy cows.

    Science.gov (United States)

    Bossaert, P; Leroy, J L M R; De Vliegher, S; Opsomer, G

    2008-09-01

    High-yielding dairy cows are more susceptible to metabolic and reproductive disorders than low-yielding cows. Insulin plays a pivotal role in the development of both problems. In the present study, we aimed to assess the glucose-induced insulin responses of dairy cows at different time points relative to calving and to relate this to the metabolic status and the time of first ovulation. Twenty-three healthy, multiparous Holstein-Friesian cows with a high genetic merit for milk yield were studied from 14 d prepartum to 42 d postpartum. Intravenous glucose tolerance tests were performed on -14, 14, and 42 d relative to calving to evaluate the plasma insulin and glucose responses to a glucose load, as estimated by the peak concentration, the area under the curve (AUC), and the clearance rates of insulin and glucose. Blood samples were obtained at 3-d intervals and analyzed for glucose, insulin, and nonesterified fatty acids (NEFA). The time of first ovulation was defined by transrectal ultrasonography and plasma progesterone analysis. Glucose-induced insulin AUC and peak concentration decreased and glucose clearance increased during lactation compared with the dry period. Plasma NEFA concentrations were negatively related to insulin AUC and peak concentrations. Fourteen cows ovulated within 42 d postpartum, and the remaining 9 cows suffered from delayed resumption of ovarian function. Survival analysis demonstrated that cows with lower NEFA concentrations during the dry period tended to have earlier resumption of ovarian activity. In conclusion, our data suggest a decreased plasma insulin response to glucose postpartum in high-yielding dairy cows, possibly contributing to metabolic stress during the early postpartum period. It is hypothesized that NEFA impair glucose-induced insulin secretion in dairy cows. Additionally, our results suggest the importance of lipolysis during the transition period as a risk factor for delayed ovulation.

  20. Insulin-like growth factor-1 signaling in renal cell carcinoma

    International Nuclear Information System (INIS)

    Tracz, Adam F.; Szczylik, Cezary; Porta, Camillo; Czarnecka, Anna M.

    2016-01-01

    Renal cell carcinoma (RCC) incidence is highest in highly developed countries and it is the seventh most common neoplasm diagnosed. RCC management include nephrectomy and targeted therapies. Type 1 insulin-like growth factor (IGF-1) pathway plays an important role in cell proliferation and apoptosis resistance. IGF-1 and insulin share overlapping downstream signaling pathways in normal and cancer cells. IGF-1 receptor (IGF1R) stimulation may promote malignant transformation promoting cell proliferation, dedifferentiation and inhibiting apoptosis. Clear cell renal cell carcinoma (ccRCC) patients with IGF1R overexpression have 70 % increased risk of death compared to patients who had tumors without IGF1R expression. IGF1R signaling deregulation may results in p53, WT, BRCA1, VHL loss of function. RCC cells with high expression of IGF1R are more resistant to chemotherapy than cells with low expression. Silencing of IGF1R increase the chemosensitivity of ccRCC cells and the effect is greater in VHL mutated cells. Understanding the role of IGF-1 signaling pathway in RCC may result in development of new targeted therapeutic interventions. First preclinical attempts with anti-IGF-1R monoclonal antibodies or fragment antigen-binding (Fab) fragments alone or in combination with an mTOR inhibitor were shown to inhibit in vitro growth and reduced the number of colonies formed by of RCC cells

  1. Effect of Insulin Therapy using Hyper-insulinemic Normoglycemic Clamp on Inflammatory Response in Brain Dead Organ Donors.

    Science.gov (United States)

    Aljiffry, M; Hassanain, M; Schricker, T; Shaheen, M; Nouh, T; Lattermann, R; Salman, A; Wykes, L; Metrakos, P

    2016-05-01

    Brain death is a major stress that is associated with a massive inflammatory response and systemic hyperglycemia. Severe inflammation leads to increased graft immunogenicity and risk of graft dysfunction; while acute hyperglycemia aggravates the inflammatory response and increases the risk of morbidity and mortality. Insulin therapy not only controls hyperglycemia but also suppresses inflammation. The present study is to investigate the anti-inflammatory properties and the normoglycemia maintenance of high dose insulin on brain dead organ donors. 15 brain dead organ donors were divided into 2 groups, insulin treated (n=6) and controls (n=9). Insulin was provided for a minimum of 6 h using the hyperinsulinemic normoglycemic clamp technique. The changes of serum cytokines, including IL-6, IL-10, IL-1β, IL-8, TNFα, TGFα and MCP-1, were measured by suspension bead array immunoassay and glucose by a glucose monitor. Compared to controls, insulin treated donors had a significant lower blood glucose 4.8 (4-6.9) vs. 9 (5.6-11.7) mmol/L, pinsulin treated donors compared with those in controls. High dose insulin therapy decreases the concentrations of inflammatory cytokines in brain dead donors and preserves normoglycemia. High dose of insulin may have anti-inflammatory effects in brain dead organ donors and therefore, improve the quality of donor organs and potentially improve outcomes. © Georg Thieme Verlag KG Stuttgart · New York.

  2. Insulin signaling displayed a differential tissue-specific response to low-dose dihydrotestosterone in female mice.

    Science.gov (United States)

    Andrisse, Stanley; Billings, Katelyn; Xue, Ping; Wu, Sheng

    2018-04-01

    Hyperandrogenemia and hyperinsulinemia are believed to play prominent roles in polycystic ovarian syndrome (PCOS). We explored the effects of low-dose dihydrotestosterone (DHT), a model of PCOS, on insulin signaling in metabolic and reproductive tissues in a female mouse model. Insulin resistance in the energy storage tissues is associated with type 2 diabetes. Insulin signaling in the ovaries and pituitary either directly or indirectly stimulates androgen production. Energy storage and reproductive tissues were isolated and molecular assays were performed. Livers and white adipose tissue (WAT) from DHT mice displayed lower mRNA and protein expression of insulin signaling intermediates. However, ovaries and pituitaries of DHT mice exhibited higher expression levels of insulin signaling genes/proteins. Insulin-stimulated p-AKT levels were blunted in the livers and WAT of the DHT mice but increased or remained the same in the ovaries and pituitaries compared with controls. Glucose uptake decreased in liver and WAT but was unchanged in pituitary and ovary of DHT mice. Plasma membrane GLUTs were decreased in liver and WAT but increased in ovary and pituitary of DHT mice. Skeletal muscle insulin-signaling genes were not lowered in DHT mice compared with control. DHT mice did not display skeletal muscle insulin resistance. Insulin-stimulated glucose transport increased in skeletal muscles of DHT mice compared with controls. DHT mice were hyperinsulinemic. However, the differential mRNA and protein expression pattern was independent of hyperinsulinemia in cultured hepatocytes and pituitary cells. These findings demonstrate a differential effect of DHT on the insulin-signaling pathway in energy storage vs. reproductive tissues independent of hyperinsulinemia.

  3. Insulin: its binding to specific receptors and its stimulation of DNA synthesis and 2',3'-cyclic nucleotide phosphohydrolase in embryonic mouse brain cell cultures

    International Nuclear Information System (INIS)

    Shanker, G.; Pieringer, R.A.

    1986-01-01

    Previously, the authors demonstrated that ornithine decarboxylase was stimulated by insulin in cultures of embryonic mouse brain cells. In the present work, they have investigated the presence and specificity of insulin receptors in these cultures. A time study showed that maximum binding of 125 [I] labelled insulin was around 75 min. Other studies measured the influence of concentration and age on insulin binding. A displacement study using increasing concentrations of cold insulin, glucagon or growth hormone demonstrated that the specificity of the receptors for insulin was rather high. It was also found that insulin displayed a clear dose-dependent stimulation of thymidine incorporation into the brain cells. Insulin also stimulated the glial enzyme 2':3'-cyclic nucleotide phosphohydrolase (CNP-ase). The results suggest a dual role for insulin; it regulates both cell proliferation as well as differentiation

  4. The effect of insulin on amino acid incorporation into exocrine pancreatic cells of the rat

    International Nuclear Information System (INIS)

    Kramer, M.F.; Poort, C.

    1975-01-01

    The rate of incorporation of radioactive leucine per cell in the acinar pancreatic cells of the rat increases by 50 per cent within one hour after subcutaneous administration of insulin, an effect that lasts for at least one more hour. The rate of incorporation has been measured by quantitative radioautography and by determination of the radioactivity per μg DNA in TCA-precipitable material from tissue homogenates. The capacity for amino acid (leucine and lysine) incorporation as measured by incubating pancreatic fragments in vitro is not enhanced by insulin treatment of the rat in vivo during one or more hours. Insulin was found to lower the serum concentration of most amino acids significantly, leucine by 50 per cent. The apparent effect of insulin on the incorporation of radioactive leucine in vivo can be explained by the difference in the specific radioactivity of the circulating amino acid in the treated rats as compared to the untreated ones. A change in amino acid concentration in the serum may likewise be the explanation of the decrease in amino acid incorporation rate in alloxan diabetic rats. (orig./GSE) [de

  5. Brain delivery of insulin boosted by intranasal coadministration with cell-penetrating peptides.

    Science.gov (United States)

    Kamei, Noriyasu; Takeda-Morishita, Mariko

    2015-01-10

    Intranasal administration is considered as an alternative route to enable effective drug delivery to the central nervous system (CNS) by bypassing the blood-brain barrier. Several reports have proved that macromolecules can be transferred directly from the nasal cavity to the brain. However, strategies to enhance the delivery of macromolecules from the nasal cavity to CNS are needed because of their low delivery efficiencies via this route in general. We hypothesized that the delivery of biopharmaceuticals to the brain parenchyma can be facilitated by increasing the uptake of drugs by the nasal epithelium including supporting and neuronal cells to maximize the potentiality of the intranasal pathway. To test this hypothesis, the CNS-related model peptide insulin was intranasally coadministered with the cell-penetrating peptide (CPP) penetratin to mice. As a result, insulin coadministered with l- or d-penetratin reached the distal regions of the brain from the nasal cavity, including the cerebral cortex, cerebellum, and brain stem. In particular, d-penetratin could intranasally deliver insulin to the brain with a reduced risk of systemic insulin exposure. Thus, the results obtained in this study suggested that CPPs are potential tools for the brain delivery of peptide- and protein-based pharmaceuticals via intranasal administration. Copyright © 2014 Elsevier B.V. All rights reserved.

  6. Relaxation response induces temporal transcriptome changes in energy metabolism, insulin secretion and inflammatory pathways.

    Directory of Open Access Journals (Sweden)

    Manoj K Bhasin

    Full Text Available The relaxation response (RR is the counterpart of the stress response. Millennia-old practices evoking the RR include meditation, yoga and repetitive prayer. Although RR elicitation is an effective therapeutic intervention that counteracts the adverse clinical effects of stress in disorders including hypertension, anxiety, insomnia and aging, the underlying molecular mechanisms that explain these clinical benefits remain undetermined. To assess rapid time-dependent (temporal genomic changes during one session of RR practice among healthy practitioners with years of RR practice and also in novices before and after 8 weeks of RR training, we measured the transcriptome in peripheral blood prior to, immediately after, and 15 minutes after listening to an RR-eliciting or a health education CD. Both short-term and long-term practitioners evoked significant temporal gene expression changes with greater significance in the latter as compared to novices. RR practice enhanced expression of genes associated with energy metabolism, mitochondrial function, insulin secretion and telomere maintenance, and reduced expression of genes linked to inflammatory response and stress-related pathways. Interactive network analyses of RR-affected pathways identified mitochondrial ATP synthase and insulin (INS as top upregulated critical molecules (focus hubs and NF-κB pathway genes as top downregulated focus hubs. Our results for the first time indicate that RR elicitation, particularly after long-term practice, may evoke its downstream health benefits by improving mitochondrial energy production and utilization and thus promoting mitochondrial resiliency through upregulation of ATPase and insulin function. Mitochondrial resiliency might also be promoted by RR-induced downregulation of NF-κB-associated upstream and downstream targets that mitigates stress.

  7. High-level expression of human insulin receptor cDNA in mouse NIH 3T3 cells

    International Nuclear Information System (INIS)

    Whittaker, J.; Okamoto, A.K.; Thys, R.; Bell, G.I.; Steiner, D.F.; Hofmann, C.A.

    1987-01-01

    In order to develop a simple, efficient system for the high-level expression of human insulin receptors in eukaryotic cells, a full-length human kidney insulin receptor cDNA was inserted into a bovine papilloma virus vector under the control of the mouse metallothionein promoter. After transfection of mouse NIH 3T3 cells with this construct, seven cell lines expressing insulin receptors were isolated; two cell lines had more than 10 6 receptors per cell. The cell line with the highest 125 I-insulin binding (NIH 3T3 HIR3.5) had 6 x 10 6 receptors with a K/sub d/ of 10 -9 M. This level was not dependent on exposure to metals but could be increased further to 2 x 10 7 receptors per cell by addition of sodium butyrate to the culture medium. The α and β subunits had apparent molecular weights of 147,000 and 105,000, respectively (compared to 135,000 and 95,000 in IM-9 human lymphocytes), values identical to those of the α and β subunits of the insulin receptors of nontransformed NIH 3T3 cells. This size difference was due to altered carbohydrate composition, as N-glycanase digestion reduced the apparent receptor subunit size of the transfected cells and IM-9 lymphocytes to identical values. The alteration in N-linked oligosaccharide composition could not be ascribed to differences in the kinetics of posttranslational processing of the insulin receptors, which was comparable to that of other cells studied. The basal rate of glycogen synthesis in the cells overexpressing insulin receptors was increased 4- to 5-fold compared with controls. Low levels of added insulin (0.1 nM) caused a 50% increase in the rate of glycogen synthesis

  8. In vitro generation of functional insulin-producing cells from lipoaspirated human adipose tissue-derived stem cells.

    Science.gov (United States)

    Mohamad Buang, Mohamad Lizan; Seng, Heng Kien; Chung, Lee Han; Saim, Aminuddin Bin; Idrus, Ruszymah Bt Hj

    2012-01-01

    Tissue engineering strategy has been considered as an alternative treatment for diabetes mellitus due to lack of permanent pharmaceutical treatment and islet donors for transplantation. Various cell lines have been used to generate functional insulin-producing cells (IPCs) including progenitor pancreatic cell lines, embryonic stem cells (ESCs), umbilical cord blood stem cells (UCB-SCs), adult bone marrow stem cells (BMSCs), and adipose tissue-derived stem cells (ADSCs). Human ADSCs from lipoaspirated abdominal fat tissue was differentiated into IPCs following a two-step induction protocol based on a combination of alternating high and low glucose, nicotinamide, activin A and glucagon-like peptide 1 (GLP-1) for a duration of 3 weeks. During differentiation, histomorphological changes of the stem cells towards pancreatic β-islet characteristics were observed via light microscope and transmission electron microscope (TEM). Dithizone (DTZ) staining, which is selective towards IPCs, was used to stain the new islet-like cells. Production of insulin hormone by the cells was analyzed via enzyme-linked immunosorbent assay (ELISA), whereas its hormonal regulation was tested via a glucose challenge test. Histomorphological changes of the differentiated cells were noted to resemble pancreatic β-cells, whereas DTZ staining positively stained the cells. The differentiated cells significantly produced human insulin as compared to the undifferentiated ADSCs, and its production was increased with an increase of glucose concentration in the culture medium. These initial data indicate that human lipoaspirated ADSCs have the potential to differentiate into functional IPCs, and could be used as a therapy to treat diabetes mellitus in the future. Copyright © 2012 IMSS. Published by Elsevier Inc. All rights reserved.

  9. Huntingtin-interacting protein 14 is a type 1 diabetes candidate protein regulating insulin secretion and β-cell apoptosis

    DEFF Research Database (Denmark)

    Berchtold, Lukas Adrian; Størling, Zenia Marian; Ortis, Fernanda

    2011-01-01

    Type 1 diabetes (T1D) is a complex disease characterized by the loss of insulin-secreting β-cells. Although the disease has a strong genetic component, and several loci are known to increase T1D susceptibility risk, only few causal genes have currently been identified. To identify disease...... genes in T1D, including the INS gene. An unexpected top-scoring candidate gene was huntingtin-interacting protein (HIP)-14/ZDHHC17. Immunohistochemical analysis of pancreatic sections demonstrated that HIP14 is almost exclusively expressed in insulin-positive cells in islets of Langerhans. RNAi...... knockdown experiments established that HIP14 is an antiapoptotic protein required for β-cell survival and glucose-stimulated insulin secretion. Proinflammatory cytokines (IL-1β and IFN-γ) that mediate β-cell dysfunction in T1D down-regulated HIP14 expression in insulin-secreting INS-1 cells and in isolated...

  10. Insulin utilizes the PI 3-kinase pathway to inhibit SP-A gene expression in lung epithelial cells

    Directory of Open Access Journals (Sweden)

    Snyder Jeanne M

    2002-10-01

    Full Text Available Abstract Background It has been proposed that high insulin levels may cause delayed lung development in the fetuses of diabetic mothers. A key event in lung development is the production of adequate amounts of pulmonary surfactant. Insulin inhibits the expression of surfactant protein A (SP-A, the major surfactant-associated protein, in lung epithelial cells. In the present study, we investigated the signal transduction pathways involved in insulin inhibition of SP-A gene expression. Methods H441 cells, a human lung adenocarcinoma cell line, or human fetal lung explants were incubated with or without insulin. Transcription run-on assays were used to determine SP-A gene transcription rates. Northern blot analysis was used to examine the effect of various signal transduction inhibitors on SP-A gene expression. Immunoblot analysis was used to evaluate the levels and phosphorylation states of signal transduction protein kinases. Results Insulin decreased SP-A gene transcription in human lung epithelial cells within 1 hour. Insulin did not affect p44/42 mitogen-activated protein kinase (MAPK phosphorylation and the insulin inhibition of SP-A mRNA levels was not affected by PD98059, an inhibitor of the p44/42 MAPK pathway. In contrast, insulin increased p70 S6 kinase Thr389 phosphorylation within 15 minutes. Wortmannin or LY294002, both inhibitors of phosphatidylinositol 3-kinase (PI 3-kinase, or rapamycin, an inhibitor of the activation of p70 S6 kinase, a downstream effector in the PI 3-kinase pathway, abolished or attenuated the insulin-induced inhibition of SP-A mRNA levels. Conclusion Insulin inhibition of SP-A gene expression in lung epithelial cells probably occurs via the rapamycin-sensitive PI 3-kinase signaling pathway.

  11. GLP-1 responses are heritable and blunted in acquired obesity with high liver fat and insulin resistance

    DEFF Research Database (Denmark)

    Matikainen, Niina; Bogl, Leonie H; Hakkarainen, Antti

    2014-01-01

    OBJECTIVE Impaired incretin response represents an early and uniform defect in type 2 diabetes, but the contributions of genes and the environment are poorly characterized. RESEARCH DESIGN AND METHODS We studied 35 monozygotic (MZ) and 75 dizygotic (DZ) twin pairs (discordant and concordant for o...... Whereas the GLP-1 response to the OGTT is heritable, an acquired unhealthy pattern of obesity characterized by liver fat accumulation and insulin resistance is closely related to impaired GLP-1 response in young adults....... under the curve was 67% (95% CI 45-80). Cotwins from weight-concordant MZ and DZ pairs and weight-discordant MZ pairs but concordant for liver fat content demonstrated similar glucose, insulin, and incretin profiles after the OGTT and meal tests. In contrast, higher insulin responses and blunted 60-min...... GLP-1 responses during the OGTT were observed in the heavier as compared with leaner MZ cotwins discordant for BMI, liver fat, and insulin sensitivity. Blunted GLP-1 response to OGTT was observed in heavier as compared with leaner DZ cotwins discordant for obesity and insulin sensitivity. CONCLUSIONS...

  12. Mesenchymal stem cells derived in vitro transdifferentiated insulin-producing cells: A new approach to treat type 1 diabetes

    Directory of Open Access Journals (Sweden)

    Shruti Dave

    2014-01-01

    Full Text Available The pathophysiology of type 1 diabetes mellitus (T1DM is largely related to an innate defect in the immune system culminating in a loss of self-tolerance and destruction of the insulin-producing β-cells. Currently, there is no definitive cure for T1DM. Insulin injection does not mimic the precise regulation of β-cells on glucose homeostasis, leading long term to the development of complications. Stem cell therapy is a promising approach and specifically mesenchymal stem cells (MSCs offer a promising possibility that deserves to be explored further. MSCs are multipotent, nonhematopoietic progenitors. They have been explored as an treatment option in tissue regeneration as well as potential of in vitro transdifferentiation into insulin-secreting cells. Thus, the major therapeutic goals for T1DM have been achieved in this way. The regenerative capabilities of MSCs have been a driving force to initiate studies testing their therapeutic effectiveness; their immunomodulatory properties have been equally exciting; which would appear capable of disabling immune dysregulation that leads to β-cell destruction in T1DM. Furthermore, MSCs can be cultured under specially defined conditions, their transdifferentiation can be directed toward the β-cell phenotype, and the formation of insulin-producing cells (IPCs can be targeted. To date, the role of MSCs-derived IPC in T1DM-a unique approach with some positive findings-have been unexplored, but it is still in its very early phase. In this study, a new approach of MSCs-derived IPCs, as a potential therapeutic benefit for T1DM in experimental animal models as well as in humans has been summarized.

  13. Absence of Carbohydrate Response Element Binding Protein in Adipocytes Causes Systemic Insulin Resistance and Impairs Glucose Transport

    Directory of Open Access Journals (Sweden)

    Archana Vijayakumar

    2017-10-01

    Full Text Available Lower adipose-ChREBP and de novo lipogenesis (DNL are associated with insulin resistance in humans. Here, we generated adipose-specific ChREBP knockout (AdChREBP KO mice with negligible sucrose-induced DNL in adipose tissue (AT. Chow-fed AdChREBP KO mice are insulin resistant with impaired insulin action in the liver, muscle, and AT and increased AT inflammation. HFD-fed AdChREBP KO mice are also more insulin resistant than controls. Surprisingly, adipocytes lacking ChREBP display a cell-autonomous reduction in insulin-stimulated glucose transport that is mediated by impaired Glut4 translocation and exocytosis, not lower Glut4 levels. AdChREBP KO mice have lower levels of palmitic acid esters of hydroxy stearic acids (PAHSAs in serum, and AT. 9-PAHSA supplementation completely rescues their insulin resistance and AT inflammation. 9-PAHSA also normalizes impaired glucose transport and Glut4 exocytosis in ChREBP KO adipocytes. Thus, loss of adipose-ChREBP is sufficient to cause insulin resistance, potentially by regulating AT glucose transport and flux through specific lipogenic pathways.

  14. Postreceptor defects causing insulin resistance in normoinsulinemic non-insulin-dependent diabetes mellitus

    International Nuclear Information System (INIS)

    Bolinder, J.; Ostman, J.; Arner, P.

    1982-01-01

    The mechanisms of the diminished hypoglycemic response to insulin in non-insulin-dependent diabetes mellitus (NIDDM) with normal levels of circulating plasma insulin were investigated. Specific binding of mono- 125 I (Tyr A14)-insulin to isolated adipocytes and effects of insulin (5--10,000 microunits/ml) on glucose oxidation and lipolysis were determined simultaneously in subcutaneous adipose tissue of seven healthy subjects of normal weight and seven untreated NIDDM patients with normal plasma insulin levels. The two groups were matched for age, sex, and body weight. Insulin binding, measured in terms of receptor number and affinity, was normal in NIDDM, the total number of receptors averaging 350,000 per cell. Neither sensitivity nor the maximum antilipolytic effect of insulin was altered in NIDDM patients as compared with control subjects; the insulin concentration producing half the maximum effect (ED50) was 10 microunits/ml. As regards the effect of insulin on glucose oxidation, for the control subjects ED50 was 30 microunits/ml, whereas in NIDDM patients, insulin exerted no stimulatory effect. The results obtained suggest that the effect of insulin on glucose utilization in normoinsulinemic NIDDM may be diminished in spite of normal insulin binding to receptors. The resistance may be due solely to postreceptor defects, and does not involve antilipolysis

  15. Comparative evaluation of optical methods and conventional isotope techniques for the detection of insulin receptors in heterogenous cell systems

    International Nuclear Information System (INIS)

    Thun, C.

    1984-01-01

    The findings of studies using radioactively labelled (I-125) insulin to characterise its binding to various heterogenous cell systems had led to a classification of the relevant receptors with those of high affinity and low capacity or vice versa. This, in turn, raised questions as to the binding properties of each individual cell or cell material of a heterogenous nature. Apparently homogenous (lymphocytes) and heterogenous (blood and islet cells) cell populations were investigated on the basis of various techniques for the separate evaluation of individual cells, which were cytofluorometry using FITC insulin and the analysis of gold insulin under the electron microscope. For the association kinetics and equilibration analysis or affinity and receptor quantity a radioactive tracer and light microscope were used. Insulin was shown to bind to erythrocytes, reticulocytes, monocytes and lymphocytes and this result finds confirmation in the relevant literature. Furthermore, binding parameters could be determined for isolated islet cells. Cytofluorometry pointed to the fact that the insulin receptors of an apparently homogenous cell system differed in affinity and number and permitted the use of a multiple parameter procedure. Thus, it holds out promise as a method to be routinely used in the clinical diagnosis of binding parameters, without requiring previous separation procedures that are complicated or involve a loss of material. Transmission electron microscopy permitted conclusions to be drawn as to the type of cell to which insulin is attached. Owing to the use of gold insulin it was possible to throw some light on the factors determining the fate of membrane-bound insulin during its uptake into the cell. (TRV) [de

  16. Effect of an extruded pea or rice diet on postprandial insulin and cardiovascular responses in dogs.

    Science.gov (United States)

    Adolphe, J L; Drew, M D; Silver, T I; Fouhse, J; Childs, H; Weber, L P

    2015-08-01

    Peas are increasing in popularity as a source of carbohydrate, protein and fibre in extruded canine diets. The aim of this study was to test the health effects of two canine diets with identical macronutrient profiles, but containing either yellow field peas or white rice as the carbohydrate source on metabolism, cardiovascular outcomes and adiposity. First, the acute glycemic, insulinemic and cardiovascular responses to the pea- or rice-based diets were determined in normal weight beagles (n = 7 dogs). The glycemic index did not differ between the pea diet (56 ± 12) and rice diet (63 ± 9). Next, obese beagles (n = 9) were fed the yellow field pea diet or white rice diet ad libitum for 12 weeks in a crossover study. Adiposity (measured using computed tomography), metabolic (oral glucose tolerance test, plasma leptin, adiponectin, C-reactive protein) and cardiovascular assessments (echocardiography and blood pressure) were performed before and after each crossover study period. After 12 weeks on each diet, peak insulin (p = 0.05) and area under the curve (AUC) for insulin after a 10 g oral glucose tolerance test (p = 0.05) were lower with the pea than the rice diet. Diet did not show a significant effect on body weight, fat distribution, cardiovascular variables, adiponectin or leptin. In conclusion, a diet containing yellow field peas reduced the postprandial insulin response after glucose challenge in dogs despite continued obesity, indicating improved metabolic health. Journal of Animal Physiology and Animal Nutrition © 2014 Blackwell Verlag GmbH.

  17. Involvement of insulin-degrading enzyme in insulin- and atrial natriuretic peptide-sensitive internalization of amyloid-β peptide in mouse brain capillary endothelial cells.

    Science.gov (United States)

    Ito, Shingo; Ohtsuki, Sumio; Murata, Sho; Katsukura, Yuki; Suzuki, Hiroya; Funaki, Miho; Tachikawa, Masanori; Terasaki, Tetsuya

    2014-01-01

    Cerebral clearance of amyloid-β peptide (Aβ), which is implicated in Alzheimer's disease, involves elimination across the blood-brain barrier (BBB), and we previously showed that an insulin-sensitive process is involved in the case of Aβ1-40. The purpose of this study was to clarify the molecular mechanism of the insulin-sensitive Aβ1-40 elimination across mouse BBB. An in vivo cerebral microinjection study demonstrated that [125I]hAβ1-40 elimination from mouse brain was inhibited by human natriuretic peptide (hANP), and [125I]hANP elimination was inhibited by hAβ1-40, suggesting that hAβ1-40 and hANP share a common elimination process. Internalization of [125I]hAβ1-40 into cultured mouse brain capillary endothelial cells (TM-BBB4) was significantly inhibited by either insulin, hANP, other natriuretic peptides or insulin-degrading enzyme (IDE) inhibitors, but was not inhibited by phosphoramidon or thiorphan. Although we have reported the involvement of natriuretic peptide receptor C (Npr-C) in hANP internalization, cells stably expressing Npr-C internalized [125I]hANP but not [125I]hAβ1-40, suggesting that there is no direct interaction between Npr-C and hAβ1-40. IDE was detected in plasma membrane of TM-BBB4 cells, and internalization of [125I]hAβ1-40 by TM-BBB4 cells was reduced by IDE-targeted siRNAs. We conclude that elimination of hAβ1-40 from mouse brain across the BBB involves an insulin- and ANP-sensitive process, mediated by IDE expressed in brain capillary endothelial cells.

  18. The level of menadione redox-cycling in pancreatic β-cells is proportional to the glucose concentration: Role of NADH and consequences for insulin secretion

    Energy Technology Data Exchange (ETDEWEB)

    Heart, Emma [Cellular Dynamics Program, Marine Biological Laboratory, Woods Hole, MA, 02543 (United States); Palo, Meridith; Womack, Trayce [Department of Science, United States Coast Guard Academy, New London, CT, 06320 (United States); Smith, Peter J.S. [Cellular Dynamics Program, Marine Biological Laboratory, Woods Hole, MA, 02543 (United States); Institute for Life Sciences, University of Southampton (United Kingdom); Gray, Joshua P., E-mail: Joshua.p.gray@uscga.edu [Cellular Dynamics Program, Marine Biological Laboratory, Woods Hole, MA, 02543 (United States); Department of Science, United States Coast Guard Academy, New London, CT, 06320 (United States)

    2012-01-15

    Pancreatic β-cells release insulin in response to elevation of glucose from basal (4–7 mM) to stimulatory (8–16 mM) levels. Metabolism of glucose by the β-cell results in the production of low levels of reactive oxygen intermediates (ROI), such as hydrogen peroxide (H{sub 2}O{sub 2}), a newly recognized coupling factor linking glucose metabolism to insulin secretion. However, high and toxic levels of H{sub 2}O{sub 2} inhibit insulin secretion. Menadione, which produces H{sub 2}O{sub 2} via redox cycling mechanism in a dose-dependent manner, was investigated for its effect on β-cell metabolism and insulin secretion in INS-1 832/13, a rat β-cell insulinoma cell line, and primary rodent islets. Menadione-dependent redox cycling and resulting H{sub 2}O{sub 2} production under stimulatory glucose exceeded several-fold those reached at basal glucose. This was paralleled by a differential effect of menadione (0.1–10 μM) on insulin secretion, which was enhanced at basal, but inhibited at stimulatory glucose. Redox cycling of menadione and H{sub 2}O{sub 2} formation was dependent on glycolytically-derived NADH, as inhibition of glycolysis and application of non-glycogenic insulin secretagogues did not support redox cycling. In addition, activity of plasma membrane electron transport, a system dependent in part on glycolytically-derived NADH, was also inhibited by menadione. Menadione-dependent redox cycling was sensitive to the NQO1 inhibitor dicoumarol and the flavoprotein inhibitor diphenylene iodonium, suggesting a role for NQO1 and other oxidoreductases in this process. These data may explain the apparent dichotomy between the stimulatory and inhibitory effects of H{sub 2}O{sub 2} and menadione on insulin secretion. -- Highlights: ► Menadione stimulation or inhibition of insulin secretion is dependent upon applied glucose levels. ► Menadione-dependent H{sub 2}O{sub 2} production is proportional to applied glucose levels. ► Quinone-mediated redox cycling

  19. The level of menadione redox-cycling in pancreatic β-cells is proportional to the glucose concentration: Role of NADH and consequences for insulin secretion

    International Nuclear Information System (INIS)

    Heart, Emma; Palo, Meridith; Womack, Trayce; Smith, Peter J.S.; Gray, Joshua P.

    2012-01-01

    Pancreatic β-cells release insulin in response to elevation of glucose from basal (4–7 mM) to stimulatory (8–16 mM) levels. Metabolism of glucose by the β-cell results in the production of low levels of reactive oxygen intermediates (ROI), such as hydrogen peroxide (H 2 O 2 ), a newly recognized coupling factor linking glucose metabolism to insulin secretion. However, high and toxic levels of H 2 O 2 inhibit insulin secretion. Menadione, which produces H 2 O 2 via redox cycling mechanism in a dose-dependent manner, was investigated for its effect on β-cell metabolism and insulin secretion in INS-1 832/13, a rat β-cell insulinoma cell line, and primary rodent islets. Menadione-dependent redox cycling and resulting H 2 O 2 production under stimulatory glucose exceeded several-fold those reached at basal glucose. This was paralleled by a differential effect of menadione (0.1–10 μM) on insulin secretion, which was enhanced at basal, but inhibited at stimulatory glucose. Redox cycling of menadione and H 2 O 2 formation was dependent on glycolytically-derived NADH, as inhibition of glycolysis and application of non-glycogenic insulin secretagogues did not support redox cycling. In addition, activity of plasma membrane electron transport, a system dependent in part on glycolytically-derived NADH, was also inhibited by menadione. Menadione-dependent redox cycling was sensitive to the NQO1 inhibitor dicoumarol and the flavoprotein inhibitor diphenylene iodonium, suggesting a role for NQO1 and other oxidoreductases in this process. These data may explain the apparent dichotomy between the stimulatory and inhibitory effects of H 2 O 2 and menadione on insulin secretion. -- Highlights: ► Menadione stimulation or inhibition of insulin secretion is dependent upon applied glucose levels. ► Menadione-dependent H 2 O 2 production is proportional to applied glucose levels. ► Quinone-mediated redox cycling is dependent on glycolysis

  20. Role of adipose tissue derived stem cells differentiated into insulin producing cells in the treatment of type I diabetes mellitus.

    Science.gov (United States)

    Amer, Mona G; Embaby, Azza S; Karam, Rehab A; Amer, Marwa G

    2018-05-15

    Generation of new β cells is an important approach in the treatment of type 1 diabetes mellitus (type 1 DM). Adipose tissue-derived stem cells (ADSCs) might be one of the best sources for cell replacement therapy for diabetes. Therefore, this work aimed to test the possible role of transplanted insulin-producing cells (IPCs) differentiated from ADSCs in treatment of streptozotocin (STZ) induced type I DM in rats. Type 1 DM was induced by single intra peritoneal injection with STZ (50 mg/kg BW). Half of the diabetic rats were left without treatment and the other half were injected with differentiated IPCs directly into the pancreas. ADSCs were harvested, cultured and identified by testing their phenotypes through flow cytometry. They were further subjected to differentiation into IPCs using differentiation medium. mRNA expression of pancreatic transcription factors (pdx1), insulin and glucose transporter-2 genes by real time PCR was done to detect the cellular differentiation and confirmed by stimulated insulin secretion. The pancreatic tissues from all groups were examined 2 months after IPC transplantation and were subjected to histological, Immunohistochemical and morphometric study. The differentiated IPCs showed significant expression of pancreatic β cell markers and insulin secretion in glucose dependent manner. Treatment with IPCs induced apparent regeneration, diffused proliferated islet cells and significant increase in C-peptide immune reaction. We concluded that transplantation of differentiated IPCs improved function and morphology of Islet cells in diabetic rats. Consequently, this therapy option may be a promising therapeutic approach to patient with type 1 DM if proven to be effective and safe. Copyright © 2018 Elsevier B.V. All rights reserved.

  1. Reversible immortalization of Nestin-positive precursor cells from pancreas and differentiation into insulin-secreting cells

    Energy Technology Data Exchange (ETDEWEB)

    Wei, Pei; Li, Li; Qi, Hui [The Clinical Medical Research Center, The Second Clinical Medical College (Shenzhen People' s Hospital), Jinan University, 518020 Shenzhen (China); Zhou, Han-xin [Department of General Surgery, First Hospital (Shenzhen Second People' s Hospital) of Shenzhen University, 518020 Shenzhen (China); Deng, Chun-yan [The Clinical Medical Research Center, The Second Clinical Medical College (Shenzhen People' s Hospital), Jinan University, 518020 Shenzhen (China); Li, Fu-rong, E-mail: frli62@yahoo.com [The Clinical Medical Research Center, The Second Clinical Medical College (Shenzhen People' s Hospital), Jinan University, 518020 Shenzhen (China); Shenzhen Institution of Gerontology, 518020 Shenzhen (China)

    2012-02-10

    Highlights: Black-Right-Pointing-Pointer The NPPCs from mouse pancreas were isolated. Black-Right-Pointing-Pointer Tet-on system for SV40 large in NPPCs was used to get RINPPCs. Black-Right-Pointing-Pointer The RINPPCs can undergo at least 80 population doublings without senescence. Black-Right-Pointing-Pointer The RINPPCs can be induced to differentiate into insulin-producing cells. Black-Right-Pointing-Pointer The combination of GLP-1 and sodium butyrate promoted the differentiation process. -- Abstract: Pancreatic stem cells or progenitor cells posses the ability of directed differentiation into pancreatic {beta} cells. However, these cells usually have limited proliferative capacity and finite lifespan in vitro. In the present study, Nestin-positive progenitor cells (NPPCs) from mouse pancreas that expressed the pancreatic stem cells or progenitor cell marker Nestin were isolated to obtain a sufficient number of differentiated pancreatic {beta} cells. Tet-on system for SV40 large T-antigen expression in NPPCs was used to achieve reversible immortalization. The reversible immortal Nestin-positive progenitor cells (RINPPCs) can undergo at least 80 population doublings without senescence in vitro while maintaining their biological and genetic characteristics. RINPPCs can be efficiently induced to differentiate into insulin-producing cells that contain a combination of glucagon-like peptide-1 (GLP-1) and sodium butyrate. The results of the present study can be used to explore transplantation therapy of type I diabetes mellitus.

  2. The effect of different protein hydrolysate/carbohydrate mixtures on postprandial glucagon and insulin responses in healthy subjects.

    Science.gov (United States)

    Claessens, M; Calame, W; Siemensma, A D; van Baak, M A; Saris, W H M

    2009-01-01

    To study the effect of four protein hydrolysates from vegetable (pea, gluten, rice and soy) and two protein hydrolysates from animal origin (whey and egg) on glucagon and insulin responses. Eight healthy normal-weight male subjects participated in this study. The study employed a repeated-measures design with Latin square randomization and single-blind trials. Protein hydrolysates used in this study (pea, rice, soy, gluten, whey and egg protein hydrolysate) consisted of 0.2 g hydrolysate per kg body weight (bw) and 0.2 g maltodextrin per kg bw and were compared to maltodextrin alone. Postprandial plasma glucose, glucagon, insulin and amino acids were determined over 2 h. All protein hydrolysates induced an enhanced insulin secretion compared to maltodextrin alone and a correspondingly low plasma glucose response. A significant difference was observed in area under the curve (AUC) for plasma glucagon between protein hydrolysates and the maltodextrin control drink (Pprotein hydrolysate induced the lowest glucagon response. High amino-acid-induced glucagon response does not necessarily go together with low insulin response. Protein hydrolysate source affects AUC for glucagon more profoundly than for insulin, although the protein load used in this study seemed to be at lower level for significant physiological effects.

  3. Direct Sensing of Nutrients via a LAT1-like Transporter in Drosophila Insulin-Producing Cells

    Directory of Open Access Journals (Sweden)

    Gérard Manière

    2016-09-01

    Full Text Available Dietary leucine has been suspected to play an important role in insulin release, a hormone that controls satiety and metabolism. The mechanism by which insulin-producing cells (IPCs sense leucine and regulate insulin secretion is still poorly understood. In Drosophila, insulin-like peptides (DILP2 and DILP5 are produced by brain IPCs and are released in the hemolymph after leucine ingestion. Using Ca2+-imaging and ex vivo cultured larval brains, we demonstrate that IPCs can directly sense extracellular leucine levels via minidiscs (MND, a leucine transporter. MND knockdown in IPCs abolished leucine-dependent changes, including loss of DILP2 and DILP5 in IPC bodies, consistent with the idea that MND is necessary for leucine-dependent DILP release. This, in turn, leads to a strong increase in hemolymph sugar levels and reduced growth. GDH knockdown in IPCs also reduced leucine-dependent DILP release, suggesting that nutrient sensing is coupled to the glutamate dehydrogenase pathway.

  4. Longitudinal Changes in Insulin Resistance, Beta-Cell Function and Glucose Regulation Status in Prediabetes.

    Science.gov (United States)

    Kim, Chul-Hee; Kim, Hong-Kyu; Kim, Eun-Hee; Bae, Sung-Jin; Choe, Jaewon; Park, Joong-Yeol

    2018-01-01

    The changes in insulin resistance and insulin secretion and their association with changes in glucose regulation status in Asians with prediabetes remain uncertain. We included Korean adults (aged 20-79 years) with prediabetes who underwent routine medical check-ups at a mean interval of 5 years. Prediabetes was defined as fasting plasma glucose (FPG) 5.6-6.9mmol/l or HbA1c 5.7-6.4% (39-46mmol/mol). Insulin resistance (HOMA-IR) and beta-cell function (HOMA-%B) indices were assessed by homeostasis model assessment. Incident diabetes was defined as FPG ≥ 7.0mmol/l, HbA1c ≥ 6.5% (48mmol/mol), or initiation of antidiabetic medications. Among the 7,208 participants with prediabetes, 4,410 (61.2%) remained as prediabetes (control group), 2,123 (29.5%) reverted to normal glucose regulation (regressors), and 675 (9.4%) progressed to type 2 diabetes (progressors) after 5 years. Compared with the control group, the progressors had higher baseline HOMA-IR (2.48 ± 1.45 versus 2.06 ± 1.20, P prediabetes, longitudinal change in insulin resistance was the predominant factor in Koreans. Copyright © 2018 Southern Society for Clinical Investigation. Published by Elsevier Inc. All rights reserved.

  5. Superoxide generation is diminished during glucose-stimulated insulin secretion in INS-1E cells

    Czech Academy of Sciences Publication Activity Database

    Ježek, Petr; Hlavatá, Lydie; Špaček, Tomáš

    2008-01-01

    Roč. 275, Suppl.1 (2008), s. 310-310 ISSN 1742-464X. [FEBS Congress /33./ and IUBMB Conference /11./. 28.06.2008-03.07.2008, Athens] R&D Projects: GA MZd(CZ) NR7917; GA AV ČR(CZ) IAA500110701 Institutional research plan: CEZ:AV0Z50110509 Keywords : cpo1 * superoxide production * glucose-stimulated insulin secretion * INS-1E cells Subject RIV: ED - Physiology

  6. Minor long-term changes in weight have beneficial effects on insulin sensitivity and beta-cell function in obese subjects

    DEFF Research Database (Denmark)

    Rosenfalck, A M; Hendel, Helle Westergren; Rasmussen, M H

    2002-01-01

    To evaluate the long-term effect of changes in body composition induced by weight loss on insulin sensitivity (SI), non-insulin mediated glucose disposal, glucose effectiveness (SG)and beta-cell function.......To evaluate the long-term effect of changes in body composition induced by weight loss on insulin sensitivity (SI), non-insulin mediated glucose disposal, glucose effectiveness (SG)and beta-cell function....

  7. Calpain Determines the Propensity of Adult Hippocampal Neural Stem Cells to Autophagic Cell Death Following Insulin Withdrawal.

    Science.gov (United States)

    Chung, Kyung Min; Park, Hyunhee; Jung, Seonghee; Ha, Shinwon; Yoo, Seung-Jun; Woo, Hanwoong; Lee, Hyang Ju; Kim, Seong Who; Kim, Eun-Kyoung; Moon, Cheil; Yu, Seong-Woon

    2015-10-01

    Programmed cell death (PCD) has significant effects on the function of neural stem cells (NSCs) during brain development and degeneration. We have previously reported that adult rat hippocampal neural stem (HCN) cells underwent autophagic cell death (ACD) rather than apoptosis following insulin withdrawal despite their intact apoptotic capabilities. Here, we report a switch in the mode of cell death in HCN cells with calpain as a critical determinant. In HCN cells, calpain 1 expression was barely detectable while calpain 2 was predominant. Inhibition of calpain in insulin-deprived HCN cells further augmented ACD. In contrast, expression of calpain 1 switched ACD to apoptosis. The proteasome inhibitor lactacystin blocked calpain 2 degradation and elevated the intracellular Ca(2+) concentration. In combination, these effects potentiated calpain activity and converted the mode of cell death to apoptosis. Our results indicate that low calpain activity, due to absence of calpain 1 and degradation of calpain 2, results in a preference for ACD over apoptosis in insulin-deprived HCN cells. On the other hand, conditions leading to high calpain activity completely switch the mode of cell death to apoptosis. This is the first report on the PCD mode switching mechanism in NSCs. The dynamic change in calpain activity through the proteasome-mediated modulation of the calpain and intracellular Ca(2+) levels may be the critical contributor to the demise of NSCs. Our findings provide a novel insight into the complex mechanisms interconnecting autophagy and apoptosis and their roles in the regulation of NSC death. © 2015 AlphaMed Press.

  8. Characterization of the growth of murine fibroblasts that express human insulin receptors. I. The effect of insulin in the absence of other growth factors

    International Nuclear Information System (INIS)

    Randazzo, P.A.; Morey, V.A.; Polishook, A.K.; Jarett, L.

    1990-01-01

    The effect of insulin on the growth of murine fibroblasts transfected with an expression vector containing human insulin receptor cDNA (NIH 3T3/HIR) and the parental cells (NIH/3T3) was characterized. Insulin in the absence of other mitogens increased the rate of incorporation of thymidine into NIH 3T3/HIR cells with a half-maximal response occurring at an insulin concentration of 35 ng/ml and a maximal response that was equivalent to that elicited by 10% fetal calf serum. The thymidine incorporation rate was increased by 12 h, was maximal at approximately 16 h, and returned to basal rates at 24 h after the addition of insulin. Insulin induced a maximum of 65% of cells to incorporate thymidine. The increased DNA synthesis was accompanied by net growth. Addition of insulin to the NIH 3T3/HIR cells resulted in increased DNA content with a half-maximal response occurring at approximately 30 ng/ml insulin and a maximal response equivalent to that elicited by serum. An increase in cell number detected after the addition of insulin to the NIH 3T3/HIR suggests that the cells had progressed through mitosis. Insulin did not increase the rate of thymidine incorporation, DNA content, or number of the parental NIH 3T3 cells. These data show that insulin, in the absence of a second mitogen, is able to induce NIH 3T3/HIR fibroblasts to traverse the cell cycle

  9. Local and systemic response to intramammary lipopolysaccharide challenge during long-term manipulated plasma glucose and insulin concentrations in dairy cows.

    Science.gov (United States)

    Vernay, M C M B; Wellnitz, O; Kreipe, L; van Dorland, H A; Bruckmaier, R M

    2012-05-01

    The metabolic load during periods of high milk production in dairy cows causes a variety of changes of metabolite blood concentrations including dramatically decreased glucose levels. These changes supposedly impair the immune system. The goal of this study was, therefore, to evaluate adaptations of the cow's immune system in response to an intramammary lipopolysaccharide (LPS) stimulation during a 3-d modification of plasma glucose and insulin induced by different clamp infusions. Seventeen midlactating dairy cows received a hypoglycemic hyperinsulinemic clamp induced by insulin infusion (HypoG; n=5), a euglycemic hyperinsulinemic clamp induced by insulin and glucose infusion (EuG; n=6), or infusion of saline solution (NaCl; n=6) for 56 h. At 48 h of infusion, 2 udder quarters were challenged with 200 μg of Escherichia coli LPS. At 48 h of infusion (immediately before LPS challenge), tumor necrosis factor α, lactoferrin, and serum amyloid A (SAA) mRNA abundance was increased in HypoG and Il-1β mRNA abundance was decreased in EuG. After LPS challenge, plasma glucose concentration did not decrease, although plasma insulin increased simultaneously in all groups either due to enhanced endogenous release (NaCl) or due to increased insulin infusion rate (HypoG; EuG). Plasma cortisol, rectal temperatures, and milk somatic cell count of challenged quarters increased, whereas plasma nonesterified fatty acid concentrations were similarly decreased across treatments. In mammary biopsies, increased mRNA expression of tumor necrosis factor α, IL-1β, IL-8, and IL-10, and SAA were observed in LPS-treated quarters of all groups, with a more pronounced increase in IL-1β, IL-10, and SAA expression in EuG. Nuclear factor-κB mRNA expression was upregulated in NaCl and EuG but not in HypoG in response to LPS. Lactoferrin, toll-like receptor 4, and cyclooxygenase-2 mRNA expression was increased in LPS-treated quarters of EuG only, and 5-lipoxygenase mRNA expression was decreased

  10. Ionizing Radiation Potentiates High Fat Diet-Induced Insulin Resistance and Reprograms Skeletal Muscle and Adipose Progenitor Cells

    DEFF Research Database (Denmark)

    Nylander, Vibe; Ingerslev, Lars R; Andersen, Emil

    2016-01-01

    Exposure to ionizing radiation increases the risk of chronic metabolic disorders such as insulin resistance and type 2 diabetes later in life. We hypothesized that irradiation reprograms the epigenome of metabolic progenitor cells, which could account for impaired metabolism after cancer treatment...... mice. Mice subjected to total body irradiation showed alterations in glucose metabolism and, when challenged with HFD, marked hyperinsulinemia. Insulin signaling was chronically disrupted in skeletal muscle and adipose progenitor cells collected from irradiated mice and differentiated in culture...

  11. Expression of transient receptor potential ankyrin 1 (TRPA1 and its role in insulin release from rat pancreatic beta cells.

    Directory of Open Access Journals (Sweden)

    De-Shou Cao

    Full Text Available Several transient receptor potential (TRP channels are expressed in pancreatic beta cells and have been proposed to be involved in insulin secretion. However, the endogenous ligands for these channels are far from clear. Here, we demonstrate the expression of the transient receptor potential ankyrin 1 (TRPA1 ion channel in the pancreatic beta cells and its role in insulin release. TRPA1 is an attractive candidate for inducing insulin release because it is calcium permeable and is activated by molecules that are produced during oxidative glycolysis.Immunohistochemistry, RT-PCR, and Western blot techniques were used to determine the expression of TRPA1 channel. Ca²⁺ fluorescence imaging and electrophysiology (voltage- and current-clamp techniques were used to study the channel properties. TRPA1-mediated insulin release was determined using ELISA.TRPA1 is abundantly expressed in a rat pancreatic beta cell line and freshly isolated rat pancreatic beta cells, but not in pancreatic alpha cells. Activation of TRPA1 by allyl isothiocyanate (AITC, hydrogen peroxide (H₂O₂, 4-hydroxynonenal (4-HNE, and cyclopentenone prostaglandins (PGJ₂ and a novel agonist methylglyoxal (MG induces membrane current, depolarization, and Ca²⁺ influx leading to generation of action potentials in a pancreatic beta cell line and primary cultured pancreatic beta cells. Activation of TRPA1 by agonists stimulates insulin release in pancreatic beta cells that can be inhibited by TRPA1 antagonists such as HC030031 or AP-18 and by RNA interference. TRPA1-mediated insulin release is also observed in conditions of voltage-gated Na⁺ and Ca²⁺ channel blockade as well as ATP sensitive potassium (K(ATP channel activation.We propose that endogenous and exogenous ligands of TRPA1 cause Ca²⁺ influx and induce basal insulin release and that TRPA1-mediated depolarization acts synergistically with K(ATP channel blockade to facilitate insulin release.

  12. Role of growth hormone, insulin-like growth factor-I, and insulin-like growth factor binding proteins in the catabolic response to injury and infection.

    Science.gov (United States)

    Lang, Charles H; Frost, Robert A

    2002-05-01

    The erosion of lean body mass resulting from protracted critical illness remains a significant risk factor for increased morbidity and mortality in this patient population. Previous studies have documented the well known impairment in nitrogen balance results from both an increase in muscle protein degradation as well as a decreased rate of both myofibrillar and sacroplasmic protein synthesis. This protein imbalance may be caused by an increased presence or activity of various catabolic agents, such as tumor necrosis factor-alpha, interleukin-1 beta, interleukin-6 or glucocorticoids, or may be mediated via a decreased concentration or responsiveness to various anabolic hormones, such as growth hormone or insulin-like growth factor-I. This review focuses on recent developments pertaining to the importance of alterations in the growth hormone-insulin-like growth factor-I axis as a mechanism for the observed defects in muscle protein balance.

  13. Effects of intranasal insulin application on the hypothalamic BOLD response to glucose ingestion

    DEFF Research Database (Denmark)

    van Opstal, Anna M.; Akintola, Abimbola A.; Elst, Marjan van der

    2017-01-01

    The hypothalamus is a crucial structure in the brain that responds to metabolic cues and regulates energy homeostasis. Patients with type 2 diabetes demonstrate a lack of hypothalamic neuronal response after glucose ingestion, which is suggested to be an underlying cause of the disease. In this s......The hypothalamus is a crucial structure in the brain that responds to metabolic cues and regulates energy homeostasis. Patients with type 2 diabetes demonstrate a lack of hypothalamic neuronal response after glucose ingestion, which is suggested to be an underlying cause of the disease...... effect. Our data provide proof of concept for future experiments testing the potential of intranasal application of insulin to ameliorate defective homeostatic control in patients with type 2 diabetes....

  14. Antihyperglycemic Activity of Eucalyptus tereticornis in Insulin-Resistant Cells and a Nutritional Model of Diabetic Mice

    Directory of Open Access Journals (Sweden)

    Alis Guillén

    2015-01-01

    Full Text Available Eucalyptus tereticornis is a plant used in traditional medicine to control diabetes, but this effect has not been proved scientifically. Here, we demonstrated through in vitro assays that E. tereticornis extracts increase glucose uptake and inhibit their production in insulin-resistant C2C12 and HepG2 cells, respectively. Furthermore, in a nutritional model using diabetic mice, the administration of ethyl acetate extract of E. tereticornis reduced fasting glycaemia, improved tolerance to glucose, and reduced resistance to insulin. Likewise, this extract had anti-inflammatory effects in adipose tissue when compared to control diabetic mice. Via bioguided assays and sequential purification of the crude extract, a triterpenoid-rich fraction from ethyl acetate extracts was shown to be responsible for the biological activity. Similarly, we identified the main compound responsible for the antihyperglycemic activity in this extract. This study shows that triterpenes found in E. tereticornis extracts act as hypoglycemic/antidiabetic compounds and contribute to the understanding of their use in traditional medicine.

  15. The F-actin modifier villin regulates insulin granule dynamics and exocytosis downstream of islet cell autoantigen 512

    Directory of Open Access Journals (Sweden)

    Hassan Mziaut

    2016-08-01

    Full Text Available Objective: Insulin release from pancreatic islet β cells should be tightly controlled to avoid hypoglycemia and insulin resistance. The cortical actin cytoskeleton is a gate for regulated exocytosis of insulin secretory granules (SGs by restricting their mobility and access to the plasma membrane. Prior studies suggest that SGs interact with F-actin through their transmembrane cargo islet cell autoantigen 512 (Ica512 (also known as islet antigen 2/Ptprn. Here we investigated how Ica512 modulates SG trafficking and exocytosis. Methods: Transcriptomic changes in Ica512−/− mouse islets were analyzed. Imaging as well as biophysical and biochemical methods were used to validate if and how the Ica512-regulated gene villin modulates insulin secretion in mouse islets and insulinoma cells. Results: The F-actin modifier villin was consistently downregulated in Ica512−/− mouse islets and in Ica512-depleted insulinoma cells. Villin was enriched at the cell cortex of β cells and dispersed villin−/− islet cells were less round and less deformable. Basal mobility of SGs in villin-depleted cells was enhanced. Moreover, in cells depleted either of villin or Ica512 F-actin cages restraining cortical SGs were enlarged, basal secretion was increased while glucose-stimulated insulin release was blunted. The latter changes were reverted by overexpressing villin in Ica512-depleted cells, but not vice versa. Conclusion: Our findings show that villin controls the size of the F-actin cages restricting SGs and, thus, regulates their dynamics and availability for exocytosis. Evidence that villin acts downstream of Ica512 also indicates that SGs directly influence the remodeling properties of the cortical actin cytoskeleton for tight control of insulin secretion. Keywords: F-actin, Granules, Ica512, Insulin, Secretion, Villin

  16. Collagen and Stretch Modulate Autocrine Secretion of Insulin-like Growth Factor-1 and Insulin-like Growth Factor Binding Proteins from Differentiated Skeletal Muscle Cells

    Science.gov (United States)

    Perrone, Carmen E.; Fenwick-Smith, Daniela; Vandenburgh, Herman H.

    1995-01-01

    Stretch-induced skeletal muscle growth may involve increased autocrine secretion of insulin-like growth factor-1 (IGF-1) since IGF-1 is a potent growth factor for skeletal muscle hypertrophy, and stretch elevates IGF-1 mRNA levels in vivo. In tissue cultures of differentiated avian pectoralis skeletal muscle cells, nanomolar concentrations of exogenous IGF-1 stimulated growth in mechanically stretched but not static cultures. These cultures released up to 100 pg of endogenously produced IGF-1/micro-g of protein/day, as well as three major IGF binding proteins of 31, 36, and 43 kilodaltons (kDa). IGF-1 was secreted from both myofibers and fibroblasts coexisting in the muscle cultures. Repetitive stretch/relaxation of the differentiated skeletal muscle cells stimulated the acute release of IGF-1 during the first 4 h after initiating mechanical activity, but caused no increase in the long-term secretion over 24-72 h of IGF-1, or its binding proteins. Varying the intensity and frequency of stretch had no effect on the long-term efflux of IGF-1. In contrast to stretch, embedding the differentiated muscle cells in a three-dimensional collagen (Type I) matrix resulted in a 2-5-fold increase in long-term IGF-1 efflux over 24-72 h. Collagen also caused a 2-5-fold increase in the release of the IGF binding proteins. Thus, both the extracellular matrix protein type I collagen and stretch stimulate the autocrine secretion of IGF-1, but with different time kinetics. This endogenously produced growth factor may be important for the growth response of skeletal myofibers to both types of external stimuli.

  17. Bioluminescence imaging of β cells and intrahepatic insulin gene activity under normal and pathological conditions.

    Directory of Open Access Journals (Sweden)

    Tokio Katsumata

    Full Text Available In diabetes research, bioluminescence imaging (BLI has been applied in studies of β-cell impairment, development, and islet transplantation. To develop a mouse model that enables noninvasive imaging of β cells, we generated a bacterial artificial chromosome (BAC transgenic mouse in which a mouse 200-kbp genomic fragment comprising the insulin I gene drives luciferase expression (Ins1-luc BAC transgenic mouse. BLI of mice was performed using the IVIS Spectrum system after intraperitoneal injection of luciferin, and the bioluminescence signal from the pancreatic region analyzed. When compared with MIP-Luc-VU mice [FVB/N-Tg(Ins1-lucVUPwrs/J] expressing luciferase under the control of the 9.2-kbp mouse insulin I promoter (MIP, the bioluminescence emission from Ins1-luc BAC transgenic mice was enhanced approximately 4-fold. Streptozotocin-treated Ins1-luc BAC transgenic mice developed severe diabetes concomitant with a sharp decline in the BLI signal intensity in the pancreas. Conversely, mice fed a high-fat diet for 8 weeks showed an increase in the signal, reflecting a decrease or increase in the β-cell mass. Although the bioluminescence intensity of the islets correlated well with the number of isolated islets in vitro, the intensity obtained from a living mouse in vivo did not necessarily reflect an absolute quantification of the β-cell mass under pathological conditions. On the other hand, adenovirus-mediated gene transduction of β-cell-related transcription factors in Ins1-luc BAC transgenic mice generated luminescence from the hepatic region for more than 1 week. These results demonstrate that BLI in Ins1-luc BAC transgenic mice provides a noninvasive method of imaging islet β cells and extrapancreatic activity of the insulin gene in the liver under normal and pathological conditions.

  18. Hematopoietic Kit Deficiency, rather than Lack of Mast Cells, Protects Mice from Obesity and Insulin Resistance.

    Science.gov (United States)

    Gutierrez, Dario A; Muralidhar, Sathya; Feyerabend, Thorsten B; Herzig, Stephan; Rodewald, Hans-Reimer

    2015-05-05

    Obesity, insulin resistance, and related pathologies are associated with immune-mediated chronic inflammation. Kit mutant mice are protected from diet-induced obesity and associated co-morbidities, and this phenotype has previously been attributed to their lack of mast cells. We performed a comprehensive metabolic analysis of Kit-dependent Kit(W/Wv) and Kit-independent Cpa3(Cre/+) mast-cell-deficient mouse strains, employing diet-induced or genetic (Lep(Ob/Ob) background) models of obesity. Our results show that mast cell deficiency, in the absence of Kit mutations, plays no role in the regulation of weight gain or insulin resistance. Moreover, we provide evidence that the metabolic phenotype observed in Kit mutant mice, while independent of mast cells, is immune regulated. Our data underscore the value of definitive mast cell deficiency models to conclusively test the involvement of this enigmatic cell in immune-mediated pathologies and identify Kit as a key hematopoietic factor in the pathogenesis of metabolic syndrome. Copyright © 2015 Elsevier Inc. All rights reserved.

  19. Membrane-associated insulin-like growth factor (IGF binding structures in placental cells

    Directory of Open Access Journals (Sweden)

    ROMANA MASNIKOSA

    2003-11-01

    Full Text Available The biological activities of IGF-I and –II are mediated mainly by the type 1 IGF receptor (IGF 1R and controlled by their interaction with soluble proteins, the IGF binding proteins (IGFBPs. Although there is a growing body of evidence that some IGFBPs may be cell surface-bound, published data concerning cell association of IGFBP-1 are scarce and none of them concern placental cells. The cell membranes used in this study were isolated from term human placentae. Detergent-solubilized membranes were shown to contain two types of IGF binding structures that were separated by gel filtration on a Sephadex G-100 column. Proteins in the first peak were eluted at V0 (Mr > 100 kD and they bound IGF-I with greater specificity and affinity than IGF-II and insulin. Most likely, they represented the IGF 1R. Small proteins (Mr ~ 45 kD were eluted with the membrane proteins in the second maximum. They were able to bind IGF-I and IGF-II, but not insulin. The identity of these proteins was shown to be IGFBP-1 on the basis of their reaction with specific anti-IGFBP-1 antibodies. To the best of our knowledge, the existence of IGFBP-1 associated with human placental cell membranes has not been reported in the literature before. Colocalisation of IGFBP-1 with IGF 1R in cell membranes could provide efficient modulation of IGF 1R receptor-ligand interactions.

  20. Insulin-like Growth Factor Binding Protein 7 Mediates Glioma Cell Growth and Migration

    Directory of Open Access Journals (Sweden)

    Wei Jiang

    2008-12-01

    Full Text Available Insulin-like growth factor binding protein 7 (IGFBP-7 is the only member of the IGFBP superfamily that binds strongly to insulin, suggesting that IGFBP-7 may have different functions from other IGFBPs. Unlike other IGFBPs, the expression and functions of IGFBP-7 in glioma tumors have not been reported. Using cDNA microarray analysis, we found that expression of IGFBP-7 correlated with the grade of glioma tumors and the overall patient survival. This finding was further validated by real-time reverse transcription-polymerase chain reaction and Western blot analysis. We used RNAi to examine the role of IGFBP-7 in glioma cells, inhibiting IGFBP-7 expression by short interfering RNA transfection. Cell proliferation was suppressed after IGFBP-7 expression was inhibited for 5 days, and glioma cell growth was stimulated consistently by the addition of recombinant IGFBP-7 protein. Moreover, glioma cell migration was attenuated by IGFBP-7 depletion but enhanced by IGFBP-7 overexpression and addition. Overexpression of AKT1 in IGFBP-7-overxpressed cells attenuated the IGFBP-7-promoted migration and further enhanced inhibition of IGFBP-7 depletion on the migration. Phosphorylation of AKT and Erk1/2 was also inversely regulated by IGFBP-7 expression. These two factors together suggest that IGFBP-7 can regulate glioma cell migration through the AKT-ERK pathway, thereby playing an important role in glioma growth and migration.

  1. Mathematical model of the glucose-insulin regulatory system: From the bursting electrical activity in pancreatic β-cells to the glucose dynamics in the whole body

    Science.gov (United States)

    Han, Kyungreem; Kang, Hyuk; Choi, M. Y.; Kim, Jinwoong; Lee, Myung-Shik

    2012-10-01

    A theoretical approach to the glucose-insulin regulatory system is presented. By means of integrated mathematical modeling and extensive numerical simulations, we probe the cell-level dynamics of the membrane potential, intracellular Ca2+ concentration, and insulin secretion in pancreatic β-cells, together with the whole-body level glucose-insulin dynamics in the liver, brain, muscle, and adipose tissues. In particular, the three oscillatory modes of insulin secretion are reproduced successfully. Such comprehensive mathematical modeling may provide a theoretical basis for the simultaneous assessment of the β-cell function and insulin resistance in clinical examination.

  2. Analyses of insulin-potentiating fragments of human growth hormone by computative simulation; essential unit for insulin-involved biological responses.

    Science.gov (United States)

    Ohkura, K; Hori, H

    2000-07-01

    these results we considered that the macrocyclic structure in human growth hormonal peptides is regarded with the modification of insulin action, and hGH(8-13) is an essential sequence for the modification of insulin action. This hGH(8-13) peptide modifies the insulin action via stabilizing the cell membrane, and does not directly act on the insulin-involved glucose metabolism.

  3. Impact of diet-induced obesity on intestinal stem cells: hyperproliferation but impaired intrinsic function that requires insulin/IGF1.

    Science.gov (United States)

    Mah, Amanda T; Van Landeghem, Laurianne; Gavin, Hannah E; Magness, Scott T; Lund, P Kay

    2014-09-01

    Nutrient intake regulates intestinal epithelial mass and crypt proliferation. Recent findings in model organisms and rodents indicate nutrient restriction impacts intestinal stem cells (ISC). Little is known about the impact of diet-induced obesity (DIO), a model of excess nutrient intake on ISC. We used a Sox9-EGFP reporter mouse to test the hypothesis that an adaptive response to DIO or associated hyperinsulinemia involves expansion and hyperproliferation of ISC. The Sox9-EGFP reporter mouse allows study and isolation of ISC, progenitors, and differentiated lineages based on different Sox9-EGFP expression levels. Sox9-EGFP mice were fed a high-fat diet for 20 weeks to induce DIO and compared with littermates fed low-fat rodent chow. Histology, fluorescence activated cell sorting, and mRNA analyses measured impact of DIO on jejunal crypt-villus morphometry, numbers, and proliferation of different Sox9-EGFP cell populations and gene expression. An in vitro culture assay directly assessed functional capacity of isolated ISC. DIO mice exhibited significant increases in body weight, plasma glucose, insulin, and insulin-like growth factor 1 (IGF1) levels and intestinal Igf1 mRNA. DIO mice had increased villus height and crypt density but decreased intestinal length and decreased numbers of Paneth and goblet cells. In vivo, DIO resulted in a selective expansion of Sox9-EGFP(Low) ISC and percentage of ISC in S-phase. ISC expansion significantly correlated with plasma insulin levels. In vitro, isolated ISC from DIO mice formed fewer enteroids in standard 3D Matrigel culture compared to controls, indicating impaired ISC function. This decreased enteroid formation in isolated ISC from DIO mice was rescued by exogenous insulin, IGF1, or both. We conclude that DIO induces specific increases in ISC and ISC hyperproliferation in vivo. However, isolated ISC from DIO mice have impaired intrinsic survival and growth in vitro that can be rescued by exogenous insulin or IGF1.

  4. Glycaemic responses after ingestion of some local foods by non-insulin dependent diabetic subjects.

    Science.gov (United States)

    Ayuo, P O; Ettyang, G A

    1996-12-01

    Fifteen non-insulin dependent diabetic volunteers, aged 51 +/- 3.9 years, were studied over a two month period to determine their glycaemic responses to various local foods. They were all on chlorpropamide and one subject was removed from analysis due to concurrent use of insulin. They received on separate occasions two servings of white bread, one serving of: brown bread, white lice, English potatoes, maize meal, millet and cassava each. Each meal contained 50 g of carbohydrate. A total of 107 glucose tolerance tests (GTTs) were performed and the glycaemic index (GI) for each food calculated. The mean blood sugars at 0,60 and 120 minutes were comparable for each food, and the peak rise occurred at 60 minutes. The highest rise (4.0 mmol/I) was seen with millet porridge. The highest GI was seen with white rice and the lowest with English potatoes (159.9 and 34.3 respectively). Overall, the cereals conferred higher GIs than the root vegetables. The GIs of English potatoes, maize meal, millet and cassava significantly differed from that of white bread. It is concluded that, using GIs, dietary guidelines comprising locally available and affordable foods can be made.

  5. Insulin-like growth factor-II receptors in cultured rat hepatocytes: regulation by cell density

    International Nuclear Information System (INIS)

    Scott, C.D.; Baxter, R.C.

    1987-01-01

    Insulin-like growth factor-II (IGF-II) receptors in primary cultures of adult rat hepatocytes were characterized and their regulation by cell density examined. In hepatocytes cultured at 5 X 10(5) cells per 3.8 cm2 plate [ 125 I]IGF-II bound to specific, high affinity receptors (Ka = 4.4 +/- 0.5 X 10(9) l/mol). Less than 1% cross-reactivity by IGF-I and no cross-reactivity by insulin were observed. IGF-II binding increased when cells were permeabilized with 0.01% digitonin, suggesting the presence of an intracellular receptor pool. Determined by Scatchard analysis and by polyacrylamide gel electrophoresis after affinity labeling, the higher binding was due solely to an increase in binding sites present on 220 kDa type II IGF receptors. In hepatocytes cultured at low densities, the number of cell surface receptors increased markedly, from 10-20,000 receptors per cell at a culture density of 6 X 10(5) cells/well to 70-80,000 receptors per cell at 0.38 X 10(5) cells/well. The increase was not due simply to the exposure of receptors from the intracellular pool, as a density-related increase in receptors was also seen in cells permeabilized with digitonin. There was no evidence that IGF binding proteins, either secreted by hepatocytes or present in fetal calf serum, had any effect on the measurement of receptor concentration or affinity. We conclude that rat hepatocytes in primary culture contain specific IGF-II receptors and that both cell surface and intracellular receptors are regulated by cell density

  6. Analysis of the effect of diabetes type 2 duration on beta cell secretory function and insulin resistance

    Directory of Open Access Journals (Sweden)

    Popović Ljiljana

    2006-01-01

    Full Text Available Diabetes type 2 is a chronic metabolic disorder. Pathogenesis of diabetes type 2 results from the impaired insulin secretion, impaired insulin action and increased endogenous glucose production. Diabetes evolves through several phases characterized by qualitative and quantitative changes of beta cell secretory function. The aim of our study was to analyze the impact of diabetes duration on beta cell secretory function and insulin resistance. The results indicated significant negative correlation of diabetes duration and fasting insulinemia, as well as beta cell secretory function assessed by HOMA β index. Our study also found significant negative correlation of diabetes duration and insulin resistance assessed by HOMA IR index. Significant positive correlation was established between beta cell secretory capacity (fasting insulinemia and HOMA β and insulin resistance assessed by HOMA IR index, independently of diabetes duration. These results indicate that: beta cell secretory capacity, assessed by HOMA β index, significantly decreases with diabetes duration. In parallel with decrease of fasting insulinemia, reduction of insulin resistance assessed by HOMA IR index was found as well.

  7. Novel covalently linked insulin dimer engineered to investigate the function of insulin dimerization.

    Directory of Open Access Journals (Sweden)

    Tine N Vinther

    Full Text Available An ingenious system evolved to facilitate insulin binding to the insulin receptor as a monomer and at the same time ensure sufficient stability of insulin during storage. Insulin dimer is the cornerstone of this system. Insulin dimer is relatively weak, which ensures dissociation into monomers in the circulation, and it is stabilized by hexamer formation in the presence of zinc ions during storage in the pancreatic β-cell. Due to the transient nature of insulin dimer, direct investigation of this important form is inherently difficult. To address the relationship between insulin oligomerization and insulin stability and function, we engineered a covalently linked insulin dimer in which two monomers were linked by a disulfide bond. The structure of this covalent dimer was identical to the self-association dimer of human insulin. Importantly, this covalent dimer was capable of further oligomerization to form the structural equivalent of the classical hexamer. The covalently linked dimer neither bound to the insulin receptor, nor induced a metabolic response in vitro. However, it was extremely thermodynamically stable and did not form amyloid fibrils when subjected to mechanical stress, underlining the importance of oligomerization for insulin stability.

  8. Energy expenditure, body composition and insulin response to glucose in male twins discordant for the Trp64Arg polymorphism of the β3-adrenergic receptor gene

    DEFF Research Database (Denmark)

    Højlund, Kurt; Christiansen, Christian; Bjørnsbo, K.S.

    2006-01-01

    AIM: The tryptophan to arginine change in position 64 (Trp64Arg) polymorphism of the beta3-adrenergic receptor (beta3AR) gene has been associated with an increased prevalence of obesity, insulin resistance and type 2 diabetes. In this, decreased rates of energy expenditure and impaired insulin...... and environmental background, the Trp64Arg polymorphism of the beta3AR gene is associated with lower fat mass, fasting insulin levels and an appropriate insulin response to glucose. Thus, heterozygosity for the Trp64Arg variant is unlikely to increase the risk of obesity, insulin resistance or type 2 diabetes....

  9. Retinoblastoma protein co-purifies with proteasomal insulin-degrading enzyme: Implications for cell proliferation control

    Energy Technology Data Exchange (ETDEWEB)

    Radulescu, Razvan T., E-mail: ratura@gmx.net [Molecular Concepts Research (MCR), Muenster (Germany); Duckworth, William C. [Department of Medicine, Phoenix VA Health Care System, Phoenix, AZ (United States); Levy, Jennifer L. [Research Service, Phoenix VA Health Care System, Phoenix, AZ (United States); Fawcett, Janet, E-mail: janet.fawcett@va.gov [Research Service, Phoenix VA Health Care System, Phoenix, AZ (United States)

    2010-04-30

    Previous investigations on proteasomal preparations containing insulin-degrading enzyme (IDE; EC 3.4.24.56) have invariably yielded a co-purifying protein with a molecular weight of about 110 kDa. We have now found both in MCF-7 breast cancer and HepG2 hepatoma cells that this associated molecule is the retinoblastoma tumor suppressor protein (RB). Interestingly, the amount of RB in this protein complex seemed to be lower in HepG2 vs. MCF-7 cells, indicating a higher (cytoplasmic) protein turnover in the former vs. the latter cells. Moreover, immunofluorescence showed increased nuclear localization of RB in HepG2 vs. MCF-7 cells. Beyond these subtle differences between these distinct tumor cell types, our present study more generally suggests an interplay between RB and IDE within the proteasome that may have important growth-regulatory consequences.

  10. The Nutrient-Responsive Hormone CCHamide-2 Controls Growth by Regulating Insulin-like Peptides in the Brain of Drosophila melanogaster.

    Science.gov (United States)

    Sano, Hiroko; Nakamura, Akira; Texada, Michael J; Truman, James W; Ishimoto, Hiroshi; Kamikouchi, Azusa; Nibu, Yutaka; Kume, Kazuhiko; Ida, Takanori; Kojima, Masayasu

    2015-05-01

    The coordination of growth with nutritional status is essential for proper development and physiology. Nutritional information is mostly perceived by peripheral organs before being relayed to the brain, which modulates physiological responses. Hormonal signaling ensures this organ-to-organ communication, and the failure of endocrine regulation in humans can cause diseases including obesity and diabetes. In Drosophila melanogaster, the fat body (adipose tissue) has been suggested to play an important role in coupling growth with nutritional status. Here, we show that the peripheral tissue-derived peptide hormone CCHamide-2 (CCHa2) acts as a nutrient-dependent regulator of Drosophila insulin-like peptides (Dilps). A BAC-based transgenic reporter revealed strong expression of CCHa2 receptor (CCHa2-R) in insulin-producing cells (IPCs) in the brain. Calcium imaging of brain explants and IPC-specific CCHa2-R knockdown demonstrated that peripheral-tissue derived CCHa2 directly activates IPCs. Interestingly, genetic disruption of either CCHa2 or CCHa2-R caused almost identical defects in larval growth and developmental timing. Consistent with these phenotypes, the expression of dilp5, and the release of both Dilp2 and Dilp5, were severely reduced. Furthermore, transcription of CCHa2 is altered in response to nutritional levels, particularly of glucose. These findings demonstrate that CCHa2 and CCHa2-R form a direct link between peripheral tissues and the brain, and that this pathway is essential for the coordination of systemic growth with nutritional availability. A mammalian homologue of CCHa2-R, Bombesin receptor subtype-3 (Brs3), is an orphan receptor that is expressed in the islet β-cells; however, the role of Brs3 in insulin regulation remains elusive. Our genetic approach in Drosophila melanogaster provides the first evidence, to our knowledge, that bombesin receptor signaling with its endogenous ligand promotes insulin production.

  11. Acute effect of meal glycemic index and glycemic load on blood glucose and insulin responses in humans

    Directory of Open Access Journals (Sweden)

    Díaz Erik

    2006-09-01

    Full Text Available Abstract Objective Foods with contrasting glycemic index when incorporated into a meal, are able to differentially modify glycemia and insulinemia. However, little is known about whether this is dependent on the size of the meal. The purposes of this study were: i to determine if the differential impact on blood glucose and insulin responses induced by contrasting GI foods is similar when provided in meals of different sizes, and; ii to determine the relationship between the total meal glycemic load and the observed serum glucose and insulin responses. Methods Twelve obese women (BMI 33.7 ± 2.4 kg/m2 were recruited. Subjects received 4 different meals in random order. Two meals had a low glycemic index (40–43% and two had a high-glycemic index (86–91%. Both meal types were given as two meal sizes with energy supply corresponding to 23% and 49% of predicted basal metabolic rate. Thus, meals with three different glycemic loads (95, 45–48 and 22 g were administered. Blood samples were taken before and after each meal to determine glucose, free-fatty acids, insulin and glucagon concentrations over a 5-h period. Results An almost 2-fold higher serum glucose and insulin incremental area under the curve (AUC over 2 h for the high- versus low-glycemic index same sized meals was observed (p Conclusion This study showed that foods of contrasting glycemic index induced a proportionally comparable difference in serum insulin response when provided in both small and large meals. The same was true for the serum glucose response but only in large meals. Glycemic load was useful in predicting the acute impact on blood glucose and insulin responses within the context of mixed meals.

  12. Insulin response of the glucose and fatty acid metabolism in dry dairy cows across a range of body condition scores.

    Science.gov (United States)

    De Koster, J; Hostens, M; Van Eetvelde, M; Hermans, K; Moerman, S; Bogaert, H; Depreester, E; Van den Broeck, W; Opsomer, G

    2015-07-01

    The objective of the present research was to determine the insulin response of the glucose and fatty acid metabolism in dry dairy cows with a variable body condition score (BCS). Ten pregnant Holstein Friesian dairy cows (upcoming parity 2 to 5) were selected based on BCS at the beginning of the study (2mo before expected parturition date). During the study, animals were monitored weekly for BCS and backfat thickness and in the last 2wk, blood samples were taken for determination of serum nonesterified fatty acid (NEFA) concentration. Animals underwent a hyperinsulinemic euglycemic clamp test in the third week before the expected parturition date. The hyperinsulinemic euglycemic clamp test consisted of 4 consecutive insulin infusions with increasing insulin doses: 0.1, 0.5, 2, and 5mIU/kg per minute. For each insulin infusion period, a steady state was defined as a period of 30min where no or minor changes of the glucose infusion were necessary to keep the blood glucose concentration constant and near basal levels. During the steady state, the glucose infusion rate [steady state glucose infusion rate (SSGIR) in µmol/kg per minute] and NEFA concentration [steady state NEFA concentration (SSNEFA) in mmol/L] were determined and reflect the insulin response of the glucose and fatty acid metabolism. Dose response curves were created based on the insulin concentrations during the steady state and the SSGIR or SSNEFA. The shape of the dose response curves is determined by the concentration of insulin needed to elicit the half maximal effect (EC50) and the maximal SSGIR or the minimal SSNEFA for the glucose or fatty acid metabolism, respectively. The maximal SSGIR was negatively associated with variables reflecting adiposity of the cows (BCS, backfat thickness, NEFA concentration during the dry period, and absolute weight of the different adipose depots determined after euthanasia and dissection of the different depots), whereas the EC50 of the glucose metabolism was

  13. Studies on Several Hormone Responses Following Intravenous Alimentation: Insulin and growth hormone responses following oral or intravenous alimentation in patient with far advanced gastric cancer

    Energy Technology Data Exchange (ETDEWEB)

    Sung, H K; Koh, J H; Ryu, Y W; Lee, J O; Lee, C W; Kim, J Y; Lee, J K [Korea Atomic Energy Research Institute, Seoul (Korea, Republic of)

    1975-09-15

    Glucose tolerance, insulin and growth hormone responses following glucose for amino acids administration by means of parenteral or oral load were studied in patients with far advanced gastric cancer. Hormone responses following nutrients load showed in patients with gastric cancer were compared to those of healthy subjects. Results were as follows:1) Blood sugar appearance following oral glucose administration was diminished in patients with far advanced gastric cancer. 2) The insulin responses of gastric cancer following oral glucose were also diminished as compared to that of normal subjects and were identical with parenteral route. 3) Parenteral administration of glucose or amino acids to patients with gastric cancer resulted in a increase of plasma growth hormone level. 4) Lower insulin response to amino acids was observed on parenteral administration in patient with gastric cancer as in healthy subjects. 5) Author discussed that the low insulin response after oral glucose administration showed in gastric cancer, and any additional insulin requirement arise when longer periods of parenteral amino acid administration are necessary, as in the patient with malnutrition.

  14. Studies on Several Hormone Responses Following Intravenous Alimentation: Insulin and growth hormone responses following oral or intravenous alimentation in patient with far advanced gastric cancer

    International Nuclear Information System (INIS)

    Sung, H. K.; Koh, J. H.; Ryu, Y. W.; Lee, J. O.; Lee, C. W.; Kim, J. Y.; Lee, J. K.

    1975-01-01

    Glucose tolerance, insulin and growth hormone responses following glucose for amino acids administration by means of parenteral or oral load were studied in patients with far advanced gastric cancer. Hormone responses following nutrients load showed in patients with gastric cancer were compared to those of healthy subjects. Results were as follows:1) Blood sugar appearance following oral glucose administration was diminished in patients with far advanced gastric cancer. 2) The insulin responses of gastric cancer following oral glucose were also diminished as compared to that of normal subjects and were identical with parenteral route. 3) Parenteral administration of glucose or amino acids to patients with gastric cancer resulted in a increase of plasma growth hormone level. 4) Lower insulin response to amino acids was observed on parenteral administration in patient with gastric cancer as in healthy subjects. 5) Author discussed that the low insulin response after oral glucose administration showed in gastric cancer, and any additional insulin requirement arise when longer periods of parenteral amino acid administration are necessary, as in the patient with malnutrition.

  15. Clustering effects on postprandial insulin secretion and sensitivity in response to meals with different fatty acid compositions.

    Science.gov (United States)

    Bermudez, Beatriz; Ortega-Gomez, Almudena; Varela, Lourdes M; Villar, Jose; Abia, Rocio; Muriana, Francisco J G; Lopez, Sergio

    2014-07-25

    Dietary fatty acids play a role in glucose homeostasis. The aim of this study was to assess the individual relationship between dietary saturated (SFA), monounsaturated (MUFA) and polyunsaturated (PUFA) fatty acids with postprandial β-cell function and insulin sensitivity in subjects with normal and high fasting triglycerides. We assessed postprandial β-cell function (by the insulinogenic index and the ratio of the insulin to glucose areas under the time-concentration curve) and insulin sensitivity (by the oral glucose and the minimal model insulin sensitivity indices) over four nonconsecutive, randomly assigned, high-fat meals containing a panel of SFA (palmitic and stearic acids), MUFA (palmitoleic and oleic acids) and PUFA (linoleic and α-linolenic acids) in 14 subjects with normal and in 14 subjects with high fasting triglycerides. The proportions of each fatty acid in the meals and the values for surrogate measures of postprandial β-cell function and insulin sensitivity were subjected to a Pearson correlation and hierarchical cluster analysis, which revealed two classes of dietary fatty acids for regulating postprandial glucose homeostasis. We successfully discriminated the adverse effects of SFA palmitic acid from the beneficial effects of MUFA oleic acid on postprandial β-cell function (r ≥ 0.84 for SFA palmitic acid and r ≥ -0.71 for MUFA oleic acid; P < 0.05) and insulin sensitivity (r ≥ -0.92 for SFA palmitic acid and r ≥ 0.89 for MUFA oleic acid; P < 0.001) both in subjects with normal and high fasting triglycerides. In conclusion, dietary MUFA oleic acid, in contrast to SFA palmitic acid, favours the tuning towards better postprandial glycaemic control in subjects with normal and high fasting triglycerides.

  16. Metformin downregulates the insulin/IGF-I signaling pathway and inhibits different uterine serous carcinoma (USC) cells proliferation and migration in p53-dependent or -independent manners.

    Science.gov (United States)

    Sarfstein, Rive; Friedman, Yael; Attias-Geva, Zohar; Fishman, Ami; Bruchim, Ilan; Werner, Haim

    2013-01-01

    Accumulating epidemiological evidence shows that obesity is associated with an increased risk of several types of adult cancers, including endometrial cancer. Chronic hyperinsulinemia, a typical hallmark of diabetes, is one of the leading factors responsible for the obesity-cancer connection. Numerous cellular and circulating factors are involved in the biochemical chain of events leading from hyperinsulinemia and insulin resistance to increased cancer risk and, eventually, tumor development. Metformin is an oral anti-diabetic drug of the biguanide family used for treatment of type 2 diabetes. Recently, metformin was shown to exhibit anti-proliferative effects in ovarian and Type I endometrial cancer, although the mechanisms responsible for this non-classical metformin action remain unclear. The insulin-like growth factors (IGFs) play a prominent role in cancer biology and their mechanisms of action are tightly interconnected with the insulin signaling pathways. Given the cross-talk between the insulin and IGF signaling pathways, the aim of this study was to examine the hypothesis that the anti-proliferative actions of metformin in uterine serous carcinoma (USC) are potentially mediated via suppression of the IGF-I receptor (IGF-IR) pathway. Our results show that metformin interacts with the IGF pathway, and induces apoptosis and inhibition of proliferation and migration of USC cell lines with both wild type and mutant p53. Taken together, our results suggest that metformin therapy could be a novel and attractive therapeutic approach for human USC, a highly aggressive variant of endometrial cancer.

  17. American Ginseng Stimulates Insulin Production and Prevents Apoptosis through Regulation of Uncoupling Protein-2 in Cultured β Cells

    Directory of Open Access Journals (Sweden)

    John Zeqi Luo

    2006-01-01

    Full Text Available American ginseng root displays the ability to achieve glucose homeostasis both experimentally and clinically but the unknown mechanism used by ginseng to achieve its therapeutic effects on diabetes limits its application. Disruption in the insulin secretion of pancreatic β cells is considered the major cause of diabetes. A mitochondrial protein, uncoupling protein-2 (UCP-2 has been found to play a critical role in insulin synthesis and β cell survival. Our preliminary studies found that the extracts of American ginseng inhibit UCP-2 expression which may contribute to the ability of ginseng protecting β cell death and improving insulin synthesis. Therefore, we hypothesized that ginseng extracts suppress UCP-2 in the mitochondria of pancreatic β cells, promoting insulin synthesis and anti-apoptosis (a programmed cell-death mechanism. To test the hypothesis, the serum-deprived quiescent β cells were cultured with or without interleukin-1β (IL-1β, (200 pg ml−1, a cytokine to induce β cell apoptosis and water extracts of American ginseng (25 μg per 5 μl administered to wells of 0.5 ml culture for 24 h. We evaluated effects of ginseng on UCP-2 expression, insulin production, anti-/pro-apoptotic factors Bcl-2/caspase-9 expression and cellular ATP levels. We found that ginseng suppresses UCP-2, down-regulates caspase-9 while increasing ATP and insulin production/secretion and up-regulates Bcl-2, reducing apoptosis. These findings suggest that stimulation of insulin production and prevention of β cell loss by American ginseng extracts can occur via the inhibition of mitochondrial UCP-2, resulting in increase in the ATP level and the anti-apoptotic factor Bcl-2, while down-regulation of pro-apoptotic factor caspase-9 occurs, lowering the occurrence of apoptosis, which support the hypothesis.

  18. Fumosorinone, a novel PTP1B inhibitor, activates insulin signaling in insulin-resistance HepG2 cells and shows anti-diabetic effect in diabetic KKAy mice

    International Nuclear Information System (INIS)

    Liu, Zhi-Qin; Liu, Ting; Chen, Chuan; Li, Ming-Yan; Wang, Zi-Yu; Chen, Ruo-song; Wei, Gui-xiang; Wang, Xiao-yi; Luo, Du-Qiang

    2015-01-01

    Insulin resistance is a characteristic feature of type 2 diabetes mellitus (T2DM) and is characterized by defects in insulin signaling. Protein tyrosine phosphatase 1B (PTP1B) is a key negative regulator of the insulin signaling pathways, and its increased activity and expression are implicated in the pathogenesis of insulin resistance. Therefore, the inhibition of PTP1B is anticipated to become a potential therapeutic strategy to treat T2DM. Fumosorinone (FU), a new natural product isolated from insect fungi Isaria fumosorosea, was found to inhibit PTP1B activity in our previous study. Herein, the effects of FU on insulin resistance and mechanism in vitro and in vivo were investigated. FU increased the insulin-provoked glucose uptake in insulin-resistant HepG2 cells, and also reduced blood glucose and lipid levels of type 2 diabetic KKAy mice. FU decreased the expression of PTP1B both in insulin-resistant HepG2 cells and in liver tissues of diabetic KKAy mice. Furthermore, FU increased the phosphorylation of IRβ, IRS-2, Akt, GSK3β and Erk1/2 in insulin-resistant HepG2 cells, as well as the phosphorylation of IRβ, IRS-2, Akt in liver tissues of diabetic KKAy mice. These results showed that FU increased glucose uptake and improved insulin resistance by down-regulating the expression of PTP1B and activating the insulin signaling pathway, suggesting that it may possess antidiabetic properties. - Highlights: • Fumosorinone is a new PTP1B inhibitor isolated from insect pathogenic fungi. • Fumosorinone attenuated the insulin resistance both in vitro and in vivo. • Fumosorinone decreased the expression of PTP1B both in vitro and in vivo. • Fumosorinone activated the insulin signaling pathway both in vitro and in vivo

  19. Fumosorinone, a novel PTP1B inhibitor, activates insulin signaling in insulin-resistance HepG2 cells and shows anti-diabetic effect in diabetic KKAy mice

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Zhi-Qin [College of Life Sciences, Key Laboratory of Medicinal Chemistry and Molecular Diagnosis of Ministry of Education, Hebei University, Baoding 071002 (China); College of Pharmaceutical Sciences, key laboratory of pharmaceutical quality control of Hebei province, Hebei University, Baoding 071002 (China); Liu, Ting; Chen, Chuan [College of Life Sciences, Key Laboratory of Medicinal Chemistry and Molecular Diagnosis of Ministry of Education, Hebei University, Baoding 071002 (China); Li, Ming-Yan; Wang, Zi-Yu; Chen, Ruo-song; Wei, Gui-xiang; Wang, Xiao-yi [College of Pharmaceutical Sciences, key laboratory of pharmaceutical quality control of Hebei province, Hebei University, Baoding 071002 (China); Luo, Du-Qiang, E-mail: duqiangluo999@126.com [College of Life Sciences, Key Laboratory of Medicinal Chemistry and Molecular Diagnosis of Ministry of Education, Hebei University, Baoding 071002 (China)

    2015-05-15

    Insulin resistance is a characteristic feature of type 2 diabetes mellitus (T2DM) and is characterized by defects in insulin signaling. Protein tyrosine phosphatase 1B (PTP1B) is a key negative regulator of the insulin signaling pathways, and its increased activity and expression are implicated in the pathogenesis of insulin resistance. Therefore, the inhibition of PTP1B is anticipated to become a potential therapeutic strategy to treat T2DM. Fumosorinone (FU), a new natural product isolated from insect fungi Isaria fumosorosea, was found to inhibit PTP1B activity in our previous study. Herein, the effects of FU on insulin resistance and mechanism in vitro and in vivo were investigated. FU increased the insulin-provoked glucose uptake in insulin-resistant HepG2 cells, and also reduced blood glucose and lipid levels of type 2 diabetic KKAy mice. FU decreased the expression of PTP1B both in insulin-resistant HepG2 cells and in liver tissues of diabetic KKAy mice. Furthermore, FU increased the phosphorylation of IRβ, IRS-2, Akt, GSK3β and Erk1/2 in insulin-resistant HepG2 cells, as well as the phosphorylation of IRβ, IRS-2, Akt in liver tissues of diabetic KKAy mice. These results showed that FU increased glucose uptake and improved insulin resistance by down-regulating the expression of PTP1B and activating the insulin signaling pathway, suggesting that it may possess antidiabetic properties. - Highlights: • Fumosorinone is a new PTP1B inhibitor isolated from insect pathogenic fungi. • Fumosorinone attenuated the insulin resistance both in vitro and in vivo. • Fumosorinone decreased the expression of PTP1B both in vitro and in vivo. • Fumosorinone activated the insulin signaling pathway both in vitro and in vivo.

  20. [Associations of insulin resistance and pancreatic beta-cell function with plasma glucose level in type 2 diabetes].

    Science.gov (United States)

    Nian, Xiaoping; Sun, Gaisheng; Dou, Chunmei; Hou, Hongbo; Fan, Xiuping; Yu, Hongmei; Ma, Ling; He, Bingxian

    2002-06-10

    To investigate the influence of insulin resistance and pancreatic beta-cell function on plasma glucose level in type 2 diabetes so as to provide theoretical basis for reasonable selection of hypoglycemic agents. The plasma non-specific insulin (NSINS), true insulin (TI) and glucose in eight-one type 2 diabetics, 38 males and 43 females, with a mean age of 53 years, were examined 0, 30, 60 and 120 minutes after they had 75 grams of instant noodles. The patients were divided into two groups according to their fasting plasma glucose (FPG): group A (FPG = 8.89 mmol/L). The insulin resistance was evaluated by HOMA-IR, the beta-cell function was evaluated by HOMA-beta formula and the formula deltaI(30)/deltaG(30) = (deltaI(30)-deltaI(0))/(deltaG(30)-deltaG(0)). The insulin area under curve (INSAUC) was evaluated by the formula INSAUC=FINS/2+INS(30)+INS(60)+INS(120)/2. The mean FPG was 6.23 mmol/L in group A and 12.6 mmol/L in group B. PG2H was 11.7 mmol/L in group A and 19.2 mmol/L in group B. The TI levels in group B at 0, 30, 60, 120 min during standard meal test were significantly higher than those in group A: 6.15 +/- 1.06 vs 4.77 +/- 1.06, 9.76 +/- 1.1 vs 5.88 +/- 1.1,14.68 +/- 1.11 vs 6.87 +/- 1.1 and 17.13 +/- 1.12 vs 8.0 +/- 1.1 microU/dl (all P< 0.01). The NSINS showed the same trend. The insulin resistance in group B was 1.5 times that in group A. With the insulin resistance adjusted, the beta cell function in group A was 5 to 6 times that in group B. The INSAUC in group A was 1.66 times larger than that in group B, especially the INSAUC for true insulin (2 times larger). The contribution of insulin resistance and beta cell function to PG2H was half by half in group A and 1:8 in group B. beta cell function calculated by insulin (Homa-beta) explained 41% of the plasma glucose changes in group A and 54% of the plasma glucose changes in group B. The contribution of insulin deficiency to plasma glocose was 3.3.times that of insulin resistance in group A and was 9