Science.gov

Sample records for premixed human insulin

  1. PROGENS-HbA1c study: safety and effectiveness of premixed recombinant human insulin (Gensulin M30)

    PubMed Central

    Walicka, Magdalena; Jóźwiak, Jacek; Rzeszotarski, Jacek; Zarzycka-Lindner, Grażyna; Zonenberg, Anna; Bijoś, Paweł; Masierek, Małgorzata

    2016-01-01

    Introduction Insulin analogues have gained widespread popularity. However, in many countries the use of these drugs is limited by their relatively high cost, so there is still a need for more cost-effective human insulin therapies. The aim of the study was to assess the effectiveness and safety of the premixed recombinant human insulin (rhuI) Gensulin M30 in a real-life setting. Material and methods The study group consisted of 4257 patients (2196 female, 2061 male) with type 2 diabetes, aged 63.7 ±9.4, with body mass index (BMI) 30.3 ±4.5 kg/m2 and diabetes duration 9 ±5.5 years. All patients were treated with premixed rhuI Gensulin M30. In 91.7% of patients, insulin was used in combination with metformin. In 3.7% of patients, it was used with sulphonylureas. The patients were observed for a period of 6 months. Results The total insulin dose on visit 1 was 36.1 ±18.7 U (0.42 ±0.22 U/kg), and by the end of the study it reached 40.3 ±18.9 U (0.48 ±0.22 U/kg). A significant, continuous decrease of the levels of glycated hemoglobin (HbA1c), along with fasting and postprandial plasma glucose, was observed during the study period. The frequency of hypoglycemia increased slightly during the study, although these figures remained low, especially with regard to severe hypoglycemic episodes (0.02 episodes/patient/year). The lowest number of hypoglycemic episodes occurred in patients treated with insulin and metformin, while the highest number of episodes was observed in patients treated with insulin alone. No weight changes were noted in the patients during the study. Conclusions This study shows rhuI Gensulin M30 to be effective and safe in a real-life setting.

  2. Interactions of short-acting, intermediate-acting and pre-mixed human insulins with free radicals--Comparative EPR examination.

    PubMed

    Olczyk, Paweł; Komosinska-Vassev, Katarzyna; Ramos, Paweł; Mencner, Łukasz; Olczyk, Krystyna; Pilawa, Barbara

    2015-07-25

    Electron paramagnetic resonance (EPR) spectroscopy was used to examine insulins interactions with free radicals. Human recombinant DNA insulins of three groups were studied: short-acting insulin (Insuman Rapid); intermediate-acting insulins (Humulin N, Insuman Basal), and pre-mixed insulins (Humulin M3, Gensulin M50, Gensulin M40, Gensulin M30). The aim of an X-band (9.3GHz) study was comparative analysis of antioxidative properties of the three groups of human insulins. DPPH was used as a stable free radical model. Amplitudes of EPR lines of DPPH as the paramagnetic free radical reference, and DPPH interacting with the individual tested insulins were compared. For all the examined insulins kinetics of their interactions with free radicals up to 60 min were obtained. The strongest interactions with free radicals were observed for the short-acting insulin - Insuman Rapid. The lowest interactions with free radicals were characteristic for intermediate-acting insulin - Insuman Basal. The pre-mixed insulins i.e. Humulin M3 and Gensulin M50 revealed the fastest interactions with free radicals. The short acting, intermediate acting and premixed insulins have been found to be effective agents in reducing free radical formation in vitro and should be further considered as potential useful tools in attenuation of oxidative stress in diabetic patients.

  3. Interactions of short-acting, intermediate-acting and pre-mixed human insulins with free radicals--Comparative EPR examination.

    PubMed

    Olczyk, Paweł; Komosinska-Vassev, Katarzyna; Ramos, Paweł; Mencner, Łukasz; Olczyk, Krystyna; Pilawa, Barbara

    2015-07-25

    Electron paramagnetic resonance (EPR) spectroscopy was used to examine insulins interactions with free radicals. Human recombinant DNA insulins of three groups were studied: short-acting insulin (Insuman Rapid); intermediate-acting insulins (Humulin N, Insuman Basal), and pre-mixed insulins (Humulin M3, Gensulin M50, Gensulin M40, Gensulin M30). The aim of an X-band (9.3GHz) study was comparative analysis of antioxidative properties of the three groups of human insulins. DPPH was used as a stable free radical model. Amplitudes of EPR lines of DPPH as the paramagnetic free radical reference, and DPPH interacting with the individual tested insulins were compared. For all the examined insulins kinetics of their interactions with free radicals up to 60 min were obtained. The strongest interactions with free radicals were observed for the short-acting insulin - Insuman Rapid. The lowest interactions with free radicals were characteristic for intermediate-acting insulin - Insuman Basal. The pre-mixed insulins i.e. Humulin M3 and Gensulin M50 revealed the fastest interactions with free radicals. The short acting, intermediate acting and premixed insulins have been found to be effective agents in reducing free radical formation in vitro and should be further considered as potential useful tools in attenuation of oxidative stress in diabetic patients. PMID:25975232

  4. PROGENS-HbA1c study: safety and effectiveness of premixed recombinant human insulin (Gensulin M30)

    PubMed Central

    Walicka, Magdalena; Jóźwiak, Jacek; Rzeszotarski, Jacek; Zarzycka-Lindner, Grażyna; Zonenberg, Anna; Bijoś, Paweł; Masierek, Małgorzata

    2016-01-01

    Introduction Insulin analogues have gained widespread popularity. However, in many countries the use of these drugs is limited by their relatively high cost, so there is still a need for more cost-effective human insulin therapies. The aim of the study was to assess the effectiveness and safety of the premixed recombinant human insulin (rhuI) Gensulin M30 in a real-life setting. Material and methods The study group consisted of 4257 patients (2196 female, 2061 male) with type 2 diabetes, aged 63.7 ±9.4, with body mass index (BMI) 30.3 ±4.5 kg/m2 and diabetes duration 9 ±5.5 years. All patients were treated with premixed rhuI Gensulin M30. In 91.7% of patients, insulin was used in combination with metformin. In 3.7% of patients, it was used with sulphonylureas. The patients were observed for a period of 6 months. Results The total insulin dose on visit 1 was 36.1 ±18.7 U (0.42 ±0.22 U/kg), and by the end of the study it reached 40.3 ±18.9 U (0.48 ±0.22 U/kg). A significant, continuous decrease of the levels of glycated hemoglobin (HbA1c), along with fasting and postprandial plasma glucose, was observed during the study period. The frequency of hypoglycemia increased slightly during the study, although these figures remained low, especially with regard to severe hypoglycemic episodes (0.02 episodes/patient/year). The lowest number of hypoglycemic episodes occurred in patients treated with insulin and metformin, while the highest number of episodes was observed in patients treated with insulin alone. No weight changes were noted in the patients during the study. Conclusions This study shows rhuI Gensulin M30 to be effective and safe in a real-life setting. PMID:27695488

  5. Role of premixed insulin analogues in the treatment of patients with type 2 diabetes mellitus: A narrative review

    PubMed Central

    Elizarova, Svetlana; Galstyan, Gagik R; Wolffenbuttel, Bruce HR

    2014-01-01

    Because of the progressive nature of type 2 diabetes mellitus (T2DM), insulin therapy will eventually become necessary in most patients. Recent evidence suggests that maintaining optimal glycemic control by early insulin therapy can reduce the risk of microvascular and macrovascular complications in patients with T2DM. The present review focuses on relevant clinical evidence supporting the use of premixed insulin analogues in T2DM when intensifying therapy, and as starter insulins in insulin-naïve patients. Our aim is to provide relevant facts and clinical evidence useful in the decision-making process of treatment selection and individualized treatment goal setting to obtain sustained blood glucose control. PMID:24127999

  6. Role of premixed insulin analogues in the treatment of patients with type 2 diabetes mellitus: a narrative review.

    PubMed

    Elizarova, Svetlana; Galstyan, Gagik R; Wolffenbuttel, Bruce H R

    2014-03-01

    Because of the progressive nature of type 2 diabetes mellitus (T2DM), insulin therapy will eventually become necessary in most patients. Recent evidence suggests that maintaining optimal glycemic control by early insulin therapy can reduce the risk of microvascular and macrovascular complications in patients with T2DM. The present review focuses on relevant clinical evidence supporting the use of premixed insulin analogues in T2DM when intensifying therapy, and as starter insulins in insulin-naïve patients. Our aim is to provide relevant facts and clinical evidence useful in the decision-making process of treatment selection and individualized treatment goal setting to obtain sustained blood glucose control. PMID:24127999

  7. Protein Crystal Recombinant Human Insulin

    NASA Technical Reports Server (NTRS)

    1994-01-01

    The comparison of protein crystal, Recombiant Human Insulin; space-grown (left) and earth-grown (right). On STS-60, Spacehab II indicated that space-grown crystals are larger and of greater optical clarity than their earth-grown counterparts. Recombiant Human Insulin facilitates the incorporation of glucose into cells. In diabetics, there is either a decrease in or complete lack of insulin, thereby leading to several harmful complications. Principal Investigator is Larry DeLucas.

  8. Insulin Human Inhalation

    MedlinePlus

    ... insulin and therefore cannot control the amount of sugar in the blood). It is also used in ... normally and, therefore, cannot control the amount of sugar in the blood) who need insulin to control ...

  9. [Comparison of biosynthetic human insulin and purified pork insulin. Studies in insulin-resistant obese patients using the insulin suppression test].

    PubMed

    Richard, J L; Rodier, M; Cavalie, G; Lachkar, H; Orsetti, A; Monnier, L; Mirouze, J

    1986-02-01

    An insulin suppression test performed in random order with either biosynthetic human insulin or purified pork insulin was used to compare biological activity of these two insulins in obese patients suffering from varying degrees of glucose intolerance. Blood glucose curve, steady-state blood glucose levels, insulin sensitivity indices and steady-state plasma insulin levels were identical during the two sets of tests. Furthermore endogenous insulin and glucagon secretion were similarly suppressed. The insulin suppression test is a simple and rapid procedure to compare the biological activity of fast-acting insulins. Our results confirm the insulin-resistance in obesity and clearly show that biosynthetic human and porcine insulins have similar biological potency.

  10. Insulin is ubiquitous in extrapancreatic tissues of rats and humans.

    PubMed Central

    Rosenzweig, J L; Havrankova, J; Lesniak, M A; Brownstein, M; Roth, J

    1980-01-01

    Insulin has been detected, at levels higher than those in plasma, in a broad range of extrapancreatic tissues in both rats and humans. Rat liver insulin was shown to be indistinguishable from genuine insulin by radioimmunoassay, Sephadex chromatography, bioassay, and antibody neutralization. Liver insulin (like brain insulin) was unchanged in ob/ob mice, in rats treated with streptozotocin, or in fasted rats, despite marked alterations in pancreatic secretion of insulin and in liver content of insulin receptors. Insulin was found in cultured human IM-9 lymphocytes and cultured fibroblasts at concentrations greater than 100 times the levels in the media. IM-9 lymphocyte insulin also was shown to be indistinguishable from genuine insulin, by the same criteria used for liver insulin. The insulin concentration in cultured human cells was unaffected by depletion of insulin from the culture medium or by addition of beef insulin to the medium. The data suggest that a part, if not all, of the extrapancreatic tissue insulin is independent of plasma insulin and may be synthesized by the tissues themselves. PMID:6987656

  11. Electrochemically triggered release of human insulin from an insulin-impregnated reduced graphene oxide modified electrode.

    PubMed

    Teodorescu, Florina; Rolland, Laure; Ramarao, Viswanatha; Abderrahmani, Amar; Mandler, Daniel; Boukherroub, Rabah; Szunerits, Sabine

    2015-09-28

    An electrochemical insulin-delivery system based on reduced graphene oxide impregnated with insulin is described. Upon application of a potential pulse of -0.8 V for 30 min, up to 70 ± 4% of human insulin was released into a physiological medium while preserving its biological activity.

  12. Insulin action in the human brain: evidence from neuroimaging studies.

    PubMed

    Kullmann, S; Heni, M; Fritsche, A; Preissl, H

    2015-06-01

    Thus far, little is known about the action of insulin in the human brain. Nonetheless, recent advances in modern neuroimaging techniques, such as functional magnetic resonance imaging (fMRI) or magnetoencephalography (MEG), have made it possible to investigate the action of insulin in the brain in humans, providing new insights into the pathogenesis of brain insulin resistance and obesity. Using MEG, the clinical relevance of the action of insulin in the brain was first identified, linking cerebral insulin resistance with peripheral insulin resistance, genetic predisposition and weight loss success in obese adults. Although MEG is a suitable tool for measuring brain activity mainly in cortical areas, fMRI provides high spatial resolution for cortical as well as subcortical regions. Thus, the action of insulin can be detected within all eating behaviour relevant regions, which include regions deeply located within the brain, such as the hypothalamus, midbrain and brainstem, as well as regions within the striatum. In this review, we outline recent advances in the field of neuroimaging aiming to investigate the action of insulin in the human brain using different routes of insulin administration. fMRI studies have shown a significant insulin-induced attenuation predominantly in the occipital and prefrontal cortical regions and the hypothalamus, successfully localising insulin-sensitive brain regions in healthy, mostly normal-weight individuals. However, further studies are needed to localise brain areas affected by insulin resistance in obese individuals, which is an important prerequisite for selectively targeting brain insulin resistance in obesity.

  13. Insulin-like substance and insulin-degrading complex of hemolysate of human erythrocytes

    SciTech Connect

    Matulyavichyus, V.A.; Vareikis, E.I.; Lashas, L.V.

    1986-08-20

    A lysate of human erythrocytes was fractionated on gel-filtration resins of different types and immunoreactive insulin, the insulinase activity and the effect of individual fractions on the insulinase activity was determined in the fractions obtained. It was established that the hemolysate contains a complex of insulin-metabolizing compounds, including an insulin-like substance, insulinase, and an inhibitor and activator of the insulinase activity. The insulin-like substance coincided with native insulin in site of elution from a column of Sephadex G-50 and its concentration in the lysate exceeded that of insulin in the blood plasma. Insulinase, which has a molecular weight of about 100,000, cleaved (/sup 125/I) insulin to fragments soluble in trichloroacetic acid, but had no effect on hypophyseal proteins and glycoprotein hormones. The insulinase activity was inhibited by low temperatures, atropine, and a newly discovered intraerythrocytic proteinase inhibitor, which also inhibits the serine proteinases trypsin and chymotrypsin. A substance eluted from a column of Sephadex G-100 in the region of low-molecular-weight substances increased the insulinase activity. The elution curve of substances with proteinase-inhibiting and insulinase-activating activities indicates that there is more than one inhibitory and activating factor. The results of the studies suggest that the insulin-degrading complex in human erythrocytes acts as a regulator of the insulin level in the blood plasma. It is also possible that the insulin-like substance is produced in the cytosol of the erythrocytes.

  14. Mechanisms of human insulin resistance and thiazolidinedione-mediated insulin sensitization

    PubMed Central

    Sears, D. D.; Hsiao, G.; Hsiao, A.; Yu, J. G.; Courtney, C. H.; Ofrecio, J. M.; Chapman, J.; Subramaniam, S.

    2009-01-01

    Cellular and tissue defects associated with insulin resistance are coincident with transcriptional abnormalities and are improved after insulin sensitization with thiazolidinedione (TZD) PPARγ ligands. We characterized 72 human subjects by relating their clinical phenotypes with functional pathway alterations. We transcriptionally profiled 364 biopsies harvested before and after hyperinsulinemic-euglycemic clamp studies, at baseline and after 3-month TZD treatment. We have identified molecular and functional characteristics of insulin resistant subjects and distinctions between TZD treatment responder and nonresponder subjects. Insulin resistant subjects exhibited alterations in skeletal muscle (e.g., glycolytic flux and intramuscular adipocytes) and adipose tissue (e.g., mitochondrial metabolism and inflammation) that improved relative to TZD-induced insulin sensitization. Pre-TZD treatment expression of MLXIP in muscle and HLA-DRB1 in adipose tissue from insulin resistant subjects was linearly predictive of post-TZD insulin sensitization. We have uniquely characterized coordinated cellular and tissue functional pathways that are characteristic of insulin resistance, TZD-induced insulin sensitization, and potential TZD responsiveness. PMID:19841271

  15. Pancreatic expression of human insulin gene in transgenic mice.

    PubMed

    Bucchini, D; Ripoche, M A; Stinnakre, M G; Desbois, P; Lorès, P; Monthioux, E; Absil, J; Lepesant, J A; Pictet, R; Jami, J

    1986-04-01

    We have investigated the possibility of obtaining integration and expression of a native human gene in transgenic mice. An 11-kilobase (kb) human chromosomal DNA fragment including the insulin gene (1430 base pairs) was microinjected into fertilized mouse eggs. This fragment was present in the genomic DNA of several developing animals. One transgenic mouse and its progeny were analyzed for expression of the foreign gene. Synthesis and release of human insulin was revealed by detection of the human C-peptide in the plasma and urine. Human insulin mRNA was found in pancreas but not in other tissues. These findings indicate that the 11-kb human DNA fragment carries the sequences necessary for tissue-specific expression of the insulin gene and the human regulatory sequences react to homologous signals in the mouse.

  16. Molecular basis of catalytic chamber-assisted unfolding and cleavage of human insulin by human insulin-degrading enzyme.

    PubMed

    Manolopoulou, Marika; Guo, Qing; Malito, Enrico; Schilling, Alexander B; Tang, Wei-Jen

    2009-05-22

    Insulin is a hormone vital for glucose homeostasis, and insulin-degrading enzyme (IDE) plays a key role in its clearance. IDE exhibits a remarkable specificity to degrade insulin without breaking the disulfide bonds that hold the insulin A and B chains together. Using Fourier transform ion cyclotron resonance (FTICR) mass spectrometry to obtain high mass accuracy, and electron capture dissociation (ECD) to selectively break the disulfide bonds in gas phase fragmentation, we determined the cleavage sites and composition of human insulin fragments generated by human IDE. Our time-dependent analysis of IDE-digested insulin fragments reveals that IDE is highly processive in its initial cleavage at the middle of both the insulin A and B chains. This ensures that IDE effectively splits insulin into inactive N- and C-terminal halves without breaking the disulfide bonds. To understand the molecular basis of the recognition and unfolding of insulin by IDE, we determined a 2.6-A resolution insulin-bound IDE structure. Our structure reveals that IDE forms an enclosed catalytic chamber that completely engulfs and intimately interacts with a partially unfolded insulin molecule. This structure also highlights how the unique size, shape, charge distribution, and exosite of the IDE catalytic chamber contribute to its high affinity ( approximately 100 nm) for insulin. In addition, this structure shows how IDE utilizes the interaction of its exosite with the N terminus of the insulin A chain as well as other properties of the catalytic chamber to guide the unfolding of insulin and allowing for the processive cleavages.

  17. Molecular Basis of Catalytic Chamber-assisted Unfolding and Cleavage of Human Insulin by Human Insulin-degrading Enzyme

    SciTech Connect

    Manolopoulou, Marika; Guo, Qing; Malito, Enrico; Schilling, Alexander B.; Tang, Wei-Jen

    2009-06-02

    Insulin is a hormone vital for glucose homeostasis, and insulin-degrading enzyme (IDE) plays a key role in its clearance. IDE exhibits a remarkable specificity to degrade insulin without breaking the disulfide bonds that hold the insulin A and B chains together. Using Fourier transform ion cyclotron resonance (FTICR) mass spectrometry to obtain high mass accuracy, and electron capture dissociation (ECD) to selectively break the disulfide bonds in gas phase fragmentation, we determined the cleavage sites and composition of human insulin fragments generated by human IDE. Our time-dependent analysis of IDE-digested insulin fragments reveals that IDE is highly processive in its initial cleavage at the middle of both the insulin A and B chains. This ensures that IDE effectively splits insulin into inactive N- and C-terminal halves without breaking the disulfide bonds. To understand the molecular basis of the recognition and unfolding of insulin by IDE, we determined a 2.6-A resolution insulin-bound IDE structure. Our structure reveals that IDE forms an enclosed catalytic chamber that completely engulfs and intimately interacts with a partially unfolded insulin molecule. This structure also highlights how the unique size, shape, charge distribution, and exosite of the IDE catalytic chamber contribute to its high affinity ( approximately 100 nm) for insulin. In addition, this structure shows how IDE utilizes the interaction of its exosite with the N terminus of the insulin A chain as well as other properties of the catalytic chamber to guide the unfolding of insulin and allowing for the processive cleavages.

  18. Human gut microbes impact host serum metabolome and insulin sensitivity.

    PubMed

    Pedersen, Helle Krogh; Gudmundsdottir, Valborg; Nielsen, Henrik Bjørn; Hyotylainen, Tuulia; Nielsen, Trine; Jensen, Benjamin A H; Forslund, Kristoffer; Hildebrand, Falk; Prifti, Edi; Falony, Gwen; Le Chatelier, Emmanuelle; Levenez, Florence; Doré, Joel; Mattila, Ismo; Plichta, Damian R; Pöhö, Päivi; Hellgren, Lars I; Arumugam, Manimozhiyan; Sunagawa, Shinichi; Vieira-Silva, Sara; Jørgensen, Torben; Holm, Jacob Bak; Trošt, Kajetan; Kristiansen, Karsten; Brix, Susanne; Raes, Jeroen; Wang, Jun; Hansen, Torben; Bork, Peer; Brunak, Søren; Oresic, Matej; Ehrlich, S Dusko; Pedersen, Oluf

    2016-07-21

    Insulin resistance is a forerunner state of ischaemic cardiovascular disease and type 2 diabetes. Here we show how the human gut microbiome impacts the serum metabolome and associates with insulin resistance in 277 non-diabetic Danish individuals. The serum metabolome of insulin-resistant individuals is characterized by increased levels of branched-chain amino acids (BCAAs), which correlate with a gut microbiome that has an enriched biosynthetic potential for BCAAs and is deprived of genes encoding bacterial inward transporters for these amino acids. Prevotella copri and Bacteroides vulgatus are identified as the main species driving the association between biosynthesis of BCAAs and insulin resistance, and in mice we demonstrate that P. copri can induce insulin resistance, aggravate glucose intolerance and augment circulating levels of BCAAs. Our findings suggest that microbial targets may have the potential to diminish insulin resistance and reduce the incidence of common metabolic and cardiovascular disorders. PMID:27409811

  19. Human gut microbes impact host serum metabolome and insulin sensitivity.

    PubMed

    Pedersen, Helle Krogh; Gudmundsdottir, Valborg; Nielsen, Henrik Bjørn; Hyotylainen, Tuulia; Nielsen, Trine; Jensen, Benjamin A H; Forslund, Kristoffer; Hildebrand, Falk; Prifti, Edi; Falony, Gwen; Le Chatelier, Emmanuelle; Levenez, Florence; Doré, Joel; Mattila, Ismo; Plichta, Damian R; Pöhö, Päivi; Hellgren, Lars I; Arumugam, Manimozhiyan; Sunagawa, Shinichi; Vieira-Silva, Sara; Jørgensen, Torben; Holm, Jacob Bak; Trošt, Kajetan; Kristiansen, Karsten; Brix, Susanne; Raes, Jeroen; Wang, Jun; Hansen, Torben; Bork, Peer; Brunak, Søren; Oresic, Matej; Ehrlich, S Dusko; Pedersen, Oluf

    2016-07-21

    Insulin resistance is a forerunner state of ischaemic cardiovascular disease and type 2 diabetes. Here we show how the human gut microbiome impacts the serum metabolome and associates with insulin resistance in 277 non-diabetic Danish individuals. The serum metabolome of insulin-resistant individuals is characterized by increased levels of branched-chain amino acids (BCAAs), which correlate with a gut microbiome that has an enriched biosynthetic potential for BCAAs and is deprived of genes encoding bacterial inward transporters for these amino acids. Prevotella copri and Bacteroides vulgatus are identified as the main species driving the association between biosynthesis of BCAAs and insulin resistance, and in mice we demonstrate that P. copri can induce insulin resistance, aggravate glucose intolerance and augment circulating levels of BCAAs. Our findings suggest that microbial targets may have the potential to diminish insulin resistance and reduce the incidence of common metabolic and cardiovascular disorders.

  20. Human adipose tissue expresses intrinsic circadian rhythm in insulin sensitivity.

    PubMed

    Carrasco-Benso, Maria P; Rivero-Gutierrez, Belen; Lopez-Minguez, Jesus; Anzola, Andrea; Diez-Noguera, Antoni; Madrid, Juan A; Lujan, Juan A; Martínez-Augustin, Olga; Scheer, Frank A J L; Garaulet, Marta

    2016-09-01

    In humans, insulin sensitivity varies according to time of day, with decreased values in the evening and at night. Mechanisms responsible for the diurnal variation in insulin sensitivity are unclear. We investigated whether human adipose tissue (AT) expresses intrinsic circadian rhythms in insulin sensitivity that could contribute to this phenomenon. Subcutaneous and visceral AT biopsies were obtained from extremely obese participants (body mass index, 41.8 ± 6.3 kg/m(2); 46 ± 11 y) during gastric-bypass surgery. To assess the rhythm in insulin signaling, AKT phosphorylation was determined every 4 h over 24 h in vitro in response to different insulin concentrations (0, 1, 10, and 100 nM). Data revealed that subcutaneous AT exhibited robust circadian rhythms in insulin signaling (P < 0.00001). Insulin sensitivity reached its maximum (acrophase) around noon, being 54% higher than during midnight (P = 0.009). The amplitude of the rhythm was positively correlated with in vivo sleep duration (r = 0.53; P = 0.023) and negatively correlated with in vivo bedtime (r = -0.54; P = 0.020). No circadian rhythms were detected in visceral AT (P = 0.643). Here, we demonstrate the relevance of the time of the day for how sensitive AT is to the effects of insulin. Subcutaneous AT shows an endogenous circadian rhythm in insulin sensitivity that could provide an underlying mechanism for the daily rhythm in systemic insulin sensitivity.-Carrasco-Benso, M. P., Rivero-Gutierrez, B., Lopez-Minguez, J., Anzola, A., Diez-Noguera, A., Madrid, J. A., Lujan, J. A., Martínez-Augustin, O., Scheer, F. A. J. L., Garaulet, M. Human adipose tissue expresses intrinsic circadian rhythm in insulin sensitivity.

  1. Human Recombinant Insulin 1g - ug

    NASA Technical Reports Server (NTRS)

    2004-01-01

    Proteins are the building blocks of our bodies and the living world around us. Within our bodies proteins make it possible for red blood cells to carry oxygen throughout the body. Others help transmit nerve impulses so we can hear, smell and feel the world around us. While others play a crucial role in preventing or causing disease. If the structure of a protein is known, then companies can develop new or improved drugs to fight the disease of which the protein is a part. To determine protein structure, researchers must grow near-perfect crystals of the protein. On Earth convection currents, sedimentation and other gravity-induced phenomena hamper crystal growth efforts. In microgravity researchers can grow near-perfect crystals in an environment free of these effects. Because of the enormous potential for new pharmaceutical products the Center for Macromolecular Crystallography--the NASA Commercial Space Center responsible for commercial protein crystal growth efforts has more than fifty major industry and academic partners. Research on crystals of human insulin could lead to improved treatments for diabetes.

  2. Multiple tube premixing device

    DOEpatents

    Uhm, Jong Ho; Naidu, Balachandar; Ziminksy, Willy Steve; Kraemer, Gilbert Otto; Yilmaz, Ertan; Lacy, Benjamin; Stevenson, Christian; Felling, David

    2013-08-13

    The present application provides a premixer for a combustor. The premixer may include a fuel plenum with a number of fuel tubes and a burner tube with a number of air tubes. The fuel tubes extend about the air tubes.

  3. Multiple tube premixing device

    DOEpatents

    Uhm, Jong Ho; Varatharajan, Balachandar; Ziminsky, Willy Steve; Kraemer, Gilbert Otto; Yilmaz, Ertan; Lacy, Benjamin; Stevenson, Christian; Felling, David

    2012-12-11

    The present application provides a premixer for a combustor. The premixer may include a fuel plenum with a number of fuel tubes and a burner tube with a number of air tubes. The fuel tubes extend about the air tubes.

  4. Studies in premixed combustion

    SciTech Connect

    Sivashinsky, G.I.

    1992-01-01

    This report discusses the following topics on premixed combustion: theory of turbulent flame propagation; pattern formation in premixed flames and related problems; and pattern formation in extended systems. (LSP)

  5. Human Insulinomas Show Distinct Patterns of Insulin Secretion In Vitro.

    PubMed

    Henquin, Jean-Claude; Nenquin, Myriam; Guiot, Yves; Rahier, Jacques; Sempoux, Christine

    2015-10-01

    Insulinomas are β-cell tumors that cause hypoglycemia through inappropriate secretion of insulin. Characterization of the in vitro dynamics of insulin secretion by perifused fragments of 10 human insulinomas permitted their subdivision into three functional groups with similar insulin content. Group A (four patients with fasting and/or postprandial hypoglycemic episodes) showed qualitatively normal responses to glucose, leucine, diazoxide, tolbutamide, and extracellular CaCl2 omission or excess. The effect of glucose was concentration dependent, but, compared with normal islets, insulin secretion was excessive in both low- and high-glucose conditions. Group B (three patients with fasting hypoglycemic episodes) was mainly characterized by large insulin responses to 1 mmol/L glucose, resulting in very high basal secretion rates that were inhibited by diazoxide and restored by tolbutamide but were not further augmented by other agents except for high levels of CaCl2. Group C (three patients with fasting hypoglycemic episodes) displayed very low rates of insulin secretion and virtually no response to stimuli (including high CaCl2 concentration) and inhibitors (CaCl2 omission being paradoxically stimulatory). In group B, the presence of low-Km hexokinase-I in insulinoma β-cells (not in adjacent islets) was revealed by immunohistochemistry. Human insulinomas thus show distinct, though not completely heterogeneous, defects in insulin secretion that are attributed to the undue expression of hexokinase-I in 3 of 10 patients. PMID:26116696

  6. The Renin Angiotensin Aldosterone System and Insulin Resistance in Humans

    PubMed Central

    Underwood, Patricia C

    2012-01-01

    Alterations in the renin angiotensin aldosterone system (RAAS) contribute to the underlying pathophysiology of insulin resistance in humans; however, individual differences in the treatment response of insulin resistance to RAAS blockade persist. Thus, understanding inter-individual differences in the relationship between the RAAS and insulin resistance may provide insights into improved personalized treatments and improved outcomes. The effects of the systemic RAAS on blood pressure regulation and glucose metabolism have been studied extensively; however, recent discoveries on the influence of local tissue RAAS in the skeletal muscle, heart, vasculature, adipocytes, and pancreas have led to an improved understanding of how activated tissue RAAS influences the development of insulin resistance and diabetes in humans. Angiotensin II (ANGII) is the predominant RAAS component contributing to insulin resistance; however, other players such as aldosterone, renin, and ACE2 are also involved. This review examines the role of local ANGII activity on insulin resistance development in skeletal muscle, adipocytes, and pancreas, followed by a discussion of the other RAAS components implicated in insulin resistance, including ACE2, Ang1-7, renin, and aldosterone. PMID:23242734

  7. Bovine and human insulin adsorption at lipid monolayers: a comparison

    NASA Astrophysics Data System (ADS)

    Mauri, Sergio; Pandey, Ravindra; Rzeznicka, Izabela; Lu, Hao; Bonn, Mischa; Weidner, Tobias

    2015-07-01

    Insulin is a widely used peptide in protein research and it is utilised as a model peptide to understand the mechanics of fibril formation, which is believed to be the cause of diseases such as Alzheimer and Creutzfeld-Jakob syndrome. Insulin has been used as a model system due to its biomedical relevance, small size and relatively simple tertiary structure. The adsorption of insu lin on a variety of surfaces has become the focus of numerous studies lately. These works have helped in elucidating the consequence of surface/protein hydrophilic/hydrophobic interaction in terms of protein refolding and aggregation. Unfortunately, such model surfaces differ significantly from physiological surfaces. Here we spectroscopically investigate the adsorption of insulin at lipid monolayers, to further our understanding of the interaction of insulin with biological surfaces. In particular we study the effect of minor mutations of insulin’s primary amino acid sequence on its interaction with 1,2-Dipalmitoyl-sn-glycero-3-phosphoglycerol (DPPG) model lipid layers. We probe the structure of bovine and human insulin at the lipid/water interface using sum frequency generation spectroscopy (SFG). The SFG experiments are complemented with XPS analysis of Langmuir-Schaefer deposited lipid/insulin films. We find that bovine and human insulin, even though very similar in sequence, show a substantially different behavior when interacting with lipid films.

  8. Influence of PAMAM dendrimers on the human insulin

    NASA Astrophysics Data System (ADS)

    Nowacka, Olga; Miłowska, Katarzyna; Ionov, Maksim; Bryszewska, Maria

    2015-12-01

    Dendrimers are specific class of polymeric macromolecules with wide spectrum of properties. One of the promising activities of dendrimers involves inhibition of protein fibril formation. Aggregation and fibrillation of insulin occurs in insulin-dependent diabetic patients after repeated administration, due to these processes being very easily triggered by the conditions of drug administration. The aim of this work was to study the influence of various generations PAMAM dendrimers on human insulin zeta potential, secondary structure and dithiotreitol (DTT)-induced aggregation. We observed the dependence between the number of positive charges on the surface of the PAMAM dendrimer and the values of zeta potential. Addition of dendrimers to insulin caused insignificant changes in the secondary structure. There was a small decrease in ellipticity, but it did not result in alterations in the circular dichroism (CD) spectrum shape. Dendrimers neither induced protein aggregation nor inhibited the aggregation process induced by DTT, except for 0.01 µmol/l concentration.

  9. Interleukin-1β inhibits insulin signaling and prevents insulin-stimulated system A amino acid transport in primary human trophoblasts.

    PubMed

    Aye, Irving L M H; Jansson, Thomas; Powell, Theresa L

    2013-12-01

    Interleukin-1β (IL-1β) promotes insulin resistance in tissues such as liver and skeletal muscle; however the influence of IL-1β on placental insulin signaling is unknown. We recently reported increased IL-1β protein expression in placentas of obese mothers, which could contribute to insulin resistance. In this study, we tested the hypothesis that IL-1β inhibits insulin signaling and prevents insulin-stimulated amino acid transport in cultured primary human trophoblast (PHT) cells. Cultured trophoblasts isolated from term placentas were treated with physiological concentrations of IL-1β (10pg/ml) for 24h. IL-1β increased the phosphorylation of insulin receptor substrate-1 (IRS-1) at Ser307 (inhibitory) and decreased total IRS-1 protein abundance but did not affect insulin receptor β expression. Furthermore, IL-1β inhibited insulin-stimulated phosphorylation of IRS-1 (Tyr612, activation site) and Akt (Thr308) and prevented insulin-stimulated increase in PI3K/p85 and Grb2 protein expression. IL-1β alone stimulated cRaf (Ser338), MEK (Ser221) and Erk1/2 (Thr202/Tyr204) phosphorylation. The inflammatory pathways nuclear factor kappa B and c-Jun N-terminal kinase, which are involved in insulin resistance, were also activated by IL-1β treatment. Moreover, IL-1β inhibited insulin-stimulated System A, but not System L amino acid uptake, indicating functional impairment of insulin signaling. In conclusion, IL-1β inhibited the insulin signaling pathway by inhibiting IRS-1 signaling and prevented insulin-stimulated System A transport, thereby promoting insulin resistance in cultured PHT cells. These findings indicate that conditions which lead to increased systemic maternal or placental IL-1β levels may attenuate the effects of maternal insulin on placental function and consequently fetal growth.

  10. Effects of Dietary n-3 Fatty Acids on Hepatic and Peripheral Insulin Sensitivity in Insulin-Resistant Humans

    PubMed Central

    Lalia, Antigoni Z.; Johnson, Matthew L.; Jensen, Michael D.; Hames, Kazanna C.; Port, John D.

    2015-01-01

    OBJECTIVE Dietary n-3 polyunsaturated fatty acids, including eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), prevent insulin resistance and stimulate mitochondrial biogenesis in rodents, but the findings of translational studies in humans are thus far ambiguous. The aim of this study was to evaluate the influence of EPA and DHA on insulin sensitivity, insulin secretion, and muscle mitochondrial function in insulin-resistant, nondiabetic humans using a robust study design and gold-standard measurements. RESEARCH DESIGN AND METHODS Thirty-one insulin-resistant adults received 3.9 g/day EPA+DHA or placebo for 6 months in a randomized double-blind study. Hyperinsulinemic-euglycemic clamp with somatostatin was used to assess hepatic and peripheral insulin sensitivity. Postprandial glucose disposal and insulin secretion were measured after a meal. Measurements were performed at baseline and after 6 months of treatment. Abdominal fat distribution was evaluated by MRI. Muscle oxidative capacity was measured in isolated mitochondria using high-resolution respirometry and noninvasively by magnetic resonance spectroscopy. RESULTS Compared with placebo, EPA+DHA did not alter peripheral insulin sensitivity, postprandial glucose disposal, or insulin secretion. Hepatic insulin sensitivity, determined from the suppression of endogenous glucose production by insulin, exhibited a small but significant improvement with EPA+DHA compared with placebo. Muscle mitochondrial function was unchanged by EPA+DHA or placebo. CONCLUSIONS This study demonstrates that dietary EPA+DHA does not improve peripheral glucose disposal, insulin secretion, or skeletal muscle mitochondrial function in insulin-resistant nondiabetic humans. There was a modest improvement in hepatic insulin sensitivity with EPA+DHA, but this was not associated with any improvements in clinically meaningful outcomes. PMID:25852206

  11. Insulin

    MedlinePlus

    ... pump is connected to your body by a flexible tube that has a tip that sticks under your skin. A cartridge of insulin is put in the pump. The insulin flows through the tube into your body. The pump controls how much insulin goes into your body. The ...

  12. Sildenafil Reduces Insulin-Resistance in Human Endothelial Cells

    PubMed Central

    Mammi, Caterina; Pastore, Donatella; Lombardo, Marco F.; Ferrelli, Francesca; Caprio, Massimiliano; Consoli, Claudia; Tesauro, Manfredi; Gatta, Lucia; Fini, Massimo; Federici, Massimo; Sbraccia, Paolo; Donadel, Giulia; Bellia, Alfonso; Rosano, Giuseppe M.; Fabbri, Andrea; Lauro, Davide

    2011-01-01

    Background The efficacy of Phosphodiesterase 5 (PDE5) inhibitors to re-establish endothelial function is reduced in diabetic patients. Recent evidences suggest that therapy with PDE5 inhibitors, i.e. sildenafil, may increase the expression of nitric oxide synthase (NOS) proteins in the heart and cardiomyocytes. In this study we analyzed the effect of sildenafil on endothelial cells in insulin resistance conditions in vitro. Methodology/Principal Findings Human umbilical vein endothelial cells (HUVECs) were treated with insulin in presence of glucose 30 mM (HG) and glucosamine 10 mM (Gluc-N) with or without sildenafil. Insulin increased the expression of PDE5 and eNOS mRNA assayed by Real time-PCR. Cytofluorimetric analysis showed that sildenafil significantly increased NO production in basal condition. This effect was partially inhibited by the PI3K inhibitor LY 294002 and completely inhibited by the NOS inhibitor L-NAME. Akt-1 and eNOS activation was reduced in conditions mimicking insulin resistance and completely restored by sildenafil treatment. Conversely sildenafil treatment can counteract this noxious effect by increasing NO production through eNOS activation and reducing oxidative stress induced by hyperglycaemia and glucosamine. Conclusions/Significance These data indicate that sildenafil might improve NOS activity of endothelial cells in insulin resistance conditions and suggest the potential therapeutic use of sildenafil for improving vascular function in diabetic patients. PMID:21297971

  13. Comparison of Human and Bovine Insulin Amyloidogenesis under Uniform Shear.

    PubMed

    McBride, Samantha A; Tilger, Christopher F; Sanford, Sean P; Tessier, Peter M; Hirsa, Amir H

    2015-08-20

    A diverse range of proteins can assemble into amyloid fibrils, a process that generally results in a loss of function and an increase in toxicity. The occurrence and rate of conversion is strongly dependent on several factors including molecular structure and exposure to hydrodynamic forces. To investigate the origins of shear-induced enhancement in the rate of fibrillization, a stable rotating Couette flow was used to evaluate the kinetics of amyloid formation under uniform shear for two similar insulin species (human and bovine) that demonstrate unique fibrillization kinetics. The presence of shear-induced nuclei predicted by previous studies is supported by observations of a lag between the consumption of soluble insulin and the precipitation of amyloid aggregates. The apparent fibrillization rate generally increases with shear. However, a two-parameter kinetic model revealed that the nucleation rate has a maximum value at intermediate shear rates. The fibril elongation rate increases monotonically with shear and is similar for both insulin variants, suggesting that increased elongation rates are related to mixing. Differences between human and bovine insulin kinetics under shear are attributable to the nucleation step. PMID:26225416

  14. Isolation of the human insulin-like growth factor genes: insulin-like growth factor II and insulin genes are contiguous.

    PubMed Central

    Bell, G I; Gerhard, D S; Fong, N M; Sanchez-Pescador, R; Rall, L B

    1985-01-01

    Overlapping recombinant clones that encompass the insulin-like growth factor (IGF) I and II genes have been isolated from a human genomic DNA library. Each gene is present once per haploid genome; the IGF-I gene spans greater than 35 kilobase pairs (kbp) and the IGF-II gene is at least 15 kbp. The exon-intron organization of these genes is similar, each having four exons, which is one more than the related insulin gene. Comparison of the restriction endonuclease cleavage maps of the IGF-II and insulin genes, including their flanking regions and hybridization with an IGF-II cDNA probe, revealed that they are adjacent to one another. The IGF-II and insulin genes have the same polarity and are separated by 12.6 kbp of intergenic DNA that includes a dispersed middle repetitive Alu sequence. The order of the genes is 5'-insulin-IGF-II-3'. Images PMID:3901002

  15. Gas turbine premixing systems

    DOEpatents

    Kraemer, Gilbert Otto; Varatharajan, Balachandar; Evulet, Andrei Tristan; Yilmaz, Ertan; Lacy, Benjamin Paul

    2013-12-31

    Methods and systems are provided for premixing combustion fuel and air within gas turbines. In one embodiment, a combustor includes an upstream mixing panel configured to direct compressed air and combustion fuel through premixing zone to form a fuel-air mixture. The combustor includes a downstream mixing panel configured to mix additional combustion fuel with the fule-air mixture to form a combustion mixture.

  16. Molecular Basis of Catalytic Chamber-assisted Unfolding and Cleavage of Human Insulin by Human Insulin-degrading Enzyme*S⃞

    PubMed Central

    Manolopoulou, Marika; Guo, Qing; Malito, Enrico; Schilling, Alexander B.; Tang, Wei-Jen

    2009-01-01

    Insulin is a hormone vital for glucose homeostasis, and insulin-degrading enzyme (IDE) plays a key role in its clearance. IDE exhibits a remarkable specificity to degrade insulin without breaking the disulfide bonds that hold the insulin A and B chains together. Using Fourier transform ion cyclotron resonance (FTICR) mass spectrometry to obtain high mass accuracy, and electron capture dissociation (ECD) to selectively break the disulfide bonds in gas phase fragmentation, we determined the cleavage sites and composition of human insulin fragments generated by human IDE. Our time-dependent analysis of IDE-digested insulin fragments reveals that IDE is highly processive in its initial cleavage at the middle of both the insulin A and B chains. This ensures that IDE effectively splits insulin into inactive N- and C-terminal halves without breaking the disulfide bonds. To understand the molecular basis of the recognition and unfolding of insulin by IDE, we determined a 2.6-Å resolution insulin-bound IDE structure. Our structure reveals that IDE forms an enclosed catalytic chamber that completely engulfs and intimately interacts with a partially unfolded insulin molecule. This structure also highlights how the unique size, shape, charge distribution, and exosite of the IDE catalytic chamber contribute to its high affinity (∼100 nm) for insulin. In addition, this structure shows how IDE utilizes the interaction of its exosite with the N terminus of the insulin A chain as well as other properties of the catalytic chamber to guide the unfolding of insulin and allowing for the processive cleavages. PMID:19321446

  17. Differences in bioactivity between human insulin and insulin analogues approved for therapeutic use- compilation of reports from the past 20 years.

    PubMed

    Werner, Haim; Chantelau, Ernst A

    2011-01-01

    In order to provide comprehensive information on the differences in bioactivity between human insulin and insulin analogues, published in vitro comparisons of human insulin and the rapid acting analogues insulin lispro (Humalog®), insulin aspart ( NovoRapid®), insulin glulisine (Apidra®), and the slow acting analogues insulin glargine (Lantus®), and insulin detemir (Levemir®) were gathered from the past 20 years (except for receptor binding studies). A total of 50 reports were retrieved, with great heterogeneity among study methodology. However, various differences in bioactivity compared to human insulin were obvious (e.g. differences in effects on metabolism, mitogenesis, apoptosis, intracellular signalling, thrombocyte function, protein degradation). Whether or not these differences have clinical bearings (and among which patient populations) remains to be determined.

  18. Differences in bioactivity between human insulin and insulin analogues approved for therapeutic use- compilation of reports from the past 20 years

    PubMed Central

    2011-01-01

    In order to provide comprehensive information on the differences in bioactivity between human insulin and insulin analogues, published in vitro comparisons of human insulin and the rapid acting analogues insulin lispro (Humalog®), insulin aspart ( NovoRapid®), insulin glulisine (Apidra®), and the slow acting analogues insulin glargine (Lantus®), and insulin detemir (Levemir®) were gathered from the past 20 years (except for receptor binding studies). A total of 50 reports were retrieved, with great heterogeneity among study methodology. However, various differences in bioactivity compared to human insulin were obvious (e.g. differences in effects on metabolism, mitogenesis, apoptosis, intracellular signalling, thrombocyte function, protein degradation). Whether or not these differences have clinical bearings (and among which patient populations) remains to be determined. PMID:21714872

  19. Insulin-like growth factor I stimulates lipid oxidation, reduces protein oxidation, and enhances insulin sensitivity in humans.

    PubMed Central

    Hussain, M A; Schmitz, O; Mengel, A; Keller, A; Christiansen, J S; Zapf, J; Froesch, E R

    1993-01-01

    To elucidate the effects of insulin-like growth factor I (IGF-I) on fuel oxidation and insulin sensitivity, eight healthy subjects were treated with saline and recombinant human (IGF-I (10 micrograms/kg.h) during 5 d in a crossover, randomized fashion, while receiving an isocaloric diet (30 kcal/kg.d) throughout the study period. On the third and fourth treatment days, respectively, an L-arginine stimulation test and an intravenous glucose tolerance test were performed. A euglycemic, hyperinsulinemic clamp combined with indirect calorimetry and a glucose tracer infusion were performed on the fifth treatment day. IGF-I treatment led to reduced fasting and stimulated (glucose and/or L-arginine) insulin and growth hormone secretion. Basal and stimulated glucagon secretion remained unchanged. Intravenous glucose tolerance was unaltered despite reduced insulin secretion. Resting energy expenditure and lipid oxidation were both elevated, while protein oxidation was reduced, and glucose turnover rates were unaltered on the fifth treatment day with IGF-I as compared to the control period. Enhanced lipolysis was reflected by elevated circulating free fatty acids. Moreover, insulin-stimulated oxidative and nonoxidative glucose disposal (i.e., insulin sensitivity) were enhanced during IGF-I treatment. Thus, IGF-I treatment leads to marked changes in lipid and protein oxidation, whereas, at the dose used, carbohydrate metabolism remains unaltered in the face of reduced insulin levels and enhanced insulin sensitivity. Images PMID:8227340

  20. Insulin

    NASA Technical Reports Server (NTRS)

    2004-01-01

    The manipulation of organic materials--cells, tissues, and even living organisms--offers many exciting possibilities for the future from organic computers to improved aquaculture. Commercial researchers are using the microgravity environment to produce large near perfect protein crystals Research on insulin has yielded crystals that far surpass the quality of insulin crystals grown on the ground. Using these crystals industry partners are working to develop new and improved treatments for diabetes. Other researchers are exploring the possibility of producing antibiotics using plant cell cultures which could lead to both orbital production and the improvement of ground-based antibiotic production.

  1. Mechanisms of insulin resistance in human obesity: evidence for receptor and postreceptor defects.

    PubMed Central

    Kolterman, O G; Insel, J; Saekow, M; Olefsky, J M

    1980-01-01

    To assess the mechanisms of the insulin resistance in human obesity, we have determined, using a modification of the euglycemic glucose clamp technique, the shape of the in vivo insulin-glucose disposal dose-response curves in 7 control and 13 obese human subjects. Each subject had at least three euglycemic studies performed at insulin infusion rates of 15, 40, 120, 240, or 1,200 mU/M2/min. The glucose disposal rate was decreased in all obese subjects compared with controls (101 +/- 16 vs. 186 +/- 16 mg/M2/min) during the 40 mU/M2/min insulin infusion. The mean dose-response curve for the obese subjects was displaced to the right, i.e., the half-maximally effective insulin concentration was 270 +/- 27 microU/ml for the obese compared with 130 +/- 10 microU/ml for controls. In nine of the obese subjects, the dose-response curves were shifted to the right, and maximal glucose disposal rates (at a maximally effective insulin concentration) were markedly decreased, indicating both a receptor and a postreceptor defect. On the other hand, four obese patients had right-shifted dose-response curves but reached normal maximal glucose disposal rates, consistent with decreased insulin receptors as the only abnormality. When the individual data were analyzed, it was found that the lease hyperinsulinemic, least insulin-resistant patients displayed only the receptor defect, whereas those with the greatest hyperinsulinemia exhibited the largest post-receptor defect, suggesting a continuous spectrum of defects as one advances from mild to severe insulin resistance. When insulin's ability to suppress hepatic glucose output was assessed, hyperinsulinemia produced total suppresssion in all subjects. The dose-response curve for the obese subjects was shifted to the right, indicating a defect in insulin receptors. Insulin binding to isolated adipocytes obtained from the obese subjects was decreased, and a highly significant inverse linear relationship was demonstrated between insulin

  2. Effects of Insulin Detemir and NPH Insulin on Body Weight and Appetite-Regulating Brain Regions in Human Type 1 Diabetes: A Randomized Controlled Trial

    PubMed Central

    van Golen, Larissa W.; Veltman, Dick J.; IJzerman, Richard G.; Deijen, Jan Berend; Heijboer, Annemieke C.; Barkhof, Frederik; Drent, Madeleine L.; Diamant, Michaela

    2014-01-01

    Studies in rodents have demonstrated that insulin in the central nervous system induces satiety. In humans, these effects are less well established. Insulin detemir is a basal insulin analog that causes less weight gain than other basal insulin formulations, including the current standard intermediate-long acting Neutral Protamine Hagedorn (NPH) insulin. Due to its structural modifications, which render the molecule more lipophilic, it was proposed that insulin detemir enters the brain more readily than other insulins. The aim of this study was to investigate whether insulin detemir treatment differentially modifies brain activation in response to food stimuli as compared to NPH insulin. In addition, cerebral spinal fluid (CSF) insulin levels were measured after both treatments. Brain responses to viewing food and non-food pictures were measured using functional Magnetic Resonance Imaging in 32 type 1 diabetic patients, after each of two 12-week treatment periods with insulin detemir and NPH insulin, respectively, both combined with prandial insulin aspart. CSF insulin levels were determined in a subgroup. Insulin detemir decreased body weight by 0.8 kg and NPH insulin increased weight by 0.5 kg (p = 0.02 for difference), while both treatments resulted in similar glycemic control. After treatment with insulin detemir, as compared to NPH insulin, brain activation was significantly lower in bilateral insula in response to visual food stimuli, compared to NPH (p = 0.02 for right and p = 0.05 for left insula). Also, CSF insulin levels were higher compared to those with NPH insulin treatment (p = 0.003). Our findings support the hypothesis that in type 1 diabetic patients, the weight sparing effect of insulin detemir may be mediated by its enhanced action on the central nervous system, resulting in blunted activation in bilateral insula, an appetite-regulating brain region, in response to food stimuli. Trial Registration ClinicalTrials.gov NCT00626080

  3. Stress-impaired transcription factor expression and insulin secretion in transplanted human islets

    PubMed Central

    Dai, Chunhua; Kayton, Nora S.; Shostak, Alena; Poffenberger, Greg; Cyphert, Holly A.; Aramandla, Radhika; Thompson, Courtney; Papagiannis, Ioannis G.; Shiota, Masakazu; Stafford, John M.; Greiner, Dale L.; Herrera, Pedro L.; Shultz, Leonard D.; Stein, Roland; Powers, Alvin C.

    2016-01-01

    Type 2 diabetes is characterized by insulin resistance, hyperglycemia, and progressive β cell dysfunction. Excess glucose and lipid impair β cell function in islet cell lines, cultured rodent and human islets, and in vivo rodent models. Here, we examined the mechanistic consequences of glucotoxic and lipotoxic conditions on human islets in vivo and developed and/or used 3 complementary models that allowed comparison of the effects of hyperglycemic and/or insulin-resistant metabolic stress conditions on human and mouse islets, which responded quite differently to these challenges. Hyperglycemia and/or insulin resistance impaired insulin secretion only from human islets in vivo. In human grafts, chronic insulin resistance decreased antioxidant enzyme expression and increased superoxide and amyloid formation. In human islet grafts, expression of transcription factors NKX6.1 and MAFB was decreased by chronic insulin resistance, but only MAFB decreased under chronic hyperglycemia. Knockdown of NKX6.1 or MAFB expression in a human β cell line recapitulated the insulin secretion defect seen in vivo. Contrary to rodent islet studies, neither insulin resistance nor hyperglycemia led to human β cell proliferation or apoptosis. These results demonstrate profound differences in how excess glucose or lipid influence mouse and human insulin secretion and β cell activity and show that reduced expression of key islet-enriched transcription factors is an important mediator of glucotoxicity and lipotoxicity. PMID:27064285

  4. Lack of the architectural factor HMGA1 causes insulin resistance and diabetes in humans and mice.

    PubMed

    Foti, Daniela; Chiefari, Eusebio; Fedele, Monica; Iuliano, Rodolfo; Brunetti, Leonardo; Paonessa, Francesco; Manfioletti, Guidalberto; Barbetti, Fabrizio; Brunetti, Arturo; Croce, Carlo M; Fusco, Alfredo; Brunetti, Antonio

    2005-07-01

    Type 2 diabetes mellitus is a widespread disease, affecting millions of people globally. Although genetics and environmental factors seem to have a role, the cause of this metabolic disorder is largely unknown. Here we report a genetic flaw that markedly reduced the intracellular expression of the high mobility group A1 (HMGA1) protein, and adversely affected insulin receptor expression in cells and tissues from four subjects with insulin resistance and type 2 diabetes. Restoration of HMGA1 protein expression in subjects' cells enhanced INSR gene transcription, and restored cell-surface insulin receptor protein expression and insulin-binding capacity. Loss of Hmga1 expression, induced in mice by disrupting the Hmga1 gene, considerably decreased insulin receptor expression in the major targets of insulin action, largely impaired insulin signaling and severely reduced insulin secretion, causing a phenotype characteristic of human type 2 diabetes. PMID:15924147

  5. [Insulin resistance as a mechanism of adaptation during human evolution].

    PubMed

    Ricart, W; Fernández-Real, J M

    2010-10-01

    The recent application of concepts of evolution to human disease is proving useful to understand certain pathophysiological mechanisms of different entities that span genomic alterations of immunity, respiratory and hormone function, and the circulatory and neural systems. However, effort has concentrated on explaining the keys to adaptation that define human metabolism and, since the early 1960s, several theories have been developed. This article reviews some of the hypotheses postulated in recent years on the potential benefit of insulin resistance and discusses the most recent knowledge. The concept of the thrifty gene seems to have been definitively refuted by current knowledge. The current paradigm describes an interaction between the metabolic and the immune systems resulting from their coevolution, promoted by evolutionary pressures triggered by fasting, infection and intake of different foods. The activation and regulation of these ancient mechanisms in integrated and interdependent areas defines insulin resistance as a survival strategy that is critical during fasting and in the fight against infection. The relationship with some components of the diet and, particularly, with the symbiotic intestinal microflora points to new paradigms in understanding the pathophysiology of obesity, metabolic syndrome and type 2 diabetes mellitus.

  6. [Insulin resistance as a mechanism of adaptation during human evolution].

    PubMed

    Ricart, W; Fernández-Real, J M

    2010-10-01

    The recent application of concepts of evolution to human disease is proving useful to understand certain pathophysiological mechanisms of different entities that span genomic alterations of immunity, respiratory and hormone function, and the circulatory and neural systems. However, effort has concentrated on explaining the keys to adaptation that define human metabolism and, since the early 1960s, several theories have been developed. This article reviews some of the hypotheses postulated in recent years on the potential benefit of insulin resistance and discusses the most recent knowledge. The concept of the thrifty gene seems to have been definitively refuted by current knowledge. The current paradigm describes an interaction between the metabolic and the immune systems resulting from their coevolution, promoted by evolutionary pressures triggered by fasting, infection and intake of different foods. The activation and regulation of these ancient mechanisms in integrated and interdependent areas defines insulin resistance as a survival strategy that is critical during fasting and in the fight against infection. The relationship with some components of the diet and, particularly, with the symbiotic intestinal microflora points to new paradigms in understanding the pathophysiology of obesity, metabolic syndrome and type 2 diabetes mellitus. PMID:20675202

  7. Metal induced conformational changes in human insulin: crystal structures of Sr2+, Ni2+ and Cu2+ complexes of human insulin.

    PubMed

    Krishna, Nagampalli Raghavendra Sashi; Pattabhi, Vasantha; Rajan, S S

    2011-05-01

    Crystal structures of Sr(2+), Ni(2+) and Cu(2+) of human insulin complexes have been determined. The structures of Sr(2+) and Ni(2+) complexes are similar to Zn(2+) insulin and are in T6 conformation. (All the six monomers in the insulin hexamer are in Tensed conformation (T), which means the first eight residues of B-chain are in an extended conformation). Cu(2+) complex, though it assumes T6 conformation, has more structural differences due to lowering of crystal symmetry and space group shift from H3 (Hexagonal crystal system) to P3 (Trigonal crystal system) and a doubling of the c axis. 2Ni(2+) human insulin when compared to 4Ni(2+) Arg insulin suggests that terminal modifications may be responsible for additional metal binding. All the three metals have been shown to have a role in diabetes and hence may be therapeutically useful. PMID:21171943

  8. Analysis of alternatives for insulinizing patients to achieve glycemic control and avoid accompanying risks of hypoglycemia

    PubMed Central

    GAO, JIALIN; XIONG, QIANYIN; MIAO, JUN; ZHANG, YAO; XIA, LIBING; LU, MEIQIN; ZHANG, BINHUA; CHEN, YUEPING; ZHANG, ANSU; YU, CUI; WANG, LI-ZHUO

    2015-01-01

    The aims of the present study were to explore the efficacy of glycemic control and the risks of hypoglycemia with different methods of insulin therapy, and to provide reference data for the clinical treatment of diabetes. In this retrospective study, hospitalized patients diagnosed with type 2 diabetes between March and December 2014, in the Department of Endocrinology in the First Affiliated Hospital of Wannan Medical College, were divided into three groups, including an intensive insulin analogue therapy group, a premixed insulin analogue treatment group and a premixed human insulin therapy group. The efficacy of glycemic control and the incidence of hypoglycemia were determined in each of the insulin treatment groups. Compared with the other treatment groups, the intensive insulin analogue therapy group was associated with superior blood glucose control, shorter time to reach standard insulin regimen, shorter hospitalization time, fewer fluctuations in blood glucose levels and lower insulin dosage on discharge from hospital. However, this treatment was also associated with a high risk of hypoglycemia. In conclusion, when combined with the effective prevention of hypoglycemia and appropriate nursing care (especially in hospital care), intensive insulin analogue therapy may provide the greatest benefit to patients. PMID:26137223

  9. Insulin concentration is critical in culturing human neural stem cells and neurons.

    PubMed

    Rhee, Y-H; Choi, M; Lee, H-S; Park, C-H; Kim, S-M; Yi, S-H; Oh, S-M; Cha, H-J; Chang, M-Y; Lee, S-H

    2013-08-08

    Cell culture of human-derived neural stem cells (NSCs) is a useful tool that contributes to our understanding of human brain development and allows for the development of therapies for intractable human brain disorders. Human NSC (hNSC) cultures, however, are not commonly used, mainly because of difficulty with consistently maintaining the cells in a healthy state. In this study, we show that hNSC cultures, unlike NSCs of rodent origins, are extremely sensitive to insulin, an indispensable culture supplement, and that the previously reported difficulty in culturing hNSCs is likely because of a lack of understanding of this relationship. Like other neural cell cultures, insulin is required for hNSC growth, as withdrawal of insulin supplementation results in massive cell death and delayed cell growth. However, severe apoptotic cell death was also detected in insulin concentrations optimized to rodent NSC cultures. Thus, healthy hNSC cultures were only produced in a narrow range of relatively low insulin concentrations. Insulin-mediated cell death manifested not only in all human NSCs tested, regardless of origin, but also in differentiated human neurons. The underlying cell death mechanism at high insulin concentrations was similar to insulin resistance, where cells became less responsive to insulin, resulting in a reduction in the activation of the PI3K/Akt pathway critical to cell survival signaling.

  10. Brain Insulin Resistance at the Crossroads of Metabolic and Cognitive Disorders in Humans.

    PubMed

    Kullmann, Stephanie; Heni, Martin; Hallschmid, Manfred; Fritsche, Andreas; Preissl, Hubert; Häring, Hans-Ulrich

    2016-10-01

    Ever since the brain was identified as an insulin-sensitive organ, evidence has rapidly accumulated that insulin action in the brain produces multiple behavioral and metabolic effects, influencing eating behavior, peripheral metabolism, and cognition. Disturbances in brain insulin action can be observed in obesity and type 2 diabetes (T2D), as well as in aging and dementia. Decreases in insulin sensitivity of central nervous pathways, i.e., brain insulin resistance, may therefore constitute a joint pathological feature of metabolic and cognitive dysfunctions. Modern neuroimaging methods have provided new means of probing brain insulin action, revealing the influence of insulin on both global and regional brain function. In this review, we highlight recent findings on brain insulin action in humans and its impact on metabolism and cognition. Furthermore, we elaborate on the most prominent factors associated with brain insulin resistance, i.e., obesity, T2D, genes, maternal metabolism, normal aging, inflammation, and dementia, and on their roles regarding causes and consequences of brain insulin resistance. We also describe the beneficial effects of enhanced brain insulin signaling on human eating behavior and cognition and discuss potential applications in the treatment of metabolic and cognitive disorders.

  11. Transcriptional directionality of the human insulin-degrading enzyme promoter.

    PubMed

    Zhang, Lang; Wang, Pan; Ding, Qingyang; Wang, Zhao

    2013-10-01

    Unidirectional promoters dominate among mammalian genomes. However, the mechanism through which the transcriptional directionality of promoters is accomplished remains to be clarified. Insulin-degrading enzyme (IDE) is a ubiquitously expressed zinc metalloprotease, whose promoter contains a CpG island. We previously showed that the basal promoter region of mouse IDE has bidirectional transcriptional activity, but an upstream promoter element blocks its antisense transcription. Therefore, we wonder whether the human IDE promoter contains an analogous element. Similarly, the basal promoter region of human IDE (-102 ~ +173 and -196 ~ +173 relative to the transcription start site) showed bidirectional transcriptional activity. However, the region from -348 to +173 could only be transcribed from the normal orientation, implying that an upstream promoter element between -348 and -196 blocks the antisense transcription of the human IDE promoter. Through promoter deletion and mutagenesis analysis, we mapped this element precisely and found that the upstream promoter element locates between -318 and -304. Furthermore, the transcription-blocking elements in the mouse and human IDE promoters inhibited the transcription of the SV40 promoter when put downstream of it. In conclusion, we identify an upstream promoter element which blocks the antisense transcription of the human IDE promoter. Our studies are helpful to clarify the transcriptional directionality of promoters.

  12. New method to differentiate human peripheral blood monocytes into insulin producing cells: Human hematosphere culture.

    PubMed

    Hur, Jin; Yang, Ji Min; Choi, Jae-Il; Yun, Ji-Yeon; Jang, Jae Hee; Kim, Joonoh; Kim, Ju-Young; Oh, Il-Young; Yoon, Chang-Hwan; Cho, Hyun-Jai; Park, Young-Bae; Kim, Hyo-Soo

    2012-02-24

    Strategy to differentiate stem cells into insulin producing cells (IPCs) in vitro has been a promising one to get cell source of β-cell replacement therapy for diabetes. It has been suggested that islets and neurons share features and nestin-positive cells could differentiate into IPCs. We have recently developed a three-dimensional culture system using human peripheral blood cells named as blood-born hematosphere (BBHS). Here we showed that most of BBHS were composed of nestin-positive cells. Under the four-stage differentiation protocol for IPCs, we plated nestin-positive BBHS onto fibronectin-coated dish. These cells form islet-like clusters and most of them expressed insulin. Pancreatic specific genes were turned on, such as transcription factors (Pdx-1, Ngn3 and Nkx6.1), genes related to endocrine function (Glut-2 and PC2) or β cell function (Kir6.2, SUR1). Furthermore islet differentiation was confirmed by dithizone (DTZ) staining to detect zinc ion which binds insulin protein within the cells. Finally, IPCs derived from BBHS showed capability to secrete insulin in response to glucose stimulation. Taken together, our novel protocol successfully induced islet-like human insulin producing cells out of BBHS. This strategy of ex vivo expansion of IPCs using BBHS provides an autologous therapeutic cell source for the treatment of diabetes. PMID:22310720

  13. Myotubes derived from human-induced pluripotent stem cells mirror in vivo insulin resistance.

    PubMed

    Iovino, Salvatore; Burkart, Alison M; Warren, Laura; Patti, Mary Elizabeth; Kahn, C Ronald

    2016-02-16

    Induced pluripotent stem cells (iPS cells) represent a unique tool for the study of the pathophysiology of human disease, because these cells can be differentiated into multiple cell types in vitro and used to generate patient- and tissue-specific disease models. Given the critical role for skeletal muscle insulin resistance in whole-body glucose metabolism and type 2 diabetes, we have created a novel cellular model of human muscle insulin resistance by differentiating iPS cells from individuals with mutations in the insulin receptor (IR-Mut) into functional myotubes and characterizing their response to insulin in comparison with controls. Morphologically, IR-Mut cells differentiated normally, but had delayed expression of some muscle differentiation-related genes. Most importantly, whereas control iPS-derived myotubes exhibited in vitro responses similar to primary differentiated human myoblasts, IR-Mut myotubes demonstrated severe impairment in insulin signaling and insulin-stimulated 2-deoxyglucose uptake and glycogen synthesis. Transcriptional regulation was also perturbed in IR-Mut myotubes with reduced insulin-stimulated expression of metabolic and early growth response genes. Thus, iPS-derived myotubes from individuals with genetically determined insulin resistance demonstrate many of the defects observed in vivo in insulin-resistant skeletal muscle and provide a new model to analyze the molecular impact of muscle insulin resistance. PMID:26831110

  14. Myotubes derived from human-induced pluripotent stem cells mirror in vivo insulin resistance.

    PubMed

    Iovino, Salvatore; Burkart, Alison M; Warren, Laura; Patti, Mary Elizabeth; Kahn, C Ronald

    2016-02-16

    Induced pluripotent stem cells (iPS cells) represent a unique tool for the study of the pathophysiology of human disease, because these cells can be differentiated into multiple cell types in vitro and used to generate patient- and tissue-specific disease models. Given the critical role for skeletal muscle insulin resistance in whole-body glucose metabolism and type 2 diabetes, we have created a novel cellular model of human muscle insulin resistance by differentiating iPS cells from individuals with mutations in the insulin receptor (IR-Mut) into functional myotubes and characterizing their response to insulin in comparison with controls. Morphologically, IR-Mut cells differentiated normally, but had delayed expression of some muscle differentiation-related genes. Most importantly, whereas control iPS-derived myotubes exhibited in vitro responses similar to primary differentiated human myoblasts, IR-Mut myotubes demonstrated severe impairment in insulin signaling and insulin-stimulated 2-deoxyglucose uptake and glycogen synthesis. Transcriptional regulation was also perturbed in IR-Mut myotubes with reduced insulin-stimulated expression of metabolic and early growth response genes. Thus, iPS-derived myotubes from individuals with genetically determined insulin resistance demonstrate many of the defects observed in vivo in insulin-resistant skeletal muscle and provide a new model to analyze the molecular impact of muscle insulin resistance.

  15. Determination of insulin in humans with insulin-dependent diabetes mellitus patients by HPLC with diode array detection.

    PubMed

    Yilmaz, Bilal; Kadioglu, Yucel; Capoglu, Ilyas

    2012-08-01

    A simple and reliable high-performance liquid chromatographic method with diode array detection has been developed and validated for the determination of insulin in human plasma. A good chromatographic separation was achieved on a C18 column with a mobile phase consisting of acetonitrile and 0.2M sodium sulfate (pH 2.4), 25:75 (v/v). Its flow rate was 1.2 mL/min. Calibration curve was linear within the concentration range of 0.15-25 µg/mL. Intra-day and inter-day relative standard deviations for insulin in human plasma were less than 6.3 and 8.5%, respectively. The limits of detection and quantification of insulin were 0.10 and 0.15 µg/mL, respectively. Also, this assay was applied to determine the pharmacokinetic parameters of insulin in eight insulin-dependent diabetes mellitus patients after subcutaneous injection of 25 IU of Actrapid HM.

  16. Premixed turbulent flame calculation

    NASA Technical Reports Server (NTRS)

    El-Tahry, S.; Rutland, C. J.; Ferziger, J. H.; Rogers, M. M.

    1987-01-01

    The importance of turbulent premixed flames in a variety of applications has led to a substantial amount of effort towards improving the understanding of these flames. Although these efforts have increased the understanding, many questions still remain. The use of direct numerical simulation (DNS) in solving these questions is examined.

  17. Protein crystal growth in microgravity review of large scale temperature induction method: bovine insulin, human insulin and human alpha interferon

    NASA Astrophysics Data System (ADS)

    Long, Marianna M.; Bishop, John Bradford; Nagabhushan, Tattanahalli L.; Reichert, Paul; Smith, G. David; DeLucas, Lawrence J.

    1996-10-01

    The protein crystal growth facility (PCF) is space-flight hardware that accommodates large scale protein crystal growth experiments using temperature change as the inductive step. Recent modifications include specialized instrumentation for monitoring crystal nucleation with laser light scattering. This paper reviews results from the PCF's first seven flights on the Space Shuttle, the last with laser light scattering instrumentation. The PCF's objective is twofold: (1) production of high quality protein crystals for X-ray analysis and subsequent structure based drug design and (2) preparation of a large quantity of relatively contaminant free crystals for use as time-release protein pharmaceuticals. The first three Shuttle flights with bovine insulin constituted the PCF's proof of concept, demonstrating that the space-grown crystals were larger and diffracted to higher resolution than their earth-grown counterparts. The later four PCF missions were used to grow recombinant human insulin crystals for X-ray analysis and to continue productions trials aimed at the development of a processing facility for crystalline recombinant alpha interferon.

  18. Insulin binding properties of normal and transformed human epidermal cultured keratinocytes

    SciTech Connect

    Verrando, P.; Ortonne, J.P.

    1985-10-01

    Insulin binding to its receptors was studied in cultured normal and transformed (A431 line) human epidermal keratinocytes. The specific binding was a temperature-dependent, saturable process. Normal keratinocytes possess a mean value of about 80,000 receptors per cell. Fifteen hours exposure of the cells to insulin lowered their receptor number (about 65% loss in available sites); these reappeared when the hormone was removed from the culture medium. In the A431 epidermoid carcinoma cell line, there is a net decrease in insulin binding (84% of the initial bound/free hormone ratio in comparison with normal cells) essentially related to a loss in receptor affinity for insulin. Thus, cultured human keratinocytes which express insulin receptors may be a useful tool in understanding skin pathology related to insulin disorders.

  19. Comparison of the effects of recombinant human insulin-like growth factor-I and insulin on glucose and leucine kinetics in humans.

    PubMed Central

    Laager, R; Ninnis, R; Keller, U

    1993-01-01

    To compare the metabolic effects of elevated plasma concentrations of IGF-I and insulin, overnight-fasted normal subjects were studied twice, once receiving IGF-I and once insulin at doses that resulted in identical increases in glucose uptake during 8-h euglycemic clamping. Recombinant human IGF-I or insulin were infused in one group at high doses (30 micrograms/kg per h IGF-I or 0.23 nmol/kg per h insulin) and in another group at low doses (5 micrograms/kg per h IGF-I or 0.04 nmol/kg per h insulin). Glucose rate of disappearance (measured by [6,6-D2]-glucose infusions) increased from baseline by 239 +/- 16% during high dose IGF-I vs 197 +/- 18% during insulin (P = 0.021 vs IGF-I). Hepatic glucose production decreased by 37 +/- 6% during high dose IGF-I vs 89 +/- 13% during insulin (P = 0.0028 vs IGF-I). IGF-I suppressed whole body leucine flux ([1-13C]-leucine infusion technique) more than insulin (42 +/- 4 vs 32 +/- 3% during high doses, P = 0.0082). Leucine oxidation rate decreased during high dose IGF-I more than during insulin (55 +/- 4 vs 32 +/- 6%, P = 0.0001). The decreases of plasma concentrations of free fatty acids, acetoacetate, and beta-hydroxybutyrate after 8 h of IGF-I and insulin administration were similar. Plasma C-peptide levels decreased by 57 +/- 4% during high doses of IGF-I vs 36 +/- 6% during insulin (P = 0.005 vs IGF-I). The present data demonstrate that, compared to insulin, an acute increase in plasma IGF-I levels results in preferential enhancement of peripheral glucose utilization, diminished suppression of hepatic glucose production, augmented decrease of whole body protein breakdown (leucine flux), and of irreversible leucine catabolism but in similar antilipolytic effects. The data suggest that insulin-like effects of IGF-I in humans are mediated in part via IGF-I receptors and in part via insulin receptors. PMID:8408642

  20. Insulin Resistance Alters Islet Morphology in Nondiabetic Humans

    PubMed Central

    Mezza, Teresa; Muscogiuri, Giovanna; Sorice, Gian Pio; Clemente, Gennaro; Hu, Jiang; Pontecorvi, Alfredo; Holst, Jens J.; Giaccari, Andrea; Kulkarni, Rohit N.

    2014-01-01

    Type 2 diabetes is characterized by poor glucose uptake in metabolic tissues and manifests when insulin secretion fails to cope with worsening insulin resistance. In addition to its effects on skeletal muscle, liver, and adipose tissue metabolism, it is evident that insulin resistance also affects pancreatic β-cells. To directly examine the alterations that occur in islet morphology as part of an adaptive mechanism to insulin resistance, we evaluated pancreas samples obtained during pancreatoduodenectomy from nondiabetic subjects who were insulin-resistant or insulin-sensitive. We also compared insulin sensitivity, insulin secretion, and incretin levels between the two groups. We report an increased islet size and an elevated number of β- and α-cells that resulted in an altered β-cell–to–α-cell area in the insulin- resistant group. Our data in this series of studies suggest that neogenesis from duct cells and transdifferentiation of α-cells are potential contributors to the β-cell compensatory response to insulin resistance in the absence of overt diabetes. PMID:24215793

  1. Development of a Quantitative PCR Assay for Detection of Human Insulin-Like Growth Factor Receptor and Insulin Receptor Isoforms.

    PubMed

    Flannery, Clare A; Rowzee, Anne M; Choe, Gina H; Saleh, Farrah L; Radford, Caitlin C; Taylor, Hugh S; Wood, Teresa L

    2016-04-01

    The biological activity of insulin and the insulin-like growth factor (IGF) ligands, IGF-I and IGF-II, is based in part on the relative abundance and distribution of their target receptors: the insulin receptor (IR) splice variants A (IR-A) and B (IR-B) and IGF 1 receptor (IGF-1R). However, the relative quantity of all three receptors in human tissues has never been measured together on the same scale. Due to the high homology between insulin receptor (IR)-A and IR-B proteins and lack of antibodies that discern the two IR splice variants, their mRNA sequence is the most reliable means of distinguishing between the receptors. Hence, highly specific primers for IR-A, IR-B, and IGF-1R mRNA were designed to accurately detect all three receptors by quantitative RT-PCR and enable direct quantification of relative receptor expression levels. A standard concentration curve of cDNA from each receptor was performed. Assay specificity was tested using competition assays and postamplification analysis by gel electrophoresis and cloning. Forward and reverse primer concentrations were optimized to ensure equal efficiencies across primer pairs. This assay enables a specific molecular signature of IGF/insulin signaling receptors to be assayed in different tissues, cell types, or cancers. PMID:26862994

  2. Small human islets comprised of more β-cells with higher insulin content than large islets

    PubMed Central

    Farhat, Bilal; Almelkar, Akshay; Ramachandran, Karthik; Williams, S. Janette; Huang, Han-Hung; Zamierowksi, David; Novikova, Lesya; Stehno-Bittel, Lisa

    2013-01-01

    For the past 30 y, data have suggested that unique islet populations exist, based on morphology and glucose sensitivity. Yet little has been done to determine the mechanism of these functional differences. The purpose of this study was to determine whether human islets were comprised functionally unique populations, and to elucidate a possible mechanism. Islets or pancreatic sections from 29 human donors were analyzed. Islets were isolated and measured for insulin secretion, cell composition and organization, insulin and glucagon granule density and insulin content. Insulin secretion was significantly greater in small compared with large islets. In sectioned human pancreata, β-cells comprised a higher proportion of the total endocrine cells in small islets (63%) than large islets (39%). A higher percentage of β-cells in small islets contacted blood vessels (44%) compared with large islets (31%). Total insulin content of isolated human islets was significantly greater in the small (1323 ± 512 μIU/IE) compared with large islets (126 ± 48 μIU/IE). There was less immunostaining for insulin in the large islets from human pancreatic sections, especially in the core of the islet, compared with small islets. The results suggest that differences in insulin secretion between large and small islets may be due to a higher percentage of β-cells in small islets with more β-cells in contact with blood vessels and a higher concentration of insulin/β-cell in small islets. PMID:23648896

  3. Study of the aggregation of human insulin Langmuir monolayer.

    PubMed

    Liu, Wei; Johnson, Sheba; Micic, Miodrag; Orbulescu, Jhony; Whyte, Jeffrey; Garcia, Andrew R; Leblanc, Roger M

    2012-02-21

    The human insulin (HI) Langmuir monolayer at the air-water interface was systematically investigated in the presence and absence of Zn(II) ions in the subphase. HI samples were dissolved in acidic (pH 2) and basic (pH 9) aqueous solutions and then spread at the air-water interface. Spectroscopic data of aqueous solutions of HI show a difference in HI conformation at different pH values. Moreover, the dynamics of the insulin protein showed a dependence on the concentration of Zn(II) ions. In the absence of Zn(II) ions in the subphase, the acidic and basic solutions showed similar behavior at the air-water interface. In the presence of Zn(II) ions in the subphase, the surface pressure-area and surface potential-area isotherms suggest that HI may aggregate at the air-water interface. It was observed that increasing the concentration of Zn(II) ions in the acidic (pH 2) aqueous solution of HI led to an increase of the area at a specific surface pressure. It was also seen that the conformation of HI in the basic (pH 9) medium had a reverse effect (decrease in the surface area) with the increase of the concentration of Zn(II) ions in solution. From the compression-decompression cycles we can conclude that the aggregated HI film at air-water interface is not stable and tends to restore a monolayer of monomers. These results were confirmed from UV-vis and fluorescence spectroscopy analysis. Infrared reflection-absorption and circular dichroism spectroscopy techniques were used to determine the secondary structure and orientation changes of HI by zinc ions. Generally, the aggregation process leads to a conformation change from α-helix to β-strand and β-turn, and at the air-water interface, the aggregation process was likewise seen to induce specific orientations for HI in the acidic and basic media. A proposed surface orientation model is presented here as an explanation to the experimental data, shedding light for further research on the behavior of insulin as a Langmuir

  4. Bariatric Surgery in Morbidly Obese Insulin Resistant Humans Normalises Insulin Signalling but Not Insulin-Stimulated Glucose Disposal

    PubMed Central

    de Berker, David A. R.; May, Margaret T.; Hers, Ingeborg; Dayan, Colin M.; Andrews, Robert C.; Tavaré, Jeremy M.

    2015-01-01

    Aims Weight-loss after bariatric surgery improves insulin sensitivity, but the underlying molecular mechanism is not clear. To ascertain the effect of bariatric surgery on insulin signalling, we examined glucose disposal and Akt activation in morbidly obese volunteers before and after Roux-en-Y gastric bypass surgery (RYGB), and compared this to lean volunteers. Materials and Methods The hyperinsulinaemic euglycaemic clamp, at five infusion rates, was used to determine glucose disposal rates (GDR) in eight morbidly obese (body mass index, BMI=47.3±2.2 kg/m2) patients, before and after RYGB, and in eight lean volunteers (BMI=20.7±0.7 kg/m2). Biopsies of brachioradialis muscle, taken at fasting and insulin concentrations that induced half-maximal (GDR50) and maximal (GDR100) GDR in each subject, were used to examine the phosphorylation of Akt-Thr308, Akt-473, and pras40, in vivo biomarkers for Akt activity. Results Pre-operatively, insulin-stimulated GDR was lower in the obese compared to the lean individuals (P<0.001). Weight-loss of 29.9±4 kg after surgery significantly improved GDR50 (P=0.004) but not GDR100 (P=0.3). These subjects still remained significantly more insulin resistant than the lean individuals (p<0.001). Weight loss increased insulin-stimulated skeletal muscle Akt-Thr308 and Akt-Ser473 phosphorylation, P=0.02 and P=0.03 respectively (MANCOVA), and Akt activity towards the substrate PRAS40 (P=0.003, MANCOVA), and in contrast to GDR, were fully normalised after the surgery (obese vs lean, P=0.6, P=0.35, P=0.46, respectively). Conclusions Our data show that although Akt activity substantially improved after surgery, it did not lead to a full restoration of insulin-stimulated glucose disposal. This suggests that a major defect downstream of, or parallel to, Akt signalling remains after significant weight-loss. PMID:25876175

  5. Premixed direct injection nozzle

    DOEpatents

    Zuo, Baifang; Johnson, Thomas Edward; Lacy, Benjamin Paul; Ziminsky, Willy Steve

    2011-02-15

    An injection nozzle having a main body portion with an outer peripheral wall is disclosed. The nozzle includes a plurality of fuel/air mixing tubes disposed within the main body portion and a fuel flow passage fluidly connected to the plurality of fuel/air mixing tubes. Fuel and air are partially premixed inside the plurality of the tubes. A second body portion, having an outer peripheral wall extending between a first end and an opposite second end, is connected to the main body portion. The partially premixed fuel and air mixture from the first body portion gets further mixed inside the second body portion. The second body portion converges from the first end toward said second end. The second body portion also includes cooling passages that extend along all the walls around the second body to provide thermal damage resistance for occasional flame flash back into the second body.

  6. Cloning of a new member of the insulin gene superfamily (INSL4) expressed in human placenta

    SciTech Connect

    Chassin, D.; Laurent, A.; Janneau, J.L.

    1995-09-20

    A new member of the insulin gene superfamily was identified by screening a subtracted cDNA library of first-trimester human placenta and, hence, was tentatively named early placenta insulin-like peptide (EPIL). In this paper, we report the cloning and sequencing of the EPIL cDNA and the EPIL gene (INSL4). Comparison of the deduced amino acid sequence of the early placenta insulin-like peptide revealed significant overall and structural homologies with members of the insulin-like hormone superfamily. Moreover, the organization of the early placenta insulin-like gene, which is composed of two exons and one intron, is similiar to that of insulin and relaxin. By in situ hybridization, the INSL4 gene was assigned to band p24 of the short arm of chromosome 9. RT-PCR analysis of EPIL tissue distribution revealed that its transcripts are expressed in the placenta and uterus. 22 refs., 3 figs.

  7. High-mix insulins

    PubMed Central

    Kalra, Sanjay; Farooqi, Mohammad Hamed; El-Houni, Ali E.

    2015-01-01

    Premix insulins are commonly used insulin preparations, which are available in varying ratios of different molecules. These drugs contain one short- or rapid-acting, and one intermediate- or long-acting insulin. High-mix insulins are mixtures of insulins that contain 50% or more than 50% of short-acting insulin. This review describes the clinical pharmacology of high-mix insulins, including data from randomized controlled trials. It suggests various ways, in which high-mix insulin can be used, including once daily, twice daily, thrice daily, hetero-mix, and reverse regimes. The authors provide a rational framework to help diabetes care professionals, identify indications for pragmatic high-mix use. PMID:26425485

  8. Human islet preparations distributed for research exhibit a variety of insulin-secretory profiles

    PubMed Central

    Kayton, Nora S.; Poffenberger, Gregory; Henske, Joseph; Dai, Chunhua; Thompson, Courtney; Aramandla, Radhika; Shostak, Alena; Nicholson, Wendell; Brissova, Marcela; Bush, William S.

    2015-01-01

    Human islet research is providing new insights into human islet biology and diabetes, using islets isolated at multiple US centers from donors with varying characteristics. This creates challenges for understanding, interpreting, and integrating research findings from the many laboratories that use these islets. In what is, to our knowledge, the first standardized assessment of human islet preparations from multiple isolation centers, we measured insulin secretion from 202 preparations isolated at 15 centers over 11 years and noted five distinct patterns of insulin secretion. Approximately three quarters were appropriately responsive to stimuli, but one quarter were dysfunctional, with unstable basal insulin secretion and/or an impairment in stimulated insulin secretion. Importantly, the patterns of insulin secretion by responsive human islet preparations (stable Baseline and Fold stimulation of insulin secretion) isolated at different centers were similar and improved slightly over the years studied. When all preparations studied were considered, basal and stimulated insulin secretion did not correlate with isolation center, biological differences of the islet donor, or differences in isolation, such as Cold Ischemia Time. Dysfunctional islet preparations could not be predicted from the information provided by the isolation center and had altered expression of genes encoding components of the glucose-sensing pathway, but not of insulin production or cell death. These results indicate that insulin secretion by most preparations from multiple centers is similar but that in vitro responsiveness of human islets cannot be predicted, necessitating preexperimental human islet assessment. These results should be considered when one is designing, interpreting, and integrating experiments using human islets. PMID:25648831

  9. Large-Scale Refolding and Enzyme Reaction of Human Preproinsulin for Production of Human Insulin.

    PubMed

    Kim, Chang-Kyu; Lee, Seung-Bae; Son, Young-Jin

    2015-10-01

    Human insulin is composed of 21 amino acids of an A-chain and 30 amino acids of a B-chain. This is the protein hormone that has the role of blood sugar control. When the recombinant human proinsulin is expressed in Escherichia coli, a serious problem is the formation of an inclusion body. Therefore, the inclusion body must be denatured and refolded under chaotropic agents and suitable reductants. In this study, H27R-proinsulin was refolded from the denatured form with β-mercaptoethanol and urea. The refolding reaction was completed after 15 h at 15°C, whereas the reaction at 25°C was faster than that at 15°C. The refolding yield at 15°C was 17% higher than that at 25°C. The refolding reaction could be carried out at a high protein concentration (2 g/l) using direct refolding without sulfonation. The most economical and optimal refolding condition for human preproinsulin was 1.5 g/l protein, 10 mM glycine buffer containing 0.6 M urea, pH 10.6, and 0.3 mM β-mercaptoethanol at 15°C for 16 h. The maximum refolding yield was 74.8% at 15°C with 1.5 g/l protein. Moreover, the refolded preproinsulin could be converted into normal mature insulin with two enzymes. The average amount of human insulin was 138.2 g from 200 L of fermentation broth after enzyme reaction with H27R-proinsulin. The direct refolding process for H27R-proinsulin was successfully set up without sulfonation. The step yields for refolding and enzyme reaction were comparatively high. Therefore, our refolding process for production of recombinant insulin may be beneficial to the large-scale production of other biologically active proteins.

  10. Pharmacokinetics and Pharmacodynamics of High-Dose Human Regular U-500 Insulin Versus Human Regular U-100 Insulin in Healthy Obese Subjects

    PubMed Central

    de la Peña, Amparo; Riddle, Matthew; Morrow, Linda A.; Jiang, Honghua H.; Linnebjerg, Helle; Scott, Adam; Win, Khin M.; Hompesch, Marcus; Mace, Kenneth F.; Jacobson, Jennie G.; Jackson, Jeffrey A.

    2011-01-01

    OBJECTIVE Human regular U-500 (U-500R) insulin (500 units/mL) is increasingly being used clinically, yet its pharmacokinetics (PK) and pharmacodynamics (PD) have not been well studied. Therefore, we compared PK and PD of clinically relevant doses of U-500R with the same doses of human regular U-100 (U-100R) insulin (100 units/mL). RESEARCH DESIGN AND METHODS This was a single-site, randomized, double-blind, crossover euglycemic clamp study. Single subcutaneous injections of 50- and 100-unit doses of U-500R and U-100R were administered to 24 healthy obese subjects. RESULTS Both overall insulin exposure (area under the serum insulin concentration versus time curve from zero to return to baseline [AUC0-t’]) and overall effect (total glucose infused during a clamp) were similar between formulations at both 50- and 100-unit doses (90% [CI] of ratios contained within [0.80, 1.25]). However, peak concentration and effect were significantly lower for U-500R at both doses (P < 0.05). Both formulations produced relatively long durations of action (18.3 to 21.5 h). Time-to-peak concentration and time to maximum effect were significantly longer for U-500R than U-100R at the 100-unit dose (P < 0.05). Time variables reflective of duration of action (late tRmax50, tRlast) were prolonged for U-500R versus U-100R at both doses (P < 0.05). CONCLUSIONS Overall exposure to and action of U-500R insulin after subcutaneous injection were no different from those of U-100R insulin. For U-500R, peaks of concentration and action profiles were blunted and the effect after the peak was prolonged. These findings may help guide therapy with U-500R insulin for highly insulin-resistant patients with diabetes. PMID:21994429

  11. Adipose Cell Size and Regional Fat Deposition as Predictors of Metabolic Response to Overfeeding in Insulin-Resistant and Insulin-Sensitive Humans.

    PubMed

    McLaughlin, Tracey; Craig, Colleen; Liu, Li-Fen; Perelman, Dalia; Allister, Candice; Spielman, Daniel; Cushman, Samuel W

    2016-05-01

    Obesity is associated with insulin resistance, but significant variability exists between similarly obese individuals, pointing to qualitative characteristics of body fat as potential mediators. To test the hypothesis that obese, insulin-sensitive (IS) individuals possess adaptive adipose cell/tissue responses, we measured subcutaneous adipose cell size, insulin suppression of lipolysis, and regional fat responses to short-term overfeeding in BMI-matched overweight/obese individuals classified as IS or insulin resistant (IR). At baseline, IR subjects exhibited significantly greater visceral adipose tissue (VAT), intrahepatic lipid (IHL), plasma free fatty acids, adipose cell diameter, and percentage of small adipose cells. With weight gain (3.1 ± 1.4 kg), IR subjects demonstrated no significant change in adipose cell size, VAT, or insulin suppression of lipolysis and only 8% worsening of insulin-mediated glucose uptake (IMGU). Alternatively, IS subjects demonstrated significant adipose cell enlargement; decrease in the percentage of small adipose cells; increase in VAT, IHL, and lipolysis; 45% worsening of IMGU; and decreased expression of lipid metabolism genes. Smaller baseline adipose cell size and greater enlargement with weight gain predicted decline in IMGU, as did increase in IHL and VAT and decrease in insulin suppression of lipolysis. Weight gain in IS humans causes maladaptive changes in adipose cells, regional fat distribution, and insulin resistance. The correlation between development of insulin resistance and changes in adipose cell size, VAT, IHL, and insulin suppression of lipolysis highlight these factors as potential mediators between obesity and insulin resistance. PMID:26884438

  12. Adipose Cell Size and Regional Fat Deposition as Predictors of Metabolic Response to Overfeeding in Insulin-Resistant and Insulin-Sensitive Humans.

    PubMed

    McLaughlin, Tracey; Craig, Colleen; Liu, Li-Fen; Perelman, Dalia; Allister, Candice; Spielman, Daniel; Cushman, Samuel W

    2016-05-01

    Obesity is associated with insulin resistance, but significant variability exists between similarly obese individuals, pointing to qualitative characteristics of body fat as potential mediators. To test the hypothesis that obese, insulin-sensitive (IS) individuals possess adaptive adipose cell/tissue responses, we measured subcutaneous adipose cell size, insulin suppression of lipolysis, and regional fat responses to short-term overfeeding in BMI-matched overweight/obese individuals classified as IS or insulin resistant (IR). At baseline, IR subjects exhibited significantly greater visceral adipose tissue (VAT), intrahepatic lipid (IHL), plasma free fatty acids, adipose cell diameter, and percentage of small adipose cells. With weight gain (3.1 ± 1.4 kg), IR subjects demonstrated no significant change in adipose cell size, VAT, or insulin suppression of lipolysis and only 8% worsening of insulin-mediated glucose uptake (IMGU). Alternatively, IS subjects demonstrated significant adipose cell enlargement; decrease in the percentage of small adipose cells; increase in VAT, IHL, and lipolysis; 45% worsening of IMGU; and decreased expression of lipid metabolism genes. Smaller baseline adipose cell size and greater enlargement with weight gain predicted decline in IMGU, as did increase in IHL and VAT and decrease in insulin suppression of lipolysis. Weight gain in IS humans causes maladaptive changes in adipose cells, regional fat distribution, and insulin resistance. The correlation between development of insulin resistance and changes in adipose cell size, VAT, IHL, and insulin suppression of lipolysis highlight these factors as potential mediators between obesity and insulin resistance.

  13. Sustained prolonged topical delivery of bioactive human insulin for potential treatment of cutaneous wounds.

    PubMed

    Hrynyk, Michael; Martins-Green, Manuela; Barron, Annelise E; Neufeld, Ronald J

    2010-10-15

    Skin damaged by heat, radiation, or chemical exposure is difficult to treat and slow to heal. Indeed full restoration of the tissue is difficult to obtain. Sub-dermal insulin injection was recently shown to stimulate wound healing of the skin by accelerating wound closure, stimulating angiogenesis and inducing a regenerative process of healing. We have developed a topical delivery vehicle that is capable of releasing therapeutic levels of bioactive insulin for several weeks with the potential to stimulate and sustain healing. By encapsulating the crystalline form of insulin within poly(d,l-lactide-co-glycolide) microspheres, we succeeded in stabilizing and then releasing bioactive insulin for up to 25 days. To measure bioactivity we used Rat L6 myofibroblasts, stimulated them with this slow release insulin and determined activation of the receptors on the cell surface by quantifying AKT phosphorylation. There was only a minor and gradual decrease in AKT phosphorylation over time. To determine whether the slow release insulin could stimulate keratinocyte migration, wounding was simulated by scratching confluent cultures of human keratinocytes (HaCaT). Coverage of the scratch "wounds" was significantly faster in the presence of insulin released from microspheres than in the insulin-free control. Extended and sustained topical delivery of active insulin from a stable protein crystal-based reservoir shows promise in promoting tissue healing.

  14. Sustained prolonged topical delivery of bioactive human insulin for potential treatment of cutaneous wounds.

    PubMed

    Hrynyk, Michael; Martins-Green, Manuela; Barron, Annelise E; Neufeld, Ronald J

    2010-10-15

    Skin damaged by heat, radiation, or chemical exposure is difficult to treat and slow to heal. Indeed full restoration of the tissue is difficult to obtain. Sub-dermal insulin injection was recently shown to stimulate wound healing of the skin by accelerating wound closure, stimulating angiogenesis and inducing a regenerative process of healing. We have developed a topical delivery vehicle that is capable of releasing therapeutic levels of bioactive insulin for several weeks with the potential to stimulate and sustain healing. By encapsulating the crystalline form of insulin within poly(d,l-lactide-co-glycolide) microspheres, we succeeded in stabilizing and then releasing bioactive insulin for up to 25 days. To measure bioactivity we used Rat L6 myofibroblasts, stimulated them with this slow release insulin and determined activation of the receptors on the cell surface by quantifying AKT phosphorylation. There was only a minor and gradual decrease in AKT phosphorylation over time. To determine whether the slow release insulin could stimulate keratinocyte migration, wounding was simulated by scratching confluent cultures of human keratinocytes (HaCaT). Coverage of the scratch "wounds" was significantly faster in the presence of insulin released from microspheres than in the insulin-free control. Extended and sustained topical delivery of active insulin from a stable protein crystal-based reservoir shows promise in promoting tissue healing. PMID:20691251

  15. Diabetes and Insulin

    MedlinePlus

    ... years, but may eventually need insulin to maintain glucose control. What are the different types of insulin? Different ... glulisine • Short-acting: regular human insulin Basal insulin. Controls blood glucose levels between meals and throughout the night. This ...

  16. Insulin in human milk and the use of hormones in infant formulas.

    PubMed

    Shamir, Raanan; Shehadeh, Naim

    2013-01-01

    Human milk contains a substantial number of hormones and growth factors. Studies in animal models show that some of these peptides (e.g. insulin, insulin-like growth factor 1, IGF-1, epidermal growth factors) have an effect on the small intestine after orogastric administration. Recently, two efforts were made to incorporate growth factors into infant formulas. One of these efforts included the incorporation of IGF-1, and the second is an ongoing effort to evaluate the safety and efficacy of incorporating insulin into infant formulas. The rational and current evidence for adding insulin to infant formulas (presence in human milk, effects of orally administrated insulin on gut maturation, intestinal permeability, systemic effects and preliminary encouraging results of supplementing insulin to a preterm infant formula) is detailed in this review. If the addition of insulin to preterm infant formulas indeed results in better growth and accelerated intestinal maturation, future studies will need to address the supplementation of insulin in term infants and assess the efficacy of such supplementation in enhancing gut maturation and prevention of later noncommunicable diseases such as allergy, autoimmune diseases and obesity. PMID:24107496

  17. Insulin in human milk and the use of hormones in infant formulas.

    PubMed

    Shamir, Raanan; Shehadeh, Naim

    2013-01-01

    Human milk contains a substantial number of hormones and growth factors. Studies in animal models show that some of these peptides (e.g. insulin, insulin-like growth factor 1, IGF-1, epidermal growth factors) have an effect on the small intestine after orogastric administration. Recently, two efforts were made to incorporate growth factors into infant formulas. One of these efforts included the incorporation of IGF-1, and the second is an ongoing effort to evaluate the safety and efficacy of incorporating insulin into infant formulas. The rational and current evidence for adding insulin to infant formulas (presence in human milk, effects of orally administrated insulin on gut maturation, intestinal permeability, systemic effects and preliminary encouraging results of supplementing insulin to a preterm infant formula) is detailed in this review. If the addition of insulin to preterm infant formulas indeed results in better growth and accelerated intestinal maturation, future studies will need to address the supplementation of insulin in term infants and assess the efficacy of such supplementation in enhancing gut maturation and prevention of later noncommunicable diseases such as allergy, autoimmune diseases and obesity.

  18. Effect of insulin and glucose on adenosine metabolizing enzymes in human B lymphocytes.

    PubMed

    Kocbuch, Katarzyna; Sakowicz-Burkiewicz, Monika; Grden, Marzena; Szutowicz, Andrzej; Pawelczyk, Tadeusz

    2009-01-01

    In diabetes several aspects of immunity are altered, including the immunomodulatory action of adenosine. Our study was undertaken to investigate the effect of different glucose and insulin concentrations on activities of adenosine metabolizing enzymes in human B lymphocytes line SKW 6.4. The activity of adenosine deaminase in the cytosolic fraction was very low and was not affected by different glucose concentration, but in the membrane fraction of cells cultured with 25 mM glucose it was decreased by about 35% comparing to the activity in cells maintained in 5 mM glucose, irrespective of insulin concentration. The activities of 5'-nucleotidase (5'-NT) and ecto-5'-NT in SKW 6.4 cells depended on insulin concentration, but not on glucose. Cells cultured with 10(-8) M insulin displayed an about 60% lower activity of cytosolic 5'-NT comparing to cells maintained at 10(-11) M insulin. The activity of ecto-5'-NT was decreased by about 70% in cells cultured with 10(-8) M insulin comparing to cells grown in 10(-11) M insulin. Neither insulin nor glucose had an effect on adenosine kinase (AK) activity in SKW 6.4 cells or in human B cells isolated from peripheral blood. The extracellular level of adenosine and inosine during accelerated catabolism of cellular ATP depended on glucose, but not on insulin concentration. Concluding, our study demonstrates that glucose and insulin differentially affect the activities of adenosine metabolizing enzymes in human B lymphocytes, but changes in those activities do not correlate with the adenosine level in cell media during accelerated ATP catabolism, implying that nucleoside transport is the primary factor determining the extracellular level of adenosine.

  19. Effects of pasteurization on adiponectin and insulin concentrations in donor human milk.

    PubMed

    Ley, Sylvia H; Hanley, Anthony J; Stone, Debbie; O'Connor, Deborah L

    2011-09-01

    Although pasteurization is recommended before distributing donor human milk in North America, limited data are available on its impact on metabolic hormones in milk. We aimed to investigate the effects of pasteurization on adiponectin and insulin concentrations in donor human milk. The study investigates concentrations of components in donor human milk before and after Holder pasteurization. After the guidelines of the Human Milk Bank Association of North America, human milk samples were pooled to produce 17 distinct batches (4 individuals per batch) and pasteurized at 62.5°C for 30 min. Adiponectin, insulin, energy, fat, total protein, and glucose concentrations were measured pre- and postpasteurization. Pasteurization reduced milk adiponectin and insulin by 32.8 and 46.1%, respectively (both p < 0.0001). Adiponectin and insulin were significantly correlated with energy and fat milk composition (r = 0.36-0.47; all p < 0.05). Pasteurization effects on milk hormone concentrations remained significant after adjusting for fat and energy (beta ± SEE: -4.11 ± 1.27, p = 0.003 for adiponectin; -70.0 ± 15.0, p < 0.0001 for insulin). Holder pasteurization reduced adiponectin and insulin concentrations in donor human milk. In view of emerging knowledge on the importance of milk components, continued work to find the optimal pasteurization process that mitigates risks but promotes retention of bioactive components is needed.

  20. Human insulin fibril-assisted synthesis of fluorescent gold nanoclusters in alkaline media under physiological temperature.

    PubMed

    Garcia, Andrew R; Rahn, Ivy; Johnson, Sheba; Patel, Ravi; Guo, Jingru; Orbulescu, Jhony; Micic, Miodrag; Whyte, Jeffrey D; Blackwelder, Patricia; Leblanc, Roger M

    2013-05-01

    Fluorescent insulin fibrils gold nanoclusters (Au NCs) have been synthesized through the reduction of gold by human insulin in fibrillated form. Likewise, nanocluster formation has been regulated by insulin, working as a protein-based template. Environment- and surface-controlled experiments have shown the optimized synthesis conditions is comprised of a pure aqueous alkaline solvent for insulin under constant heat at physiological temperature (37°C) prior to addition of the Au precursor (HAuCl4), followed by subsequent heating (37°C) and vigorous stirring after the addition of HAuCl4 until the completion of the synthetic approach. Microscopy experiments detected the presence of primordial fibril structures in samples of heated human insulin in the alkaline medium prior to addition of HAuCl4, while encountering more developed insulin fibrils in the terminal production of Au NCs. This investigation provides insight to the development of a novel synthesis of Au NCs in the alkaline medium, while providing a graphical description of the environmental and surface-dependent effects that were presented in the synthesis of human insulin nanoclusters. The study provides pertinent information for future synthetic procedures, as the protein state of several protein-nanoparticle systems may reflect on the results that were obtained herein.

  1. Crystallization of Enantiomerically Pure Proteins from Quasi-Racemic Mixtures: Structure Determination by X-Ray Diffraction of Isotope-Labeled Ester Insulin and Human Insulin.

    PubMed

    Mandal, Kalyaneswar; Dhayalan, Balamurugan; Avital-Shmilovici, Michal; Tokmakoff, Andrei; Kent, Stephen B H

    2016-03-01

    As a part of a program aimed towards the study of the dynamics of human insulin-protein dimer formation using two-dimensional infrared spectroscopy, we used total chemical synthesis to prepare stable isotope labeled [(1-(13) C=(18) O)Phe(B24) )] human insulin, via [(1-(13) C=(18) O)Phe(B24) )] ester insulin as a key intermediate product that facilitates folding of the synthetic protein molecule (see preceding article). Here, we describe the crystal structure of the synthetic isotope-labeled ester insulin intermediate and the product synthetic human insulin. Additionally, we present our observations on hexamer formation with these two proteins in the absence of phenol derivatives and/or Zn metal ions. We also describe and discuss the fractional crystallization of quasi-racemic protein mixtures containing each of these two synthetic proteins.

  2. Glucose responsive insulin production from human embryonic germ (EG) cell derivatives

    SciTech Connect

    Clark, Gregory O.; Yochem, Robert L.; Axelman, Joyce; Sheets, Timothy P.; Kaczorowski, David J.; Shamblott, Michael J. . E-mail: mshambl1@jhmi.edu

    2007-05-11

    Type 1 diabetes mellitus subjects millions to a daily burden of disease management, life threatening hypoglycemia and long-term complications such as retinopathy, nephropathy, heart disease, and stroke. Cell transplantation therapies providing a glucose-regulated supply of insulin have been implemented clinically, but are limited by safety, efficacy and supply considerations. Stem cells promise a plentiful and flexible source of cells for transplantation therapies. Here, we show that cells derived from human embryonic germ (EG) cells express markers of definitive endoderm, pancreatic and {beta}-cell development, glucose sensing, and production of mature insulin. These cells integrate functions necessary for glucose responsive regulation of preproinsulin mRNA and expression of insulin C-peptide in vitro. Following transplantation into mice, cells become insulin and C-peptide immunoreactive and produce plasma C-peptide in response to glucose. These findings suggest that EG cell derivatives may eventually serve as a source of insulin producing cells for the treatment of diabetes.

  3. Insulin stimulates synthesis and release of human chorionic gonadotropin by choriocarcinoma cell lines

    SciTech Connect

    Ren, S.G.; Braunstein, G.D. )

    1991-03-01

    Recent studies have shown that insulin regulates placental lactogen, progesterone, and estrogen production from human trophoblast cells. This study was performed to examine whether insulin also regulates the production of hCG by this type of cell. After 24-36 h of preincubation, JEG-3 and JAR cells (2-3 x 10(5) cells/ml.well) or human term trophoblast cells (1 x 10(6) cells/ml.well) were exposed to the test hormone in serum-free Dulbecco's Modified Eagle's Medium for 24-96 h. Secretion of hCG from JEG-3 cells was stimulated by human insulin, human proinsulin, or porcine insulin in a dose-dependent manner, with lowest effective doses of 6.7, 96, and 53 mg/L, respectively. Time-course studies showed that hCG secretion peaked at 72-96 h with insulin exposure; in contrast, no decernable peak was seen without insulin in serum-free media. Exposure of JEG-3 cells for 24 h to 209 mg/liter insulin stimulated hCG synthesis, with 40 +/- 3% more immunoreactive intracellular hCG (P less than 0.05). Cells grown in the presence of insulin and (35S)methionine had 47 +/- 21% more labeled intracellular hCG and 56 +/- 13% more immunoprecipitable (35S)methionine-hCG secreted into the medium than the control cultures (P less than 0.05). During this time period, human placental lactogen release and total trichloroacetice acid-precipitable (35S)methionine protein were not increased. The insulin-induced stimulation of hCG synthesis was inhibited by cycloheximide. Additionally, insulin did not significantly affect total intracellular protein during 24-96 h of incubation. Insulin also increased hCG release from JAR cells, but not from human term trophoblast cells. A mouse monoclonal antibody to the IGF-I receptor inhibited the stimulation of insulin in JEG-3 cells.

  4. Root and shoot parts of strawberry: factories for production of functional human pro-insulin.

    PubMed

    Tavizi, Ashkan; Javaran, Mokhtar Jalali; Moieni, Ahmad; Mohammadi-Dehcheshmeh, Manijeh; Mohebodini, Mehdi; Ebrahimie, Esmaeil

    2015-05-01

    Diabetes, a disease caused by excessive blood sugar, is caused by the lack of insulin. For commercial production, insulin is made in bacteria or yeast by protein recombinant technology. The focus of this research is evaluating another resource and producing of recombinant insulin protein in as strawberry as this plant has high potential in production of pharmaceutical proteins. Strawberry is a suitable bioreactor for production of recombinant proteins especially edible vaccines. In this research, human pro-insulin gene was cloned in pCAMBIA1304 vector under CaMV35S promoter and NOS terminator. Agrobacterium tumefaciens LBA4404, AGL1, EHA105, EHA101, C58, C58 (pGV2260) and C58 (pGV3101) strains were used for transformation of pro-insulin gene into strawberry cv. Camarosa, Selva, Sarian Hybrid, Pajaro, Paros, Gaviota, Alpine. Additionally, Agrobacterium rhizogenes K599, R1000, A4 and MSU440 strains were utilized for gene transformation into hairy roots. PCR analysis indicated the presence of transformed human pro-insulin gene in the strawberry and hairy roots. Also, its transcription was confirmed using RT-PCR. Furthermore, the analysis of plants, fruits and hairy roots at the level of proteins using dot blot, ELISA, SDS-PAGE and ECL tests re-confirmed the expression of this protein in the transgenic plants as well as hairy roots. Protein purification of human pro-insulin from transgenic tissues was performed using affinity chromatography. Finally, the bioassay of recombinant pro-insulin was performed. The analysis of second generations of transgenic plants (T1) at DNA and protein levels was also performed as a complementary experiment. This study opens a new avenue in molecular farming of human pro-insulin through its mass production in roots and shoots of strawberry.

  5. Altered Skeletal Muscle Lipase Expression and Activity Contribute to Insulin Resistance in Humans

    PubMed Central

    Badin, Pierre-Marie; Louche, Katie; Mairal, Aline; Liebisch, Gerhard; Schmitz, Gerd; Rustan, Arild C.; Smith, Steven R.; Langin, Dominique; Moro, Cedric

    2011-01-01

    OBJECTIVE Insulin resistance is associated with elevated content of skeletal muscle lipids, including triacylglycerols (TAGs) and diacylglycerols (DAGs). DAGs are by-products of lipolysis consecutive to TAG hydrolysis by adipose triglyceride lipase (ATGL) and are subsequently hydrolyzed by hormone-sensitive lipase (HSL). We hypothesized that an imbalance of ATGL relative to HSL (expression or activity) may contribute to DAG accumulation and insulin resistance. RESEARCH DESIGN AND METHODS We first measured lipase expression in vastus lateralis biopsies of young lean (n = 9), young obese (n = 9), and obese-matched type 2 diabetic (n = 8) subjects. We next investigated in vitro in human primary myotubes the impact of altered lipase expression/activity on lipid content and insulin signaling. RESULTS Muscle ATGL protein was negatively associated with whole-body insulin sensitivity in our population (r = −0.55, P = 0.005), whereas muscle HSL protein was reduced in obese subjects. We next showed that adenovirus-mediated ATGL overexpression in human primary myotubes induced DAG and ceramide accumulation. ATGL overexpression reduced insulin-stimulated glycogen synthesis (−30%, P < 0.05) and disrupted insulin signaling at Ser1101 of the insulin receptor substrate-1 and downstream Akt activation at Ser473. These defects were fully rescued by nonselective protein kinase C inhibition or concomitant HSL overexpression to restore a proper lipolytic balance. We show that selective HSL inhibition induces DAG accumulation and insulin resistance. CONCLUSIONS Altogether, the data indicate that altered ATGL and HSL expression in skeletal muscle could promote DAG accumulation and disrupt insulin signaling and action. Targeting skeletal muscle lipases may constitute an interesting strategy to improve insulin sensitivity in obesity and type 2 diabetes. PMID:21498783

  6. Computational model of cellular metabolic dynamics: effect of insulin on glucose disposal in human skeletal muscle.

    PubMed

    Li, Yanjun; Solomon, Thomas P J; Haus, Jacob M; Saidel, Gerald M; Cabrera, Marco E; Kirwan, John P

    2010-06-01

    Identifying the mechanisms by which insulin regulates glucose metabolism in skeletal muscle is critical to understanding the etiology of insulin resistance and type 2 diabetes. Our knowledge of these mechanisms is limited by the difficulty of obtaining in vivo intracellular data. To quantitatively distinguish significant transport and metabolic mechanisms from limited experimental data, we developed a physiologically based, multiscale mathematical model of cellular metabolic dynamics in skeletal muscle. The model describes mass transport and metabolic processes including distinctive processes of the cytosol and mitochondria. The model simulated skeletal muscle metabolic responses to insulin corresponding to human hyperinsulinemic-euglycemic clamp studies. Insulin-mediated rate of glucose disposal was the primary model input. For model validation, simulations were compared with experimental data: intracellular metabolite concentrations and patterns of glucose disposal. Model variations were simulated to investigate three alternative mechanisms to explain insulin enhancements: Model 1 (M.1), simple mass action; M.2, insulin-mediated activation of key metabolic enzymes (i.e., hexokinase, glycogen synthase, pyruvate dehydrogenase); or M.3, parallel activation by a phenomenological insulin-mediated intracellular signal that modifies reaction rate coefficients. These simulations indicated that models M.1 and M.2 were not sufficient to explain the experimentally measured metabolic responses. However, by application of mechanism M.3, the model predicts metabolite concentration changes and glucose partitioning patterns consistent with experimental data. The reaction rate fluxes quantified by this detailed model of insulin/glucose metabolism provide information that can be used to evaluate the development of type 2 diabetes.

  7. Differentiation of human embryonic stem cells into insulin-producing clusters.

    PubMed

    Segev, Hanna; Fishman, Bettina; Ziskind, Anna; Shulman, Margarita; Itskovitz-Eldor, Joseph

    2004-01-01

    Type I diabetes mellitus is caused by an autoimmune destruction of the insulin-producing beta cells. The major obstacle in using transplantation for curing the disease is the limited source of insulin-producing cells. The isolation of human embryonic stem (hES) cells introduced a new prospect for obtaining a sufficient number of beta cells for transplantation. We present here a method for forming immature islet-like clusters of insulin-producing cells derived from hES cells. The protocol consisted of several steps. Embryoid bodies were first cultured and plated in insulin-transferrin-selenium-fibronectin medium, followed by medium supplemented with N2, B27, and basic fibroblast growth factor (bFGF). Next, the glucose concentration in the medium was lowered, bFGF was withdrawn, and nicotinamide was added. Dissociating the cells and growing them in suspension resulted in the formation of clusters which exhibited higher insulin secretion and had longer durability than cells grown as monolayers. Reverse transcription-polymerase chain reaction detected an enhanced expression of pancreatic genes in the differentiated cells. Immunofluorescence and in situ hybridization analyses revealed a high percentage of insulin-expressing cells in the clusters. In addition to insulin, most cells also coexpressed glucagon or somatostatin, indicating a similarity to immature pancreatic cells. Further improvement of this insulin-producing cell protocol may lead to the formation of an unlimited source of cells suitable for transplantation.

  8. Insulin sensitivity of protein and glucose metabolism in human forearm skeletal muscle.

    PubMed Central

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

    1992-01-01

    Physiologic increases of insulin promote net amino acid uptake and protein anabolism in forearm skeletal muscle by restraining protein degradation. The sensitivity of this process to insulin is not known. Using the forearm perfusion method, we infused insulin locally in the brachial artery at rates of 0.00 (saline control), 0.01, 0.02, 0.035, or 0.05 mU/min per kg for 150 min to increase local forearm plasma insulin concentration by 0, approximately 20, approximately 35, approximately 60, and approximately 120 microU/ml (n = 35). L-[ring-2,6-3H]phenylalanine and L-[1-14C]leucine were infused systemically, and the net forearm balance, rate of appearance (Ra) and rate of disposal (R(d)) of phenylalanine and leucine, and forearm glucose balance were measured basally and in response to insulin infusion. Compared to saline, increasing rates of insulin infusion progressively increased net forearm glucose uptake from 0.9 mumol/min per 100 ml (saline) to 1.0, 1.8, 2.4, and 4.7 mumol/min per 100 ml forearm, respectively. Net forearm balance for phenylalanine and leucine was significantly less negative than basal (P < 0.01 for each) in response to the lowest dose insulin infusion, 0.01 mU/min per kg, and all higher rates of insulin infusion. Phenylalanine and leucine R(a) declined by approximately 38 and 40% with the lowest dose insulin infusion. Higher doses of insulin produced no greater effect (decline in R(a) varied between 26 and 42% for phenylalanine and 30-50% for leucine). In contrast, R(d) for phenylalanine and leucine did not change with insulin. We conclude that even modest increases of plasma insulin can markedly suppress proteolysis, measured by phenylalanine R(a), in human forearm skeletal muscle. Further increments of insulin within the physiologic range augment glucose uptake but have little additional effect on phenylalanine R(a) or balance. These results suggest that proteolysis in human skeletal muscle is more sensitive than glucose uptake to physiologic

  9. Insulin Regulates the Unfolded Protein Response in Human Adipose Tissue

    PubMed Central

    Boden, Guenther; Cheung, Peter; Salehi, Sajad; Homko, Carol; Loveland-Jones, Catherine; Jayarajan, Senthil; Stein, T. Peter; Williams, Kevin Jon; Liu, Ming-Lin; Barrero, Carlos A.; Merali, Salim

    2014-01-01

    Endoplasmic reticulum (ER) stress is increased in obesity and is postulated to be a major contributor to many obesity-related pathologies. Little is known about what causes ER stress in obese people. Here, we show that insulin upregulated the unfolded protein response (UPR), an adaptive reaction to ER stress, in vitro in 3T3-L1 adipocytes and in vivo, in subcutaneous (sc) adipose tissue of nondiabetic subjects, where it increased the UPR dose dependently over the entire physiologic insulin range (from ∼35 to ∼1,450 pmol/L). The insulin-induced UPR was not due to increased glucose uptake/metabolism and oxidative stress. It was associated, however, with increased protein synthesis, with accumulation of ubiquitination associated proteins, and with multiple posttranslational protein modifications (acetylations, methylations, nitrosylations, succinylation, and ubiquitinations), some of which are potential causes for ER stress. These results reveal a new physiologic role of insulin and provide a putative mechanism for the development of ER stress in obesity. They may also have clinical and therapeutic implications, e.g., in diabetic patients treated with high doses of insulin. PMID:24130338

  10. Insulin increases phosphorylation of mitochondrial proteins in human skeletal muscle in vivo.

    PubMed

    Zhao, Xiaolu; Bak, Steffen; Pedersen, Andreas J T; Jensen, Ole Nørregaard; Højlund, Kurt

    2014-05-01

    There is increasing evidence that multiple proteins involved in key regulatory processes in mitochondria are phosphorylated in mammalian tissues. Insulin regulates glucose metabolism by phosphorylation-dependent signaling and has been shown to stimulate ATP synthesis in human skeletal muscle. Here, we investigated the effect of insulin on the phosphorylation of mitochondrial proteins in human skeletal muscle in vivo. Using a combination of TiO(2) phosphopeptide-enrichment, HILIC fractionation, and LC-MS/MS, we compared the phosphoproteomes of isolated mitochondria from skeletal muscle samples obtained from healthy individuals before and after 4 h of insulin infusion. In total, we identified 207 phosphorylation sites in 95 mitochondrial proteins. Of these phosphorylation sites, 45% were identified in both basal and insulin-stimulated samples. Insulin caused a 2-fold increase in the number of different mitochondrial phosphopeptides (87 ± 7 vs 40 ± 7, p = 0.015) and phosphoproteins (46 ± 2 vs 26 ± 3, p = 0.005) identified in each mitochondrial preparation. Almost half of the mitochondrial phosphorylation sites (n = 94) were exclusively identified in the insulin-stimulated state and included the majority of novel sites. Phosphorylation sites detected more often or exclusively in insulin-stimulated samples include multiple sites in mitochondrial proteins involved in oxidative phosphorylation, tricarboxylic acid cycle, and fatty acid metabolism, as well as several components of the newly defined mitochondrial inner membrane organizing system (MINOS). In conclusion, the present study demonstrates that insulin increases the phosphorylation of several mitochondrial proteins in human skeletal muscle in vivo and provides a first step in the understanding of how insulin potentially regulates mitochondrial processes by phosphorylation-dependent mechanisms.

  11. Reduced DPP4 activity improves insulin signaling in primary human adipocytes.

    PubMed

    Röhrborn, Diana; Brückner, Julia; Sell, Henrike; Eckel, Jürgen

    2016-03-11

    DPP4 is a ubiquitously expressed cell surface protease which is also released to the circulation as soluble DPP4 (sDPP4). Recently, we identified DPP4 as a novel adipokine oversecreted in obesity and thus potentially linking obesity to the metabolic syndrome. Furthermore, sDPP4 impairs insulin signaling in an autocrine and paracrine fashion in different cell types. However, it is still unknown which functional role DPP4 might play in adipocytes. Therefore, primary human adipocytes were treated with a specific DPP4 siRNA. Adipocyte differentiation was not affected by DPP4 silencing. Interestingly, DPP4 reduction improved insulin responsiveness of adipocytes at the level of insulin receptor, proteinkinase B (Akt) and Akt substrate of 160 kDa. To investigate whether the observed effects could be attributed to the enzymatic activity of DPP4, human adipocytes were treated with the DPP4 inhibitors sitagliptin and saxagliptin. Our data show that insulin-stimulated activation of Akt is augmented by DPP4 inhibitor treatment. Based on our previous observation that sDPP4 induces insulin resistance in adipocytes, and that adipose DPP4 levels are higher in obese insulin-resistant patients, we now suggest that the abundance of DPP4 might be a regulator of adipocyte insulin signaling. PMID:26872429

  12. Fibroblast Growth Factor 21 Improves Insulin Sensitivity and Synergizes with Insulin in Human Adipose Stem Cell-Derived (hASC) Adipocytes

    PubMed Central

    Lee, Darwin V.; Li, Dongmei; Yan, Qingyun; Zhu, Yimin; Goodwin, Bryan; Calle, Roberto; Brenner, Martin B.; Talukdar, Saswata

    2014-01-01

    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. PMID:25365322

  13. Impairment of glucose-induced insulin secretion in human pancreatic islets transplanted to diabetic nude mice.

    PubMed

    Jansson, L; Eizirik, D L; Pipeleers, D G; Borg, L A; Hellerström, C; Andersson, A

    1995-08-01

    Hyperglycemia-induced beta-cell dysfunction may be an important component in the pathogenesis of non-insulin-dependent diabetes mellitus. However, most available data in this field were obtained from rodent islets. To investigate the relevance of this hypothesis for human beta-cells in vivo, human pancreatic islets were transplanted under the renal capsule of nude mice. Experimental groups were chosen so that grafted islets were exposed to either hyper- or normoglycemia or combinations of these for 4 or 6 wk. Grafts of normoglycemic recipients responded with an increased insulin release to a glucose stimulus during perfusion, whereas grafts of hyperglycemic recipients failed to respond to glucose. The insulin content of the grafts in the latter groups was only 10% of those observed in controls. Recipients initially hyperglycemic (4 wk), followed by 2 wk of normoglycemia regained a normal graft insulin content, but a decreased insulin response to glucose remained. No ultrastructural signs of beta-cell damage were observed, with the exception of increased glycogen deposits in animals hyperglycemic at the time of killing. It is concluded that prolonged exposure to a diabetic environment induces a long-term secretory defect in human beta-cells, which is not dependent on the size of the islet insulin stores.

  14. Serum Autotaxin/ENPP2 Correlates with Insulin Resistance in Older Humans with Obesity

    PubMed Central

    Reeves, Valerie L.; Trybula, Joy S.; Wills, Rachel C.; Goodpaster, Bret H.; Dubé, John J.; Kienesberger, Petra C.; Kershaw, Erin E.

    2015-01-01

    Objective Autotaxin (ATX) is an adipocyte-derived lysophospholipase D that generates the lipid signaling molecule lysophosphatidic acid (LPA). The ATX/LPA pathway in adipose tissue has recently been implicated in obesity and insulin resistance in animal models, but the role of circulating ATX in humans remains unclear. The aim of the present study was to determine the relationship between serum ATX and insulin resistance. Methods In this retrospective study, older (60–75 years), non-diabetic human participants with overweight or obesity (BMI 25–37 kg/m2), were characterized for metabolic phenotype including measures of energy, glucose, and lipid homeostasis. The relationship between serum ATX and metabolic parameters was then determined using correlative and predictive statistics. Results Serum ATX was higher in females than in males. After controlling for sex, serum ATX correlated with multiple measures of adiposity and glucose homeostasis/insulin action. Serum ATX and BMI also independently predicted glucose infusion rate during a hyperinsulinemic euglycemic clamp and homeostatic model assessment of insulin resistance after controlling for sex and medication use. Conclusion Serum ATX correlates with and predicts measures of glucose homeostasis and insulin sensitivity in older humans, suggesting that it may be a potential pathogenic factor and/or diagnostic/therapeutic target for insulin resistance in this population. PMID:26727116

  15. Differential interaction of Apolipoprotein-E isoforms with insulin receptors modulates brain insulin signaling in mutant human amyloid precursor protein transgenic mice.

    PubMed

    Chan, Elizabeth S; Chen, Christopher; Cole, Gregory M; Wong, Boon-Seng

    2015-09-08

    It is unclear how human apolipoprotein E4 (ApoE4) increases the risk for Alzheimer's disease (AD). Although Aβ levels can lead to insulin signaling impairment, these experiments were done in the absence of human ApoE. To examine ApoE role, we crossed the human ApoE-targeted replacement mice with mutant human amyloid precursor protein (APP) mice. In 26 week old mice with lower Aβ levels, the expression and phosphorylation of insulin signaling proteins remained comparable among APP, ApoE3xAPP and ApoE4xAPP mouse brains. When the mice aged to 78 weeks, these proteins were markedly reduced in APP and ApoE4xAPP mouse brains. While Aβ can bind to insulin receptor, how ApoE isoforms modulate this interaction remains unknown. Here, we showed that ApoE3 had greater association with insulin receptor as compared to ApoE4, regardless of Aβ42 concentration. In contrast, ApoE4 bound more Aβ42 with increasing peptide levels. Using primary hippocampal neurons, we showed that ApoE3 and ApoE4 neurons are equally sensitive to physiological levels of insulin. However, in the presence of Aβ42, insulin failed to elicit a downstream response only in ApoE4 hippocampal neurons. Taken together, our data show that ApoE genotypes can modulate this Aβ-mediated insulin signaling impairment.

  16. Brown adipose tissue improves whole-body glucose homeostasis and insulin sensitivity in humans.

    PubMed

    Chondronikola, Maria; Volpi, Elena; Børsheim, Elisabet; Porter, Craig; Annamalai, Palam; Enerbäck, Sven; Lidell, Martin E; Saraf, Manish K; Labbe, Sebastien M; Hurren, Nicholas M; Yfanti, Christina; Chao, Tony; Andersen, Clark R; Cesani, Fernando; Hawkins, Hal; Sidossis, Labros S

    2014-12-01

    Brown adipose tissue (BAT) has attracted scientific interest as an antidiabetic tissue owing to its ability to dissipate energy as heat. Despite a plethora of data concerning the role of BAT in glucose metabolism in rodents, the role of BAT (if any) in glucose metabolism in humans remains unclear. To investigate whether BAT activation alters whole-body glucose homeostasis and insulin sensitivity in humans, we studied seven BAT-positive (BAT(+)) men and five BAT-negative (BAT(-)) men under thermoneutral conditions and after prolonged (5-8 h) cold exposure (CE). The two groups were similar in age, BMI, and adiposity. CE significantly increased resting energy expenditure, whole-body glucose disposal, plasma glucose oxidation, and insulin sensitivity in the BAT(+) group only. These results demonstrate a physiologically significant role of BAT in whole-body energy expenditure, glucose homeostasis, and insulin sensitivity in humans, and support the notion that BAT may function as an antidiabetic tissue in humans.

  17. Premixed direct injection disk

    SciTech Connect

    York, William David; Ziminsky, Willy Steve; Johnson, Thomas Edward; Lacy, Benjamin; Zuo, Baifang; Uhm, Jong Ho

    2013-04-23

    A fuel/air mixing disk for use in a fuel/air mixing combustor assembly is provided. The disk includes a first face, a second face, and at least one fuel plenum disposed therebetween. A plurality of fuel/air mixing tubes extend through the pre-mixing disk, each mixing tube including an outer tube wall extending axially along a tube axis and in fluid communication with the at least one fuel plenum. At least a portion of the plurality of fuel/air mixing tubes further includes at least one fuel injection hole have a fuel injection hole diameter extending through said outer tube wall, the fuel injection hole having an injection angle relative to the tube axis. The invention provides good fuel air mixing with low combustion generated NOx and low flow pressure loss translating to a high gas turbine efficiency, that is durable, and resistant to flame holding and flash back.

  18. Lean premixed/prevaporized combustion

    NASA Technical Reports Server (NTRS)

    Lefebvre, A. H. (Editor)

    1977-01-01

    Recommendations were formulated on the status and application of lean premixed/prevaporized combustion to the aircraft gas turbine for the reduction of pollutant emissions. The approach taken by the NASA Stratospheric Cruise Emission Reduction Program (SCERP) in pursuing the lean premixed/prevaporized combustion technique was also discussed. The proceedings contains an overview of the SCERP program, the discussions and recommendations of the participants, and an overall summary.

  19. Rates and tissue sites of non-insulin- and insulin-mediated glucose uptake in humans

    SciTech Connect

    Baron, A.D.; Brechtel, G.; Wallace, P.; Edelman, S.V.

    1988-12-01

    In vivo glucose uptake can occur via two mechanisms, namely, insulin-mediated glucose uptake (IMGU) and non-insulin-mediated glucose uptake (NIMGU). Although the principal tissue sites for IMGU are skeletal muscle, the tissue sites for NIMGU at a given serum glucose concentration are not known. To examine this issue, rates of whole body glucose uptake (Rd) were measured at basal and during glucose clamp studies performed at euglycemia (approximately 90 mg/dl) and hyperglycemia (approximately 220 mg/dl) in six lean healthy men. Studies were performed during hyperinsulinemia (approximately 70 microU/ml) and during somatostatin-induced insulinopenia to measure IMGU and NIMGU, respectively. During each study, leg glucose balance (arteriovenous catheter technique) was also measured. With this approach, rates of whole body skeletal muscle IMGU and NIMGU can be estimated, and the difference between overall Rd and skeletal muscle glucose uptake represents non-skeletal muscle Rd. The results indicate that approximately 20% of basal Rd is into skeletal muscle. During insulinopenia approximately 86% of body NIMGU occurs in non-skeletal muscle tissues at euglycemia. When hyperglycemia was created, whole body NIMGU increased from 128 +/- 6 to 213 +/- 18 mg/min (P less than 0.01); NIMGU into non-skeletal muscle tissues was 134 +/- 11 and 111 +/- 6 mg/min at hyperglycemia and euglycemia, respectively, P = NS. Therefore, virtually all the hyperglycemia induced increment in NIMGU occurred in skeletal muscle. During hyperinsulinemia, IMGU in skeletal muscle represented 75 and 95% of body Rd, at euglycemia and hyperglycemia, respectively.

  20. Gestational Diabetes Reduces Adenosine Transport in Human Placental Microvascular Endothelium, an Effect Reversed by Insulin

    PubMed Central

    Salomón, Carlos; Westermeier, Francisco; Puebla, Carlos; Arroyo, Pablo; Guzmán-Gutiérrez, Enrique; Pardo, Fabián; Leiva, Andrea; Casanello, Paola; Sobrevia, Luis

    2012-01-01

    Gestational diabetes mellitus (GDM) courses with increased fetal plasma adenosine concentration and reduced adenosine transport in placental macrovascular endothelium. Since insulin modulates human equilibrative nucleoside transporters (hENTs) expression/activity, we hypothesize that GDM will alter hENT2-mediated transport in human placental microvascular endothelium (hPMEC), and that insulin will restore GDM to a normal phenotype involving insulin receptors A (IR-A) and B (IR-B). GDM effect on hENTs expression and transport activity, and IR-A/IR-B expression and associated cell signalling cascades (p42/44 mitogen-activated protein kinases (p42/44mapk) and Akt) role in hPMEC primary cultures was assayed. GDM associates with elevated umbilical whole and vein, but not arteries blood adenosine, and reduced hENTs adenosine transport and expression. IR-A/IR-B mRNA expression and p42/44mapk/Akt ratios (‘metabolic phenotype’) were lower in GDM. Insulin reversed GDM-reduced hENT2 expression/activity, IR-A/IR-B mRNA expression and p42/44mapk/Akt ratios to normal pregnancies (‘mitogenic phenotype’). It is suggested that insulin effects required IR-A and IR-B expression leading to differential modulation of signalling pathways restoring GDM-metabolic to a normal-mitogenic like phenotype. Insulin could be acting as protecting factor for placental microvascular endothelial dysfunction in GDM. PMID:22808198

  1. Human umbilical cord-derived mesenchymal stem cells can secrete insulin in vitro and in vivo.

    PubMed

    Boroujeni, Zahra Niki; Aleyasin, Ahmad

    2014-01-01

    Diabetes mellitus is characterized by autoimmune destruction of pancreatic beta cells, leading to decreased insulin production. Differentiation of mesenchymal stem cells (MSCs) into insulin-producing cells offers novel ways of diabetes treatment. MSCs can be isolated from the human umbilical cord tissue and differentiate into insulin-secreting cells. Human umbilical cord-derived stem cells (hUDSCs) were obtained after birth, selected by plastic adhesion, and characterized by flow cytometric analysis. hUDSCs were transduced with nonintegrated lentivirus harboring PDX1 (nonintegrated LV-PDX1) and was cultured in differentiation medium in 21 days. Pancreatic duodenum homeobox protein-1 (PDX1) is a transcription factor in pancreatic development. Significant expressions of PDX1, neurogenin3 (Ngn3), glucagon, glucose transporter2 (Glut2), and somatostatin were detected by quantitative RT-PCR (P < 0.05). PDX1 and insulin proteins were shown by immunocytochemistry analysis. Insulin secretion of hUDSCs(PDX1+) in the high-glucose medium was 1.8 μU/mL. They were used for treatment of diabetic rats and could decrease the blood glucose level from 400 mg/dL to a normal level in 4 days. In conclusion, our results demonstrated that hUDSCs are able to differentiate into insulin-producing cells by transduction with nonintegrated LV-PDX1. These hUDSCs(PDX1+) have the potential to be used as a viable resource in cell-based gene therapy of type 1 diabetes.

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

    PubMed

    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.

  3. A unique human blood-derived cell population displays high potential for producing insulin.

    PubMed

    Zhao, Yong; Huang, Zhihua; Lazzarini, Ping; Wang, Yong; Di, Anke; Chen, Meiling

    2007-08-17

    Blood can provide a valuable source for the generation of stem cells. Herein we identified a novel cell population from adult human blood, designated peripheral blood insulin-producing cells (PB-IPC). Phenotypic analysis demonstrated that PB-IPC displayed the embryonic stem (ES) cell-associated transcription factors including Oct-4 and Nanog, along with the hematopoietic markers CD9, CD45, and CD117; but lacked expression of the hematopoietic stem cell marker CD34 as well as lymphocyte and monocyte/macrophage markers. Notably, in vitro and in vivo characterization revealed that PB-IPC demonstrated characteristics of islet beta cell progenitors including the expression of beta cell-specific insulin gene transcription factors and prohormone convertases, production of insulin, formation of insulin granules, and the ability to reduce hyperglycemia and migrate into pancreatic islets after transplantation into the diabetic mice. These findings may open up new avenues for autologous blood stem cell-based therapies for diabetes.

  4. Human amnion epithelial cells can be induced to differentiate into functional insulin-producing cells.

    PubMed

    Hou, Yanan; Huang, Qin; Liu, Tianjin; Guo, Lihe

    2008-09-01

    Pancreatic islet transplantation has demonstrated that long-term insulin independence may be achieved in patients suffering from diabetes mellitus type 1. However, limited availability of islet tissue means that new sources of insulin-producing cells that are responsive to glucose are required. Here, we show that human amnion epithelial cells (HAEC) can be induced to differentiate into functional insulin-producing cells in vitro. After induction of differentiation, HAEC expressed multiple pancreatic beta-cell genes, including insulin, pancreas duodenum homeobox-1, paired box gene 6, NK2 transcription factor-related locus 2, Islet 1, glucokinase, and glucose transporter-2, and released C-peptide in a glucose-regulated manner in response to other extracellular stimulations. The transplantation of induced HAEC into streptozotocin-induced diabetic C57 mice reversed hyperglycemia, restored body weight, and maintained euglycemia for 30 d. These findings indicated that HAEC may be a new source for cell replacement therapy in type 1 diabetes.

  5. Effects of human insulin and insulin aspart preparations on levels of IGF-I, IGFBPs and IGF bioactivity in patients with type 1 diabetes

    PubMed Central

    2014-01-01

    Background Insulin aspart (IAsp) and its biphasic preparations BIAsp50 and BIAsp70 (containing 50% and 70% IAsp, respectively) have distinct glucose-lowering properties as compared to human insulin (HI). We investigated whether this affected the circulating IGF-system which depends on the hepatic insulin exposure. Methods In a randomized, four-period crossover study, 19 patients with type 1 diabetes received identical doses (0.2 U/kg sc) of IAsp, BIAsp70, BIAsp50 and HI together with a standardized meal. Serum total IGF-I and IGFBP-1 to -3 were measured by immunoassays for nine hours post-prandially. Bioactive IGF was determined by an in-house, cell-based IGF-I receptor kinase activation (KIRA) assay. Results Despite marked differences in peripheral insulin concentrations and plasma glucose, the four insulin preparations resulted in parallel decreases in IGFBP-1 levels during the first 3 hours, and parallel increases during the last part of the study (3–9 hours). Thus, only minor significances were seen. Insulin aspart and human insulin resulted in a lower area under the curve (AUC) during the first 3 hours as compared to BIAsp70 (p = 0.009), and overall, human insulin resulted in a lower IGFBP-1 AUC than BIAsp70 (p = 0.025). Nevertheless, responses and AUCs of bioactive IGF were similar for all four insulin preparations. Changes in levels of bioactive IGF were inversely correlated to those of IGFBP-1, increasing during the first 3 hours, whereafter levels declined (-0.83 ≤ r ≤ -0.30; all p-values <0.05). Total IGF-I and IGFBP-3 remained stable during the 9 hours, whereas IGFBP-2 changed opposite of IGFBP-1, increasing after 3–4 hours whereafter levels gradually declined. The four insulin preparations resulted in similar profiles and AUCs of total IGF-I, IGFBP-2 and IGFBP-3. Conclusions Despite distinct glucose-lowering properties, the tested insulin preparations had similar effects on IGF-I concentration and IGF bioactivity, IGFBP-2

  6. The human insulin gene displays transcriptionally active epigenetic marks in islet-derived mesenchymal precursor cells in the absence of insulin expression.

    PubMed

    Mutskov, Vesco; Raaka, Bruce M; Felsenfeld, Gary; Gershengorn, Marvin C

    2007-12-01

    Human islet-derived precursor cells (hIPCs), mesenchymal cells derived in vitro from adult pancreas, proliferate freely and do not express insulin but can be differentiated to epithelial cells that express insulin. hIPCs have been studied with the goal of obtaining large quantities of insulin-producing cells suitable for transplantation into patients suffering from type 1 diabetes. It appeared that undifferentiated hIPCs are "committed" to a pancreatic endocrine phenotype through multiple cell divisions, suggesting that epigenetic modifications at the insulin locus could be responsible. We determined patterns of histone modifications over the insulin gene in human islets and hIPCs and compared them with HeLa and human bone marrow-derived mesenchymal stem cells (hBM-MSCs), neither of which expresses insulin. The insulin gene in islets displays high levels of histone modifications (H4 hyperacetylation and dimethylation of H3 lysine 4) typical of active genes. These are not present in HeLa and hBM-MSCs, which instead have elevated levels of H3 lysine 9 dimethylation, a mark of inactive genes. hIPCs, in contrast, show significant levels of active chromatin modifications, as much as half those seen in islets, and show no measurable H3 K9 methylation. Cells expanded from a minor population of mesenchymal stromal cells found in islets exhibit the same histone modifications as established hIPCs. We conclude that hIPCs, which do not express the insulin gene, nonetheless uniquely exhibit epigenetic marks that could poise them for activation of insulin expression. This epigenetic signature may be a general mechanism whereby tissue-derived precursor cells are committed to a distinct specification. Disclosure of potential conflicts of interest is found at the end of this article.

  7. Reversal of diabetes with insulin-producing cells derived in vitro from human pluripotent stem cells.

    PubMed

    Rezania, Alireza; Bruin, Jennifer E; Arora, Payal; Rubin, Allison; Batushansky, Irina; Asadi, Ali; O'Dwyer, Shannon; Quiskamp, Nina; Mojibian, Majid; Albrecht, Tobias; Yang, Yu Hsuan Carol; Johnson, James D; Kieffer, Timothy J

    2014-11-01

    Transplantation of pancreatic progenitors or insulin-secreting cells derived from human embryonic stem cells (hESCs) has been proposed as a therapy for diabetes. We describe a seven-stage protocol that efficiently converts hESCs into insulin-producing cells. Stage (S) 7 cells expressed key markers of mature pancreatic beta cells, including MAFA, and displayed glucose-stimulated insulin secretion similar to that of human islets during static incubations in vitro. Additional characterization using single-cell imaging and dynamic glucose stimulation assays revealed similarities but also notable differences between S7 insulin-secreting cells and primary human beta cells. Nevertheless, S7 cells rapidly reversed diabetes in mice within 40 days, roughly four times faster than pancreatic progenitors. Therefore, although S7 cells are not fully equivalent to mature beta cells, their capacity for glucose-responsive insulin secretion and rapid reversal of diabetes in vivo makes them a promising alternative to pancreatic progenitor cells or cadaveric islets for the treatment of diabetes.

  8. Mitochondrial Respiratory Capacity and Content Are Normal in Young Insulin-Resistant Obese Humans

    PubMed Central

    Fisher-Wellman, Kelsey H.; Weber, Todd M.; Cathey, Brook L.; Brophy, Patricia M.; Gilliam, Laura A.A.; Kane, Constance L.; Maples, Jill M.; Gavin, Timothy P.; Houmard, Joseph A.; Neufer, P. Darrell

    2014-01-01

    Considerable debate exists about whether alterations in mitochondrial respiratory capacity and/or content play a causal role in the development of insulin resistance during obesity. The current study was undertaken to determine whether such alterations are present during the initial stages of insulin resistance in humans. Young (∼23 years) insulin-sensitive lean and insulin-resistant obese men and women were studied. Insulin resistance was confirmed through an intravenous glucose tolerance test. Measures of mitochondrial respiratory capacity and content as well as H2O2 emitting potential and the cellular redox environment were performed in permeabilized myofibers and primary myotubes prepared from vastus lateralis muscle biopsy specimens. No differences in mitochondrial respiratory function or content were observed between lean and obese subjects, despite elevations in H2O2 emission rates and reductions in cellular glutathione. These findings were apparent in permeabilized myofibers as well as in primary myotubes. The results suggest that reductions in mitochondrial respiratory capacity and content are not required for the initial manifestation of peripheral insulin resistance. PMID:23974920

  9. Cavitation-enhanced delivery of insulin in agar and porcine models of human skin.

    PubMed

    Feiszthuber, Helga; Bhatnagar, Sunali; Gyöngy, Miklós; Coussios, Constantin-C

    2015-03-21

    Ultrasound-assisted transdermal insulin delivery offers a less painful and less invasive alternative to subcutaneous insulin injections. However, ultrasound-based drug delivery, otherwise known as sonophoresis, is a highly variable phenomenon, in part dependent on cavitation. The aim of the current work is to investigate the role of cavitation in transdermal insulin delivery. Fluorescently stained, soluble Actrapid insulin was placed on the surface of human skin-mimicking materials subjected to 265 kHz, 10% duty cycle focused ultrasound. A confocally and coaxially aligned 5 MHz broadband ultrasound transducer was used to detect cavitation. Two different skin models were used. The first model, 3% agar hydrogel, was insonated with a range of pressures (0.25-1.40 MPa peak rarefactional focal pressure-PRFP), with and without cavitation nuclei embedded within the agar at a concentration of 0.05% w/v. The second, porcine skin was insonated at 1.00 and 1.40 MPa PRFP. In both models, fluorescence measurements were used to determine penetration depth and concentration of delivered insulin. Results show that in agar gel, both insulin penetration depth and concentration only increased significantly in the presence of inertial cavitation, with up to a 40% enhancement. In porcine skin the amount of fluorescent insulin was higher in the epidermis of those samples that were exposed to ultrasound compared to the control samples, but there was no significant increase in penetration distance. The results underline the importance of instigating and monitoring inertial cavitation during transdermal insulin delivery. PMID:25716689

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

    SciTech Connect

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

    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.

  11. Cavitation-enhanced delivery of insulin in agar and porcine models of human skin

    NASA Astrophysics Data System (ADS)

    Feiszthuber, Helga; Bhatnagar, Sunali; Gyöngy, Miklós; Coussios, Constantin-C.

    2015-03-01

    Ultrasound-assisted transdermal insulin delivery offers a less painful and less invasive alternative to subcutaneous insulin injections. However, ultrasound-based drug delivery, otherwise known as sonophoresis, is a highly variable phenomenon, in part dependent on cavitation. The aim of the current work is to investigate the role of cavitation in transdermal insulin delivery. Fluorescently stained, soluble Actrapid insulin was placed on the surface of human skin-mimicking materials subjected to 265 kHz, 10% duty cycle focused ultrasound. A confocally and coaxially aligned 5 MHz broadband ultrasound transducer was used to detect cavitation. Two different skin models were used. The first model, 3% agar hydrogel, was insonated with a range of pressures (0.25-1.40 MPa peak rarefactional focal pressure—PRFP), with and without cavitation nuclei embedded within the agar at a concentration of 0.05% w/v. The second, porcine skin was insonated at 1.00 and 1.40 MPa PRFP. In both models, fluorescence measurements were used to determine penetration depth and concentration of delivered insulin. Results show that in agar gel, both insulin penetration depth and concentration only increased significantly in the presence of inertial cavitation, with up to a 40% enhancement. In porcine skin the amount of fluorescent insulin was higher in the epidermis of those samples that were exposed to ultrasound compared to the control samples, but there was no significant increase in penetration distance. The results underline the importance of instigating and monitoring inertial cavitation during transdermal insulin delivery.

  12. Cavitation-enhanced delivery of insulin in agar and porcine models of human skin.

    PubMed

    Feiszthuber, Helga; Bhatnagar, Sunali; Gyöngy, Miklós; Coussios, Constantin-C

    2015-03-21

    Ultrasound-assisted transdermal insulin delivery offers a less painful and less invasive alternative to subcutaneous insulin injections. However, ultrasound-based drug delivery, otherwise known as sonophoresis, is a highly variable phenomenon, in part dependent on cavitation. The aim of the current work is to investigate the role of cavitation in transdermal insulin delivery. Fluorescently stained, soluble Actrapid insulin was placed on the surface of human skin-mimicking materials subjected to 265 kHz, 10% duty cycle focused ultrasound. A confocally and coaxially aligned 5 MHz broadband ultrasound transducer was used to detect cavitation. Two different skin models were used. The first model, 3% agar hydrogel, was insonated with a range of pressures (0.25-1.40 MPa peak rarefactional focal pressure-PRFP), with and without cavitation nuclei embedded within the agar at a concentration of 0.05% w/v. The second, porcine skin was insonated at 1.00 and 1.40 MPa PRFP. In both models, fluorescence measurements were used to determine penetration depth and concentration of delivered insulin. Results show that in agar gel, both insulin penetration depth and concentration only increased significantly in the presence of inertial cavitation, with up to a 40% enhancement. In porcine skin the amount of fluorescent insulin was higher in the epidermis of those samples that were exposed to ultrasound compared to the control samples, but there was no significant increase in penetration distance. The results underline the importance of instigating and monitoring inertial cavitation during transdermal insulin delivery.

  13. Computational model of cellular metabolic dynamics: effect of insulin on glucose disposal in human skeletal muscle

    PubMed Central

    Li, Yanjun; Solomon, Thomas P. J.; Haus, Jacob M.; Saidel, Gerald M.; Cabrera, Marco E.

    2010-01-01

    Identifying the mechanisms by which insulin regulates glucose metabolism in skeletal muscle is critical to understanding the etiology of insulin resistance and type 2 diabetes. Our knowledge of these mechanisms is limited by the difficulty of obtaining in vivo intracellular data. To quantitatively distinguish significant transport and metabolic mechanisms from limited experimental data, we developed a physiologically based, multiscale mathematical model of cellular metabolic dynamics in skeletal muscle. The model describes mass transport and metabolic processes including distinctive processes of the cytosol and mitochondria. The model simulated skeletal muscle metabolic responses to insulin corresponding to human hyperinsulinemic-euglycemic clamp studies. Insulin-mediated rate of glucose disposal was the primary model input. For model validation, simulations were compared with experimental data: intracellular metabolite concentrations and patterns of glucose disposal. Model variations were simulated to investigate three alternative mechanisms to explain insulin enhancements: Model 1 (M.1), simple mass action; M.2, insulin-mediated activation of key metabolic enzymes (i.e., hexokinase, glycogen synthase, pyruvate dehydrogenase); or M.3, parallel activation by a phenomenological insulin-mediated intracellular signal that modifies reaction rate coefficients. These simulations indicated that models M.1 and M.2 were not sufficient to explain the experimentally measured metabolic responses. However, by application of mechanism M.3, the model predicts metabolite concentration changes and glucose partitioning patterns consistent with experimental data. The reaction rate fluxes quantified by this detailed model of insulin/glucose metabolism provide information that can be used to evaluate the development of type 2 diabetes. PMID:20332360

  14. Effects of Insulin and High Glucose on Human Meibomian Gland Epithelial Cells

    PubMed Central

    Ding, Juan; Liu, Yang; Sullivan, David A.

    2015-01-01

    Purpose Type 2 diabetes is a risk factor for meibomian gland dysfunction (MGD). We hypothesize that this diabetic impact is due, at least in part, to the effects of insulin resistance/deficiency and hyperglycemia on human meibomian gland epithelial cells (HMGECs). To begin to test this hypothesis, we examined whether insulin and high glucose influence immortalized (I) HMGECs. Methods Immortalized HMGECs were cultured in serum-containing or -free media and treated with insulin, insulin-like growth factor–1 (IGF-1), IGF-1 receptor (R) blocking antibody, and glucose or mannitol for varying time periods. Specific proteins were detected by Western blots, cell proliferation was evaluated by manual cell counting and lipids were assessed with LipidTOX and high performance thin layer chromatography. Results We found that insulin induces a dose-dependent increase in phosphatidylinositide 3-kinase/Akt (AKT) signaling in IHMGECs. This effect involves the IGF-1R, but not the insulin receptor (IR), and is associated with a stimulation of cell proliferation and neutral lipid accumulation. In contrast, high glucose exposure alters cell morphology, causes a progressive cell loss, and significantly reduces the levels of IGF-1R, phospho (p)-AKT, Foxhead box protein O1 (FOXO1), and sterol-regulatory element binding protein (SREBP-1) in IHMGECs. Conclusions Our data show that insulin stimulates, and that high glucose is toxic for, IHMGECs. These results support our hypothesis that insulin resistance/deficiency and hyperglycemia are deleterious for HMGECs and may help explain why type II diabetes is a risk factor for MGD. PMID:26658502

  15. Is Dynamic Autocrine Insulin Signaling Possible? A Mathematical Model Predicts Picomolar Concentrations of Extracellular Monomeric Insulin within Human Pancreatic Islets

    PubMed Central

    Wang, Minghu; Li, Jiaxu; Lim, Gareth E.; Johnson, James D.

    2013-01-01

    Insulin signaling is essential for -cell survival and proliferation in vivo. Insulin also has potent mitogenic and anti-apoptotic actions on cultured -cells, with maximum effect in the high picomolar range and diminishing effect at high nanomolar doses. In order to understand whether these effects of insulin are constitutive or can be subjected to physiological modulation, it is essential to estimate the extracellular concentration of monomeric insulin within an intact islet. Unfortunately, the in vivo concentration of insulin monomers within the islet cannot be measured directly with current technology. Here, we present the first mathematical model designed to estimate the levels of monomeric insulin within the islet extracellular space. Insulin is released as insoluble crystals that exhibit a delayed dissociation into hexamers, dimers, and eventually monomers, which only then can act as signaling ligands. The rates at which different forms of insulin dissolve in vivo have been estimated from studies of peripheral insulin injection sites. We used this and other information to formulate a mathematical model to estimate the local insulin concentration within a single islet as a function of glucose. Model parameters were estimated from existing literature. Components of the model were validated using experimental data, if available. Model analysis predicted that the majority of monomeric insulin in the islet is that which has been returned from the periphery, and the concentration of intra-islet monomeric insulin varies from 50–300 pM when glucose is in the physiological range. Thus, our results suggest that the local concentration of monomeric insulin within the islet is in the picomolar ‘sweet spot’ range of insulin doses that activate the insulin receptor and have the most potent effects on -cells in vitro. Together with experimental data, these estimations support the concept that autocrine/paracrine insulin signalling within the islet is dynamic, rather

  16. The human insulin receptor substrate-1 gene (IRS1) is localized on 2q36

    SciTech Connect

    Nishiyama, Masaki; Matsufuji, Senya; Hayashi, Shin-ichi; Furusaka, Akihiro; Tanaka, Teruji ); Inazawa, J.; Nakamura, Yusuke ); Ariyama, Takeshi ); Wands, J.R. )

    1994-03-01

    The chromosomal localization of some of the genes participating in the insulin signaling pathway is known. The insulin and insulin receptor genes have been mapped to chromosomes 11 and 19, respectively. To identify the chromosomal localization of the human IRS1 gene, the fluorescence in situ hybridization technique was employed with Genomic Clone B-10. A total of 50 metaphase cells exhibiting either single or double spots of hybridization signals were examined. Among them, 32 showed the specific signals on 2q36. Therefore, the authors assigned the human IRS1 gene to 2q36. The genes for homeobox sequence (HOX4), fibronectin 1, alkaline phosphatase (intestinal), transition protein 1, villin 1, collagen (type IV), Waardenburg syndrome (type 1), alanine-glyoxylate aminotransferase, and glucagon have been localized in the vicinity of the IRS1 gene.

  17. TWEAK prevents TNF-α-induced insulin resistance through PP2A activation in human adipocytes.

    PubMed

    Vázquez-Carballo, Ana; Ceperuelo-Mallafré, Victòria; Chacón, Matilde R; Maymó-Masip, Elsa; Lorenzo, Margarita; Porras, Almudena; Vendrell, Joan; Fernández-Veledo, Sonia

    2013-07-01

    Visceral fat is strongly associated with insulin resistance. Obesity-associated adipose tissue inflammation and inflammatory cytokine production are considered key mediators of insulin signaling inhibition. TWEAK is a relatively new member of the TNF cytokine superfamily, which can exist as full length membrane-associated (mTWEAK) and soluble (sTWEAK) isoforms. Although TWEAK has been shown to have important functions in chronic inflammatory diseases its physiological role in adipose tissue remains unresolved. In this study, we explore the molecular mechanisms involved in the modulation of TNF-α-induced effects on insulin sensitivity by sTWEAK in a human visceral adipose cell line and also in primary human adipocytes obtained from visceral fat depots. Our data reveal that sTWEAK ameliorates TNF-α-induced insulin resistance on glucose uptake, GLUT4 translocation and insulin signaling without affecting other metabolic effects of TNF-α such as lipolysis or apoptotis. Co-immunoprecipitation experiments in adipose cells revealed that pretreatment with sTWEAK specifically inhibits TRAF2 association with TNFR1, but not with TNFR2, which mediates insulin resistance. However, sTWEAK does not affect other downstream molecules activated by TNF-α, such as TAK1. Rather, sTWEAK abolishes the stimulatory effect of TNF-α on JNK1/2, which is directly involved in the development of insulin resistance. This is associated with an increase in PP2A activity upon sTWEAK treatment. Silencing of the PP2A catalytic subunit gene overcomes the dephosphorylation effect of sTWEAK on JNK1/2, pointing to PP2A as a relevant mediator of sTWEAK-induced JNK inactivation. Overall, our data reveal a protective role of TWEAK in glucose homeostasis and identify PP2A as a new driver in the modulation of TNF-α signaling by sTWEAK.

  18. Synergistic effects of C-peptide and insulin on low O2-induced ATP release from human erythrocytes

    PubMed Central

    Stephenson, Alan H.; Ellsworth, Mary L.; Sprague, Randy S.

    2013-01-01

    Erythrocytes participate in the matching of oxygen (O2) delivery with local need in skeletal muscle via the release of O2 and the vasodilator, ATP. It was reported that a concentration of insulin found in humans with insulin resistance inhibits low O2-induced ATP release. However, in vivo, insulin is coreleased with connecting peptide (C-peptide) at equimolar concentrations, but because of the shorter insulin half-life, the peptides circulate at ratios of C-peptide to insulin ranging from 1:1 to 6:1. Here, we investigate the hypothesis that C-peptide and insulin work synergistically to maintain low O2-induced ATP release from human erythrocytes. Using a thin-film tonometer to alter O2 tension, we determined that either C-peptide or insulin alone inhibits low O2-induced ATP release in a concentration-dependent manner; however, coadministration of the peptides at a 1:1 ratio does not (n = 5; P < 0.05). Because this ratio of C-peptide to insulin is not present in vivo for extended periods, we also investigated the effect of additional physiological ratios on ATP release. In the presence of insulin concentrations that would be found in fasting humans (0.05 nM), C-peptide to insulin ratios of 4:1 and 6:1 did not adversely affect low O2-induced ATP release. However, at a concentration of insulin found in the peripheral circulation of humans under postprandial conditions (0.5 nM), a ratio of C-peptide to insulin of 6:1 inhibited low O2-induced ATP release (n = 5). These findings demonstrate a heretofore unrecognized synergism between C-peptide and insulin that could have physiological importance in the regulation of perfusion distribution in skeletal muscle. PMID:24089376

  19. Small molecules induce efficient differentiation into insulin-producing cells from human induced pluripotent stem cells.

    PubMed

    Kunisada, Yuya; Tsubooka-Yamazoe, Noriko; Shoji, Masanobu; Hosoya, Masaki

    2012-03-01

    Human induced pluripotent stem (hiPS) cells have potential uses for drug discovery and cell therapy, including generation of pancreatic β-cells for diabetes research and treatment. In this study, we developed a simple protocol for generating insulin-producing cells from hiPS cells. Treatment with activin A and a GSK3β inhibitor enhanced efficient endodermal differentiation, and then combined treatment with retinoic acid, a bone morphogenic protein inhibitor, and a transforming growth factor-β (TGF-β) inhibitor induced efficient differentiation of pancreatic progenitor cells from definitive endoderm. Expression of the pancreatic progenitor markers PDX1 and NGN3 was significantly increased at this step and most cells were positive for anti-PDX1 antibody. Moreover, several compounds, including forskolin, dexamethasone, and a TGF-β inhibitor, were found to induce the differentiation of insulin-producing cells from pancreatic progenitor cells. By combined treatment with these compounds, more than 10% of the cells became insulin positive. The differentiated cells secreted human c-peptide in response to various insulin secretagogues. In addition, all five hiPS cell lines that we examined showed efficient differentiation into insulin-producing cells with this protocol.

  20. A sensitive chemiluminescent enzyme immunoassay for the bioanalysis of carboxyl-terminal B-chain analogues of human insulin.

    PubMed

    Cao, Y; Smith, W C; Bowsher, R R

    2001-08-01

    Quantification of analogues of human insulin in biological matrices is complicated by differences in their immunoreactivity and the presence of both the analogue and endogenous concentrations of insulin in test samples. To facilitate pharmacokinetic comparisons of carboxyl-terminal B-chain analogues of human insulin, we undertook development of a sensitive ELISA. The ELISA detection method was optimized systematically to permit routine analysis of 10-microl serum samples. Accordingly, a noncompetitive 'sandwich' chemiluminescent ELISA was validated for the quantification of carboxyl-terminal B-chain insulin analogues in human serum over a concentration range from 5 to 3125 pM. The mean bias (RE%) within the validated range varied from -10.3 to 4.3%, with an intermediate precision (inter-assay CV%) from 4.2 to 11.5%. The two-sided 90% expectation tolerance interval for total measurement error was within +/-25% of the nominal concentration for all levels of validation samples. Insulin lispro, human insulin, proinsulin, despentapeptide insulin (DPI) and porcine insulin displayed comparable crossreactivity in the ELISA. Potential utility of the new assay for insulin bioanalysis in nonhuman species was investigated by assessing the pharmacokinetic profile of DPI in rats following administration of a single subcutaneous dose. The sensitive chemiluminescent detection method is simple to perform and should be readily adaptable for ELISAs of other therapeutic proteins.

  1. Human chorionic somatommamotropin (HCS) and pregnancy. Its relation with insulin.

    PubMed

    Prieto Villapun, J C; Cifuentes de Castro, I; Serrano Rios, M

    1976-01-01

    Plasma HCS levels have been measured in normal and pathological pregnant women. In the normal group HCS levels increased from 6--8 weeks till 33-34 weeks and then felt significantly. HCS pattern in prediabetic and chemical diabetic pregnant women was similar to the normal group. However HCS levels in chemical diabetics were significantly higher during the first two trimesters. HCS levels increased in twin pregnancy, diminished in cases of eclampsia, hypertension, fetal growth retardation, mole and blighted ovum, and disappeared after intrauterine death. Nothing could be deduced from the obese and Rh-isoimmunization groups. It is confirmed the value of HCS determination as an index of placental maturation. Also, insulin/HCS ratio may be of some aid in the study of carbohydrate intolerance in pregnancy.

  2. Human insulin polymorphism upon ligand binding and pH variation: the case of 4-ethylresorcinol

    PubMed Central

    Fili, S.; Valmas, A.; Norrman, M.; Schluckebier, G.; Beckers, D.; Degen, T.; Wright, J.; Fitch, A.; Gozzo, F.; Giannopoulou, A. E.; Karavassili, F.; Margiolaki, I.

    2015-01-01

    This study focuses on the effects of the organic ligand 4-ethylresorcinol on the crystal structure of human insulin using powder X-ray crystallography. For this purpose, systematic crystallization experiments have been conducted in the presence of the organic ligand and zinc ions within the pH range 4.50–8.20, while observing crystallization behaviour around the isoelectric point of insulin. High-throughput crystal screening was performed using a laboratory X-ray diffraction system. The most representative samples were selected for synchrotron X-ray diffraction measurements, which took place at the European Synchrotron Radiation Facility (ESRF) and the Swiss Light Source (SLS). Four different crystalline polymorphs have been identified. Among these, two new phases with monoclinic symmetry have been found, which are targets for the future development of microcrystalline insulin drugs. PMID:26306195

  3. Human insulin/IGF-1 and familial longevity at middle age

    PubMed Central

    Rozing, Maarten P.; Westendorp, Rudi G.J.; Frölich, Marijke; de Craen, Anton J.M.; Beekman, Marian; Heijmans, Bastiaan T.; Mooijaart, Simon P.; Blauw, Gerard-Jan; Slagboom, P. Eline; van Heemst, Diana; Group, on behalf of the Leiden Longevity Study (LLS)

    2009-01-01

    Recently, we have shown that compared to controls, long-lived familial nonagenarians (mean age: 93.4 years) from the Leiden Longevity Study displayed a lower mortality rate, and their middle-aged offspring displayed a lower prevalence of cardio-metabolic diseases, including diabetes mellitus. The evolutionarily conserved insulin/IGF-1 signaling (IIS) pathway has been implicated in longevity in model organisms, but its relevance for human longevity has generated much controversy. Here, we show that compared to their partners, the offspring of familial nonagenarians displayed similar non-fasted serum levels of IGF-1, IGFBP3 and insulin but lower non-fasted serum levels of glucose, indicating that familial longevity is associated with differences in insulin sensitivity. PMID:20157552

  4. Endodermal differentiation of human pluripotent stem cells to insulin-producing cells in 3D culture.

    PubMed

    Takeuchi, Hiroki; Nakatsuji, Norio; Suemori, Hirofumi

    2014-03-27

    Insulin-producing cells (IPCs) derived from human pluripotent stem cells (hPSCs) may be useful in cell therapy and drug discovery for diabetes. Here, we examined various growth factors and small molecules including those previously reported to develop a robust differentiation method for induction of mature IPCs from hPSCs. We established a protocol that induced PDX1-positive pancreatic progenitor cells at high efficiency, and further induced mature IPCs by treatment with forskolin, dexamethasone, Alk5 inhibitor II and nicotinamide in 3D culture. The cells that differentiated into INSULIN-positive and C-PEPTIDE-positive cells secreted insulin in response to glucose stimulation, indicating a functional IPC phenotype. We also found that this method was applicable to different types of hPSCs.

  5. Effect of phorbol and glucose on insulin secretion from the human fetal pancreas.

    PubMed

    Tuch, B E; Williams, P F; Handelsman, D; Dunlop, M; Grigoriou, S; Turtle, J R

    1987-04-01

    It has been reported previously that 12-0-tetradecanoylphorbol-13-acetate is capable of stimulating the release of insulin from adult and neonatal pancreatic tissue. The data from this study show that this agent at a concentration of 1.3 uM, in the presence of 2.8 mM glucose, was unable to cause significant secretion of insulin from cultured human fetal pancreatic explants. By contrast 20 mM glucose was able to cause a small but significant immediate increase in secretion of insulin, but was unable to maintain this response beyond ten minutes. When the two agents were combined, a synergistic effect was seen throughout the entire 50 minute period of stimulation. The reason for this synergism is unclear since, whilst both secretagogues were able to cause a rise in the levels of diacylglycerol, together no extra effect was observed.

  6. The human insulin gene linked polymorphic region exhibits an altered DNA structure.

    PubMed Central

    Hammond-Kosack, M C; Dobrinski, B; Lurz, R; Docherty, K; Kilpatrick, M W

    1992-01-01

    Regulation of transcription of the human insulin gene appears to involve a series of DNA sequences in the 5' region. Hypersensitivity to DNA structural probes has previously been demonstrated in regulatory regions of cloned genomic DNA fragments, and been correlated with gene activity. To investigate the structure of the DNA in the human insulin gene, bromoacetaldehyde and S1 nuclease were reacted with a supercoiled plasmid containing a 5kb genomic insulin fragment. Both probes revealed the human insulin gene linked polymorphic region (ILPR), a region (-363) upstream of the transcriptional start site which contains multiple repeats of a 14-15mer oligonucleotide with the consensus sequence ACAGGGGT(G/C)(T/C)GGGG, as the major hypersensitive site. Fine mapping and electron microscopic analysis both show a very different behaviour of the two DNA strands in the region of the ILPR and suggest the G-rich strand may be adopting a highly structured conformation with the complementary strand remaining largely single stranded. Images PMID:1741248

  7. RFX6 regulates insulin secretion by modulating Ca2+ homeostasis in human β cells.

    PubMed

    Chandra, Vikash; Albagli-Curiel, Olivier; Hastoy, Benoit; Piccand, Julie; Randriamampita, Clotilde; Vaillant, Emmanuel; Cavé, Hélène; Busiah, Kanetee; Froguel, Philippe; Vaxillaire, Martine; Rorsman, Patrik; Polak, Michel; Scharfmann, Raphael

    2014-12-24

    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 Ca(2+)-channel genes resulting in the reduction in L-type Ca(2+)-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 Ca(2+)-channel expression. Our data therefore provide insights for understanding certain forms of neonatal diabetes. PMID:25497100

  8. Insulin enhances the gain of arterial baroreflex control of muscle sympathetic nerve activity in humans.

    PubMed

    Young, Colin N; Deo, Shekhar H; Chaudhary, Kunal; Thyfault, John P; Fadel, Paul J

    2010-09-15

    Recent animal studies indicate that insulin increases arterial baroreflex control of lumbar sympathetic nerve activity; however, the extent to which these findings can be extrapolated to humans is unknown. To begin to address this, muscle sympathetic nerve activity (MSNA) and arterial blood pressure were measured in 19 healthy subjects (27 ± 1 years) before, and for 120 min following, two common methodologies used to evoke sustained increases in plasma insulin: a mixed meal and a hyperinsulinaemic euglycaemic clamp. Weighted linear regression analysis between MSNA and diastolic blood pressure was used to determine the gain (i.e. sensitivity) of arterial baroreflex control of MSNA. Plasma insulin was significantly elevated within 30 min following meal intake (34 ± 6 uIU ml(1); P < 0.05) and remained above baseline for up to 120 min. Similarly, after meal intake, arterial baroreflex-MSNA gain for burst incidence and total MSNA was increased and remained elevated for the duration of the protocol (e.g. burst incidence gain: 3.29 ± 0.54 baseline vs. 5.64 ± 0.67 bursts (100 heart beats)(1) mmHg(1) at 120 min; P < 0.05). During the hyperinsulinaemic euglycaemic clamp, in which insulin was elevated to postprandial concentrations (42 ± 6 μIU ml(1); P < 0.05), while glucose was maintained constant, arterial baroreflex-MSNA gain was similarly enhanced (e.g. burst incidence gain: 2.44 ± 0.29 baseline vs. 4.74 ± 0.71 bursts (100 heart beats)(1) mmHg(1) at 120 min; P < 0.05). Importantly, during time control experiments, with sustained fasting insulin concentrations, the arterial baroreflex-MSNA gain remained unchanged. These findings demonstrate, for the first time in healthy humans, that increases in plasma insulin enhance the gain of arterial baroreflex control of MSNA.

  9. Generation of insulin-producing islet-like clusters from human embryonic stem cells.

    PubMed

    Jiang, Jianjie; Au, Melinda; Lu, Kuanghui; Eshpeter, Alana; Korbutt, Gregory; Fisk, Greg; Majumdar, Anish S

    2007-08-01

    Recent success in pancreatic islet transplantation has energized the field to discover an alternative source of stem cells with differentiation potential to beta cells. Generation of glucose-responsive, insulin-producing beta cells from self-renewing, pluripotent human ESCs (hESCs) has immense potential for diabetes treatment. We report here the development of a novel serum-free protocol to generate insulin-producing islet-like clusters (ILCs) from hESCs grown under feeder-free conditions. In this 36-day protocol, hESCs were treated with sodium butyrate and activin A to generate definitive endoderm coexpressing CXCR4 and Sox17, and CXCR4 and Foxa2. The endoderm population was then converted into cellular aggregates and further differentiated to Pdx1-expressing pancreatic endoderm in the presence of epidermal growth factor, basic fibroblast growth factor, and noggin. Soon thereafter, expression of Ptf1a and Ngn3 was detected, indicative of further pancreatic differentiation. The aggregates were finally matured in the presence of insulin-like growth factor II and nicotinamide. The temporal pattern of pancreas-specific gene expression in the hESC-derived ILCs showed considerable similarity to in vivo pancreas development, and the final population contained representatives of the ductal, exocrine, and endocrine pancreas. The hESC-derived ILCs contained 2%-8% human C-peptide-positive cells, as well as glucagon- and somatostatin-positive cells. Insulin content as high as 70 ng of insulin/mug of DNA was measured in the ILCs, representing levels higher than that of human fetal islets. In addition, the hESC-derived ILCs contained numerous secretory granules, as determined by electron microscopy, and secreted human C-peptide in a glucose-dependent manner. Disclosure of potential conflicts of interest is found at the end of this article.

  10. The Effects of Insulin Resistance on Individual Tissues: An Application of a Mathematical Model of Metabolism in Humans.

    PubMed

    Pearson, Taliesin; Wattis, Jonathan A D; King, John R; MacDonald, Ian A; Mazzatti, Dawn J

    2016-06-01

    Whilst the human body expends energy constantly, the human diet consists of a mix of carbohydrates and fats delivered in a discontinuous manner. To deal with this sporadic supply of energy, there are transport, storage and utilisation mechanisms, for both carbohydrates and fats, around all tissues of the body. Insulin-resistant states such as type 2 diabetes and obesity are characterised by reduced efficiency of these mechanisms. Exactly how these insulin-resistant states develop, for example whether there is an order in which tissues become insulin resistant, is an active area of research with the hope of gaining a better overall understanding of insulin resistance. In this paper, we use a previously derived system of 12 first-order coupled differential equations that describe the transport between, and storage in, different tissues of the human body. We briefly revisit the derivation of the model before parametrising the model to account for insulin resistance. We then solve the model numerically, separately simulating each individual tissue as insulin resistant, and discuss and compare these results, drawing three main conclusions. The implications of these results are in accordance with biological intuition. First, insulin resistance in a tissue creates a knock-on effect on the other tissues in the body, whereby they attempt to compensate for the reduced efficiency of the insulin-resistant tissue. Second, insulin resistance causes a fatty liver, and the insulin resistance of tissues other than the liver can cause fat to accumulate in the liver. Finally, although insulin resistance in individual tissues can cause slightly reduced skeletal muscle metabolic flexibility, it is when the whole body is insulin resistant that the biggest effect on skeletal muscle flexibility is seen. PMID:27306890

  11. Protein-tyrosine phosphatase activity in human adipocytes is strongly correlated with insulin-stimulated glucose uptake and is a target of insulin-induced oxidative inhibition.

    PubMed

    Wu, Xiangdong; Hardy, V Elise; Joseph, Jeffrey I; Jabbour, Serge; Mahadev, Kalyankar; Zhu, Li; Goldstein, Barry J

    2003-06-01

    Protein-tyrosine phosphatases (PTPases), in particular PTP1B, have been shown to modulate insulin signal transduction in liver and skeletal muscle in animal models; however, their role in human adipose tissue remains unclear. The uptake of (14)C-D-glucose in response to 10 or 100 nmol/L insulin was measured in isolated subcutaneous adipocytes from subjects with a mean age of 44 years (range, 26 to 58) and mean body mass index (BMI) of 35.6 (range, 29.7 to 45.5). The endogenous activity of total PTPases and specifically of PTP1B in immunoprecipitates was measured in cell lysates under an inert atmosphere with and without added reducing agents. Using nonlinear regression analysis, higher BMI was significantly correlated with lower adipocyte glucose uptake (r = 0.73, P =.01) and with increased endogenous total PTPase activity (r = 0.64, P =.04). Correlation with waist circumference gave similar results. The endogenous total PTPase activity also strongly correlated with insulin-stimulated glucose uptake (R =.89, P <.0001); however, the activity of PTP1B was unrelated to the level of glucose uptake. Consistent with the insulin-stimulated oxidative inhibition of thiol-dependent PTPases reported for 3T3-L1 adipocytes and hepatoma cells, treatment of human adipocytes with 100 nmol/L insulin for 5 minutes lowered endogenous PTPase activity to 37% of control (P <.001), which was increased 25% by subsequent treatment with dithiothreitol in vitro. Cellular treatment with diphenyleneiodonium (DPI), an NADPH oxidase inhibitor that blocks the cellular generation of H(2)O(2) and reduces the insulin-induced reduction of cellular PTPase activity, also diminished insulin-stimulated glucose uptake by 82% (P =.001). These data suggest that total cellular PTPase activity, but not the activity of PTP1B, is higher in more obese subjects and is negatively associated with insulin-stimulated glucose transport. The insulin-stimulated oxidative inhibition of PTPases may also have an important

  12. Statistics of premixed flame cells

    NASA Technical Reports Server (NTRS)

    Noever, David A.

    1991-01-01

    The statistics of random cellular patterns in premixed flames are analyzed. Agreement is found with a variety of topological relations previously found for other networks, namely, Lewis's law and Aboav's law. Despite the diverse underlying physics, flame cells are shown to share a broad class of geometric properties with other random networks-metal grains, soap foams, bioconvection, and Langmuir monolayers.

  13. Premixed Prevaporized Combustor Technology Forum

    NASA Technical Reports Server (NTRS)

    1979-01-01

    The Forum was held to present the results of recent and current work intended to provide basic information required for demonstration of lean, premixed prevaporized combustors for aircraft gas turbine engine application. Papers are presented which deal with the following major topics: (1) engine interfaces; (2) fuel-air preparation; (3) autoignition; (4) lean combustion; and (5) concept design studies.

  14. Statistics of premixed flame cells

    SciTech Connect

    Noever, D.A. )

    1991-07-15

    The statistics of random cellular patterns in premixed flames are analyzed. Agreement is found with a variety of topological relations previously found for other networks, namely, Lewis's law and Aboav's law. Despite the diverse underlying physics, flame cells are shown to share a broad class of geometric properties with other random networks---metal grains, soap foams, bioconvection, and Langmuir monolayers.

  15. GLUCOSE METABOLISM IN PIGS EXPRESSING HUMAN GENES UNDER AN INSULIN PROMOTER

    PubMed Central

    Wijkstrom, M.; Bottino, R.; Iwase, H.; Hara, H.; Ekser, B.; van der Windt, D.J.; Long, C.; Toledo, F.; Phelps, C.; Trucco, M.; Cooper, D.K.C.; Ayares, D.

    2014-01-01

    Background Xenotransplantation of porcine islets can reverse diabetes in nonhuman primates. The remaining hurdles for clinical application include safe and effective T-cell directed immunosuppression, but protection against the innate immune system and coagulation dysfunction may be more difficult to achieve. Islet-targeted genetic manipulation of islet-source pigs represents a powerful tool to protect against graft loss. However, whether these genetic alterations would impair islet function is unknown. Methods On a background of α1,3-galactosyltransferase gene-knockout (GTKO)/human (h) CD46, additional genes (hCD39, human tissue factor pathway inhibitor, porcine CTLA4-Ig) were inserted in different combinations under an insulin promoter to promote expression in islets (confirmed by immunofluorescence). Seven pigs were tested for baseline and glucose/arginine-challenged levels of glucose, insulin, C-peptide, and glucagon. Results This preliminary study did not show definite evidence of β-cell deficiencies, even when 3 transgenes were expressed under the insulin promoter. Of 7 animals, all were normoglycemic at fasting, and 5 of 7 had normal glucose disposal rates after challenge. All animals exhibited insulin, C-peptide, and glucagon responses to both glucose and arginine challenge; however, significant interindividual variation was observed. Conclusions Multiple islet-targeted transgenic expression was not associated with an overtly detrimental effect on islet function, suggesting that complex genetic constructs designed for islet protection warrants further testing in islet xenotransplantation models. PMID:25382150

  16. Analysis of the regions flanking the human insulin gene and sequence of an Alu family member.

    PubMed Central

    Bell, G I; Pictet, R; Rutter, W J

    1980-01-01

    The regions around the human insulin gene have been studied by heteroduplex, hybridization and sequence analysis. These studies indicated that there is a region of heterogeneous length located approximately 700 bp before the 5' end of the gene; and that the 19 kb of cloned DNA which includes the 1430 bp insulin gene as well as 5650 bp before and 11,500 bp after the gene is single copy sequence except for 500 bp located 6000 bp from the 3' end of the gene. This 500 bp segment contains a member of the Alu family of dispersed middle repetitive sequences as well as another less highly repeated homopolymeric segment. The sequence of this region was determined. This Alu repeat is bordered by 19 bp direct repeats and also contains an 83 bp sequence which is present twice. The regions flanking the human and rat I insulin genes were compared by heteroduplex analysis to localize homologous sequences in the flanking regions which could be involved in the regulation of insulin biosynthesis. The homology between the two genes is restricted to the region encoding preproinsulin and a short region of approximately 60 bp flanking the 5' side of the genes. Images PMID:6253909

  17. Distribution of insulin mRNA transcripts within the human body.

    PubMed

    Bell, Glenn D; Reddy, Shiva; Sun, Xueying; Yang, Yi; Krissansen, Geoffrey W

    2014-08-29

    Here we sought evidence for the existence of insulin mRNA-producing cells outside the human pancreas. Commercially available complementary DNA (cDNA) arrays prepared from 72 different types of adult human tissues were screened by PCR for transcripts encoding insulin, and other classic pancreatic hormones. Insulin mRNA transcripts were detected by standard PCR in the pancreas, stomach, pylorus region of the stomach, and the duodenum; and additionally by nested PCR in the jejunum, ileum and cecum, but not in other body tissues including the brain and colon. Most of these tissues also variably expressed mRNA transcripts for amylase α2B, amylin, glucagon, somatostatin, and pancreatic polypeptide. In summary, using sensitive PCR methods we have provided evidence for the presence of rare insulin mRNA-expressing cells within the stomach, small intestine, and cecum. Their role at these sites may be to support classical enteroendocrine cells as sentinels to sense and monitor gastric contents passing into and through the bowel.

  18. Omega-3 Fatty Acids Reduce Adipose Tissue Macrophages in Human Subjects With Insulin Resistance

    PubMed Central

    Spencer, Michael; Finlin, Brian S.; Unal, Resat; Zhu, Beibei; Morris, Andrew J.; Shipp, Lindsey R.; Lee, Jonah; Walton, R. Grace; Adu, Akosua; Erfani, Rod; Campbell, Marilyn; McGehee, Robert E.; Peterson, Charlotte A.; Kern, Philip A.

    2013-01-01

    Fish oils (FOs) have anti-inflammatory effects and lower serum triglycerides. This study examined adipose and muscle inflammatory markers after treatment of humans with FOs and measured the effects of ω-3 fatty acids on adipocytes and macrophages in vitro. Insulin-resistant, nondiabetic subjects were treated with Omega-3-Acid Ethyl Esters (4 g/day) or placebo for 12 weeks. Plasma macrophage chemoattractant protein 1 (MCP-1) levels were reduced by FO, but the levels of other cytokines were unchanged. The adipose (but not muscle) of FO-treated subjects demonstrated a decrease in macrophages, a decrease in MCP-1, and an increase in capillaries, and subjects with the most macrophages demonstrated the greatest response to treatment. Adipose and muscle ω-3 fatty acid content increased after treatment; however, there was no change in insulin sensitivity or adiponectin. In vitro, M1-polarized macrophages expressed high levels of MCP-1. The addition of ω-3 fatty acids reduced MCP-1 expression with no effect on TNF-α. In addition, ω-3 fatty acids suppressed the upregulation of adipocyte MCP-1 that occurred when adipocytes were cocultured with macrophages. Thus, FO reduced adipose macrophages, increased capillaries, and reduced MCP-1 expression in insulin-resistant humans and in macrophages and adipocytes in vitro; however, there was no measureable effect on insulin sensitivity. PMID:23328126

  19. Detection of the single-chain precursor in the production and purification process of recombinant human insulin.

    PubMed

    Leng, Chunsheng; Li, Qingwei; Wu, Fenfang; Chen, Liyong; Su, Peng

    2013-08-01

    High quality recombinant insulin requires being free of single-chain precursor (proinsulin), a task that depends on the selectivity and sensitivity of the monitoring process for detecting proinsulin. In this study we developed an enzyme-linked immunosorbent assay (ELISA) system that was specifically tailored to detect recombinant proinsulin. The proinsulin consists of six components: an initiating methionine, 48 amino acids from human growth hormones (HGH, used as the protection peptide), first connecting Arg-residue, B-chain of insulin, and second connecting Arg-peptide and A-chain of insulin. This form of proinsulin is more stable and can be efficiently expressed by E. coli than insulin. Herein, we evaluated the specificity, precision, recovery, sensitivity, and detection range of the proinsulin ELISA kit. The results showed that the ELISA kit is a very useful tool for monitoring the proinsulin yield in early stages of insulin production as well as the residual proinsulin in the final product, insulin.

  20. Biology of insulin-like factor 3 in human reproduction.

    PubMed

    Ivell, Richard; Anand-Ivell, Ravinder

    2009-01-01

    BACKGROUND Insulin-like factor 3 (INSL3) is a neohormone that has evolved to address specific mammalian traits, in particular, the first phase of testicular descent towards the scrotum during mid-gestation. METHODS A thorough literature search was made in PubMed using the terms INSL3, as well as the older synonyms RLF and Ley-IL. RESULTS INSL3 is a major secretory product of the testicular Leydig cells in the fetus and in adult men, and in rodent models, reduction in fetal INSL3 expression is an early marker of the testicular dysgenesis syndrome. In women, it is produced in lower amounts by ovarian theca and luteal cells, and circulating levels are increased in women with polycystic ovarian syndrome. During pregnancy, there is evidence for an interaction regulating the feto-placental unit. The presence of INSL3 in amniocentesis samples taken at 12-14 weeks gestation is absolutely specific for male gender, and levels are predictive of subsequent pre-eclampsia and/or birthweight. INSL3 is also involved in adult traits, such as spermatogenesis and bone metabolism. In adult men, INSL3 is constitutively expressed and secreted into the bloodstream at a constant level, reflecting the number and/or functional capacity of the Leydig cells. In complete contrast, testosterone is highly variable within individuals, is acutely responsive to fluctuations in the hypothalamic-pituitary-gonadal axis and appears to have marginal diagnostic value. INSL3 declines consistently with age in adult men. CONCLUSIONS INSL3 promises to become an important new diagnostic tool to characterize those men with late-onset hypogonadism and to add clinical diagnostic value at amniocentesis.

  1. Acute and chronic effects of glyceryl trinitrate therapy on insulin and glucose regulation in humans.

    PubMed

    Jedrzkiewicz, Sean; Parker, John D

    2013-05-01

    This study examined the effect of acute and sustained transdermal glyceryl trinitrate (GTN) therapy on insulin and glucose regulation. Totally, 12 males (18-30 years) underwent a glucose tolerance test at baseline (visit 1), 90 minutes after acute transdermal GTN 0.6 mg/h (visit 2), following 7 days of continuous GTN (visit 3), and 2 to 3 days after stopping GTN (visit 4). At each visit, plasma glucose and insulin concentrations were measured before and 30, 60, 90, and 120 minutes after a 75-g oral glucose load. Indices of glucose metabolism that were examined included the insulin sensitivity index, the homeostasis model assessment of insulin resistance (HOMA-IR), and the insulinogenic index. The acute administration of GTN had no effect on glucose and insulin responses (visit 2). However, after 7 days of GTN exposure (visit 3) there was an increase in the mean glucose concentration measured after the oral glucose load. On visit 1, the mean glucose concentration (± standard deviation) following the 75 g oral glucose challenge was 5.7 ± 0.5 µmol/L. On visit 3, after 7 days of transdermal GTN therapy, the mean glucose concentration after the oral glucose was significantly higher; 6.2 ± 0.5 µmol/L (P < .015; 95% confidence intervals 0.25-0.77). There was also an increase in the HOMA-IR index; on visit 1, the median HOMA-IR (interquartile range) was 5.2 (3.9) versus 6.9 (6.8) on visit 3 (P < .015). Other indices of glucose metabolism did not change. These observations document that GTN therapy modifies glucose metabolism causing evidence of increased insulin resistance during sustained therapy in normal humans.

  2. Comparative 2D NMR studies of human insulin and des-pentapeptide insulin: Sequential resonance assignment and implications for protein dynamics and receptor recognition

    SciTech Connect

    Hua, Qingxin ); Weiss, M.A. Massachusetts General Hospital, Boston, MA )

    1991-06-04

    The solution structure and dynamics of human insulin are ivestigated by 2D {sup 1}H NMR spectroscopy in reference to a previously analyzed analogue, des-pentapeptide (B26-B30) insulin. This spectroscopic comparison is of interest since (i) the structure of the C-terminal region of the B-chain has not been determined in the monomeric state and (ii) the role of this region in binding to the insulin receptor has been the subject of long-standing speculation. The present NMR studies are conducted in the presence of an organic cosolvent (20% acetic acid), under which conditions both proteins are monomeric and stably folded. Complete sequential assignment of human insulin is obtained and leads to the following conclusions. (1) The secondary structure of the insulin monomer (three {alpha}-helices and B-chain {beta}-turn) is similar to that observed in the 2-Zn crustal state. (2) The folding of DPI is essentially the same as the corresponding portion of intact insulin, in accord with the similarities between their respective crystal structues. (3) residues B24-B28 adopt an extended configuration in the monomer and pack against the hydrophobic core as in crystallographic dimers; residues B29 and B30 are largely disordered. (4) The insulin fold is shown to provide a model for collective motions in a protein with implications for the mechanism of protein-protein recognition. To their knowledge, this paper describes the first detailed analysis of a protein NMR spectrum under conditions of extensive conformational broadening.

  3. Influence of obesity and insulin sensitivity on insulin signaling genes in human omental and subcutaneous adipose tissue.

    PubMed

    MacLaren, R; Cui, W; Simard, S; Cianflone, K

    2008-02-01

    Obesity and insulin resistance are independent risk factors for metabolic syndrome, diabetes, and cardiovascular disease. Adipose tissue samples from nonobese (NO), insulin-sensitive obese (ISO), and insulin-resistant obese (IRO) subjects from subcutaneous (SC) and omental (OM) adipose tissue (n = 28) were analyzed by microarray and confirmed by real-time PCR. Insulin signaling gene expression changes were greater in OM than in SC tissue and were related to insulin resistance rather than to obesity; few genes correlated with body mass index. Insulin receptor and insulin receptor substrate 1 (IRS-1) increased in the IRO versus pooled insulin-sensitive (NO+ISO) subjects. In glucose transport, PI3Kalpha and PDK2 decreased in IRO subjects, whereas PI3Kgamma, Akt2, GLUT4, and GLUT1 increased. IRS-1 regulators Jnk and IKK increased in IRO (P < 0.01 and P < 0.001 respectively). In protein synthesis, most genes examined were downregulated in IRO subjects, including mTor, Rheb, and 4EBP and eIF members (all P < 0.05). In proliferation, SHC, SOS, and Raf1 (P < 0.05) were increased, whereas Ras and MEK1/2 kinase 1 (P < 0.05) were decreased, in IRO subjects. Finally, in differentiation, PPARgamma, CEBPalpha, and CEBPbeta decreased, whereas PPARdelta, CEBPgamma, and CEBPepsilon increased, in IRO subjects (P < 0.05). Together, microarray and real-time PCR data demonstrate that insulin resistance rather than obesity is associated with altered gene expression of insulin signaling genes, especially in OM adipose tissue. PMID:17986714

  4. Proteasome Dysfunction Associated to Oxidative Stress and Proteotoxicity in Adipocytes Compromises Insulin Sensitivity in Human Obesity

    PubMed Central

    Díaz-Ruiz, Alberto; Guzmán-Ruiz, Rocío; Moreno, Natalia R.; García-Rios, Antonio; Delgado-Casado, Nieves; Membrives, Antonio; Túnez, Isaac; El Bekay, Rajaa; Fernández-Real, José M.; Tovar, Sulay; Diéguez, Carlos; Tinahones, Francisco J.; Vázquez-Martínez, Rafael; López-Miranda, José

    2015-01-01

    Abstract Aims: Obesity is characterized by a low-grade systemic inflammatory state and adipose tissue (AT) dysfunction, which predispose individuals to the development of insulin resistance (IR) and metabolic disease. However, a subset of obese individuals, referred to as metabolically healthy obese (MHO) individuals, are protected from obesity-associated metabolic abnormalities. Here, we aim at identifying molecular factors and pathways in adipocytes that are responsible for the progression from the insulin-sensitive to the insulin-resistant, metabolically unhealthy obese (MUHO) phenotype. Results: Proteomic analysis of paired samples of adipocytes from subcutaneous (SC) and omental (OM) human AT revealed that both types of cells are altered in the MUHO state. Specifically, the glutathione redox cycle and other antioxidant defense systems as well as the protein-folding machinery were dysregulated and endoplasmic reticulum stress was increased in adipocytes from IR subjects. Moreover, proteasome activity was also compromised in adipocytes of MUHO individuals, which was associated with enhanced accumulation of oxidized and ubiquitinated proteins in these cells. Proteasome activity was also impaired in adipocytes of diet-induced obese mice and in 3T3-L1 adipocytes exposed to palmitate. In line with these data, proteasome inhibition significantly impaired insulin signaling in 3T3-L1 adipocytes. Innovation: This study provides the first evidence of the occurrence of protein homeostasis deregulation in adipocytes in human obesity, which, together with oxidative damage, interferes with insulin signaling in these cells. Conclusion: Our results suggest that proteasomal dysfunction and impaired proteostasis in adipocytes, resulting from protein oxidation and/or misfolding, constitute major pathogenic mechanisms in the development of IR in obesity. Antioxid. Redox Signal. 23, 597–612. PMID:25714483

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

    SciTech Connect

    Timper, Katharina; Seboek, Dalma; Eberhardt, Michael; Linscheid, Philippe; Christ-Crain, Mirjam; Keller, Ulrich; Mueller, Beat; Zulewski, Henryk . E-mail: henryk.zulewski@unibas.ch

    2006-03-24

    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.

  6. Structure Based Discovery of Small Molecules to Regulate the Activity of Human Insulin Degrading Enzyme

    PubMed Central

    Çakir, Bilal; Dağliyan, Onur; Dağyildiz, Ezgi; Bariş, İbrahim; Kavakli, Ibrahim Halil; Kizilel, Seda; Türkay, Metin

    2012-01-01

    Background Insulin-degrading enzyme (IDE) is an allosteric Zn+2 metalloprotease involved in the degradation of many peptides including amyloid-β, and insulin that play key roles in Alzheimer's disease (AD) and type 2 diabetes mellitus (T2DM), respectively. Therefore, the use of therapeutic agents that regulate the activity of IDE would be a viable approach towards generating pharmaceutical treatments for these diseases. Crystal structure of IDE revealed that N-terminal has an exosite which is ∼30 Å away from the catalytic region and serves as a regulation site by orientation of the substrates of IDE to the catalytic site. It is possible to find small molecules that bind to the exosite of IDE and enhance its proteolytic activity towards different substrates. Methodology/Principal Findings In this study, we applied structure based drug design method combined with experimental methods to discover four novel molecules that enhance the activity of human IDE. The novel compounds, designated as D3, D4, D6, and D10 enhanced IDE mediated proteolysis of substrate V, insulin and amyloid-β, while enhanced degradation profiles were obtained towards substrate V and insulin in the presence of D10 only. Conclusion/Significance This paper describes the first examples of a computer-aided discovery of IDE regulators, showing that in vitro and in vivo activation of this important enzyme with small molecules is possible. PMID:22355395

  7. The role of adipose cell size and adipose tissue insulin sensitivity in the carbohydrate intolerance of human obesity.

    PubMed

    Salans, L B; Knittle, J L; Hirsch, J

    1968-01-01

    Glucose metabolism and insulin sensitivity of isolated human adipose tissue was studied as a function of adipose cell size and number. Glucose metabolism by these tissues was closely related to the number of cells in the fragment, irrespective of cell size. Adipose cells of obese individuals metabolized glucose to carbon dioxide and triglyceride at rates similar to adipose cells of nonobese subjects. In contrast, insulin responsiveness of adipose tissue was dependent upon adipose cell size. The larger its adipose cells the less insulin sensitive was the tissue. Thus, adipose tissue of obese subjects, with enlarged cells, showed a diminished response to insulin. After weight loss and reduction in adipose cell size, insulin sensitivity of the adipose tissue of obese patients was restored to normal. When adipose tissue of obese individuals showed impaired responsiveness to insulin, their plasma insulin levels, after oral glucose, were elevated. Weight loss and reduction in adipose cell size restored plasma insulin concentration to normal, concomitant with the return of normal tissue insulin sensitivity.

  8. Transplantation of insulin-secreting cells differentiated from human adipose tissue-derived stem cells into type 2 diabetes mice.

    PubMed

    Nam, Ji Sun; Kang, Hyun Mi; Kim, Jiyoung; Park, Seah; Kim, Haekwon; Ahn, Chul Woo; Park, Jin Oh; Kim, Kyung Rae

    2014-01-10

    Currently, there are limited ways to preserve or recover insulin secretory capacity in human pancreas. We evaluated the efficacy of cell therapy using insulin-secreting cells differentiated from human eyelid adipose tissue-derived stem cells (hEAs) into type 2 diabetes mice. After differentiating hEAs into insulin-secreting cells (hEA-ISCs) in vitro, cells were transplanted into a type 2 diabetes mouse model. Serum levels of glucose, insulin and c-peptide were measured, and changes of metabolism and inflammation were assessed in mice that received undifferentiated hEAs (UDC group), differentiated hEA-ISCs (DC group), or sham operation (sham group). Human gene expression and immunohistochemical analysis were done. DC group mice showed improved glucose level, and survival up to 60 days compared to those of UDC and sham group. Significantly increased levels of human insulin and c-peptide were detected in sera of DC mice. RT-PCR and immunohistochemical analysis showed human gene expression and the presence of human cells in kidneys of DC mice. When compared to sham mice, DC mice exhibited lower levels of IL-6, triglyceride and free fatty acids as the control mice. Transplantation of hEA-ISCs lowered blood glucose level in type 2 diabetes mice by increasing circulating insulin level, and ameliorating metabolic parameters including IL-6.

  9. Label-free detection of insulin and glucagon within human islets of Langerhans using Raman spectroscopy.

    PubMed

    Hilderink, Janneke; Otto, Cees; Slump, Cees; Lenferink, Aufried; Engelse, Marten; van Blitterswijk, Clemens; de Koning, Eelco; Karperien, Marcel; van Apeldoorn, Aart

    2013-01-01

    Intrahepatic transplantation of donor islets of Langerhans is a promising therapy for patients with type 1 diabetes. It is of critical importance to accurately monitor islet quality before transplantation, which is currently done by standard histological methods that are performed off-line and require extensive sample preparation. As an alternative, we propose Raman spectroscopy which is a non-destructive and label-free technique that allows continuous real-time monitoring of the tissue to study biological changes as they occur. By performing Raman spectroscopic measurements on purified insulin and glucagon, we showed that the 520 cm(-1) band assigned to disulfide bridges in insulin, and the 1552 cm(-1) band assigned to tryptophan in glucagon are mutually exclusive and could therefore be used as indirect markers for the label-free distinction between both hormones. High-resolution hyperspectral Raman imaging for these bands showed the distribution of disulfide bridges and tryptophan at sub-micrometer scale, which correlated with the location of insulin and glucagon as revealed by conventional immunohistochemistry. As a measure for this correlation, quantitative analysis was performed comparing the Raman images with the fluorescence images, resulting in Dice coefficients (ranging between 0 and 1) of 0.36 for insulin and 0.19 for glucagon. Although the use of separate microscope systems with different spatial resolution and the use of indirect Raman markers cause some image mismatch, our findings indicate that Raman bands for disulfide bridges and tryptophan can be used as distinctive markers for the label-free detection of insulin and glucagon in human islets of Langerhans. PMID:24167603

  10. Probiotic supplementation prevents high-fat, overfeeding-induced, insulin resistance in humans

    PubMed Central

    Hulston, Carl J.; Churnside, Amelia A.; Venables, Michelle C.

    2015-01-01

    The purpose of this study was to determine whether probiotic supplementation (Lactobacillus casei Shirota [LcS]) prevents diet-induced insulin resistance in humans. Seventeen healthy individuals were randomised to probiotic (n = 8) or control (n = 9) groups. The probiotic group consumed an LcS-fermented milk drink twice daily for 4 weeks whereas the control group received no supplementation. Subjects maintained their normal diet for the first 3 weeks of the study, after which they consumed a high-fat (65% energy) high-energy (+50% kcal) diet for 7 days. Whole body insulin sensitivity was assessed via an oral glucose tolerance test conducted before and after overfeeding. Body mass increased by 0.6 ± 0.2 kg in the control group (p < 0.05) and 0.3 ± 0.2 kg in the probiotic group (p > 0.05). Fasting plasma glucose concentrations increased following 7 days of overeating (control group only; 5.3 ± 0.1 vs. 5.6 ± 0.2 mmol/L, p < 0.05) whereas fasting serum insulin concentrations were maintained in both groups. Glucose AUC increased by 10% (from 817 ± 45 to 899 ± 39 mmol/L/120 min, p < 0.05) and whole body insulin sensitivity decreased by 27% (from 5.3 ± 1.4 to 3.9 ± 0.9, p < 0.05) in the control group, whereas normal insulin sensitivity was maintained in the probiotic group (4.4 ± 0.8 and 4.5 ± 0.9 before and after overeating, respectively, p > 0.05). These results suggest that probiotic supplementation may be useful in the prevention of diet-induced metabolic diseases such as type II diabetes. PMID:25630516

  11. Label-Free Detection of Insulin and Glucagon within Human Islets of Langerhans Using Raman Spectroscopy

    PubMed Central

    Hilderink, Janneke; Otto, Cees; Slump, Cees; Lenferink, Aufried; Engelse, Marten; van Blitterswijk, Clemens; de Koning, Eelco; Karperien, Marcel; van Apeldoorn, Aart

    2013-01-01

    Intrahepatic transplantation of donor islets of Langerhans is a promising therapy for patients with type 1 diabetes. It is of critical importance to accurately monitor islet quality before transplantation, which is currently done by standard histological methods that are performed off-line and require extensive sample preparation. As an alternative, we propose Raman spectroscopy which is a non-destructive and label-free technique that allows continuous real-time monitoring of the tissue to study biological changes as they occur. By performing Raman spectroscopic measurements on purified insulin and glucagon, we showed that the 520 cm-1 band assigned to disulfide bridges in insulin, and the 1552 cm-1 band assigned to tryptophan in glucagon are mutually exclusive and could therefore be used as indirect markers for the label-free distinction between both hormones. High-resolution hyperspectral Raman imaging for these bands showed the distribution of disulfide bridges and tryptophan at sub-micrometer scale, which correlated with the location of insulin and glucagon as revealed by conventional immunohistochemistry. As a measure for this correlation, quantitative analysis was performed comparing the Raman images with the fluorescence images, resulting in Dice coefficients (ranging between 0 and 1) of 0.36 for insulin and 0.19 for glucagon. Although the use of separate microscope systems with different spatial resolution and the use of indirect Raman markers cause some image mismatch, our findings indicate that Raman bands for disulfide bridges and tryptophan can be used as distinctive markers for the label-free detection of insulin and glucagon in human islets of Langerhans. PMID:24167603

  12. Protein crystal growth in microgravity review of large scale temperature induction method: Bovine insulin, human insulin and human α-interferon

    NASA Astrophysics Data System (ADS)

    Long, Marianna M.; Bishop, John Bradford; Delucas, Lawrence J.; Nagabhushan, Tattanhalli L.; Reichert, Paul; Smith, G. David

    1997-01-01

    The Protein Crystal Growth Facility (PCF) is space-flight hardware that accommodates large scale protein crystal growth experiments using temperature change as the inductive step. Recent modifications include specialized instrumentation for monitoring crystal nucleation with laser light scattering. This paper reviews results from its first seven flights on the Space Shuttle, the last with laser light scattering instrumentation in place. The PCF's objective is twofold: (1) the production of high quality protein crystals for x-ray analysis and subsequent structure-based drug design and (2) preparation of a large quantity of relatively contaminant free crystals for use as time-release protein pharmaceuticals. The first three Shuttle flights with bovine insulin constituted the PCF's proof of concept, demonstrating that the space-grown crystals were larger and diffracted to higher resolution than their earth-grown counterparts. The later four PCF missions were used to grow recombinant human insulin crystals for x-ray analysis and continue productions trials aimed at the development of a processing facility for crystalline recombinant a-interferon.

  13. Reversal of diabetes following transplantation of an insulin-secreting human liver cell line: Melligen cells.

    PubMed

    Lawandi, Janet; Tao, Chang; Ren, Binhai; Williams, Paul; Ling, Dora; Swan, M Anne; Nassif, Najah T; Torpy, Fraser R; O'Brien, Bronwyn A; Simpson, Ann M

    2015-01-01

    As an alternative to the transplantation of islets, a human liver cell line has been genetically engineered to reverse type 1 diabetes (TID). The initial liver cell line (Huh7ins) commenced secretion of insulin in response to a glucose concentration of 2.5 mmol/l. After transfection of the Huh7ins cells with human islet glucokinase, the resultant Melligen cells secreted insulin in response to glucose within the physiological range; commencing at 4.25 mmol/l. Melligen cells exhibited increased glucokinase enzymatic activity in response to physiological glucose concentrations, as compared with Huh7ins cells. When transplanted into diabetic immunoincompetent mice, Melligen cells restored normoglycemia. Quantitative real-time polymerase chain reaction (qRT-PCR) revealed that both cell lines expressed a range of β-cell transcription factors and pancreatic hormones. Exposure of Melligen and Huh7ins cells to proinflammatory cytokines (TNF-α, IL-1β, and IFN-γ) affected neither their viability nor their ability to secrete insulin to glucose. Gene expression (microarray and qRT-PCR) analyses indicated the survival of Melligen cells in the presence of known β-cell cytotoxins was associated with the expression of NF-κB and antiapoptotic genes (such as BIRC3). This study describes the successful generation of an artificial β-cell line, which, if encapsulated to avoid allograft rejection, may offer a clinically applicable cure for T1D. PMID:26029722

  14. Glucose-dependent insulinotropic polypeptide: effects on insulin and glucagon secretion in humans.

    PubMed

    Christensen, Mikkel Bring

    2016-04-01

    hypoglycaemia. The results from the three studies indicate that GIP has effects on insulin and glucagon responses highly dependent upon the blood glucose levels. At fasting glycaemia and lower levels of glycaemia, GIP acts to increase glucagon with little effect on insulin release. At hyperglycaemia the insulin releasing effect of GIP prevail, which lead to an increase in glucose disposal by approximately 75% in healthy subjects (Study 1) and 25% in patients with Type 2 diabetes (Study 2) relative to placebo. After insulin-induced hypoglycaemia in patients with type 1 diabetes (Study 3), GIP increase glucagon release, which probably augments endogenous glucose production. This was associated with a reduced need for exogenously added glucose to prevent hypoglycaemia. In conclusion, the studies position GIP as a bifunctional blood glucose stabilising hormone that glucose-dependently regulates insulin and glucagon responses in humans. PMID:27034187

  15. Preparation of Tyr-C-peptide from genetically altered human insulin precursor.

    PubMed

    Sun, H; Tang, J; Hu, M

    1995-12-01

    C-peptide radioimmunoassay (C-peptide RIA) is widely used in determination of pancreatic B-cell secretion activity. 125I labeled Tyr-C-peptide is indispensable in C-peptide RIA kit. Herein we discuss a way of obtaining recombinant Tyr-C-peptide. Arg32Tyr human pro-insulin mutant (R32Y-proinsulin) gene was constructed by site-directed mutagenesis and overexpressed in Escherichia coli. Purified R32Y-proinsulin was converted to insulin and Tyr-C-peptide by trypsin and carboxypeptidase B codigestion. Tyr-C-peptide was isolated through reverse-phase HPLC (RP-HPLC) and identified by C-peptide RIA and amino acid analysis.

  16. Regulation of various proteolytic pathways by insulin and amino acids in human fibroblasts.

    PubMed

    Esteban, Inmaculada; Aguado, Carmen; Sánchez, Maribel; Knecht, Erwin

    2007-07-24

    Intracellular protein degradation is a regulated process with several proteolytic pathways. Although regulation of macroautophagy has been investigated in some detail in hepatocytes and in few other cells, less is known on this regulation in other cells and proteolytic pathways. We show that in human fibroblasts insulin and amino acids reduce protein degradation by different signalling pathways and that this inhibition proceeds in part via the mammalian target of rapamycin, especially with amino acids, which probably increase lysosomal pH. Moreover, the regulatory amino acids (Phe, Arg, Met, Tyr, Trp and Cys) are partially different from other cells. Finally, and in addition to macroautophagy, insulin and amino acids modify, to different extents and sometimes in opposite directions, the activities of other proteolytic pathways.

  17. Pancreatic Transdifferentiation and Glucose-Regulated Production of Human Insulin in the H4IIE Rat Liver Cell Line.

    PubMed

    Ren, Binhai; Tao, Chang; Swan, Margaret Anne; Joachim, Nichole; Martiniello-Wilks, Rosetta; Nassif, Najah T; O'Brien, Bronwyn A; Simpson, Ann M

    2016-01-01

    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. PMID:27070593

  18. Pancreatic Transdifferentiation and Glucose-Regulated Production of Human Insulin in the H4IIE Rat Liver Cell Line

    PubMed Central

    Ren, Binhai; Tao, Chang; Swan, Margaret Anne; Joachim, Nichole; Martiniello-Wilks, Rosetta; Nassif, Najah T.; O’Brien, Bronwyn A.; Simpson, Ann M.

    2016-01-01

    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. PMID:27070593

  19. Effects of recombinant insulin-like growth factor I on insulin secretion and renal function in normal human subjects.

    PubMed Central

    Guler, H P; Schmid, C; Zapf, J; Froesch, E R

    1989-01-01

    Insulin-like growth factor I (IGF-I) is an important mediator of growth hormone (GH) action and it appeared tempting to evaluate possible clinical applications. Recombinant IGF-I was infused s.c. at a dose of 20 micrograms/kg of body weight per hour during 6 days in two healthy adult subjects. Blood glucose and fasting insulin levels remained within normal limits and IGF-II levels were suppressed. In contrast to insulin, fasting C peptide levels were decreased. GH secretion was also suppressed by IGF-I. Our preliminary data allow us to distinguish between the effects of GH per se and those of IGF-I: GH causes hyperinsulinism, whereas IGF-I leads to decreased insulin secretion. Glomerular filtration rate, as estimated by creatinine clearance, increased to 130% of preinfusion values during the IGF-I infusion. Total creatinine and urea excretion remained unchanged. We conclude that IGF-I influences kidney function and, in contrast to GH, exerts an insulin-sparing effect. It may be speculated that the therapeutic spectrum of IGF-I is quite different from that of GH. Images PMID:2649897

  20. A general flamelet transformation useful for distinguishing between premixed and non-premixed modes of combustion

    SciTech Connect

    Knudsen, E.; Pitsch, H.

    2009-03-15

    The flame index was originally proposed by Yamashita et al. as a method of locally distinguishing between premixed and non-premixed combustion. Although this index has been applied both passively in the analysis of direct numerical simulation data, and actively using single step combustion models, certain limitations restrict its use in more detailed combustion models. In this work a general flamelet transformation that holds in the limits of both premixed and non-premixed combustion is developed. This transformation makes use of two statistically independent variables: a mixture fraction and a reaction progress parameter. The transformation is used to produce a model for distinguishing between premixed and non-premixed combustion regimes. The new model locally examines the term budget of the general flamelet transformation. The magnitudes of each of the terms in the budget are calculated and compared to the chemical source term. Determining whether a flame burns in a premixed or a non-premixed regime then amounts to determining which sets of these terms most significantly contribute to balancing the source term. The model is tested in a numerical simulation of a laminar triple flame, and is compared to a recent manifestation of the flame index approach. Additionally, the model is applied in a presumed probability density function (PDF) large eddy simulation (LES) of a lean premixed swirl burner. The model is used to locally select whether tabulated premixed or tabulated non-premixed chemistry should be referenced in the LES. Results from the LES are compared to experiments. (author)

  1. [Differentiation of human amniotic mesenchymal stem cells into insulin-secreting cells induced by regenerating pancreatic extract].

    PubMed

    Zhang, Yanmei; Wang, Dianliang; Zeng, Hongyan; Wang, Lieming; Sun, Jinwei; Zhang, Zhen; Dong, Shasha

    2012-02-01

    In this study, the natural biological inducer, rat regenerating pancreatic extract (RPE), was used to induce human amniotic mesenchymal stem cells (hAMSCs) into insulin-secreting cells. We excised 60% of rat pancreas in order to stimulate pancreatic regeneration. RPE was extracted and used to induce hAMSCs at a final concentration of 20 microg/mL. The experiment methods used were as follows: morphological-identification, dithizone staining, immumofluorescence analysis, reverse transcription-PCR (RT-PCR) and insulin secretion stimulated by high glucose. The results show that the cell morphology of passge3 hAMSCs changed significantly after the induction of RPE, resulting in cluster shape after induction for 15 days. Dithizone staining showed that there were scarlet cell masses in RPE-treated culture. Immumofluorescence analysis indicated that induced cells were insulin-positive expression. RT-PCR showed the positive expression of human islet-related genes Pdx1 and insulin in the induced cells. The result of insulin secretion stimulated by high glucose indicated that insulin increasingly secreted and then kept stable with prolongation of high glucose stimulation. In conclusion, hAMSCs had the potential to differentiate into insulin-secreting cells induced by RPE in vitro. PMID:22667123

  2. SIRT3 Deficiency Induces Endothelial Insulin Resistance and Blunts Endothelial-Dependent Vasorelaxation in Mice and Human with Obesity

    PubMed Central

    Yang, Lu; Zhang, Julei; Xing, Wenjuan; Zhang, Xing; Xu, Jie; Zhang, Haifeng; Chen, Li; Ning, Xiaona; Ji, Gang; Li, Jia; Zhao, Qingchuan; Gao, Feng

    2016-01-01

    Recent evidence implicates the critical role of Sirtuin 3 (SIRT3) in the development of many metabolic diseases, but the contribution of SIRT3 to vascular homeostasis remains largely unknown. The aim of this study was to investigate the role of SIRT3 in endothelial insulin resistance and vascular dysfunction in obesity. We found an impaired insulin-induced mesenteric vasorelaxation and concomitant reduced vascular SIRT3 expression in morbid obese human subjects compared with the non-obese subjects. Downregulation of SIRT3 in cultured human endothelial cells increased mitochondrial reactive oxygen species (mtROS) and impaired insulin signaling as evidenced by decreased phosphorylation of Akt and endothelial nitric oxide synthase and subsequent reduced nitric oxide (NO) release. In addition, obese mice induced by 24-week high-fat diet (HFD) displayed an impaired endothelium-dependent vasorelaxation to both insulin and acetylcholine, which was further exacerbated by the gene deletion of Sirt3. Scavenging of mtROS not only restored insulin-stimulated NO production in SIRT3 knockdown cells, but also improved insulin-induced vasorelaxation in SIRT3 knockout mice fed with HFD. Taken together, our findings suggest that SIRT3 positively regulates endothelial insulin sensitivity and show that SIRT3 deficiency and resultant increased mtROS contribute to vascular dysfunction in obesity. PMID:27000941

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

    PubMed

    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.

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

    PubMed Central

    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

    2013-01-01

    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 PMID:24252877

  5. Lipoprotein lipase in human milk: compartmentalization and effect of fasting, insulin, and glucose.

    PubMed

    Neville, M C; Waxman, L J; Jensen, D; Eckel, R H

    1991-02-01

    The object of this study was to investigate the effect of maternal metabolic state on the activity of lipoprotein lipase (LPL) in human milk. Although the total LPL activity in milk was not significantly affected by up to three cycles of freezing and thawing, the amount of LPL associated with the cream fraction of the milk increased from an average of less than 10% to about 70% after this treatment. The enzyme was relatively stable when the milk was stored on ice, losing activity at a rate of about 1% per hour. At 37 degrees C degradation was more rapid, about 7% per hour. When LPL activity was measured in samples taken at hourly intervals by breast pump, using oxytocin to achieve a complete letdown at each pumping, activity was found to double from the first to the third pumping. Thereafter the activity was stable under fasting conditions. Hyperglycemic and euglycemic, hyperinsulinemic glucose clamp protocols were used to evaluate the effects of glucose and insulin. Both high plasma glucose and high plasma insulin in the presence of normal glucose significantly increased LPL activity within 4 hours. We conclude that, like adipose, tissue LPL, mammary LPL is regulated by plasma insulin.

  6. Gas turbine premixer with internal cooling

    DOEpatents

    York, William David; Johnson, Thomas Edward; Lacy, Benjamin Paul; Stevenson, Christian Xavier

    2012-12-18

    A system that includes a turbine fuel nozzle comprising an air-fuel premixer. The air-fuel premixed includes a swirl vane configured to swirl fuel and air in a downstream direction, wherein the swirl vane comprises an internal coolant path from a downstream end portion in an upstream direction through a substantial length of the swirl vane.

  7. Oscillating combustion from a premix fuel nozzle

    SciTech Connect

    Richards, G.A.; Yip, M.J.

    1995-08-01

    Stringent emissions requirements for stationary gas turbines have produced new challenges in combustor design. In the past, very low NOx pollutant emissions have been achieved through various combustion modifications, such as steam or water injection, or post-combustion cleanup methods such as selective catalytic reduction (SCR). An emerging approach to NOx abatement is lean premix combustion. Lean premix combustion avoids the cost and operational problems associated with other NOx control methods. By premixing fuel and air at very low equivalence ratios, the high temperatures which produce NOx are avoided. The challenges of premix combustion include avoiding flashback, and ensuring adequate fuel/air premixing. In addition, the combustion must be stable. The combustor should not operate so close to extinction that a momentary upset will extinguish the flame (static stability), and the flame should not oscillate (dynamic stability). Oscillations are undesirable because the associated pressure fluctuations can shorten component lifetime. Unfortunately, experience has shown that premix fuel nozzles burning natural gas are susceptible to oscillations. Eliminating these oscillations can be a costly and time consuming part of new engine development. As part of the U.S. Department of Energy`s Advanced Turbine Systems Program, the Morgantown Energy Technology Center (METC) is investigating the issue of combustion oscillations produced by lean premix fuel nozzles. METC is evaluating various techniques to stabilize oscillating combustion in gas turbines. Tests results from a premix fuel nozzle using swirl stabilization and a pilot flame are reported here.

  8. Flashback resistant pre-mixer assembly

    DOEpatents

    Laster, Walter R.; Gambacorta, Domenico

    2012-02-14

    A pre-mixer assembly associated with a fuel supply system for mixing of air and fuel upstream from a main combustion zone in a gas turbine engine. The pre-mixer assembly includes a swirler assembly disposed about a fuel injector of the fuel supply system and a pre-mixer transition member. The swirler assembly includes a forward end defining an air inlet and an opposed aft end. The pre-mixer transition member has a forward end affixed to the aft end of the swirler assembly and an opposed aft end defining an outlet of the pre-mixer assembly. The aft end of the pre-mixer transition member is spaced from a base plate such that a gap is formed between the aft end of the pre-mixer transition member and the base plate for permitting a flow of purge air therethrough to increase a velocity of the air/fuel mixture exiting the pre-mixer assembly.

  9. Insulin Stimulates Translocation of Human GLUT4 to the Membrane in Fat Bodies of Transgenic Drosophila melanogaster

    PubMed Central

    Crivat, Georgeta; Lizunov, Vladimir A.; Li, Caroline R.; Stenkula, Karin G.; Zimmerberg, Joshua; Cushman, Samuel W.; Pick, Leslie

    2013-01-01

    The fruit fly Drosophila melanogaster is an excellent model system for studies of genes controlling development and disease. However, its applicability to physiological systems is less clear because of metabolic differences between insects and mammals. Insulin signaling has been studied in mammals because of relevance to diabetes and other diseases but there are many parallels between mammalian and insect pathways. For example, deletion of Drosophila Insulin-Like Peptides resulted in ‘diabetic’ flies with elevated circulating sugar levels. Whether this situation reflects failure of sugar uptake into peripheral tissues as seen in mammals is unclear and depends upon whether flies harbor the machinery to mount mammalian-like insulin-dependent sugar uptake responses. Here we asked whether Drosophila fat cells are competent to respond to insulin with mammalian-like regulated trafficking of sugar transporters. Transgenic Drosophila expressing human glucose transporter-4 (GLUT4), the sugar transporter expressed primarily in insulin-responsive tissues, were generated. After expression in fat bodies, GLUT4 intracellular trafficking and localization were monitored by confocal and total internal reflection fluorescence microscopy (TIRFM). We found that fat body cells responded to insulin with increased GLUT4 trafficking and translocation to the plasma membrane. While the amplitude of these responses was relatively weak in animals reared on a standard diet, it was greatly enhanced in animals reared on sugar-restricted diets, suggesting that flies fed standard diets are insulin resistant. Our findings demonstrate that flies are competent to mobilize translocation of sugar transporters to the cell surface in response to insulin. They suggest that Drosophila fat cells are primed for a response to insulin and that these pathways are down-regulated when animals are exposed to constant, high levels of sugar. Finally, these studies are the first to use TIRFM to monitor insulin

  10. Differential effects of prednisone and growth hormone on fuel metabolism and insulin antagonism in humans

    SciTech Connect

    Horber, F.F.; Marsh, H.M.; Haymond, M.W. )

    1991-01-01

    Human growth hormone (hGH) and prednisone cause insulin resistance and glucose intolerance. However, it is unknown whether hGH and prednisone antagonize insulin action on protein, fat, and carbohydrate metabolism by a common or independent mechanism. Therefore, protein, fat, and carbohydrate metabolism was assessed simultaneously in four groups of eight subjects each after 7 days of placebo, recombinant DNA hGH (rhGH; 0.1 mg.kg-1.day-1), prednisone (0.8 mg.kg-1.day-1), or rhGH and prednisone administration after an 18-h fast and during gut infusion of glucose and amino acids (fed state). Fasting plasma glucose concentrations were similar during placebo and rhGH but elevated (P less than 0.001) during combined treatment, whereas plasma insulin concentrations were higher (237 +/- 57 pmol/ml, P less than 0.001) during combined than during placebo, rhGH, or prednisone treatment (34, 52, and 91 pM, respectively). In the fed state, plasma glucose concentrations were elevated only during combined treatment (11.3 +/- 2.1 mM, P less than 0.001). Plasma insulin concentrations were elevated during therapy with prednisone alone and rhGH alone (667 +/- 72 and 564 +/- 65 pmol/ml, respectively, P less than 0.001) compared with placebo (226 +/- 44 pmol/ml) but lower than with the combined rhGH and prednisone treatment (1249 +/- 54 pmol/ml, P less than 0.01). Protein oxidation {sup 14}C leucine increased (P less than 0.001) with prednisone therapy, decreased (P less than 0.001) with rhGH treatment, and was normal during the combined treatment.

  11. AMP-activated protein kinase (AMPK) regulates the insulin-induced activation of the nitric oxide synthase in human platelets.

    PubMed

    Fleming, Ingrid; Schulz, Christian; Fichtlscherer, Birgit; Kemp, Bruce E; Fisslthaler, Beate; Busse, Rudi

    2003-11-01

    Little is known about the signaling cascades that eventually regulate the activity of the endothelial nitric oxide synthase (eNOS) in platelets. Here, we investigated the effects of insulin on the phosphorylation and activation of eNOS in washed human platelets and in endothelial cells. Insulin activated the protein kinase Akt in cultured endothelial cells and increased the phosphorylation of eNOS on Ser(1177) but failed to increase endothelial cyclic GMP levels or to elicit the relaxation of endothelium-intact porcine coronary arteries. In platelets, insulin also elicited the activation of Akt as well as the phosphorylation of eNOS and initiated NO production which was associated with increased cyclic GMP levels and the inhibition of thrombin-induced aggregation. The insulin-induced inhibition of aggregation was accompanied by a decreased Ca(2+) response to thrombin and was also prevented by N(omega) nitro-L-arginine. In platelets, but not in endothelial cells, insulin induced the activation of the AMP-activated protein kinase (AMPK), a metabolic stress-sensing kinase which was sensitive to the phosphatidylinositol 3-kinase (PI3-K) inhibitor wortmannin and the AMPK inhibitor iodotubercidin. Moreover, the insulin-mediated inhibition of thrombin-induced aggregation was prevented by iodotubercidin. Insulin-independent activation of the AMPK using 5-aminoimidazole-4-carboxamide ribonucleoside, increased platelet eNOS phosphorylation, increased cyclic GMP levels and attenuated platelet aggregation. These results highlight the differences in the signal transduction cascade activated by insulin in endothelial cells and platelets, and demonstrate that insulin stimulates the formation of NO in human platelets, in the absence of an increase in Ca(2+), by acti-vating PI3-K and AMPK which phosphorylates eNOS on Ser(1177).

  12. High-level expression of human insulin receptor cDNA in mouse NIH 3T3 cells

    SciTech Connect

    Whittaker, J.; Okamoto, A.K.; Thys, R.; Bell, G.I.; Steiner, D.F.; Hofmann, C.A.

    1987-08-01

    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/sup 6/ receptors per cell. The cell line with the highest /sup 125/I-insulin binding (NIH 3T3 HIR3.5) had 6 x 10/sup 6/ receptors with a K/sub d/ of 10/sup -9/ M. This level was not dependent on exposure to metals but could be increased further to 2 x 10/sup 7/ receptors per cell by addition of sodium butyrate to the culture medium. The ..cap alpha.. and ..beta.. 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 ..cap alpha.. and ..beta.. 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

  13. Inhibition of human insulin gene transcription and MafA transcriptional activity by the dual leucine zipper kinase

    PubMed Central

    Stahnke, Marie-Jeannette; Dickel, Corinna; Schröder, Sabine; Kaiser, Diana; Blume, Roland; Stein, Roland; Pouponnot, Celio; Oetjen, Elke

    2016-01-01

    Insulin biosynthesis is an essential β-cell function and inappropriate insulin secretion and biosynthesis contribute to the pathogenesis of diabetes mellitus type 2. Previous studies showed that the dual leucine zipper kinase (DLK) induces β-cell apoptosis. Since β-cell dysfunction precedes β-cell loss, in the present study the effect of DLK on insulin gene transcription was investigated in the HIT-T15 β-cell line. Downregulation of endogenous DLK increased whereas overexpression of DLK decreased human insulin gene transcription. 5′- and 3′-deletion human insulin promoter analyses resulted in the identification of a DLK responsive element that mapped to the DNA binding-site for the β-cell specific transcription factor MafA. Overexpression of DLK wild-type but not its kinase-dead mutant inhibited MafA transcriptional activity conferred by its transactivation domain. Furthermore, in the non-β-cell line JEG DLK inhibited MafA overexpression-induced human insulin promoter activity. Overexpression of MafA and DLK or its kinase-dead mutant into JEG cells revealed that DLK but not its mutant reduced MafA protein content. Inhibition of the down-stream DLK kinase c-Jun N-terminal kinase (JNK) by SP600125 attenuated DLK-induced MafA loss. Furthermore, mutation of the serine 65 to alanine, shown to confer MafA protein stability, increased MafA-dependent insulin gene transcription and prevented DLK-induced MafA loss in JEG cells. These data suggest that DLK by activating JNK triggers the phosphorylation and degradation of MafA thereby attenuating insulin gene transcription. Given the importance of MafA for β-cell function, the inhibition of DLK might preserve β-cell function and ultimately retard the development of diabetes mellitus type 2. PMID:24726898

  14. Pseudoislet formation enhances gene expression, insulin secretion and cytoprotective mechanisms of clonal human insulin-secreting 1.1B4 cells.

    PubMed

    Green, Alastair D; Vasu, Srividya; McClenaghan, Neville H; Flatt, Peter R

    2015-10-01

    We have studied the effects of cell communication on human beta cell function and resistance to cytotoxicity using the novel human insulin-secreting cell line 1.1B4 configured as monolayers and pseudoislets. Incubation with the incretin gut hormones GLP-1 and GIP caused dose-dependent stimulation of insulin secretion from 1.1B4 cell monolayers and pseudoislets. The secretory responses were 1.5-2.7-fold greater than monolayers. Cell viability (MTT), DNA damage (comet assay) and apoptosis (acridine orange/ethidium bromide staining) were investigated following 2-h exposure of 1.1B4 monolayers and pseudoislets to ninhydrin, H2O2, streptozotocin, glucose, palmitate or cocktails of proinflammatory cytokines. All agents tested decreased viability and increased DNA damage and apoptosis in both 1.1B4 monolayers and pseudoislets. However, pseudoislets exhibited significantly greater resistance to cytotoxicity (1.5-2.7-fold increases in LD50) and lower levels of DNA damage (1.3-3.4-fold differences in percentage tail DNA and olive tail moment) and apoptosis (1.3-1.5-fold difference) compared to monolayers. Measurement of gene expression by reverse-transcription, real-time PCR showed that genes involved with insulin secretion (INS, PDX1, PCSK1, PCSK2, GLP1R and GIPR), cell-cell communication (GJD2, GJA1 and CDH1) and antioxidant defence (SOD1, SOD2, GPX1 and CAT) were significantly upregulated in pseudoislets compared to monolayers, whilst the expression of proapoptotic genes (NOS2, MAPK8, MAPK10 and NFKB1) showed no significant differences. In summary, these data indicate cell-communication associated with three-dimensional islet architecture is important both for effective insulin secretion and for protection of human beta cells against cytotoxicity. PMID:25559846

  15. Fuel premixing module for gas turbine engine combustor

    NASA Technical Reports Server (NTRS)

    Chin, Jushan (Inventor); Rizk, Nader K. (Inventor); Razdan, Mohan K. (Inventor); Marshall, Andre W. (Inventor)

    2005-01-01

    A fuel-air premixing module is designed to reduce emissions from a gas turbine engine. In one form, the premixing module includes a central pilot premixer module with a main premixer module positioned thereround. Each of the portions of the fuel-air premixing module include an axial inflow swirler with a plurality of fixed swirler vanes. Fuel is injected into the main premixer module between the swirler vanes of the axial inflow swirler and at an acute angle relative to the centerline of the premixing module.

  16. β-Cell Sensitivity to GLP-1 in Healthy Humans Is Variable and Proportional to Insulin Sensitivity

    PubMed Central

    Aulinger, Benedikt A.; Vahl, Torsten P.; Wilson-Pérez, Hilary E.; Prigeon, Ron L.

    2015-01-01

    Context: Glucagon-like peptide-1 (GLP-1) is an insulinotropic factor made in the gastrointestinal tract that is essential for normal glucose tolerance. Infusion of GLP-1 increases insulin secretion in both diabetic and nondiabetic humans. However, the degree to which people vary in their β-cell sensitivity to GLP-1 and the factors contributing to this variability have not been reported. Objective: The objective was to measure the sensitivity of insulin secretion to GLP-1 in cohorts of lean and obese subjects across a broad range of insulin sensitivity. Methods: Insulin secretion was measured during clamped hyperglycemia (7.2 mmol/L) and graded GLP-1 infusion in young, healthy subjects, and GLP-1 sensitivity was computed from the insulin secretion rate (ISR) during progressive increases in plasma GLP-1. Results: All subjects had fasting glucose values <5.2 mm. The obese subjects were insulin resistant compared to the lean group (homeostasis model of assessment 2 for insulin resistance: obese, 2.6 ± 0.5; lean, 0.8 ± 0.1; P < .001). ISR increased linearly in both cohorts with escalating doses of GLP-1, but the slope of ISR in response to GLP-1 was greater in the obese than in the lean subjects (obese, 0.17 ± 0.03 nmol/min/pm; lean, 0.05 ± 0.01 nmol/min/pm; P < .001). There was a significant association of β-cell GLP-1 sensitivity and insulin resistance (r = 0.83; P < .001), and after correction for homeostasis model of assessment 2 for insulin resistance, the slopes of ISR vs GLP-1 concentration did not differ in the two cohorts (obese, 0.08 ± 0.01; lean, 0.08 ± 0.01; P = .98). However, within the entire study group, β-cell GLP-1 sensitivity corrected for insulin resistance varied nearly 10-fold. Conclusions: Insulin secretion in response to GLP-1 is proportional to insulin resistance in healthy subjects. However, there is considerable variability in the sensitivity of the β-cell to GLP-1 that is independent of insulin sensitivity. PMID:25825945

  17. Leptin rapidly suppresses insulin release from insulinoma cells, rat and human islets and, in vivo, in mice.

    PubMed Central

    Kulkarni, R N; Wang, Z L; Wang, R M; Hurley, J D; Smith, D M; Ghatei, M A; Withers, D J; Gardiner, J V; Bailey, C J; Bloom, S R

    1997-01-01

    Obesity is associated with diabetes, and leptin is known to be elevated in obesity. To investigate whether leptin has a direct effect on insulin secretion, isolated rat and human islets and cultured insulinoma cells were studied. In all cases, mouse leptin inhibited insulin secretion at concentrations within the plasma range reported in humans. Insulin mRNA expression was also suppressed in the cultured cells and rat islets. The long form of the leptin receptor (OB-Rb) mRNA was present in the islets and insulinoma cell lines. To determine the significance of these findings in vivo, normal fed mice were injected with two doses of leptin. A significant decrease in plasma insulin and associated rise in glucose concentration were observed. Fasted normal and leptin receptor-deficient db/db mice showed no response to leptin. A dose of leptin, which mimicked that found in normal mice, was administered to leptin-deficient, hyperinsulinemic ob/ob mice. This caused a marked lowering of plasma insulin concentration and a doubling of plasma glucose. Thus, leptin has a powerful acute inhibitory effect on insulin secretion. These results suggest that the action of leptin may be one mechanism by which excess adipose tissue could acutely impair carbohydrate metabolism. PMID:9389736

  18. Alterations in skeletal muscle protein-tyrosine phosphatase activity and expression in insulin-resistant human obesity and diabetes.

    PubMed Central

    Ahmad, F; Azevedo, J L; Cortright, R; Dohm, G L; Goldstein, B J

    1997-01-01

    Obese human subjects have increased protein-tyrosine phosphatase (PTPase) activity in adipose tissue that can dephosphorylate and inactivate the insulin receptor kinase. To extend these findings to skeletal muscle, we measured PTPase activity in the skeletal muscle particulate fraction and cytosol from a series of lean controls, insulin-resistant obese (body mass index > 30) nondiabetic subjects, and obese individuals with non-insulin-dependent diabetes. PTPase activities in subcellular fractions from the nondiabetic obese subjects were increased to 140-170% of the level in lean controls (P < 0.05). In contrast, PTPase activity in both fractions from the obese subjects with non-insulin-dependent diabetes was significantly decreased to 39% of the level in controls (P < 0.05). By immunoblot analysis, leukocyte antigen related (LAR) and protein-tyrosine phosphatase 1B had the greatest increase (threefold) in the particulate fraction from obese, nondiabetic subjects, and immunodepletion of this fraction using an affinity-purified antibody directed at the cytoplasmic domain of leukocyte antigen related normalized the PTPase activity when compared to the activity from control subjects. These findings provide further support for negative regulation of insulin action by specific PTPases in the pathogenesis of insulin resistance in human obesity, while other regulatory mechanisms may be operative in the diabetic state. PMID:9218523

  19. Studies on human insulin adsorption kinetics at an organic-aqueous interface determined using a label-free electroanalytical approach.

    PubMed

    Thomsen, Anne Engelbrecht; Jensen, Henrik; Jorgensen, Lene; van de Weert, Marco; Ostergaard, Jesper

    2008-06-01

    Protein adsorption represents a considerable challenge in the development and production of macromolecular drugs. From an analytical point of view the adsorption process is difficult to study in an efficient way using currently available techniques. In this work potential and time dependent adsorption and adsorption kinetics of human insulin at an 1,2-dichloroethane-aqueous interface were studied using a novel electroanalytical approach based on measurements of interfacial capacitance. Two different types of measurements were performed; potential scans and time scans. In the potential scans, the capacitance was measured over a range of applied potential differences across the interface. The interfacial potential difference is linked to the charge at the interface. Adsorption of human insulin was detectable at a bulk phase insulin concentration as low as 0.1 microM as a negative shift in the potential of zero charge (pzc). Adsorption kinetics were further studied using time scans in which the interfacial capacitance was measured at a fixed applied interfacial potential difference. Using this approach it was possible to study how the adsorption kinetics and the shape of the time scan curves were related to the bulk concentration of insulin and the interfacial potential difference. The changes in capacitance could be described phenomenologically by pseudo-first-order kinetics at low concentrations of insulin except at positive interfacial potential differences and high insulin concentrations (> or =0.25 microM) where a more complex shape of the time scans curves was observed.

  20. Effect of insulin on human skeletal muscle mitochondrial ATP production, protein synthesis, and mRNA transcripts

    NASA Astrophysics Data System (ADS)

    Stump, Craig S.; Short, Kevin R.; Bigelow, Maureen L.; Schimke, Jill M.; Sreekumaran Nair, K.

    2003-06-01

    Mitochondria are the primary site of skeletal muscle fuel metabolism and ATP production. Although insulin is a major regulator of fuel metabolism, its effect on mitochondrial ATP production is not known. Here we report increases in vastus lateralis muscle mitochondrial ATP production capacity (32-42%) in healthy humans (P < 0.01) i.v. infused with insulin (1.5 milliunits/kg of fat-free mass per min) while clamping glucose, amino acids, glucagon, and growth hormone. Increased ATP production occurred in association with increased mRNA levels from both mitochondrial (NADH dehydrogenase subunit IV) and nuclear [cytochrome c oxidase (COX) subunit IV] genes (164-180%) encoding mitochondrial proteins (P < 0.05). In addition, muscle mitochondrial protein synthesis, and COX and citrate synthase enzyme activities were increased by insulin (P < 0.05). Further studies demonstrated no effect of low to high insulin levels on muscle mitochondrial ATP production for people with type 2 diabetes mellitus, whereas matched nondiabetic controls increased 16-26% (P < 0.02) when four different substrate combinations were used. In conclusion, insulin stimulates mitochondrial oxidative phosphorylation in skeletal muscle along with synthesis of gene transcripts and mitochondrial protein in human subjects. Skeletal muscle of type 2 diabetic patients has a reduced capacity to increase ATP production with high insulin levels. cytochrome c oxidase | NADH dehydrogenase subunit IV | amino acids | citrate synthase

  1. In vivo activation of ROCK1 by insulin is impaired in skeletal muscle of humans with type 2 diabetes.

    PubMed

    Chun, Kwang-Hoon; Choi, Kang-Duk; Lee, Dae-Ho; Jung, Yoonshin; Henry, Robert R; Ciaraldi, Theodore P; Kim, Young-Bum

    2011-03-01

    To determine whether serine/threonine ROCK1 is activated by insulin in vivo in humans and whether impaired activation of ROCK1 could play a role in the pathogenesis of insulin resistance, we measured the activity of ROCK1 and the protein content of the Rho family in vastus lateralis muscle of lean, obese nondiabetic, and obese type 2 diabetic subjects. Biopsies were taken after an overnight fast and after a 3-h hyperinsulinemic euglycemic clamp. Insulin-stimulated GDR was reduced 38% in obese nondiabetic subjects compared with lean, 62% in obese diabetic subjects compared with lean, and 39% in obese diabetic compared with obese nondiabetic subjects (all comparisons P < 0.001). Insulin-stimulated IRS-1 tyrosine phosphorylation is impaired 41-48% in diabetic subjects compared with lean or obese subjects. Basal activity of ROCK1 was similar in all groups. Insulin increased ROCK1 activity 2.1-fold in lean and 1.7-fold in obese nondiabetic subjects in muscle. However, ROCK1 activity did not increase in response to insulin in muscle of obese type 2 diabetic subjects without change in ROCK1 protein levels. Importantly, insulin-stimulated ROCK1 activity was positively correlated with insulin-mediated GDR in lean subjects (P < 0.01) but not in obese or type 2 diabetic subjects. Moreover, RhoE GTPase that inhibits the catalytic activity of ROCK1 by binding to the kinase domain of the enzyme is notably increased in obese type 2 diabetic subjects, accounting for defective ROCK1 activity. Thus, these data suggest that ROCK1 may play an important role in the pathogenesis of resistance to insulin action on glucose disposal in muscle of obese type 2 diabetic subjects.

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

    NASA Astrophysics Data System (ADS)

    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.

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

    PubMed

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

    2003-06-10

    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. PMID:12756298

  4. Characterization of lipid metabolism in insulin-sensitive adipocytes differentiated from immortalized human mesenchymal stem cells

    SciTech Connect

    Prawitt, Janne; Niemeier, Andreas; Kassem, Moustapha; Beisiegel, Ulrike; Heeren, Joerg

    2008-02-15

    There is a great demand for cell models to study human adipocyte function. Here we describe the adipogenic differentiation of a telomerase-immortalized human mesenchymal stem cell line (hMSC-Tert) that maintains numerous features of terminally differentiated adipocytes even after prolonged withdrawal of the peroxisome proliferator activated receptor {gamma} (PPAR{gamma}) agonist rosiglitazone. Differentiated hMSC-Tert developed the characteristic monolocular phenotype of mature adipocytes. The expression of adipocyte specific markers was highly increased during differentiation. Most importantly, the presence of the PPAR{gamma} agonist rosiglitazone was not required for the stable expression of lipoprotein lipase, adipocyte fatty acid binding protein and perilipin on mRNA and protein levels. Adiponectin expression was post-transcriptionally down-regulated in the absence of rosiglitazone. Insulin sensitivity as measured by insulin-induced phosphorylation of Akt and S6 ribosomal protein was also independent of rosiglitazone. In addition to commonly used adipogenic markers, we investigated further PPAR{gamma}-stimulated proteins with a role in lipid metabolism. We observed an increase of lipoprotein receptor (VLDLR, LRP1) and apolipoprotein E expression during differentiation. Despite this increased expression, the receptor-mediated endocytosis of lipoproteins was decreased in differentiated adipocytes, suggesting that these proteins may have an additional function in adipose tissue beyond lipoprotein uptake.

  5. Temperature-Acclimated Brown Adipose Tissue Modulates Insulin Sensitivity in Humans

    PubMed Central

    Lee, Paul; Smith, Sheila; Linderman, Joyce; Courville, Amber B.; Brychta, Robert J.; Dieckmann, William; Werner, Charlotte D.; Chen, Kong Y.

    2014-01-01

    In rodents, brown adipose tissue (BAT) regulates cold- and diet-induced thermogenesis (CIT; DIT). Whether BAT recruitment is reversible and how it impacts on energy metabolism have not been investigated in humans. We examined the effects of temperature acclimation on BAT, energy balance, and substrate metabolism in a prospective crossover study of 4-month duration, consisting of four consecutive blocks of 1-month overnight temperature acclimation (24°C [month 1] → 19°C [month 2] → 24°C [month 3] → 27°C [month 4]) of five healthy men in a temperature-controlled research facility. Sequential monthly acclimation modulated BAT reversibly, boosting and suppressing its abundance and activity in mild cold and warm conditions (P < 0.05), respectively, independent of seasonal fluctuations (P < 0.01). BAT acclimation did not alter CIT but was accompanied by DIT (P < 0.05) and postprandial insulin sensitivity enhancement (P < 0.05), evident only after cold acclimation. Circulating and adipose tissue, but not skeletal muscle, expression levels of leptin and adiponectin displayed reciprocal changes concordant with cold-acclimated insulin sensitization. These results suggest regulatory links between BAT thermal plasticity and glucose metabolism in humans, opening avenues to harnessing BAT for metabolic benefits. PMID:24954193

  6. Whole and fractionated yellow pea flours reduce fasting insulin and insulin resistance in hypercholesterolaemic and overweight human subjects.

    PubMed

    Marinangeli, Christopher P F; Jones, Peter J H

    2011-01-01

    The objective of the present study was to compare whole pea flour (WPF) to fractionated pea flour (FPF; hulls only) for their ability to reduce risk factors associated with CVD and diabetes in overweight hypercholesterolaemic individuals. Using a cross-over design, twenty-three hypercholesterolaemic overweight men and women received two-treatment muffins/d containing WPF, FPF or white wheat flour (WF) for 28 d, followed by 28 d washout periods. Daily doses of WPF and FPF complied with the United States Department of Agriculture's recommended level of intake of half a cup of pulses/d (approximately 50 g/d). Dietary energy requirements were calculated for each study subject, and volunteers were only permitted to eat food supplied by the study personnel. Fasting insulin, body composition, urinary enterolactone levels, postprandial glucose response, as well as fasting lipid and glucose concentrations, were assessed at the beginning and at the end of each treatment. Insulin concentrations for WPF (37·8 (SEM 3·4) pmol/ml, P = 0·021) and FPF (40·5 (SEM 3·4) pmol/ml, P = 0·037) were lower compared with WF (50·7 (SEM 3·4) pmol/ml). Insulin homeostasis modelling assessment showed that consumption of WPF and FPF decreased (P < 0·05) estimates of insulin resistance (IR) compared with WF. Android:gynoid fat ratios in women participants were lower (P = 0·027) in the WPF (1·01 (sem 0·01) group compared with the WF group (1·06 (SEM 0·01). Urinary enterolactone levels tended to be higher (P = 0·087) in WPF compared with WF. Neither treatment altered circulating fasting lipids or glucose concentrations. In conclusion, under a controlled diet paradigm, a daily consumption of whole and fractionated yellow pea flours at doses equivalent to half a cup of yellow peas/d reduced IR, while WPF reduced android adiposity in women.

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

    NASA Astrophysics Data System (ADS)

    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

  8. A premixed hydrogen/oxygen catalytic igniter

    NASA Technical Reports Server (NTRS)

    Green, James M.

    1989-01-01

    The catalytic ignition of hydrogen and oxygen propellants was studied using a premixing hydrogen/oxygen injector. The premixed injector was designed to eliminate problems associated with catalytic ignition caused by poor propellant mixing in the catalyst bed. Mixture ratio, mass flow rate, and propellant inlet temperature were varied parametrically in testing, and a pulse mode life test of the igniter was conducted. The results of the tests showed that the premixed injector eliminated flame flashback in the reactor and increased the life of the igniter significantly. The results of the experimental program and a comparison with data collected in a previous program are given.

  9. Adsorption of human insulin on single-crystal gold surfaces investigated by in situ scanning tunnelling microscopy and electrochemistry.

    PubMed

    Welinder, Anna C; Zhang, Jingdong; Steensgaard, Dorte B; Ulstrup, Jens

    2010-09-14

    We have explored the adsorption of zinc-free human insulin on the three low-index single-crystalline Au(111)-, Au(100)- and Au(110)-surfaces in aqueous buffer (KH(2)PO(4), pH 5) by a combination of electrochemical scanning tunnelling microscopy (in situ STM) at single-molecule resolution and linear sweep, LSV, cyclic, CV, and square wave (SQWV) voltammetry.Multifarious electrochemical patterns were observed. Most attention was given to reductive desorption caused by insulin binding to the Au-surfaces via up to three disulfide groups per insulin monomer, presumably converted to single Au-S links. SQWV suggested the Au-S bond strength order Au(111) > Au(110) > Au(100) based on the reductive desorption potentials. The voltammetric diversity was paralleled by different in situ STM insulin adsorption modes on the three surfaces. Single-molecule resolution was achieved in all cases. The coverage followed the order Au(110) > Au(100) > Au(111) and differs from the reductive desorption order that records the Au-S bonding element. Evenly distributed single molecules were scattered over large Au(111)-terraces, with intriguing molecular arrays disclosed near the terrace edges. In comparison, high-density molecular scale structures were observed both over the terraces and across terrace edges on Au(100). Larger rectangular structures also appeared (8-12% coverage). These are Au-islands from the lift of the reconstruction. Notably, 10 x 10 nm(2) patches of highly ordered much smaller structures, possibly from insulin decomposition emerged sporadically within the dense insulin adlayer. Insulin adsorbed in highest coverage on the Au(110) and followed the directional surface topology with insulin molecules aligned in the Au(110)-surface grooves, occasionally "spilling over" and merging into larger structures.Adsorption, Au-S binding, and insulin unfolding are all parts of insulin surface behaviour and reflected in both voltammetry and in situ STM. In spite of these complications

  10. Human embryonic stem cell differentiation into insulin secreting β-cells for diabetes.

    PubMed

    Bose, Bipasha; Shenoy, Sudheer P; Konda, Sudhakar; Wangikar, Pralhad

    2012-11-01

    hESC (human embryonic stem cells), when differentiated into pancreatic β ILC (islet-like clusters), have enormous potential for the cell transplantation therapy for Type 1 diabetes. We have developed a five-step protocol in which the EBs (embryoid bodies) were first differentiated into definitive endoderm and subsequently into pancreatic lineage followed by formation of functional endocrine β islets, which were finally matured efficiently under 3D conditions. The conventional cytokines activin A and RA (retinoic acid) were used initially to obtain definitive endoderm. In the last step, ILC were further matured under 3D conditions using amino acid rich media (CMRL media) supplemented with anti-hyperglycaemic hormone-Glp1 (glucagon-like peptide 1) analogue Liraglutide with prolonged t(½) and Exendin 4. The differentiated islet-like 3D clusters expressed bonafide mature and functional β-cell markers-PDX1 (pancreatic and duodenal homoeobox-1), C-peptide, insulin and MafA. Insulin synthesis de novo was confirmed by C-peptide ELISA of culture supernatant in response to varying concentrations of glucose as well as agonist and antagonist of functional 3D β islet cells in vitro. Our results indicate the presence of almost 65% of insulin producing cells in 3D clusters. The cells were also found to ameliorate hyperglycaemia in STZ (streptozotocin) induced diabetic NOD/SCID (non-obese diabetic/severe combined immunodeficiency) mouse up to 96 days of transplantation. This protocol provides a basis for 3D in vitro generation of long-term in vivo functionally viable islets from hESC.

  11. Glucose transport in human skeletal muscle cells in culture. Stimulation by insulin and metformin.

    PubMed Central

    Sarabia, V; Lam, L; Burdett, E; Leiter, L A; Klip, A

    1992-01-01

    Primary human muscle cell cultures were established and the regulation of glucose transport was investigated. Primary cultures were allowed to proceed to the stage of myotubes through fusion of myoblasts or were used for clonal selection based on fusion potential. In clonally selected cultures, hexose (2-deoxy-glucose) uptake into myotubes was linear within the time of study and inhibitable by cytochalasin B (IC50 = 400 nM). Cytochalasin B photolabeled a protein(s) of 45,000-50,000 D in a D-glucose-protectable manner, suggesting identity with the glucose transporters. In the myotube stage, the cells expressed both the GLUT1 and GLUT4 glucose transporter protein isoforms at an average molar ratio of 7:1. Preincubation in media of increasing glucose concentrations (range 5-25 mM) progressively decreased the rate of 2-deoxyglucose uptake. Insulin elevated 2-deoxyglucose uptake in a dose-dependent manner, with half maximal stimulation achieved at 3.5 nM. Insulin also stimulated the transport of the nonmetabolizable hexose 3-O-methylglucose, as well as the activity of glycogen synthase, responsible for nonoxidative glucose metabolism. The oral antihyperglycemic drug metformin stimulated the cytochalasin B-sensitive component of both 2-deoxyglucose and 3-O-methylglucose uptake. Maximal stimulation was observed at 8 h of exposure to 50 microM metformin, and this effect was not prevented by incubation with the protein-synthesis inhibitor cycloheximide. The relative effect of metformin was higher in cells incubated in 25 mM glucose than in 5 mM glucose, consistent with its selective action in hyperglycemic conditions in vivo. Metformin (50 microM for 24 h) was more effective than insulin (1 microM for 1 h) in stimulating hexose uptake and the hormone was effective on top of the stimulation caused by the biguanide, suggesting independent mechanisms of action. Images PMID:1401073

  12. Activated α2-Macroglobulin Binding to Human Prostate Cancer Cells Triggers Insulin-like Responses

    PubMed Central

    Misra, Uma Kant; Pizzo, Salvatore Vincent

    2015-01-01

    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-[14C]glucose or 1-[14C]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 [14CH3]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. PMID:25720493

  13. Crystallographic evidence for dual coordination around zinc in the T3R3 human insulin hexamer.

    PubMed

    Ciszak, E; Smith, G D

    1994-02-15

    The T3R3 human insulin hexamer (T and R referring to extended and alpha-helical conformations, respectively, of the first eight residues of the B-chain), complexed to two zinc ions, crystallizes in space group R3 with hexagonal cell constants a = 80.64 and c = 37.78 A. The structure has been refined to a residual of 0.172 using 9225 independent data to 1.6-A resolution. The asymmetric unit consists of a TR dimer, and the insulin hexamer is generated by the action of the crystallographic 3-fold axis. The conformation of one insulin trimer is nearly identical to that of the T6 hexamer, while the other trimer approximates that of the R6 hexamer, except for the three N-terminal B-chain residues that adopt an extended rather than an alpha-helical conformation. Each of the two zinc ions, which lie on the crystallographic 3-fold axis and exhibit two different, disordered coordination geometries, is coordinated by the imidazole groups of three symmetry-related B10 histidine residues. The coordination sphere of the zinc in the T3 trimer is either tetrahedral, with the fourth site filled by a chloride ion, or octahedral, completed by three water molecules. The coordination of the zinc in the 12-A narrow channel in the R3 trimer is tetrahedral, with either a second chloride ion or a water molecule completing the coordination sphere. The putative off-axial zinc binding sites that result from the T-->R transition of monomer II do not contain zinc ion, but instead are filled with clusters of ordered water molecules.(ABSTRACT TRUNCATED AT 250 WORDS)

  14. Ancestral genomic duplication of the insulin gene in tilapia: An analysis of possible implications for clinical islet xenotransplantation using donor islets from transgenic tilapia expressing a humanized insulin gene.

    PubMed

    Hrytsenko, Olga; Pohajdak, Bill; Wright, James R

    2016-07-01

    Tilapia, a teleost fish, have multiple large anatomically discrete islets which are easy to harvest, and when transplanted into diabetic murine recipients, provide normoglycemia and mammalian-like glucose tolerance profiles. Tilapia insulin differs structurally from human insulin which could preclude their use as islet donors for xenotransplantation. Therefore, we produced transgenic tilapia with islets expressing a humanized insulin gene. It is now known that fish genomes may possess an ancestral duplication and so tilapia may have a second insulin gene. Therefore, we cloned, sequenced, and characterized the tilapia insulin 2 transcript and found that its expression is negligible in islets, is not islet-specific, and would not likely need to be silenced in our transgenic fish. PMID:27222321

  15. Preclinical and first-in-human phase I studies of KW-2450, an oral tyrosine kinase inhibitor with insulin-like growth factor receptor-1/insulin receptor selectivity.

    PubMed

    Schwartz, Gary K; Dickson, Mark A; LoRusso, Patricia M; Sausville, Edward A; Maekawa, Yoshimi; Watanabe, Yasuo; Kashima, Naomi; Nakashima, Daisuke; Akinaga, Shiro

    2016-04-01

    Numerous solid tumors overexpress or have excessively activated insulin-like growth factor receptor-1 (IGF-1R). We summarize preclinical studies and the first-in-human study of KW-2450, an oral tyrosine kinase inhibitor with IGF-1R and insulin receptor (IR) inhibitory activity. Preclinical activity of KW-2450 was evaluated in various in vitro and in vivo models. It was then evaluated in a phase I clinical trial in 13 patients with advanced solid tumors (NCT00921336). In vitro, KW-2450 inhibited human IGF-1R and IR kinases (IC50 7.39 and 5.64 nmol/L, respectively) and the growth of various human malignant cell lines. KW-2450 40 mg/kg showed modest growth inhibitory activity and inhibited IGF-1-induced signal transduction in the murine HT-29/GFP colon carcinoma xenograft model. The maximum tolerated dose of KW-2450 was 37.5 mg once daily continuously; dose-limiting toxicity occurred in two of six patients at 50 mg/day (both grade 3 hyperglycemia) and in one of seven patients at 37.5 mg/day (grade 3 rash). Four of 10 evaluable patients showed stable disease. Single-agent KW-2450 was associated with modest antitumor activity in heavily pretreated patients with solid tumors and is being further investigated in combination therapy with lapatinib/letrozole in patients with human epidermal growth factor receptor 2-postive metastatic breast cancer. PMID:26850678

  16. [Relationship between PTEN mutations and protein kinase B phosphorylation caused by insulin or recombinant human epidermal growth factor stimulation].

    PubMed

    Zhong, Hailan; Hu, Xianfu; Lin, Jianhua

    2016-08-01

    Objective To study the effect of phosphatase and tensin homolog deleted on chromosome 10 (PTEN) mutations on protein kinase B (Akt) phosphorylation of CNE-1 nasopharyngeal carcinoma cell line. Methods CNE-1 cells were cultured in RPMI1640 medium containing 100 mL/L fetal calf serum, and then transfected with wild-type PTEN (wtPTEN), mutant PTEN C124S and mutant PTEN G129E plasmid separately. After overnight serum starvation, the cells were stimulated with 0.15 IU/mL insulin or 0.3 μg/mL recombinant human epidermal growth factor (rhEGF). At last, Akt phosphorylation was evaluated by Western blotting. Results Insulin or rhEGF stimulation led to Akt activation in CNE-1 cells. The wtPTEN inhibited insulin- or rhEGF-stimulated phosphorylation of Akt. PTEN C124S mutant activated insulin-stimulated phosphorylation of Akt, but not rhEGF-stimulated phosphorylation of Akt. PTEN G129E mutant inhibited insulin-stimulated phosphorylation of Akt. Conclusion The wtPTEN inhibited insulin- or rhEGF-stimulated phosphorylation of Akt, while PTEN C124S and G129E mutants failed to activate the phosphorylation of Akt consistently. This suggested PTEN mutations might not be correlated with activated Akt.

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

    PubMed Central

    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

    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. PMID:24818157

  18. Mutation of tyrosine-141 inhibits insulin-promoted tyrosine phosphorylation and increased responsiveness of the human beta 2-adrenergic receptor.

    PubMed Central

    Valiquette, M; Parent, S; Loisel, T P; Bouvier, M

    1995-01-01

    The ability of insulin to promote phosphorylation of the human beta 2-adrenergic receptor (beta 2AR) was assessed in Chinese hamster fibroblasts transfected with beta 2AR cDNA. Phosphotyrosine residues were detected in purified beta 2AR using a polyclonal anti-phosphotyrosine antibody and by phosphoamino acid analysis following metabolic labelling with inorganic 32P. Treatment of the cells with insulin induced a 2.4-fold increase in the phosphotyrosine content of the receptor. The insulin-promoted phosphorylation of the beta 2AR was accompanied by an increase in the beta-adrenergic-stimulated adenyl cyclase activity. Substitution of a phenylalanine residue for tyrosine-141 completely prevented both the increased tyrosine phosphorylation and the enhanced responsiveness of the beta 2AR promoted by insulin treatment. Mutation of three other tyrosines located in the cytoplasmic domain of the receptor, tyrosine-366, tyrosine-350 and tyrosine-354, did not abolish the insulin-promoted tyrosine phosphorylation. Taken together, these results suggest that insulin promotes phosphorylation of the beta 2AR on tyrosine-141 and that such phosphorylation leads to a supersensitization of the receptor. Images PMID:8521811

  19. Enhanced bioavailability of subcutaneously injected insulin coadministered with collagen in rats and humans

    SciTech Connect

    Hori, R.; Komada, F.; Iwakawa, S.; Seino, Y.; Okumura, K. )

    1989-09-01

    The present study was undertaken to develop an agent that stabilizes insulin injected subcutaneously. {sup 125}I-Porcine insulin with 0.2 U/kg unlabeled porcine insulin was subcutaneously injected with or without collagen in the rat under the depilated skin of the back. At various times, the radioactivity in subcutaneous tissue was assayed for insulin and its metabolites by gel filtration. The degradation and absorption rate constants of insulin at the subcutaneous injection site were estimated according to a one-compartment model. The degradation rate constant of insulin in the presence of collagen at the injection site was less than half of the control rate. The inhibition was confirmed by increases in the immunoreactive insulin plasma levels and the hypoglycemic effect in rats and healthy volunteers. We postulate that collagen prevents insulin from being degraded by inhibiting proteolytic enzymes, mainly collagenase-like peptidase, in subcutaneous tissue.

  20. Effects of 4 Weeks Recombinant Human Growth Hormone Administration on Insulin Resistance of Skeletal Muscle in Rats

    PubMed Central

    Park, Mi Jung; Jung, Su Ryun; Jung, Hyun Lyung; Craig, Bruce W.; Lee, Chong-Do

    2008-01-01

    Purpose Effect of recombinant human growth hormone (rhGH) administration on lipid storage, and its subsequent effect on insulin sensitivity have not yet been adequately examined. Thus, we investigated the effects of rhGH treatment on muscle triglyceride (TG) and ceramide content, and insulin sensitivity after 4 weeks of rhGH administration in rats. Materials and Methods Fourteen rats were randomly assigned to two groups: rhGH injection group (GH, n = 7) and saline injection group (CON, n = 7). GH received rhGH by subcutaneous injections (130 µg·kg-1·day-1, 6 days·week-1) for 4 weeks, while CON received saline injections that were equivalent in volume to GH group. Intramuscular TG and ceramide content and hepatic TG content were measured. To determine insulin sesitivity, oral glucose tolerance test (OGTT) and muscle incubation for glucose transport rate were performed in rats, and used as indicators of insulin sensitivity. We also examined plasm lipid profiles. Results After 4 weeks of rhGH treatment, the GH group had higher muscle and liver TG contents than the CON (p < 0.05). Ceramide content in GH was significantly greater than that in CON (p < 0.05). GH also had higher plasma levels of FFA (p < 0.05), glucose and insulin responses during OGTT (p < 0.05), and lower glucose transport rates in submaximal insulin concentration (p < 0.05) as compared with CON. Results indicate that rhGH treatment is associated with insulin resistance in rats. Conclusion rhGH treatment elevated muscle TG and ceramide content, and hepatic TG content. Thus, elevation of these compounde by rhGH treatment could contribute to the development of insulin resistance in rats. PMID:19108026

  1. Clinical utility of insulin and insulin analogs

    PubMed Central

    Sanlioglu, Ahter D.; Altunbas, Hasan Ali; Balci, Mustafa Kemal; Griffith, Thomas S.; Sanlioglu, Salih

    2013-01-01

    Diabetes is a pandemic disease characterized by autoimmune, genetic and metabolic abnormalities. While insulin deficiency manifested as hyperglycemia is a common sequel of both Type-1 and Type-2 diabetes (T1DM and T2DM), it does not result from a single genetic defect—rather insulin deficiency results from the functional loss of pancreatic β cells due to multifactorial mechanisms. Since pancreatic β cells of patients with T1DM are destroyed by autoimmune reaction, these patients require daily insulin injections. Insulin resistance followed by β cell dysfunction and β cell loss is the characteristics of T2DM. Therefore, most patients with T2DM will require insulin treatment due to eventual loss of insulin secretion. Despite the evidence of early insulin treatment lowering macrovascular (coronary artery disease, peripheral arterial disease and stroke) and microvascular (diabetic nephropathy, neuropathy and retinopathy) complications of T2DM, controversy exists among physicians on how to initiate and intensify insulin therapy. The slow acting nature of regular human insulin makes its use ineffective in counteracting postprandial hyperglycemia. Instead, recombinant insulin analogs have been generated with a variable degree of specificity and action. Due to the metabolic variability among individuals, optimum blood glucose management is a formidable task to accomplish despite the presence of novel insulin analogs. In this article, we present a recent update on insulin analog structure and function with an overview of the evidence on the various insulin regimens clinically used to treat diabetes. PMID:23584214

  2. [Inhaled insulin, new perspective for insulin therapy].

    PubMed

    Radermecker, R P; Sélam, J L

    2005-01-01

    Since the discovery of insulin and its use in diabetes care, patients, physicians and nurses dream of another way of insulin administration than the subcutaneous injections actually used. Different types of insulin administration have been evaluated and, particularly, that using the pulmonary route. The use of this alternative method to deliver insulin may result in improved patient compliance, facilitate intensified therapies and avoid the delay of initiating insulin administration because patient's reluctance. The different insulin pulmonary delivering devices actually studied will be presented. Preliminary data comparing this way of administration and the subcutaneous injection of human regular insulin are good, but sufficient data comparing inhaled insulin with the new short-acting insulin analogues are not yet available. Among various difficulties of the pulmonary insulin delivery, the finding of an effective promoter, capable of increasing the bioavailability of insulin, is a crucial issue. The cost of such insulin administration might also be a problem. Finally, careful studies concerning the safety of this kind of administration, particularly potential long-term pulmonary toxicity, are mandatory. Nevertheless, inhaled insulin is an attractive topic in which most important pharmaceutical companies are currently involved.

  3. Investigation of molecular interaction between cefpodoxime acid and human mixtard insulin by ultrasonic and spectral methods.

    PubMed

    Ganesh, T; Kannappan, V; Mohamed Kamil, M G; Kumar, R

    2016-09-10

    This paper deals with the extensive investigation of molecular interaction between third generation cephalosporin antibiotic, Cefpodoxime Acid (CA) and Human Mixtard Insulin (HMI) in an aqueous medium through ultrasonic, dilute solution viscometric (DSV) and spectral [UV-vis, Attenuated total reflection (ATR)-FT IR] methods at various blend compositions of the drug and insulin at three different (303K, 310K and 313K) temperatures. This is an attempt to unravel the possibility of drug induced hypoglycemic effect. The existence of solute-solute interaction in aqueous solutions of CA and HMI is established from the variation of ultrasonic velocity and other acoustical parameters with blend composition. DSV method is used to confirm the range of blend composition at which the molecular interaction is significant. The conclusions drawn from ultrasonic and DSV methods are further established by the UV-vis and ATR- FT IR spectral studies of ternary mixtures at different blend compositions. Further, the existing interactions suggest the possibility of cefpodoxime acid induced hypoglycemia which is discussed based on the structural aspects of the two components.

  4. MicroRNA-223 Expression is Upregulated in Insulin Resistant Human Adipose Tissue.

    PubMed

    Chuang, Tung-Yueh; Wu, Hsiao-Li; Chen, Chen-Chun; Gamboa, Gloria Mabel; Layman, Lawrence C; Diamond, Michael P; Azziz, Ricardo; Chen, Yen-Hao

    2015-01-01

    MicroRNAs (miRNAs) are short noncoding RNAs involved in posttranscriptional regulation of gene expression and influence many cellular functions including glucose and lipid metabolism. We previously reported that adipose tissue (AT) from women with polycystic ovary syndrome (PCOS) or controls with insulin resistance (IR) revealed a differentially expressed microRNA (miRNA) profile, including upregulated miR-93 in PCOS patients and in non-PCOS women with IR. Overexpressed miR-93 directly inhibited glucose transporter isoform 4 (GLUT4) expression, thereby influencing glucose metabolism. We have now studied the role of miR-223, which is also abnormally expressed in the AT of IR subjects. Our data indicates that miR-223 is significantly overexpressed in the AT of IR women, regardless of whether they had PCOS or not. miR-223 expression in AT was positively correlated with HOMA-IR. Unlike what is reported in cardiomyocytes, overexpression of miR-223 in human differentiated adipocytes was associated with a reduction in GLUT4 protein content and insulin-stimulated glucose uptake. In addition, our data suggests miR-223 regulates GLUT4 expression by direct binding to its 3' untranslated region (3'UTR). In conclusion, in AT miR-223 is an IR-related miRNA that may serve as a potential therapeutic target for the treatment of IR-related disorders. PMID:26273679

  5. The human insulin receptor mRNA contains a functional internal ribosome entry segment

    PubMed Central

    Spriggs, Keith A.; Cobbold, Laura C.; Ridley, Simon H.; Coldwell, Mark; Bottley, Andrew; Bushell, Martin; Willis, Anne E.; Siddle, Kenneth

    2009-01-01

    Regulation of mRNA translation is an important mechanism determining the level of expression of proteins in eukaryotic cells. Translation is most commonly initiated by cap-dependent scanning, but many eukaryotic mRNAs contain internal ribosome entry segments (IRESs), providing an alternative means of initiation capable of independent regulation. Here, we show by using dicistronic luciferase reporter vectors that the 5′-UTR of the mRNA encoding human insulin receptor (hIR) contains a functional IRES. RNAi-mediated knockdown showed that the protein PTB was required for maximum IRES activity. Electrophoretic mobility shift assays confirmed that PTB1, PTB2 and nPTB, but not unr or PTB4, bound to hIR mRNA, and deletion mapping implicated a CCU motif 448 nt upstream of the initiator AUG in PTB binding. The IR-IRES was functional in a number of cell lines, and most active in cells of neuronal origin, as assessed by luciferase reporter assays. The IRES was more active in confluent than sub-confluent cells, but activity did not change during differentiation of 3T3-L1 fibroblasts to adipocytes. IRES activity was stimulated by insulin in sub-confluent cells. The IRES may function to maintain expression of IR protein in tissues such as the brain where mRNA translation by cap-dependent scanning is less effective. PMID:19654240

  6. LEM-CF Premixed Tool Kit

    SciTech Connect

    2015-01-19

    The purpose of LEM-CF Premixed Tool Kit is to process premixed flame simulation data from the LEM-CF solver (https://fileshare.craft-tech.com/clusters/view/lem-cf) into a large-eddy simulation (LES) subgrid model database. These databases may be used with a user-defined-function (UDF) that is included in the Tool Kit. The subgrid model UDF may be used with the ANSYS FLUENT flow solver or other commercial flow solvers.

  7. Human ketone body production and utilization studied using tracer techniques: Regulation by free fatty acids, insulin, catecholamines, and thyroid hormones

    SciTech Connect

    Keller, U.; Lustenberger, M.; Mueller-Brand, J.G.; Gerber, P.P.; Stauffacher, W.

    1989-05-01

    Ketone body concentrations fluctuate markedly during physiological and pathological conditions. Tracer techniques have been developed in recent years to study production, utilization, and the metabolic clearance rate of ketone bodies. This review describes data on the roles of insulin, catecholamines, and thyroid hormones in the regulation of ketone body kinetics. The data indicate that insulin lowers ketone body concentrations by three independent mechanisms: first, it inhibits lipolysis, and thus lowers free fatty acid availability for ketogenesis; second, it restrains ketone body production within the liver; third, it enhances peripheral ketone body utilization. To assess these effects in humans in vivo, experimental models were developed to study insulin effects with controlled concentrations of free fatty acids, insulin, glucagon, and ketone bodies. Presently available data also support an important role of catecholamines in increasing ketone body concentrations. Evidence was presented that norepinephrine increases ketogenesis not only by stimulating lipolysis, and thus releasing free fatty acids, but also by increasing intrahepatic ketogenesis. Thyroid hormone availability was associated with lipolysis and ketogenesis. Ketone body concentrations after an overnight fast were only modestly elevated in hyperthyroidism resulting from increased peripheral ketone body clearance. There was a significant correlation between serum triiodothyronine levels and the ketone body metabolic clearance rate. Thus, ketone body homeostasis in human subjects resulted from the interaction of hormones such as insulin, catecholamines, and thyroid hormones regulating lipolysis, intrahepatic ketogenesis, and peripheral ketone body utilization. 58 references.

  8. Premixed turbulent flame propagation in microgravity

    NASA Technical Reports Server (NTRS)

    Menon, S.; Jagoda, J.; Sujith, R.

    1995-01-01

    To reduce pollutant formation there is, at present, an increased interest in employing premixed fuel/air mixture in combustion devices. It is well known that greater control over local temperature can be achieved with premixed flames and with lean premixed mixtures, significant reduction of pollutants such as NO(x) can be achieved. However, an issue that is still unresolved is the predictability of the flame propagation speed in turbulent premixed mixtures, especially in lean mixtures. Although substantial progress has been made in recent years, there is still no direct verification that flame speeds in turbulent premixed flows are highly predictable in complex flow fields found in realistic combustors. One of the problems associated with experimental verification is the difficulty in obtaining access to all scales of motion in typical high Reynolds number flows, since, such flows contain scales of motion that range from the size of the device to the smallest Kolmogorov scale. The overall objective of this study is to characterize the behavior of turbulent premixed flames at reasonable high Reynolds number, Re(sub L). Of particular interest here is the thin flame limit where the laminar flame thickness is much smaller than the Kolmogorov scale. Thin flames occur in many practical combustion devices and will be numerically studied using a recently developed new formulation that is briefly described.

  9. Monoclonal antibody to the type I insulin-like growth factor (IGF-I) receptor blocks IGF-I receptor-mediated DNA synthesis: clarification of the mitogenic mechanisms of IGF-I and insulin in human skin fibroblasts

    SciTech Connect

    Flier, J.S.; Usher, P.; Moses, A.C.

    1986-02-01

    Insulin and insulin-like growth factor type I (IGF-I) stimulate an overlapping spectrum of biological responses in human skin fibroblasts. Although insulin and IGF-I are known to stimulate the incorporation of (/sup 3/H)thymidine into DNA in these cells, the identify of the receptor(s) that mediates this effect has not been fully clarified. The mouse anti-human IGF-I receptor antibody ..cap alpha..IR-3 binds with specificity to IGF-I but not to insulin receptors in human placental membranes; it also specifically inhibits the binding of /sup 125/I-labeled IGF-I but not /sup 125/I-labeled insulin to suspensions of human skin fibroblasts in a dose-dependent manner. ..cap alpha..IR-3 competitively inhibits IGF-I-mediated stimulation of (/sup 3/H)thymidine incorporation into DNA. This inhibition is dependent on the concentration of ..cap alpha..IR-3 and in the presence of a fixed antibody concentration can be partially overcome by high concentrations of IGF-I. In contrast, at concentrations of < 1 ..mu..g/ml, the effect of insulin to stimulate (/sup 3/H)thymidine incorporation is not inhibited by ..cap alpha..IR-3. However, the incremental effects of higher concentrations (> 1 ..mu..g/ml) of insulin on (/sup 3/H)thymidine incorporation are inhibited by ..cap alpha..IR-3. ..cap alpha..IR-3 is a highly specific antagonist of IGF-I receptor-mediated mitogenesis in human skin fibroblasts. By using this antibody, it is shown directly that insulin can act through the IGF-I receptor to stimulate DNA synthesis but can also activate this effect through the insulin receptor itself.

  10. Preparation of Tyr-C-peptide from genetically altered human insulin presursor

    SciTech Connect

    Huaiyu Sun; Jianguo Tang; Meihao Hu

    1995-12-01

    C-peptide radiommunoassay (C-peptide RIA) is widely used in determination of pancreatic B-cell secretion activity. {sup 125}I labeled Tyr-C-peptide is indispensable in C-peptide RIA kit. Herein we discuss a way of obtaining recombinant Tyr-C-peptide. Arg32Tyr human proinsulin mutant (R32Y-proinsulin) gene was constructed by site-directed mutagenesis and overexpressed in Escherichia coli. Purified R32Y-proinsulin was converted to insulin and Tyr-C-peptide by trypsin and carboxypeptidase B codigestion. Tyr-C-peptide was isolated through reverse-phase HPLC (RP-HPLC) and identified by C-peptide RIA and amino acid analysis. 12 refs., 5 figs., 1 tab.

  11. Crystal structure of human insulin-regulated aminopeptidase with specificity for cyclic peptides.

    PubMed

    Hermans, Stefan J; Ascher, David B; Hancock, Nancy C; Holien, Jessica K; Michell, Belinda J; Chai, Siew Yeen; Morton, Craig J; Parker, Michael W

    2015-02-01

    Insulin-regulated aminopeptidase (IRAP or oxytocinase) is a membrane-bound zinc-metallopeptidase that cleaves neuroactive peptides in the brain and produces memory enhancing effects when inhibited. We have determined the crystal structure of human IRAP revealing a closed, four domain arrangement with a large, mostly buried cavity abutting the active site. The structure reveals that the GAMEN exopeptidase loop adopts a very different conformation from other aminopeptidases, thus explaining IRAP's unique specificity for cyclic peptides such as oxytocin and vasopressin. Computational docking of a series of IRAP-specific cognitive enhancers into the crystal structure provides a molecular basis for their structure-activity relationships and demonstrates that the structure will be a powerful tool in the development of new classes of cognitive enhancers for treating a variety of memory disorders such as Alzheimer's disease.

  12. High-affinity insulin binding to an atypical insulin-like growth factor-I receptor in human breast cancer cells.

    PubMed Central

    Milazzo, G; Yip, C C; Maddux, B A; Vigneri, R; Goldfine, I D

    1992-01-01

    We studied the nature of insulin receptor binding in MCF-7 breast cancer cells. In both intact cells and solubilized receptor preparations, high-affinity insulin binding was seen. However, unlabeled insulin-like growth factor-I (IGF-I) was five-fold more potent in inhibiting 125I-insulin binding than insulin itself. With monoclonal antibodies to the insulin receptor, 30% of 125I-insulin binding was inhibited. In contrast when alpha-IR3, a monoclonal antibody that recognizes typical IGF-I receptor, was employed over 60% of 125I-insulin binding was inhibited. The B29-MAB-125I-insulin photoprobe was then cross-linked to MCF-7 membranes. Cross-linking was inhibited by both unlabeled insulin and IGF-I. Further, the B29-MAB-125I-insulin photoprobe cross-linked to MCF-7 membranes was strongly immunoprecipitated by alpha-IR3. Employing sequential affinity chromatography with insulin-Affi-gel followed by insulin receptor monoclonal antibody agarose, atypical insulin binding activity was separated from insulin receptor binding activity. This atypical receptor had intrinsic tyrosine kinase activity. Both insulin and IGF-I stimulated the phosphorylation of the receptor's beta subunit. In MCF-7 cells both IGF-I and insulin stimulated [3H]thymidine incorporation; alpha-IR3 blocked all of the IGF-I effect but only 50-60% of the insulin effect. This study demonstrates in MCF-7 cells that, in addition to typical insulin and IGF-I receptors, there is another receptor that binds both insulin and IGF-I with high affinity. Images PMID:1311720

  13. CONJUGATED LINOLEIC ACID PROMOTES HUMAN ADIPOCYTE INSULIN RESISTANCE THROUGH NFκB-DEPENDENT CYTOKINE PRODUCTION

    PubMed Central

    Chung1, Soonkyu; Brown2, J. Mark; Provo1, J. Nathan; Hopkins1, Robin; McIntosh1, Michael K.

    2005-01-01

    We previously demonstrated that trans-10, cis-12 conjugated linoleic acid (CLA) reduced the triglyceride (TG) content of human adipocytes by activating mitogen-activated protein kinase kinase/extracellular signal-related kinase (MEK/ERK) signaling via interleukins-6 (IL-6) and 8 (IL-8). However, the upstream mechanism is unknown. Here we show that CLA increased (≥ 6 h) the secretion of IL-6 and IL-8 in cultures containing both differentiated adipocytes and stromal vascular (SV) cells, non-differentiated SV cells, and adipose tissue explants. CLA’s isomer-specific induction of IL-6 and tumor necrosis factor-α (TNF-α) was associated with the activation of nuclear factor κB (NFκB) as evidenced by: 1) phosphorylation of IκBα, IκBα kinase (IKK), and NFκB p65; 2) IκBα degradation; and 3) nuclear translocation of NFκB. Pretreatment with selective NFκB inhibitors and the MEK/ERK inhibitor U0126 blocked CLA-mediated IL-6 gene expression. Trans-10, cis-12 CLA’s suppression of insulin-stimulated glucose uptake at 24 h was associated with decreased total and plasma membrane glucose transporter 4 (Glut4) proteins. Inhibition of NFκB activation or depletion of NFκB by RNA interference using siNFκB p65 attenuated CLA’s suppression of Glut4 and peroxisome proliferator activated receptor gamma (PPARγ) proteins and glucose uptake. Collectively, these data demonstrate for the first time that trans-10, cis-12 CLA promotes NFκB activation and subsequent induction of IL-6 which are, at least in part, responsible for trans-10, cis-12 CLA-mediated suppression of PPARγ target gene expression and insulin sensitivity in mature human adipocytes. PMID:16155293

  14. Personalized intensification of insulin therapy in type 2 diabetes - does a basal-bolus regimen suit all patients?

    PubMed

    Giugliano, D; Sieradzki, J; Stefanski, A; Gentilella, R

    2016-08-01

    Many patients with type 2 diabetes mellitus (T2DM) require insulin therapy. If basal insulin fails to achieve glycemic control, insulin intensification is one possible treatment intensification strategy. We summarized clinical data from randomized clinical trials designed to compare the efficacy and safety of basal-bolus and premixed insulin intensification regimens. We defined a between-group difference of ≥0.3% in end-of-study glycated hemoglobin (HbA1c) as clinically meaningful. A PubMed database search supplemented by author-identified papers yielded 15 trials which met selection criteria: randomized design, patients with T2DM receiving basal-bolus (bolus injection ≤3 times/day) vs. premixed (≤3 injections/day) insulin regimens, primary/major endpoint(s) HbA1c- and/or hypoglycemia-related, and trial duration ≥12 weeks. Glycemic control improved with both basal-bolus and premixed insulin regimens with - in most cases - acceptable levels of weight gain and hypoglycemia. A clinically meaningful difference between regimens in glycemic control was recorded in only four comparisons, all of which favored basal-bolus therapy. The incidence of hypoglycemia was significantly different between regimens in only three comparisons, one of which favored premixed insulin and two basal-bolus therapy. Of the four trials that reported a significant difference between regimens in bodyweight change, two favored basal-bolus therapy and two favored premixed insulin. Thus, on a population level, neither basal-bolus therapy nor premixed insulin showed a consistent advantage in terms of glycemic control, hypoglycemic risk, or bodyweight gain. It is therefore recommended that clinicians should adopt an individualized approach to insulin intensification - taking into account the benefits and risks of each treatment approach and the attitude and preferences of each patient - in the knowledge that both basal-bolus and premixed regimens may be successful.

  15. Glucose Homeostatic Law: Insulin Clearance Predicts the Progression of Glucose Intolerance in Humans

    PubMed Central

    Uda, Shinsuke; Kubota, Hiroyuki; Iwaki, Toshinao; Fukuzawa, Hiroki; Komori, Yasunori; Fujii, Masashi; Toyoshima, Yu; Sakaguchi, Kazuhiko; Ogawa, Wataru; Kuroda, Shinya

    2015-01-01

    Homeostatic control of blood glucose is regulated by a complex feedback loop between glucose and insulin, of which failure leads to diabetes mellitus. However, physiological and pathological nature of the feedback loop is not fully understood. We made a mathematical model of the feedback loop between glucose and insulin using time course of blood glucose and insulin during consecutive hyperglycemic and hyperinsulinemic-euglycemic clamps in 113 subjects with variety of glucose tolerance including normal glucose tolerance (NGT), impaired glucose tolerance (IGT) and type 2 diabetes mellitus (T2DM). We analyzed the correlation of the parameters in the model with the progression of glucose intolerance and the conserved relationship between parameters. The model parameters of insulin sensitivity and insulin secretion significantly declined from NGT to IGT, and from IGT to T2DM, respectively, consistent with previous clinical observations. Importantly, insulin clearance, an insulin degradation rate, significantly declined from NGT, IGT to T2DM along the progression of glucose intolerance in the mathematical model. Insulin clearance was positively correlated with a product of insulin sensitivity and secretion assessed by the clamp analysis or determined with the mathematical model. Insulin clearance was correlated negatively with postprandial glucose at 2h after oral glucose tolerance test. We also inferred a square-law between the rate constant of insulin clearance and a product of rate constants of insulin sensitivity and secretion in the model, which is also conserved among NGT, IGT and T2DM subjects. Insulin clearance shows a conserved relationship with the capacity of glucose disposal among the NGT, IGT and T2DM subjects. The decrease of insulin clearance predicts the progression of glucose intolerance. PMID:26623647

  16. Human Skeletal Muscle Disuse Atrophy: Effects on Muscle Protein Synthesis, Breakdown, and Insulin Resistance—A Qualitative Review

    PubMed Central

    Rudrappa, Supreeth S.; Wilkinson, Daniel J.; Greenhaff, Paul L.; Smith, Kenneth; Idris, Iskandar; Atherton, Philip J.

    2016-01-01

    The ever increasing burden of an aging population and pandemic of metabolic syndrome worldwide demands further understanding of the modifiable risk factors in reducing disability and morbidity associated with these conditions. Disuse skeletal muscle atrophy (sometimes referred to as “simple” atrophy) and insulin resistance are “non-pathological” events resulting from sedentary behavior and periods of enforced immobilization e.g., due to fractures or elective orthopedic surgery. Yet, the processes and drivers regulating disuse atrophy and insulin resistance and the associated molecular events remain unclear—especially in humans. The aim of this review is to present current knowledge of relationships between muscle protein turnover, insulin resistance and muscle atrophy during disuse, principally in humans. Immobilization lowers fasted state muscle protein synthesis (MPS) and induces fed-state “anabolic resistance.” While a lack of dynamic measurements of muscle protein breakdown (MPB) precludes defining a definitive role for MPB in disuse atrophy, some proteolytic “marker” studies (e.g., MPB genes) suggest a potential early elevation. Immobilization also induces muscle insulin resistance (IR). Moreover, the trajectory of muscle atrophy appears to be accelerated in persistent IR states (e.g., Type II diabetes), suggesting IR may contribute to muscle disuse atrophy under these conditions. Nonetheless, the role of differences in insulin sensitivity across distinct muscle groups and its effects on rates of atrophy remains unclear. Multifaceted time-course studies into the collective role of insulin resistance and muscle protein turnover in the setting of disuse muscle atrophy, in humans, are needed to facilitate the development of appropriate countermeasures and efficacious rehabilitation protocols. PMID:27610086

  17. Human Skeletal Muscle Disuse Atrophy: Effects on Muscle Protein Synthesis, Breakdown, and Insulin Resistance-A Qualitative Review.

    PubMed

    Rudrappa, Supreeth S; Wilkinson, Daniel J; Greenhaff, Paul L; Smith, Kenneth; Idris, Iskandar; Atherton, Philip J

    2016-01-01

    The ever increasing burden of an aging population and pandemic of metabolic syndrome worldwide demands further understanding of the modifiable risk factors in reducing disability and morbidity associated with these conditions. Disuse skeletal muscle atrophy (sometimes referred to as "simple" atrophy) and insulin resistance are "non-pathological" events resulting from sedentary behavior and periods of enforced immobilization e.g., due to fractures or elective orthopedic surgery. Yet, the processes and drivers regulating disuse atrophy and insulin resistance and the associated molecular events remain unclear-especially in humans. The aim of this review is to present current knowledge of relationships between muscle protein turnover, insulin resistance and muscle atrophy during disuse, principally in humans. Immobilization lowers fasted state muscle protein synthesis (MPS) and induces fed-state "anabolic resistance." While a lack of dynamic measurements of muscle protein breakdown (MPB) precludes defining a definitive role for MPB in disuse atrophy, some proteolytic "marker" studies (e.g., MPB genes) suggest a potential early elevation. Immobilization also induces muscle insulin resistance (IR). Moreover, the trajectory of muscle atrophy appears to be accelerated in persistent IR states (e.g., Type II diabetes), suggesting IR may contribute to muscle disuse atrophy under these conditions. Nonetheless, the role of differences in insulin sensitivity across distinct muscle groups and its effects on rates of atrophy remains unclear. Multifaceted time-course studies into the collective role of insulin resistance and muscle protein turnover in the setting of disuse muscle atrophy, in humans, are needed to facilitate the development of appropriate countermeasures and efficacious rehabilitation protocols. PMID:27610086

  18. Human Skeletal Muscle Disuse Atrophy: Effects on Muscle Protein Synthesis, Breakdown, and Insulin Resistance—A Qualitative Review

    PubMed Central

    Rudrappa, Supreeth S.; Wilkinson, Daniel J.; Greenhaff, Paul L.; Smith, Kenneth; Idris, Iskandar; Atherton, Philip J.

    2016-01-01

    The ever increasing burden of an aging population and pandemic of metabolic syndrome worldwide demands further understanding of the modifiable risk factors in reducing disability and morbidity associated with these conditions. Disuse skeletal muscle atrophy (sometimes referred to as “simple” atrophy) and insulin resistance are “non-pathological” events resulting from sedentary behavior and periods of enforced immobilization e.g., due to fractures or elective orthopedic surgery. Yet, the processes and drivers regulating disuse atrophy and insulin resistance and the associated molecular events remain unclear—especially in humans. The aim of this review is to present current knowledge of relationships between muscle protein turnover, insulin resistance and muscle atrophy during disuse, principally in humans. Immobilization lowers fasted state muscle protein synthesis (MPS) and induces fed-state “anabolic resistance.” While a lack of dynamic measurements of muscle protein breakdown (MPB) precludes defining a definitive role for MPB in disuse atrophy, some proteolytic “marker” studies (e.g., MPB genes) suggest a potential early elevation. Immobilization also induces muscle insulin resistance (IR). Moreover, the trajectory of muscle atrophy appears to be accelerated in persistent IR states (e.g., Type II diabetes), suggesting IR may contribute to muscle disuse atrophy under these conditions. Nonetheless, the role of differences in insulin sensitivity across distinct muscle groups and its effects on rates of atrophy remains unclear. Multifaceted time-course studies into the collective role of insulin resistance and muscle protein turnover in the setting of disuse muscle atrophy, in humans, are needed to facilitate the development of appropriate countermeasures and efficacious rehabilitation protocols.

  19. The nature of human serum insulin-like activity (ILA): characterization of ILA in serum and serum fractions obtained by acid-ethanol extraction and adsorption chromatography

    PubMed Central

    Poffenbarger, Philip L.; Ensinck, John W.; Hepp, Dieter K.; Williams, Robert H.

    1968-01-01

    Studies were undertaken in an attempt to clarify the apparent heterogeneous nature of human serum insulin-like activity. Methods of preparative zone electrophoresis on Pevikon, acid-ethanol extraction of trichloroacetic acid serum protein precipitates, adsorption chromatography on DEAE-cellulose and Dowex 50, gel filtration chromatography, and insulin antiserum immunoreactivity were used. The results establish the presence of a substance in serum with in vitro biological properties similar to insuln but with different physicochemical properties. The major portion of serum ILA measured by bioassay techniques can be attributed to the effects of this substance. Whereas the in vitro biological effects of this substance on muscle and adipose cells were similar to those of crystalline insulin, the substance is distinguished from insulin by: (1) the failure of insulin antiserum to inhibit its in vitro biological effect; (2) a slower electrophoretic mobility (in the gamma-beta globulin zone); and (3) a larger molecular weight, between 40,000 and 50,000 in these studies. It is similar to insulin since both are soluble in acid-ethanol. The results further indicate that previously described insulin-like activity in gamma-beta globulin preparations, the major portion of total serum insulin activity described in acid-ethanol extracts of serum, “bound” insulin, “atypical” insulin, and antibody nonsuppressible insulin-like activity bioassayed in diluted serum are all one and the same substance. PMID:4170389

  20. Palmitate-induced inflammatory pathways in human adipose microvascular endothelial cells promote monocyte adhesion and impair insulin transcytosis.

    PubMed

    Pillon, Nicolas J; Azizi, Paymon M; Li, Yujin E; Liu, Jun; Wang, Changsen; Chan, Kenny L; Hopperton, Kathryn E; Bazinet, Richard P; Heit, Bryan; Bilan, Philip J; Lee, Warren L; Klip, Amira

    2015-07-01

    Obesity is associated with inflammation and immune cell recruitment to adipose tissue, muscle and intima of atherosclerotic blood vessels. Obesity and hyperlipidemia are also associated with tissue insulin resistance and can compromise insulin delivery to muscle. The muscle/fat microvascular endothelium mediates insulin delivery and facilitates monocyte transmigration, yet its contribution to the consequences of hyperlipidemia is poorly understood. Using primary endothelial cells from human adipose tissue microvasculature (HAMEC), we investigated the effects of physiological levels of fatty acids on endothelial inflammation and function. Expression of cytokines and adhesion molecules was measured by RT-qPCR. Signaling pathways were evaluated by pharmacological manipulation and immunoblotting. Surface expression of adhesion molecules was determined by immunohistochemistry. THP1 monocyte interaction with HAMEC was measured by cell adhesion and migration across transwells. Insulin transcytosis was measured by total internal reflection fluorescence microscopy. Palmitate, but not palmitoleate, elevated the expression of IL-6, IL-8, TLR2 (Toll-like receptor 2), and intercellular adhesion molecule 1 (ICAM-1). HAMEC had markedly low fatty acid uptake and oxidation, and CD36 inhibition did not reverse the palmitate-induced expression of adhesion molecules, suggesting that inflammation did not arise from palmitate uptake/metabolism. Instead, inhibition of TLR4 to NF-κB signaling blunted palmitate-induced ICAM-1 expression. Importantly, palmitate-induced surface expression of ICAM-1 promoted monocyte binding and transmigration. Conversely, palmitate reduced insulin transcytosis, an effect reversed by TLR4 inhibition. In summary, palmitate activates inflammatory pathways in primary microvascular endothelial cells, impairing insulin transport and increasing monocyte transmigration. This behavior may contribute in vivo to reduced tissue insulin action and enhanced tissue

  1. Insulin Glulisine (rDNA origin) Injection

    MedlinePlus

    ... is a short-acting, man-made version of human insulin. Insulin glulisine works by replacing the insulin ... medications for asthma and colds; certain medications for human immunodeficiency virus (HIV) including amprenavir (Agenerase), atazanavir (Reyataz), ...

  2. Engineering of a Novel Simplified Human Insulin-Like Peptide 5 Agonist.

    PubMed

    Patil, Nitin A; Hughes, Richard A; Rosengren, K Johan; Kocan, Martina; Ang, Sheng Yu; Tailhades, Julien; Separovic, Frances; Summers, Roger J; Grosse, Johannes; Wade, John D; Bathgate, Ross A D; Hossain, Mohammed Akhter

    2016-03-10

    Insulin-like peptide 5 (INSL5) has recently been discovered as only the second orexigenic gut hormone after ghrelin. As we have previously reported, INSL5 is extremely difficult to assemble and oxidize into its two-chain three-disulfide structure. The focus of this study was to generate structure-activity relationships (SARs) of INSL5 and use it to develop a potent and simpler INSL5 mimetic with RXFP4 agonist activity. A series of human and mouse INSL5 (hINSL5/mINSL5) analogues were designed and chemically synthesized, resulting in a chimeric INSL5 analogue exhibiting more than 10-fold higher potency (0.35 nM) at human RXFP4 compared with native hINSL5 (4.57 nM). The SAR study also identified a key residue (K(A15)) in the A-chain of mINSL5 that contributes to improved RXFP4 affinity and potency of mINSL5 compared with hINSL5. This knowledge ultimately led us to engineer a minimized hINSL5 mimetic agonist that retains native hINSL5-like RXFP4 affinity and potency at human RXFP4. This minimized analogue was synthesized in 17.5-fold higher yield and in less time compared with hINSL5. PMID:26824523

  3. Efficacy and safety of biphasic insulin aspart and biphasic insulin lispro mix in patients with type 2 diabetes: A review of the literature

    PubMed Central

    Kumar, Ajay

    2016-01-01

    Type 2 diabetes (T2D) represents an escalating burden worldwide, particularly in China and India. Compared with Caucasians, Asian people with diabetes have lower body mass index, increased visceral adiposity, and postprandial glucose (PPG)/insulin resistance. Since postprandial hyperglycemia contributes significantly to total glycemic burden and is associated with heightened cardiovascular risk, targeting PPG early in T2D is paramount. Premixed insulin regimens are widely used in Asia due to their convenience and effectiveness. Data from randomized controlled trials and observational studies comparing efficacy and safety of biphasic insulin aspart 30 (BIAsp 30) with biphasic insulin lispro mix (LM 25/50) and versus other insulin therapies or oral antidiabetic drugs (OADs) in T2D demonstrated that BIAsp 30 and LM 25/50 were associated with similar or greater improvements in glycemic control versus comparator regimens, such as basal–bolus insulin, in insulin-naÏve, and prior insulin users. Studies directly comparing BIAsp 30 and LM 25 provided conflicting glycemic control results. Safety data generally showed increased hypoglycemia and weight gain with premixed insulins versus basal–bolus insulin or OADs. However, large observational trials documented improvements in glycated hemoglobin, PPG, and hypoglycemia with BIAsp 30 in multi-ethnic patient populations. In summary, this literature review demonstrates that premixed insulin regimens are an appropriate and effective treatment choice in T2D. PMID:27186543

  4. Efficacy and safety of biphasic insulin aspart and biphasic insulin lispro mix in patients with type 2 diabetes: A review of the literature.

    PubMed

    Kumar, Ajay

    2016-01-01

    Type 2 diabetes (T2D) represents an escalating burden worldwide, particularly in China and India. Compared with Caucasians, Asian people with diabetes have lower body mass index, increased visceral adiposity, and postprandial glucose (PPG)/insulin resistance. Since postprandial hyperglycemia contributes significantly to total glycemic burden and is associated with heightened cardiovascular risk, targeting PPG early in T2D is paramount. Premixed insulin regimens are widely used in Asia due to their convenience and effectiveness. Data from randomized controlled trials and observational studies comparing efficacy and safety of biphasic insulin aspart 30 (BIAsp 30) with biphasic insulin lispro mix (LM 25/50) and versus other insulin therapies or oral antidiabetic drugs (OADs) in T2D demonstrated that BIAsp 30 and LM 25/50 were associated with similar or greater improvements in glycemic control versus comparator regimens, such as basal-bolus insulin, in insulin-naÏve, and prior insulin users. Studies directly comparing BIAsp 30 and LM 25 provided conflicting glycemic control results. Safety data generally showed increased hypoglycemia and weight gain with premixed insulins versus basal-bolus insulin or OADs. However, large observational trials documented improvements in glycated hemoglobin, PPG, and hypoglycemia with BIAsp 30 in multi-ethnic patient populations. In summary, this literature review demonstrates that premixed insulin regimens are an appropriate and effective treatment choice in T2D. PMID:27186543

  5. Human iPS Cell-Derived Insulin Producing Cells Form Vascularized Organoids under the Kidney Capsules of Diabetic Mice

    PubMed Central

    Raikwar, Sudhanshu P.; Kim, Eun-Mi; Sivitz, William I.; Allamargot, Chantal; Thedens, Daniel R.; Zavazava, Nicholas

    2015-01-01

    Type 1 diabetes (T1D) is caused by autoimmune disease that leads to the destruction of pancreatic β-cells. Transplantation of cadaveric pancreatic organs or pancreatic islets can restore normal physiology. However, there is a chronic shortage of cadaveric organs, limiting the treatment of the majority of patients on the pancreas transplantation waiting list. Here, we hypothesized that human iPS cells can be directly differentiated into insulin producing cells (IPCs) capable of secreting insulin. Using a series of pancreatic growth factors, we successfully generated iPS cells derived IPCs. Furthermore, to investigate the capability of these cells to secrete insulin in vivo, the differentiated cells were transplanted under the kidney capsules of diabetic immunodeficient mice. Serum glucose levels gradually declined to either normal or near normal levels over 150 days, suggesting that the IPCs were secreting insulin. In addition, using MRI, a 3D organoid appeared as a white patch on the transplanted kidneys but not on the control kidneys. These organoids showed neo-vascularization and stained positive for insulin and glucagon. All together, these data show that a pancreatic organ can be created in vivo providing evidence that iPS cells might be a novel option for the treatment of T1D. PMID:25629318

  6. Synergistic bombesin and insulin stimulation of DNA synthesis in human fetal kidney in serum-free culture.

    PubMed

    Brière, N; Chailler, P

    1993-05-01

    The respective influences of growth factors during kidney development can be directly evaluated using the chemically-defined serum-free culture system perfected in our laboratory. Since, in this culture model, conditions are minimal for growth and differentiation, DNA synthesis sharply decreases during the first 48 h. The addition of epidermal growth factor (EGF, 100 ng/ml), insulin (5 micrograms/ml) and transferrin (5 micrograms/ml) significantly restores this important cellular function. The objective of the present study was to determine the influence of bombesin, a potent mitogen, supplemented alone or in combination with insulin, transferrin and/or EGF. Cortical explants of human fetal kidneys (17-20 weeks) were maintained during 5 days in culture. When compared with 5 day controls (L-15 medium only), bombesin generated a maximal though weak effect on DNA synthesis at a concentration of 0.3 nM, corresponding to a stimulation index (SI) of 22%. When combined with either transferrin or EGF, or with transferrin plus EGF, bombesin did not alter the SI of individual factors. Insulin, in turn, greatly increased DNA synthesis (SI = 169%), while bombesin strongly potentiated this effect (SI = 275%). Transferrin also enhanced insulin SI from 169 to 240%. When added as a third factor, bombesin further potentiated the effectiveness (SI = 338%) of the combination insulin plus transferrin. These results indicate that bombesin controls cell proliferation in synergism with other regulators and hence may act as a competence growth factor during nephrogenesis.

  7. Studies of Human Adipose Tissue in Culture III INFLUENCE OF INSULIN AND MEDIUM GLUCOSE CONCENTRATION ON CELLULAR METABOLISM

    PubMed Central

    Smith, Ulf

    1974-01-01

    Explants of human adipose tissue were maintained in culture for 1 wk in different glucose concentrations with or without the addition of insulin. After this period of time the explants were carefully washed and then subjected to short-term incubations in the same glucose concentration and in the absence of insulin. With this experimental design the influence of long-term exposure to insulin and different glucose concentrations on adipose tissue metabolism could be studied. The results of these studies show that an increase in the glucose concentration of the culture medium enhanced the basal as well as the catecholamine-stimulated lipolysis in the short-term incubations. The presence of insulin in the culture medium enhanced the lipolytic process as well. Analogous results were obtained with the cellular rate of glucose conversion to triglycerides in the short-term incubations. The stimulating effects of insulin and glucose were most pronounced in the larger adipose cells possibly due to their enlarged surface areas. The data suggest that the metabolism of adipose tissue as revealed by short-term studies may be profoundly influenced by the antecedent biochemical environment. PMID:4808648

  8. Restriction fragment length polymorphism of the human insulin gene region among type II diabetic Mexican-Americans and Tunisians.

    PubMed

    Frazier, M L; Ferrell, R E; Arem, R; Chamakhi, S; Field, J B

    1986-06-01

    The human insulin gene is flanked by a polymorphic locus that is located approximately 500 base pairs (bp) from the 5' end of the point where transcription begins (Bell et al. 1981; Bell et al, 1982). Its occurrence is due to an insertion-deletion region which gives rise to two major classes of alleles: those containing small insertions of 0-600 bp and those containing larger insertions of 1,600-2,200 bp (Owerbach and Nerup, 1982). Insertions of 600-1,600 bp are rare (Rotwein et al., 1983). The larger insertions have previously been reported to be associated with type 2 diabetes (Owerbach and Nerup, 1982). We have conducted studies on a Mexican-American population in Starr County, Texas (98% Mexican-American) and a Tunisian population in Tunis, Tunisia, to determine if the frequency distribution of these classes of insulin gene alleles are similar to the previously reported frequency distributions and if any of the classes of alleles are associated with type 2 diabetes in these populations. We conclude that none of the classes of insulin gene alleles are associated with type II diabetes among Mexican-Americans or Tunisians, and that the frequency distributions of the insulin gene alleles do not vary significantly between the Tunisians, Mexican-Americans, or the aggregate data resulting from combining the insulin gene frequencies of several of the populations described thus far (Bell et al., 1984). PMID:3016846

  9. Activation of IGF-1 and insulin signaling pathways ameliorate mitochondrial function and energy metabolism in Huntington's Disease human lymphoblasts.

    PubMed

    Naia, Luana; Ferreira, I Luísa; Cunha-Oliveira, Teresa; Duarte, Ana I; Ribeiro, Márcio; Rosenstock, Tatiana R; Laço, Mário N; Ribeiro, Maria J; Oliveira, Catarina R; Saudou, Frédéric; Humbert, Sandrine; Rego, A Cristina

    2015-02-01

    Huntington's disease (HD) is an inherited neurodegenerative disease caused by a polyglutamine repeat expansion in the huntingtin protein. Mitochondrial dysfunction associated with energy failure plays an important role in this untreated pathology. In the present work, we used lymphoblasts obtained from HD patients or unaffected parentally related individuals to study the protective role of insulin-like growth factor 1 (IGF-1) versus insulin (at low nM) on signaling and metabolic and mitochondrial functions. Deregulation of intracellular signaling pathways linked to activation of insulin and IGF-1 receptors (IR,IGF-1R), Akt, and ERK was largely restored by IGF-1 and, at a less extent, by insulin in HD human lymphoblasts. Importantly, both neurotrophic factors stimulated huntingtin phosphorylation at Ser421 in HD cells. IGF-1 and insulin also rescued energy levels in HD peripheral cells, as evaluated by increased ATP and phosphocreatine, and decreased lactate levels. Moreover, IGF-1 effectively ameliorated O2 consumption and mitochondrial membrane potential (Δψm) in HD lymphoblasts, which occurred concomitantly with increased levels of cytochrome c. Indeed, constitutive phosphorylation of huntingtin was able to restore the Δψm in lymphoblasts expressing an abnormal expansion of polyglutamines. HD lymphoblasts further exhibited increased intracellular Ca(2+) levels before and after exposure to hydrogen peroxide (H2O2), and decreased mitochondrial Ca(2+) accumulation, being the later recovered by IGF-1 and insulin in HD lymphoblasts pre-exposed to H2O2. In summary, the data support an important role for IR/IGF-1R mediated activation of signaling pathways and improved mitochondrial and metabolic function in HD human lymphoblasts.

  10. Insulin-Producing Cells From Adult Human Bone Marrow Mesenchymal Stem Cells Control Streptozotocin-Induced Diabetes In Nude Mice

    PubMed Central

    Gabr, Mahmoud M.; Zakaria, Mahmoud M.; Refaie, Ayman F.; Ismail, Amani M.; Abou-El-Mahasen, Mona A.; Ashamallah, Sylvia A.; Khater, Sherry M.; El-Halawani, Sawsan M.; Ibrahim, Rana Y.; Uin, Gan Shu; Kloc, Malgorzata; Calne, Roy Y.; Ghoneim, Mohamed A.

    2013-01-01

    Harvesting, expansion and directed differentiation of human bone marrow-derived mesenchymal stem cells (BM-MSCs) could provide an autologous source of surrogate β-cells that would alleviate the limitations of availability and/or allogenic rejection following pancreatic or islet transplantation. Bone marrow cells were obtained from three adult type 2 diabetic volunteers and 3 non-diabetic donors. After 3 days in culture, adherent MSCs were expanded for 2 passages. At passage 3, differentiation was carried out in a 3-staged procedure. Cells were cultured in a glucose-rich medium containing several activation and growth factors. Cells were evaluated in-vitro by flow cytometry, immunolabelling, Rt-PCR and human insulin and c-peptide release in responses to increasing glucose concentrations. One thousand cell-clusters were inserted under the renal capsule of diabetic nude mice followed by monitoring of their diabetic status. At the end of differentiation, ~5–10% of cells were immunofluorescent for insulin, c-peptide or glucagon; insulin and c-peptide were co-expressed. Nanogold immunolabelling for electron microscopy demonstrated the presence of c-peptide in the rough endoplasmic reticulum. Insulin-producing cells (IPCs) expressed transcription factors and genes of pancreatic hormones similar to those expressed by pancreatic islets. There was a stepwise increase in human insulin and c-peptide release by IPCs in response to increasing glucose concentrations. Transplantation of IPCs into nude diabetic mice resulted in control of their diabetic status for 3 months. The sera of IPC-transplanted mice contained human insulin and c-peptide but negligible levels of mouse insulin. When the IPCs-bearing kidneys were removed, rapid return of diabetic state was noted. BM-MSCs from diabetic and non-diabetic human subjects could be differentiated without genetic manipulation to form IPCs which, when transplanted, could maintain euglycaemia in diabetic mice for 3 months

  11. Insulin secretion and interleukin-1β dependent mechanisms in human diabetes remission after metabolic surgery.

    PubMed

    Chen, Chih-Yen; Lee, Wei-Jei; Asakawa, A; Fujitsuka, N; Chong, Keong; Chen, Shu-Chun; Lee, Shou-Dong; Inui, A

    2013-01-01

    To compare endocrine, metabolic, and inflammatory changes induced by gastric bypass (GB) and sleeve gastrectomy (SG) in patients with type 2 diabetes mellitus (T2DM), and to investigate the mechanisms of success after metabolic surgery. Sixteen GB and 16 SG patients were followed up before and at 1 year after surgery. The 75-g oral glucose tolerance test (OGTT) was performed before and after surgery. Glucose homeostasis, serum interleukin-1β, plasma gut hormones and adipokines, and the United Kingdom Prospective Diabetes Study (UKPDS) ten-year cardiovascular risks were evaluated. The diabetes remission rate was significantly higher in GB than SG. Changes in the area under the curve (AUC) for glucose were greater in those with complete and partial remission after GB and remitters after SG than non-remitters after SG, whereas changes in AUC for C-peptide were higher in complete and partial remitters after GB than non-remitters after SG. Insulinogenic index was enhanced and serum interleukin-1β was reduced in complete remitters after GB and remitters after SG. Logistic regression analysis confirmed that insulinogenic index and interleukin-1β, not insulin resistance, were the factors determining the success of diabetes remission after metabolic surgeries. GB and SG significantly reduced the ten-year risk of coronary heart disease and fatal coronary heart disease in T2DM patients after surgery, while GB had the additional benefit of reduced stroke risk. Human diabetes remission after metabolic surgery is through insulin secretion and interleukin-1β dependent mechanisms. GB is superior to SG in cardiocerebral risk reduction in Asian non-morbidly obese, not well-controlled T2DM patients.

  12. Posttranslational regulation of insulin-like growth factor binding protein-4 in normal and transformed human fibroblasts. Insulin-like growth factor dependence and biological studies.

    PubMed Central

    Conover, C A; Kiefer, M C; Zapf, J

    1993-01-01

    Insulin-like growth factor binding protein-4 (IGFBP-4) is a 24-26-kD protein expressed by a variety of cell types in vivo and in vitro. Treatment of normal adult human fibroblasts with 10 nM insulin-like growth factor II (IGF-II) for 24 h resulted in an 85% decrease in endogenous IGFBP-4, as assessed by Western ligand blot analysis of the conditioned medium. Incubation of human fibroblast-conditioned medium (HFCM) with IGF-II under cell-free conditions led to a similar loss of IGFBP-4. This posttranslationally regulated decrease in IGFBP-4 appeared to be due to a protease in HFCM: (a) It could be prevented with specific protease inhibitors or incubation at 4 degrees C; (b) proteolysis of recombinant human (rh) IGFBP-4 required HFCM; (c) immunoblotting and radiolabeling confirmed cleavage of IGFBP-4 into 18- and 14-kD IGFBP-4 fragments. The protease was specific for IGFBP-4, and was strictly dependent on IGFs for activation. IGF-II was the most effective of the natural and mutant IGFs tested, inducing complete hydrolysis of rhIGFBP-4 at a molar ratio of 0.25:1 (IGF/IGFBP-4). Simian virus 40-transformed adult human fibroblasts also expressed IGFBP-4 and IGFBP-4 protease, as well as an inhibitor of IGFBP-4 proteolysis. In biological studies, intact rhIGFBP-4 potently inhibited IGF-I-stimulated [3H]aminoisobutyric acid uptake, whereas proteolyzed rhIGFBP-4 had no inhibitory effect. In conclusion, these data provide evidence for a novel IGF-dependent IGFBP-4-specific protease that modifies IGFBP-4 structure and function, and indicate a preferential role for IGF-II in its activation. Posttranslational regulation of IGFBP-4 may provide a means for cooperative control of local cell growth by IGF-I and IGF-II. Images PMID:7680662

  13. Insulin and Insulin Resistance

    PubMed Central

    2005-01-01

    As obesity and diabetes reach epidemic proportions in the developed world, the role of insulin resistance and its consequences are gaining prominence. Understanding the role of insulin in wide-ranging physiological processes and the influences on its synthesis and secretion, alongside its actions from the molecular to the whole body level, has significant implications for much chronic disease seen in Westernised populations today. This review provides an overview of insulin, its history, structure, synthesis, secretion, actions and interactions followed by a discussion of insulin resistance and its associated clinical manifestations. Specific areas of focus include the actions of insulin and manifestations of insulin resistance in specific organs and tissues, physiological, environmental and pharmacological influences on insulin action and insulin resistance as well as clinical syndromes associated with insulin resistance. Clinical and functional measures of insulin resistance are also covered. Despite our incomplete understanding of the complex biological mechanisms of insulin action and insulin resistance, we need to consider the dramatic social changes of the past century with respect to physical activity, diet, work, socialisation and sleep patterns. Rapid globalisation, urbanisation and industrialisation have spawned epidemics of obesity, diabetes and their attendant co-morbidities, as physical inactivity and dietary imbalance unmask latent predisposing genetic traits. PMID:16278749

  14. Turbulent Premixed Flames in Microgravity

    NASA Technical Reports Server (NTRS)

    Menon, Suresh

    1996-01-01

    The experimental cold-flow facility is now full operational and is currently being used to obtain baseline turbulence data in a Couette flow. The baseline turbulence data is necessary to confirm the capability of the chosen device to generate and maintain the required turbulence intensity. Subsequent reacting flow studies will assume that a similar turbulent flow field exists ahead of the premixed flame. Some modifications and refinements had to be made to enable accurate measurements. It consists of two rollers, one (driven by a motor) which drives a continuous belt and four smaller rollers used to set the belt spacing and tension to minimize belt flutter. The entire assemble is enclosed in a structure that has the dimensions to enable future drop tower experiments of the hot facility. All critical dimensions are the same as the original plans except for the pulley ratio which has been changed to enable a wider operating regime in terms of the Reynolds number. With the current setup, Reynolds numbers as low as 100 and as high as 14,000 can be achieved. This is because the in-between belt spacing can be varied from 1 cm to 7.6 cm, and the belt speed can be accurately varied from .15 m/sec to 3.1 m/sec.

  15. Design factors for stable lean premix combustion

    SciTech Connect

    Richards, G.A.; Yip, M.J.; Gemmen, R.S.

    1995-10-01

    The Advanced Turbine Systems (ATS) program includes the development of low-emission combustors. Low emissions have already been achieved by premixing fuel and air to avoid the hot gas pockets produced by nozzles without premixing. While the advantages of premixed combustion have been widely recognized, turbine developers using premixed nozzles have experienced repeated problems with combustion oscillations. Left uncontrolled, these oscillations can lead to pressure fluctuations capable of damaging engine hardware. Elimination of such oscillations is often difficult and time consuming - particularly when oscillations are discovered in the last stages of engine development. To address this issue, METC is studying oscillating combustion from lean premixing fuel nozzles. These tests are providing generic information on the mechanisms that contribute to oscillating behavior in gas turbines. METC is also investigating the use of so-called {open_quotes}active{close_quotes} control of combustion oscillations. This technique periodically injects fuel pulses into the combustor to disrupt the oscillating behavior. Recent results on active combustion control are presented in Gemmen et al. (1995) and Richards et al. (1995). This paper describes the status of METC efforts to avoid oscillations through simple design changes.

  16. Characterization of phorbol ester-stimulated serine phosphorylation of the human insulin receptor.

    PubMed Central

    Feener, E P; Shiba, T; Hu, K Q; Wilden, P A; White, M F; King, G L

    1994-01-01

    Phorbol 12-myristate 13-acetate (PMA)-stimulated phosphorylation of the human insulin receptor (IR) was characterized and compared in two cell types of different lineage: normal rat kidney epithelial (NRK) cells and Chinese hamster ovary (CHO) fibroblasts. PMA stimulation increased IR beta-subunit phosphorylation to 252 +/- 43 and 25- +/- 47% (+/- S.D.) of the unstimulated control in NRK and CHO cells respectively. Tryptic phosphopeptide analysis by Tricine/SDS/PAGE revealed significant differences in the PMA-stimulated phosphorylation of the IR in these two cell types. This phosphorylation of the IR was predominantly located in two tryptic phosphopeptides, and these phosphopeptides were absent in an IR mutant truncated by 43 C-terminal amino acids. The major PMA-stimulated tryptic phosphopeptide from in vivo-labelled CHO/IR was immunoprecipitated with an antibody against residues Ser1315 to Lys1329, and this precipitation was blocked with excess unlabelled peptide containing this sequence. Radiosequencing by manual Edman degradation revealed that this tryptic phosphopeptide was phosphorylated at Ser1315. This PMA-stimulated phosphorylation did not inhibit autophosphorylation of the IR in vivo. These results demonstrate that PMA-stimulated phosphorylation of the IR can exhibit significant differences when expressed in different cell types, and that Ser1315 is a major PMA-stimulated phosphorylation site on the human IR. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 PMID:7945263

  17. Regeneration of insulin-producing pancreatic cells using a volatile bioactive compound and human teeth.

    PubMed

    Okada, Mio; Imai, Toshio; Yaegaki, Ken; Ishkitiev, Nikolay; Tanaka, Tomoko

    2014-10-30

    Transplantation of insulin (INS)-secreting cells differentiated in vitro from stem cells promises a safer and easier treatment of severe diabetes mellitus. A volatile bioactive compound, hydrogen sulfide (H2S), promotes cell differentiation; human tooth-pulp stem cells undergo hepatic differentiation. The aim of this study is to develop a novel protocol using H2S to enhance pancreatic differentiation from the CD117(+) cell fraction of human tooth pulp. During the differentiation, the cells were exposed to 0.1 ng ml(-1) H2S. Immunocytochemistry, RT-PCR, determination of INS c-peptide content and flow cytometry of pancreatically related markers were carried out. Expression of WNT and the PI3K/AKT signaling pathway were also determined by PCR array. After differentiation, INS, glucagon (GCG), somatostatin (SST) and pancreatic polypeptide (PPY) were positive when examined by immunofluorescence. INS and GCG were also determined flow-cytometrically. Only the cells expressing INS increased after H2S exposure. The number of cells expressing GCG was significantly decreased. Genes involved in canonical WNT and the WNT/calcium pathways were highly expressed after H2S exposure. H2S accelerated INS synthesis and secretion by regenerated INS-producing cells from human teeth. All signaling pathway functions of the PI3K-AKT pathway were extremely activated by H2S exposure. The matured INS-producing cells originating in human teeth were increased by H2S in order to control blood-glucose level.

  18. Studies of Premixed Laminar and Turbulent Flames at Microgravity

    NASA Technical Reports Server (NTRS)

    Abid, M.; Aung, K.; Ronney, P. D.; Sharif, J. A.; Wu, M.-S.

    1999-01-01

    Several topics relating to combustion limits in premixed flames at reduced gravity have been studied. These topics include: (1) flame balls; (2) numerical simulation of flame ball and planar flame structure and stability; (3) experimental simulation of buoyancy effects in premixed flames using aqueous autocatalytic reactions; and (4) premixed flame propagation in Hele-Shaw cells.

  19. Lipodystrophy in human immunodeficiency virus patients impairs insulin action and induces defects in beta-cell function.

    PubMed

    Andersen, Ove; Haugaard, Steen B; Andersen, Ulrik B; Friis-Møller, Nina; Storgaard, Heidi; Vølund, Aage; Nielsen, Jens Ole; Iversen, Johan; Madsbad, Sten

    2003-10-01

    The pathophysiology of insulin resistance in human immunodeficiency virus (HIV)-associated lipodystrophy syndrome (HALS) is not fully clarified. We investigated 18 men with HALS and 18 HIV-positive males without lipodystrophy (control subjects). Duration and modality of antiretroviral therapy were similar between study groups. A hyperinsulinemic euglycemic clamp showed an impaired glucose disposal rate (GDR) in HALS patients (5.6 v 8.3 mg glucose/min. kg(FFM), P =.0006). As demonstrated by indirect calorimetry, HALS patients showed an impaired nonoxidative glucose metabolism (NOGM, 2.2 v 4.2, P =.006), whereas levels of basal and insulin-stimulated oxidative glucose metabolism (OGM) (2.4 v 2.3, P =.55, and 3.3 v 4.0, P =.064, respectively) were not significantly different between groups. Despite comparable total fat masses, dual energy x-ray absorptiometry (DEXA) scans showed that the percentage of limb fat (ie, peripheral-fat-mass/[peripheral-fat-mass + trunk-fat-mass]. 100%) was reduced in HALS patients (36% v 46%, P =.0002). Multiple linear regression analysis indicated that percentage of limb fat explained 53% of the variability of GDR and 45% of the variability of NOGM in HALS patients. In HALS patients, leg fat mass correlated positively with NOGM (r =.51, P <.05), whereas abdominal fat mass and NOGM did not correlate (P =.91). Analyzing the relationship between first phase insulin secretion and insulin sensitivity, 6 HALS patients compared with none of the control subjects exhibited impaired insulin secretion (P <.05). Our data suggest that fat redistribution independently of antiretroviral therapy is highly related to insulin resistance in HALS patients. Furthermore, in HALS patients, impaired glucose metabolism most likely relates to decreased NOGM and to defects in beta-cell function.

  20. Looking at the carcinogenicity of human insulin analogues via the intrinsic disorder prism

    PubMed Central

    Redwan, Elrashdy M.; Linjawi, Moustafa H.; Uversky, Vladimir N.

    2016-01-01

    Therapeutic insulin, in its native and biosynthetic forms as well as several currently available insulin analogues, continues to be the protein of most interest to researchers. From the time of its discovery to the development of modern insulin analogues, this important therapeutic protein has passed through several stages and product generations. Beside the well-known link between diabetes and cancer risk, the currently used therapeutic insulin analogues raised serious concerns due to their potential roles in cancer initiation and/or progression. It is possible that structural variations in some of the insulin analogues are responsible for the appearance of new oncogenic species with high binding affinity to the insulin-like growth factor 1 (IGF1) receptor. The question we are trying to answer in this work is: are there any specific features of the distribution of intrinsic disorder propensity within the amino acid sequences of insulin analogues that may provide an explanation for the carcinogenicity of the altered insulin protein? PMID:26983499

  1. Flame propagation in partially premixed conditions

    NASA Astrophysics Data System (ADS)

    Ruetsch, G.; Poinsot, T.; Veynante, D.; Trouvé, A.

    1996-11-01

    Turbulent flame propagation is studied under inhomogenously premixed conditions via data from direct numerical simulations. Departures from the premixed case are studied using four different configurations, ranging from one dimensional unsteady flames to turbulent three-dimensional simulations. Simulations are performed in these cases with various values of the mean equivalence ratio, fluctuations about the mean equivlalence ratio, correlation length scales, and probability denisty functions of the mixture composition. Propagation characteristics are described in terms of the flamelet approach, where the the main contribution of partial premixing on flame propagation is due to flame wrinkling relative to modification of the mean flamelet structure. This behavior is consistent over a broad range of conditions, with the exception being extreme departures from stoichiometric conditions where flamability limits are exceeded and flame quenching is observed.

  2. Soot Formation in Laminar Premixed Flames

    NASA Technical Reports Server (NTRS)

    Xu, F.; Krishnan, S. S.; Faeth, G. M.

    1999-01-01

    Soot processes within hydrocarbon-fueled flames affect emissions of pollutant soot, thermal loads on combustors, hazards of unwanted fires and capabilities for computational combustion. In view of these observations, the present study is considering processes of soot formation in both burner-stabilized and freely-propagating laminar premixed flames. These flames are being studied in order to simplify the interpretation of measurements and to enhance computational tractability compared to the diffusion flame environments of greatest interest for soot processes. In addition, earlier studies of soot formation in laminar premixed flames used approximations of soot optical and structure properties that have not been effective during recent evaluations, as well as questionable estimates of flow residence times). The objective of present work was to exploit methods of avoiding these difficulties developed for laminar diffusion flames to study soot growth in laminar premixed flames. The following description of these studies is brief.

  3. Colorimetric determination of selenium in mineral premixes .

    PubMed

    Hurlbut, J A; Burkepile, R G; Geisler, C A; Kijak, P J; Rummel, N G

    1997-01-01

    A method is described for determination of sodium selenite or sodium selenate in mineral-based premixes. It is based on the formation of intense-yellow piazselenol by Se(IV) and 3,3'-diaminobenzidine. Mineral premixes typically contain calcium carbonate as a base material and magnesium carbonate, silicon dioxide, and iron(III) oxide as minor components or additives. In this method, the premix is digested briefly in nitric acid, diluted with water, and filtered to remove any Iron(III) oxide. Ethylenediaminetetraacetic acid and HCl are added to the filtrate, which is heated to near boiling for 1 h to convert any selenate to selenite. After heating, the solution is buffered between pH 2 and 3 with NaOH and formic acid and treated with NH2OH and EDTA; any Se present forms a complex with 3,3'-diaminobenzidine at 60 degrees C. The solution is made basic with NH4OH, and the piazselenol is extracted into toluene. The absorbance of the complex in dried toluene is measured at 420 nm. The method was validated independently by 2 laboratories. Samples analyzed included calcium carbonate fortified with 100, 200, and 300 micrograms Se in the form of sodium selenite or sodium selenate, a calcium carbonate premix containing sodium selenite, a calcium carbonate premix containing sodium selenate, and a commercial premix; 5 replicates of each sample type were analyzed by each laboratory. Average recoveries ranged from 89 to 109% with coefficients of variation from 1.2 to 13.6%. PMID:9241835

  4. Chaos in an imperfectly premixed model combustor

    SciTech Connect

    Kabiraj, Lipika Saurabh, Aditya; Paschereit, Christian O.; Karimi, Nader; Sailor, Anna; Mastorakos, Epaminondas; Dowling, Ann P.

    2015-02-15

    This article reports nonlinear bifurcations observed in a laboratory scale, turbulent combustor operating under imperfectly premixed mode with global equivalence ratio as the control parameter. The results indicate that the dynamics of thermoacoustic instability correspond to quasi-periodic bifurcation to low-dimensional, deterministic chaos, a route that is common to a variety of dissipative nonlinear systems. The results support the recent identification of bifurcation scenarios in a laminar premixed flame combustor (Kabiraj et al., Chaos: Interdiscip. J. Nonlinear Sci. 22, 023129 (2012)) and extend the observation to a practically relevant combustor configuration.

  5. In vitro differentiation of human umbilical cord Wharton’s jelly mesenchymal stromal cells to insulin producing clusters

    PubMed Central

    Nekoei, Seideh Masoomeh; Azarpira, Negar; Sadeghi, Ladan; Kamalifar, Sulmaz

    2015-01-01

    AIM: To investigate the differentiation of human Wharton’s jelly derived mesenchymal stromal cells (WJ-MSCs) to insulin producing clusters (IPC) this study was conducted. METHODS: The umbilical cords samples were collected from full term caesarian section mothers and the WJ-MSCS were cultured from tissue explants in High glucose-Dulbecco’s Modified Eagle Medium (H-DMEM); H-DMEM supplemented with 10% fetal bovine serum (FBS) and antibiotics. The expression of CD90, CD44, CD105, CD34 and CD133 as well as osteogenic and adipogenic differentiation of cells in appropriate medium were also evaluated. The cells were differentiated toward IPC with changing the culture medium and adding the small molecules such as nicotinic acid, epidermal growth factor, and exendin-4 during 3 wk period. The gene expression of PDX1, NGN3, Glut2, insulin was monitored by reveres transcription polymerase chain reaction method. The differentiated clusters were stained with Dithizone (DTZ) which confirms the presence of insulin granules. The insulin challenge test (low and high glucose concentration in Krebs-Ringer HEPES buffer) was also used to evaluate the functional properties of differentiated clusters. RESULTS: WJ-MSCS were positive for mesenchymal surface markers (CD90, CD44, CD105), and negative for CD34 and CD133. The accumulation of lipid vacuoles and deposition of calcium mineral in cells were considered as adipogenic and osteogenic potential of WJ-MSCS. The cells also expressed the transcriptional factors such as Nanog and OCT4. During this three step differentiation, the WJ-MSCS morphology was gradually changed from spindle shaped cells in to epithelioid cells and eventually to three dimensional clusters. The clusters expressed PDX1, NGN3, Glut2, and insulin. The cells became bright red color when stained with DTZ and the insulin secretion was also confirmed. In glucose challenge test a significant increase in insulin secretion from 0.91 ± 0.04 μIu/mL (2.8 mmol/L glucose) to

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

    PubMed Central

    Xin, Ying; Jiang, Xin; Wang, Yishu; Su, Xuejin; Sun, Meiyu; Zhang, Lihong; Tan, Yi; Wintergerst, Kupper A.; Li, Yan; Li, Yulin

    2016-01-01

    Background 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. Methods 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 × 106 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. Results 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. Conclusions 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. PMID:26756576

  7. Novel Small Molecule Glucagon-Like Peptide-1 Receptor Agonist Stimulates Insulin Secretion in Rodents and From Human Islets

    PubMed Central

    Sloop, Kyle W.; Willard, Francis S.; Brenner, Martin B.; Ficorilli, James; Valasek, Kathleen; Showalter, Aaron D.; Farb, Thomas B.; Cao, Julia X.C.; Cox, Amy L.; Michael, M. Dodson; Gutierrez Sanfeliciano, Sonia Maria; Tebbe, Mark J.; Coghlan, Michael J.

    2010-01-01

    OBJECTIVE The clinical effectiveness of parenterally-administered glucagon-like peptide-1 (GLP-1) mimetics to improve glucose control in patients suffering from type 2 diabetes strongly supports discovery pursuits aimed at identifying and developing orally active, small molecule GLP-1 receptor agonists. The purpose of these studies was to identify and characterize novel nonpeptide agonists of the GLP-1 receptor. RESEARCH DESIGN AND METHODS Screening using cells expressing the GLP-1 receptor and insulin secretion assays with rodent and human islets were used to identify novel molecules. The intravenous glucose tolerance test (IVGTT) and hyperglycemic clamp characterized the insulinotropic effects of compounds in vivo. RESULTS Novel low molecular weight pyrimidine-based compounds that activate the GLP-1 receptor and stimulate glucose-dependent insulin secretion are described. These molecules induce GLP-1 receptor-mediated cAMP signaling in HEK293 cells expressing the GLP-1 receptor and increase insulin secretion from rodent islets in a dose-dependent manner. The compounds activate GLP-1 receptor signaling, both alone or in an additive fashion when combined with the endogenous GLP-1 peptide; however, these agonists do not compete with radiolabeled GLP-1 in receptor-binding assays. In vivo studies using the IVGTT and the hyperglycemic clamp in Sprague Dawley rats demonstrate increased insulin secretion in compound-treated animals. Further, perifusion assays with human islets isolated from a donor with type 2 diabetes show near-normalization of insulin secretion upon compound treatment. CONCLUSIONS These studies characterize the insulinotropic effects of an early-stage, small molecule GLP-1 receptor agonist and provide compelling evidence to support pharmaceutical optimization. PMID:20823098

  8. Pragmatic use of insulin degludec/insulin aspart co-formulation: A multinational consensus statement

    PubMed Central

    Kalra, Sanjay; Latif, Zafar A.; Comlekci, Abdurrahman; Galvez, Guillermo Gonzalez; Malik, Rached; Pathan, Md Faruque; Kumar, Ajay

    2016-01-01

    Insulin degludec/insulin aspart (IDegAsp) is a modern coformulation of ultra-long-acting basal insulin degludec, with rapid-acting insulin aspart. IDegAsp provides effective, safe, well-tolerated glycemic control, with a low risk of hypoglycemia while allowing flexibility in meal patterns and timing of administration. This consensus statement describes a pragmatic framework to identify patients who may benefit from IDegAsp therapy. It highlights the utility of IDegAsp in type 2 diabetic patients who are insulin-naive, suboptimally controlled on basal or premixed insulin, or dissatisfied with basal–bolus regimens. It also describes potential IDegAsp usage in type 1 diabetic patients. PMID:27366723

  9. Pragmatic use of insulin degludec/insulin aspart co-formulation: A multinational consensus statement.

    PubMed

    Kalra, Sanjay; Latif, Zafar A; Comlekci, Abdurrahman; Galvez, Guillermo Gonzalez; Malik, Rached; Pathan, Md Faruque; Kumar, Ajay

    2016-01-01

    Insulin degludec/insulin aspart (IDegAsp) is a modern coformulation of ultra-long-acting basal insulin degludec, with rapid-acting insulin aspart. IDegAsp provides effective, safe, well-tolerated glycemic control, with a low risk of hypoglycemia while allowing flexibility in meal patterns and timing of administration. This consensus statement describes a pragmatic framework to identify patients who may benefit from IDegAsp therapy. It highlights the utility of IDegAsp in type 2 diabetic patients who are insulin-naive, suboptimally controlled on basal or premixed insulin, or dissatisfied with basal-bolus regimens. It also describes potential IDegAsp usage in type 1 diabetic patients. PMID:27366723

  10. Effect of low temperature cultivation on insulin secretory of human pancreatic islets.

    PubMed

    Nikolic, D M; Djordjevic, P B; Lackovic, V B; Stojiljkovic, V; Stanojevic, B

    2013-01-01

    The experiment compared the physiological function (insulin secretory capacity) and membrane integrity of human adult pancreatic islets incubated in culture at 37°C and 24°C. Pancreatic tissue was digested with Collagenase XI, using a non-automated method. Cultures were incubated at 37°C and 24°C. Secretory capacity of the islets is determined by measuring of the stimulation index (SI) on the 1st, 3rd and 7th day of cultivation. Membrane integrity of the islets was determined by dithizone staining. Both groups of examined cultures show a slight increase in SI during the incubation. However islets incubated at 24°C show higher SI values than those incubated at 37°C on the 1st, 3rd and 7th day of incubation. And on the first day of incubation, this difference was statistically significant (p <0.05). Islets incubated at 37°C showed preservation of membrane integrity, the islets are regular spherical shape, while those incubated at 24°C lose such an organization. During the seven-day cultivation, islets incubated at a standard temperature of 37°C show less preserve physiological functions in relation to cultures incubated at 24°C, but islets incubated at 37°C show more regular morphological forms. PMID:23489685

  11. Neural apoptosis in the retina during experimental and human diabetes. Early onset and effect of insulin.

    PubMed Central

    Barber, A J; Lieth, E; Khin, S A; Antonetti, D A; Buchanan, A G; Gardner, T W

    1998-01-01

    This study determined whether retinal degeneration during diabetes includes retinal neural cell apoptosis. Image analysis of retinal sections from streptozotocin (STZ) diabetic rats after 7.5 months of STZ diabetes identified 22% and 14% reductions in the thickness of the inner plexiform and inner nuclear layers, respectively (P < 0. 001). The number of surviving ganglion cells was also reduced by 10% compared to controls (P < 0.001). In situ end labeling of DNA terminal dUTP nick end labeling (TUNEL) identified a 10-fold increase in the frequency of retinal apoptosis in whole-mounted rat retinas after 1, 3, 6, and 12 months of diabetes (P < 0.001, P < 0. 001, P < 0.01, and P < 0.01, respectively). Most TUNEL-positive cells were not associated with blood vessels and did not colocalize with the endothelial cell-specific antigen, von Willebrand factor. Insulin implants significantly reduced the number of TUNEL-positive cells (P < 0.05). The number of TUNEL-positive cells was also increased in retinas from humans with diabetes. These data indicate that retinal neural cell death occurs early in diabetes. This is the first quantitative report of an increase in neural cell apoptosis in the retina during diabetes, and indicates that neurodegeneration is an important component of diabetic retinopathy. PMID:9710447

  12. Inhibition of human amylin fibril formation by insulin-mimetic vanadium complexes.

    PubMed

    He, Lei; Wang, Xuesong; Zhao, Cong; Zhu, Dengsen; Du, Weihong

    2014-05-01

    The toxicity of amyloid-forming proteins can be linked to many degenerative and systemic diseases. Human islet amyloid polypeptide (hIAPP, amylin) has been associated with type II diabetes. Methods for efficient inhibition of amyloid fibril formation are highly clinically important. This study demonstrated the significant inhibitory effects of six vanadium complexes on hIAPP aggregation. Vanadium complexes, such as bis(maltolato)-oxovanadium (BMOV), have been used as insulin-mimetic agents for the treatment of diabetes for many years. Different biophysical methods were applied to investigate the interaction between V complexes and hIAPP. The results indicated that the selected compounds affected the peptide aggregation by different action modes and protected the cells from the cytotoxicity induced by hIAPP. Both the high binding affinity and the ligand spatial effect on inhibiting hIAPP aggregation are significant. Although some of these compounds undergo biotransformation under the conditions of the experiments, and the active species are not identified, it is understood that the effect results from a particular compound and its conversion products. Importantly, our work provided information on the effects of the selected V complexes on hIAPP and demonstrated multiple levels of effects of V complexes against amyloid-related diseases.

  13. Altered action of glucagon on human liver in type 2 (non-insulin-dependent) diabetes mellitus.

    PubMed

    Arner, P; Einarsson, K; Ewerth, S; Livingston, J N

    1987-05-01

    Glucagon may play a role in the metabolic derangements of overt Type 2 (non-insulin-dependent) diabetes mellitus. We therefore have evaluated the early steps in glucagon action by investigating the hormone-sensitive adenylyl cyclase system in liver membranes from seven Type 2 diabetic patients with fasting hyperglycaemia and two-fold elevations in plasma glucagon. The comparison was made with seven control subjects matched for age, sex and body weight. Glucagon receptor binding was almost identical in the two groups. There were, however, marked alterations in the adenylyl cyclase activity in membranes from the diabetic patients. This activity was reduced by 35-50% when compared to control activity. Basal cyclase activity, as well as the activity after stimulation with glucagon or with agents (i.e., sodium fluoride and forskolin) that act beyond the glucagon receptor, was significantly decreased (p less than 0.05, p less than 0.001 respectively). In conclusion, uncontrolled Type 2 diabetes in associated with an over-all loss of responsiveness of the hormone-sensitive adenylyl cyclase in human liver, which apparently results from post-receptor alterations. This change may provide a mechanism for reducing the effect of hyperglycagonaemia in Type 2 diabetes mellitus. PMID:3038642

  14. Nordihydroguaiaretic Acid Inhibits Insulin-Like Growth Factor Signaling, Growth, and Survival in Human Neuroblastoma Cells

    PubMed Central

    Meyer, Gary E.; Chesler, Louis; Liu, Dandan; Gable, Karissa; Maddux, Betty A.; Goldenberg, David D.; Youngren, Jack F.; Goldfine, Ira D.; Weiss, William A.; Matthay, Katherine K.; Rosenthal, Stephen M.

    2010-01-01

    Neuroblastoma is a common pediatric malignancy that metastasizes to the liver, bone, and other organs. Children with metastatic disease have a less than 50% chance of survival with current treatments. Insulin-like growth factors (IGFs) stimulate neuroblastoma growth, survival, and motility, and are expressed by neuroblastoma cells and the tissues they invade. Thus, therapies that disrupt the effects of IGFs on neuroblastoma tumorigenesis may slow disease progression. We show that NVP-AEW541, a specific inhibitor of the IGF-I receptor (IGF-IR), potently inhibits neuroblastoma growth in vitro. Nordihydroguaiaretic acid (NDGA), a phenolic compound isolated from the creosote bush (Larrea divaricata), has anti-tumor properties against a number of malignancies, has been shown to inhibit the phosphorylation and activation of the IGF-IR in breast cancer cells, and is currently in Phase I trials for prostate cancer. In the present study in neuroblastoma, NDGA inhibits IGF-I-mediated activation of the IGF-IR and disrupts activation of ERK and Akt signaling pathways induced by IGF-I. NDGA inhibits growth of neuroblastoma cells and induces apoptosis at higher doses, causing IGF-I-resistant activation of caspase-3 and a large increase in the fraction of sub-G0 cells. In addition, NDGA inhibits the growth of xenografted human neuroblastoma tumors in nude mice. These results indicate that NDGA may be useful in the treatment of neuroblastoma and may function in part via disruption of IGF-IR signaling. PMID:17486636

  15. Nordihydroguaiaretic acid inhibits insulin-like growth factor signaling, growth, and survival in human neuroblastoma cells.

    PubMed

    Meyer, Gary E; Chesler, Louis; Liu, Dandan; Gable, Karissa; Maddux, Betty A; Goldenberg, David D; Youngren, Jack F; Goldfine, Ira D; Weiss, William A; Matthay, Katherine K; Rosenthal, Stephen M

    2007-12-15

    Neuroblastoma is a common pediatric malignancy that metastasizes to the liver, bone, and other organs. Children with metastatic disease have a less than 50% chance of survival with current treatments. Insulin-like growth factors (IGFs) stimulate neuroblastoma growth, survival, and motility, and are expressed by neuroblastoma cells and the tissues they invade. Thus, therapies that disrupt the effects of IGFs on neuroblastoma tumorigenesis may slow disease progression. We show that NVP-AEW541, a specific inhibitor of the IGF-I receptor (IGF-IR), potently inhibits neuroblastoma growth in vitro. Nordihydroguaiaretic acid (NDGA), a phenolic compound isolated from the creosote bush (Larrea divaricata), has anti-tumor properties against a number of malignancies, has been shown to inhibit the phosphorylation and activation of the IGF-IR in breast cancer cells, and is currently in Phase I trials for prostate cancer. In the present study in neuroblastoma, NDGA inhibits IGF-I-mediated activation of the IGF-IR and disrupts activation of ERK and Akt signaling pathways induced by IGF-I. NDGA inhibits growth of neuroblastoma cells and induces apoptosis at higher doses, causing IGF-I-resistant activation of caspase-3 and a large increase in the fraction of sub-G0 cells. In addition, NDGA inhibits the growth of xenografted human neuroblastoma tumors in nude mice. These results indicate that NDGA may be useful in the treatment of neuroblastoma and may function in part via disruption of IGF-IR signaling.

  16. Conditioned insulin and blood sugar responses in humans in relation to binge eating.

    PubMed

    Overduin, J; Jansen, A

    1997-04-01

    This study proposed to demonstrate a classically conditioned blood sugar decrease in humans and to clarify its relevance for binge eating. Six conditioning trials were run in healthy females. The conditioned stimulus (CS) was a compound peppermint flavor/fragrance, whereas the unconditioned stimulus (UCS) consisted of 50 g of oral glucose. Control subjects received an aspartame drink as the UCS. Ad lib glucose intake, blood parameters, and subjective craving were monitored before and after conditioning. Results showed that the experimental group failed to show conditioned blood sugar and glucagon decreases or C-Peptide increases. Although an increased insulin response was found in the experimental group, the effect size did not exceed that of spontaneous fluctuations. No increases in craving for sweet substances were found. An impressive increase (mean: 78%) in glucose intake after conditioning was found in both conditions, as well as in a subsequently run third condition with plain water as the UCS. The increased glucose intake probably resulted from an initial neophobia to the laboratory setting that subsided as subjects had experienced more lab sessions. Importantly, because no conditioned hypoglycemia occurred in the present study, its relationship with subjectively experienced craving for sweet substance could not be determined.

  17. Insulin Resistance in Human iPS Cells Reduces Mitochondrial Size and Function

    PubMed Central

    Burkart, Alison M.; Tan, Kelly; Warren, Laura; Iovino, Salvatore; Hughes, Katelyn J.; Kahn, C. Ronald; Patti, Mary-Elizabeth

    2016-01-01

    Insulin resistance, a critical component of type 2 diabetes (T2D), precedes and predicts T2D onset. T2D is also associated with mitochondrial dysfunction. To define the cause-effect relationship between insulin resistance and mitochondrial dysfunction, we compared mitochondrial metabolism in induced pluripotent stem cells (iPSC) from 5 healthy individuals and 4 patients with genetic insulin resistance due to insulin receptor mutations. Insulin-resistant iPSC had increased mitochondrial number and decreased mitochondrial size. Mitochondrial oxidative function was impaired, with decreased citrate synthase activity and spare respiratory capacity. Simultaneously, expression of multiple glycolytic enzymes was decreased, while lactate production increased 80%. These perturbations were accompanied by an increase in ADP/ATP ratio and 3-fold increase in AMPK activity, indicating energetic stress. Insulin-resistant iPSC also showed reduced catalase activity and increased susceptibility to oxidative stress. Thus, insulin resistance can lead to mitochondrial dysfunction with reduced mitochondrial size, oxidative activity, and energy production. PMID:26948272

  18. Insulin and IGFs enhance hepatocyte differentiation from human embryonic stem cells via the PI3K/AKT pathway.

    PubMed

    Magner, Nataly L; Jung, Yunjoon; Wu, Jian; Nolta, Jan A; Zern, Mark A; Zhou, Ping

    2013-10-01

    Human embryonic stem cells (hESCs) can be progressively differentiated into definitive endoderm (DE), hepatic progenitors, and hepatocytes, and thus provide an excellent model system for the mechanistic study of hepatocyte differentiation, which is currently poorly understood. Here, we found that insulin enhanced hepatocyte differentiation from hESC-derived DE. Insulin activated the PI3K/AKT pathway, but not the mitogen-activated protein kinase pathway in the DE cells, and inhibition of the PI3K/AKT pathways by inhibitors markedly inhibited hepatocyte differentiation. In addition, insulin-like growth factor 1 (IGF1) and IGF2 also activated the PI3K/AKT pathway in DE cells and their expression was robustly upregulated during hepatocyte differentiation from DE. Furthermore, inhibition of IGF receptor 1 (IGF1R) by a small molecule inhibitor PPP or knockdown of the IGF1R by shRNA attenuated hepatocyte differentiation. Moreover, simultaneous knockdown of the IGF1R and the insulin receptor with shRNAs markedly reduced the activation of AKT and substantially impaired hepatocyte differentiation. The PI3K pathway specifically enhanced the expression of HNF1 and HNF4 to regulate hepatocyte differentiation from DE. Although inhibition of the PI3K pathway was previously shown to be required for the induction of DE from hESCs, our study revealed a positive role of the PI3K pathway in hepatocyte differentiation after the DE stage, and has advanced our understanding of hepatocyte cell fate determination.

  19. The expression of ob gene is not acutely regulated by insulin and fasting in human abdominal subcutaneous adipose tissue.

    PubMed

    Vidal, H; Auboeuf, D; De Vos, P; Staels, B; Riou, J P; Auwerx, J; Laville, M

    1996-07-15

    The regulation of ob gene expression in abdominal subcutaneous adipose tissue was investigated using a reverse transcription-competitive PCR method to quantify the mRNA level of leptin. Leptin mRNA level was highly correlated with the body mass index of 26 subjects (12 lean, 7 non-insulin-dependent diabetic, and 7 obese patients). The effect of fasting on ob gene expression was investigated in 10 subjects maintained on a hypocaloric diet (1045 KJ/d) for 5 d. While their metabolic parameters significantly changed (decrease in insulinemia, glycemia, and resting metabolic rate and increase in plasma ketone bodies), the caloric restriction did not modify the leptin mRNA level in the adipose tissue. To verify whether insulin regulates ob gene expression, six lean subjects underwent a 3-h euglycemic hyperinsulinemic (846 +/- 138 pmol/liter) clamp. Leptin and Glut 4 mRNA levels were quantified in adipose tissue biopsies taken before and at the end of the clamp. Insulin infusion produced a significant threefold increase in Glut 4 mRNA while leptin mRNA was not affected. It is concluded that ob gene expression is not acutely regulated by insulin or by metabolic factors related to fasting in human abdominal subcutaneous adipose tissue. PMID:8755631

  20. Differentiation of PDX1 gene-modified human umbilical cord mesenchymal stem cells into insulin-producing cells in vitro.

    PubMed

    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. PMID:21837359

  1. Phase changes in T(3)R(3)(f) human insulin: temperature or pressure induced?

    PubMed

    Smith, G D; Pangborn, W A; Blessing, R H

    2001-08-01

    The structure of T(3)R(3) hexameric human insulin has been determined at 100 K from two different crystals at 1.2 and 1.3 A resolution and refined to residuals of 0.169 and 0.176, respectively. Owing to a phase change, the c axis is double its room-temperature value and the asymmetric unit contains two independent TR(f) insulin dimers. Compared with the orientation in the room-temperature structure, one dimer undergoes a rotation about the c axis of -5 degrees, while the second is rotated +4 degrees. A superposition of the backbone atoms of the two independent dimers shows that the C(alpha) atoms of five residues within the R(f)-state monomers are displaced by more than 1.0 A; smaller displacements are observed for the T-state monomers. Four zinc ions lie on the crystallographic threefold axis and each forms bonds to three symmetry-related HisB10 N(varepsilon2) atoms from the T- and R(f)-state trimers. While three of the zinc ions are tetrahedrally coordinated with a chloride ion completing the coordination sphere, mixed tetrahedral/octahedral coordination is observed for one of the T-state zinc ions. The three symmetry-related "phenolic binding sites" in one hexamer contain water molecules and a glycerol molecule, but the same sites in the second hexamer are occupied by a zinc ion coordinated to an alternate conformation of HisB10, a symmetry-related HisB5 and two chloride ions. Two additional and partially occupied zinc ion sites are observed at the interface between the two independent dimers. One zinc ion is coordinated by a T-state HisB5 of one dimer, an R-state HisB5 of the second dimer and two water molecules; the second zinc ion is coordinated by an alternate side-chain conformation of the T-state HisB5 and three water molecules. The carboxyl group of one GluB13 side chain, which exists in two discrete conformations, appears to be protonated, because short contacts exist to a second carboxyl group or to a carbonyl O atom.

  2. A novel assay in vitro of human islet amyloid polypeptide amyloidogenesis and effects of insulin secretory vesicle peptides on amyloid formation.

    PubMed Central

    Kudva, Y C; Mueske, C; Butler, P C; Eberhardt, N L

    1998-01-01

    Human islet amyloid polypeptide (IAPP) is a 37-residue peptide that is co-secreted with insulin by the beta-cell and might be involved in the pathogenesis of non-insulin-dependent diabetes mellitus. We developed an improved assay in vitro based on the fluorescence of bound thioflavin T to study factors affecting amyloidogenesis. Monomeric IAPP formed amyloid fibrils, as detected by increased fluorescence and by electron microscopy. Fluorimetric analysis revealed that the initial rate of amyloid formation was: (1) proportional to the peptide monomer concentration, (2) maximal at pH 9.5, (3) maximal at 200 mMKCl, and (4) proportional to temperature from 4 to 37 degreesC. We found that 5-fold and 10-fold molar excesses of proinsulin inhibited fibril formation by 39% and 59% respectively. Insulin was somewhat more potent with 5-fold and 10-fold molar excesses inhibiting fibril formation by 69% and 73% respectively, whereas C-peptide had no effect at these concentrations. Thus at physiological ratios of IAPP to insulin, insulin and proinsulin, but not C-peptide, can retard amyloidogenesis. Because insulin resistance or hyperglycaemia increase the IAPP-to-insulin ratio, increased intracellular IAPP compared with insulin expression in genetically predisposed individuals might contribute to intracellular amyloid formation, beta-cell death and the genesis of non-insulin-dependent diabetes mellitus. PMID:9560308

  3. Blood-based biomarkers of age-associated epigenetic changes in human islets associate with insulin secretion and diabetes

    PubMed Central

    Bacos, Karl; Gillberg, Linn; Volkov, Petr; Olsson, Anders H; Hansen, Torben; Pedersen, Oluf; Gjesing, Anette Prior; Eiberg, Hans; Tuomi, Tiinamaija; Almgren, Peter; Groop, Leif; Eliasson, Lena; Vaag, Allan; Dayeh, Tasnim; Ling, Charlotte

    2016-01-01

    Aging associates with impaired pancreatic islet function and increased type 2 diabetes (T2D) risk. Here we examine whether age-related epigenetic changes affect human islet function and if blood-based epigenetic biomarkers reflect these changes and associate with future T2D. We analyse DNA methylation genome-wide in islets from 87 non-diabetic donors, aged 26–74 years. Aging associates with increased DNA methylation of 241 sites. These sites cover loci previously associated with T2D, for example, KLF14. Blood-based epigenetic biomarkers reflect age-related methylation changes in 83 genes identified in human islets (for example, KLF14, FHL2, ZNF518B and FAM123C) and some associate with insulin secretion and T2D. DNA methylation correlates with islet expression of multiple genes, including FHL2, ZNF518B, GNPNAT1 and HLTF. Silencing these genes in β-cells alter insulin secretion. Together, we demonstrate that blood-based epigenetic biomarkers reflect age-related DNA methylation changes in human islets, and associate with insulin secretion in vivo and T2D. PMID:27029739

  4. Blood-based biomarkers of age-associated epigenetic changes in human islets associate with insulin secretion and diabetes.

    PubMed

    Bacos, Karl; Gillberg, Linn; Volkov, Petr; Olsson, Anders H; Hansen, Torben; Pedersen, Oluf; Gjesing, Anette Prior; Eiberg, Hans; Tuomi, Tiinamaija; Almgren, Peter; Groop, Leif; Eliasson, Lena; Vaag, Allan; Dayeh, Tasnim; Ling, Charlotte

    2016-03-31

    Aging associates with impaired pancreatic islet function and increased type 2 diabetes (T2D) risk. Here we examine whether age-related epigenetic changes affect human islet function and if blood-based epigenetic biomarkers reflect these changes and associate with future T2D. We analyse DNA methylation genome-wide in islets from 87 non-diabetic donors, aged 26-74 years. Aging associates with increased DNA methylation of 241 sites. These sites cover loci previously associated with T2D, for example, KLF14. Blood-based epigenetic biomarkers reflect age-related methylation changes in 83 genes identified in human islets (for example, KLF14, FHL2, ZNF518B and FAM123C) and some associate with insulin secretion and T2D. DNA methylation correlates with islet expression of multiple genes, including FHL2, ZNF518B, GNPNAT1 and HLTF. Silencing these genes in β-cells alter insulin secretion. Together, we demonstrate that blood-based epigenetic biomarkers reflect age-related DNA methylation changes in human islets, and associate with insulin secretion in vivo and T2D.

  5. Blood-based biomarkers of age-associated epigenetic changes in human islets associate with insulin secretion and diabetes.

    PubMed

    Bacos, Karl; Gillberg, Linn; Volkov, Petr; Olsson, Anders H; Hansen, Torben; Pedersen, Oluf; Gjesing, Anette Prior; Eiberg, Hans; Tuomi, Tiinamaija; Almgren, Peter; Groop, Leif; Eliasson, Lena; Vaag, Allan; Dayeh, Tasnim; Ling, Charlotte

    2016-01-01

    Aging associates with impaired pancreatic islet function and increased type 2 diabetes (T2D) risk. Here we examine whether age-related epigenetic changes affect human islet function and if blood-based epigenetic biomarkers reflect these changes and associate with future T2D. We analyse DNA methylation genome-wide in islets from 87 non-diabetic donors, aged 26-74 years. Aging associates with increased DNA methylation of 241 sites. These sites cover loci previously associated with T2D, for example, KLF14. Blood-based epigenetic biomarkers reflect age-related methylation changes in 83 genes identified in human islets (for example, KLF14, FHL2, ZNF518B and FAM123C) and some associate with insulin secretion and T2D. DNA methylation correlates with islet expression of multiple genes, including FHL2, ZNF518B, GNPNAT1 and HLTF. Silencing these genes in β-cells alter insulin secretion. Together, we demonstrate that blood-based epigenetic biomarkers reflect age-related DNA methylation changes in human islets, and associate with insulin secretion in vivo and T2D. PMID:27029739

  6. [Insulinization in type 2 diabetes mellitus. Intensification options].

    PubMed

    Fuente, Graciela V; Sinay, Isaac; Costa Gil, José E; Puchulu, Félix; Dieuzeide, Guillermo; Rodríguez, Martín; Faingold, María C; Litwak, León E

    2016-01-01

    Diabetes mellitus is associated with vascular complications and high rates of morbidity and mortality. Timely insulin therapy, intensified when necessary, represent appropriate measures to prevent or delay the onset of complications. However, the incidence of hypoglycemia and difficulties in treatment adherence represent barriers to achieve therapeutic success. Premixes analogs and, specially, combinations of insulin analogues are associated with pharmacokinetic and pharmacodynamic advantages, that translate into clinical benefits such as improved metabolic control, decreased hypoglycemic events and, for their simplicity, potentially greater adherence.

  7. In vitro differentiation of human amniotic epithelial cells into insulin-producing 3D spheroids.

    PubMed

    Okere, Bernard; Alviano, Francesco; Costa, Roberta; Quaglino, Daniela; Ricci, Francesca; Dominici, Massimo; Paolucci, Paolo; Bonsi, Laura; Iughetti, Lorenzo

    2015-09-01

    Regenerative medicine and stem cell therapy may represent the solution for the treatment of non-curable human diseases such as type 1 diabetes. In this context of growing demand for functional and safe stem cells, human amniotic epithelial cells (hAECs) from term placenta have attracted increasing interest for their wide availability, stem cell properties, and differentiation plasticity, which make them a promising tool for stem cell-based therapeutic applications. We initially assayed the stemness characteristics of hAECs in serum-free conditions. Subsequently we developed a culture procedure on extracellular matrix for the formation of three-dimensional (3D) spheroids. Finally, we tested the immunomodulation and differentiation potential of hAEC spheroids: the presence of pancreatic endocrine hormones was revealed with transmission electron microscopy and immunofluorescence analyses; the release of C-peptide in hyperglycemic conditions was assayed with ELISA. The serum-free culture conditions we applied proved to maintain the basic stemness characteristics of hAECs. We also demonstrated that 3D spheroids formed by hAECs in extracellular matrix can be induced to differentiate into insulin-producing cells. Finally, we proved that control and induced cells equally inhibit the proliferation of activated mononuclear cells. The results of this study highlight the properties of amnion derived epithelial cells as promising and abundant source for cell-based therapies. In particular we are the first group to show the in vitro pancreatic induction of hAECs cultured on extracellular matrix in a 3D fashion. We accordingly propose the outcomes of this study as a novel contribution to the development of future cell replacement therapies involving placenta-derived cells.

  8. Immunohistochemical localization of components of the insulin-like growth factor system in human permanent teeth.

    PubMed

    Götz, Werner; Heinen, Michael; Lossdörfer, Stefan; Jäger, Andreas

    2006-05-01

    There is growing evidence that the insulin-like growth factor (IGF) system plays an important role in the biology of oro-dento-facial tissues and organs, including the development, homeostasis and regeneration of the periodontium. To obtain basic data on the occurrence and distribution of IGF components in human permanent teeth we immunohistochemically investigated 25 extracted, decalcified and paraffin-embedded teeth using mono and polyclonal antibodies against the ligands IGF-I and -II, the IGF1 receptor (IGF1R) and all six IGF binding proteins (IGFBP-1 to -6). In the extracellular matrix (ECM) of the adhering periodontal ligament (PDL), immunoreactivity for IGF-I, -II and IGFBP-1 and -6 was observed. PDL fibroblasts showed immunostaining for the IGF1R. For the cementum, in the acellular cementum only IGF-II could be detected, while outer cementum layers with inserting Sharpey's fibers reacted with all antibodies applied except for IGFBP-4 and -6. In the pulp, mainly fibrotic areas and areas around denticles were immunoreactive for IGF-I, IGFBP-1, -3, -5 and -6. Predentin and odontoblastic processes were stained for IGF-I and IGFBP-3. The spatially oriented occurrence of components of the IGF system in human permanent teeth indicates that specific functions of the IGFs may be localized in particular tissue compartments. In the cementum, several IGF components were found indicating roles in tissue homeostasis or attachment. The PDL may function as a reservoir for IGFs probably bound to ECM components. PDL fibroblasts could then respond in a paracrine manner. In the pulp, the IGF system may be involved in odontoblast biology, fibrosis and denticle formation. PMID:16321360

  9. Large-Eddy Simulation of Premixed and Partially Premixed Turbulent Combustion Using a Level Set Method

    NASA Astrophysics Data System (ADS)

    Duchamp de Lageneste, Laurent; Pitsch, Heinz

    2001-11-01

    Level-set methods (G-equation) have been recently used in the context of RANS to model turbulent premixed (Hermann 2000) or partially premixed (Chen 1999) combustion. By directly taking into account unsteady effects, LES can be expected to improve predictions over RANS. Since the reaction zone thickness of premixed flames in technical devices is usually much smaller than the LES grid spacing, chemical reactions completely occur on the sub-grid scales and hence have to be modeled entirely. In the level-set methodology, the flame front is represented by an arbitrary iso-surface G0 of a scalar field G whose evolution is described by the so-called G-equation. This equation is only valid at G=G_0, and hence decoupled from other G levels. Heat release is then modeled using a flamelet approach in which temperature is determined as a function of G and the mixture-fraction Z. In the present study, the proposed approach has been formulated for LES and validated using data from a turbulent Bunsen burner experiment (Chen, Peters 1996). Simulation of an experimental Lean Premixed Prevapourised (LPP) dump combustor (Besson, Bruel 1999, 2000) under different premixed or partially premixed conditions will also be presented.

  10. Differentiation of human adipose-derived mesenchymal stem cell into insulin-producing cells: an in vitro study.

    PubMed

    Moshtagh, P Rahnamay; Emami, S Hojati; Sharifi, Ali M

    2013-09-01

    Stem cells with the ability to differentiate into insulin-producing cells (IPCs) are becoming the most promising therapy for diabetes mellitus and reduce the major limitations of availability and allogeneic rejection of beta cell transplantations. Mesenchymal stem cells (MSCs) are pluripotent stromal cells with the ability to proliferate and differentiate into a variety of cell types including endocrine cells of the pancreas. This study sought to inspect the in vitro differentiation of human adipose-derived tissue stem cells into IPCs which could provide an abundant source of cells for the purpose of diabetic cell therapy in addition to avoid immunological rejection. Adipose-derived MSCs were obtained from liposuction aspirates and induced to differentiate into insulin-secreting cells under a three-stage protocol based on a combination of low-glucose DMEM medium, β-mercaptoethanol, and nicotinamide for pre-induction and high-glucose DMEM, β-mercaptoethanol, nicotinamide, and exendin-4 for induction stages of differentiation. Differentiation was evaluated by the analysis of morphology, dithizone staining, RT-PCR, and immunocytochemistry. Morphological changes including typical islet-like cell clusters were observed by phase-contrast microscope at the end of differentiation protocol. Based on dithizone staining, differentiated cells were positive and undifferentiated cells were not stained. Furthermore, RT-PCR results confirmed the expression of insulin, PDX1, Ngn3, PAX4, and GLUT2 in differentiated cells. Moreover, insulin production by the IPCs was confirmed by immunocytochemistry analysis. It is concluded that adipose-derived MSCs could differentiate into insulin-producing cells in vitro.

  11. Human marrow-derived mesodermal progenitor cells generate insulin-secreting islet-like clusters in vivo.

    PubMed

    Ai, Cuiwei; Todorov, Ivan; Slovak, Marilyn L; Digiusto, David; Forman, Stephen J; Shih, Chu-Chih

    2007-10-01

    Transplantation of pancreatic islet cells is the only known potential cure for diabetes mellitus. However, the difficulty in obtaining sufficient numbers of purified islets for transplantation severely limits its use. A renewable and clinically accessible source of stem cells capable of differentiating into insulin-secreting beta-cells might circumvent this limitation. Here, we report that human fetal bone marrow (BM)-derived mesodermal progenitor cells (MPCs) possess the potential to generate insulinsecreting islet-like clusters (ISILCs) when injected into human fetal pancreatic tissues implanted in severe combined immunodeficiency (SCID) mice. Seven essential genes involved in pancreatic endocrine development, including insulin, glucagon, somatostatin, pdx-1, glut-2, nkx 2.2, and nkx 6.1, are expressed in these BM-MPC-derived ISILCs, suggesting that ISILCs are generated through neogenesis of BM-MPCs. Our data further suggest that differentiation of BM-MPCs into ISILCs is not mediated by cell fusion. Insulin secretion from these ISILCs is regulated by glucose concentration in vitro, and transplantation of purified ISILCs normalizes hyperglycemia in streptozocin (STZ)- induced nonobese diabetic (NOD)/SCID mice.

  12. Aspartame and its constituent amino acids: effects on prolactin, cortisol, growth hormone, insulin, and glucose in normal humans.

    PubMed

    Carlson, H E; Shah, J H

    1989-03-01

    Because large doses of phenylalanine stimulate prolactin secretion in man, we studied the acute effects of oral doses of aspartame (0.534 g, equivalent to the amount of aspartame in approximately 1 L beverage), aspartic acid (0.242 g), and phenylalanine (0.3 and 1.0 g) on serum prolactin and other hormones in normal humans. Prolactin was not stimulated by any of the aspartame meals, aspartic acid, or 0.3 g phenylalanine; a small rise in serum prolactin, similar to that produced by a high-protein mixed meal, followed ingestion of 1.0 g phenylalanine. Serum growth hormone showed no statistically significant changes in response to any of the experimental meals whereas cortisol and insulin fell slightly and glucose rose slightly during each of the meals. We conclude that these doses of aspartame do not alter secretion of prolactin, cortisol, growth hormone, or insulin in normal individuals.

  13. Enterovirus infection of human islets of Langerhans affects β-cell function resulting in disintegrated islets, decreased glucose stimulated insulin secretion and loss of Golgi structure

    PubMed Central

    Hodik, M; Skog, O; Lukinius, A; Isaza-Correa, J M; Kuipers, J; Giepmans, B N G; Frisk, G

    2016-01-01

    Aims/hypothesis In type 1 diabetes (T1D), most insulin-producing β cells are destroyed, but the trigger is unknown. One of the possible triggers is a virus infection and the aim of this study was to test if enterovirus infection affects glucose stimulated insulin secretion and the effect of virus replication on cellular macromolecules and organelles involved in insulin secretion. Methods Isolated human islets were infected with different strains of coxsackievirus B (CVB) virus and the glucose-stimulated insulin release (GSIS) was measured in a dynamic perifusion system. Classical morphological electron microscopy, large-scale electron microscopy, so-called nanotomy, and immunohistochemistry were used to study to what extent virus-infected β cells contained insulin, and real-time PCR was used to analyze virus induced changes of islet specific genes. Results In islets infected with CVB, GSIS was reduced in correlation with the degree of virus-induced islet disintegration. The expression of the gene encoding insulin was decreased in infected islets, whereas the expression of glucagon was not affected. Also, in islets that were somewhat disintegrated, there were uninfected β cells. Ultrastructural analysis revealed that virus particles and virus replication complexes were only present in β cells. There was a significant number of insulin granules remaining in the virus-infected β cells, despite decreased expression of insulin mRNA. In addition, no typical Golgi apparatus was detected in these cells. Exposure of islets to synthetic dsRNA potentiated glucose-stimulated insulin secretion. Conclusions/interpretation Glucose-stimulated insulin secretion; organelles involved in insulin secretion and gene expression were all affected by CVB replication in β cells. PMID:27547409

  14. Insulin-like growth factor-I stimulates IL-10 production in human T cells.

    PubMed

    Kooijman, Ron; Coppens, Astrid

    2004-10-01

    There is vast body of evidence that the insulin-like growth factor (IGF)-I exerts immunomodulatory effects in vitro and in vivo. In vitro studies indicate that stimulatory effects of IGF-I may be exerted through augmentation of inflammatory cytokine production. To further explore the immunomodulatory effects of IGF-I through regulation of cytokine production, we tested the in vitro effects of IGF-I on the secretion of inflammatory T helper cell type 1 (Th1) and Th2 cytokines by human peripheral blood mononuclear cells (PBMC). To this end, PBMC were stimulated with the T cell mitogen phytohemagglutinin (PHA), and cytokines in the culture media were assessed after 18, 42, 66, and 80 h of culture. We found that IGF-I stimulated the secretion of the Th2 cytokine interleukin (IL)-10 by 40-70% in PHA-stimulated PBMC. In addition, we observed a small stimulatory effect (15%) on the secretion of another Th2 cytokine IL-4. The secretion of IL-2, IL-5, IL-6, interferon-gamma, and the inflammatory cytokines IL-1beta, IL-8, and tumor necrosis factor alpha was not or was hardly affected. IL-10 secretion was also stimulated in purified T cells, and we established that IGF-I also stimulated IL-10 mRNA expression by 100-150%. The monocyte-activating bacterial cell-wall product lipopolysaccharide induced IL-10 production in PBMC, but this was not affected by IGF-I. As IL-10 predominantly exerts anti-inflammatory actions and suppresses Th1-dependent immune responses, our results indicate that IGF-I may exert inhibitory actions on inflammatory and Th1-mediated cellular immune responses through stimulation of IL-10 production in T cells.

  15. MAFA and T3 Drive Maturation of Both Fetal Human Islets and Insulin-Producing Cells Differentiated From hESC

    PubMed Central

    Aguayo-Mazzucato, Cristina; DiIenno, Amanda; Hollister-Lock, Jennifer; Cahill, Christopher; Sharma, Arun; Weir, Gordon; Colton, Clark

    2015-01-01

    Context: Human embryonic stem cells (hESCs) differentiated toward β-cells and fetal human pancreatic islet cells resemble each other transcriptionally and are characterized by immaturity with a lack of glucose responsiveness, low levels of insulin content, and impaired proinsulin-to-insulin processing. However, their response to stimuli that promote functionality have not been compared. Objective: The objective of the study was to evaluate the effects of our previous strategies for functional maturation developed in rodents in these two human models of β-cell immaturity and compare their responses. Design, Settings, Participants, and Interventions: In proof-of-principle experiments using either adenoviral-mediated overexpression of V-Maf avian musculoaponeurotic fibrosarcoma oncogene homolog A (MAFA) or the physiologically driven path via thyroid hormone (T3) and human fetal islet-like cluster (ICC) functional maturity was evaluated. Then the effects of T3 were evaluated upon the functional maturation of hESCs differentiated toward β-cells. Main Outcome Measures: Functional maturation was evaluated by the following parameters: glucose responsiveness, insulin content, expression of the mature β-cell transcription factor MAFA, and proinsulin-to-insulin processing. Results: ICCs responded positively to MAFA overexpression and T3 treatment as assessed by two different maturation parameters: increased insulin secretion at 16.8 mM glucose and increased proinsulin-to-insulin processing. In hESCs differentiated toward β-cells, T3 enhanced MAFA expression, increased insulin content (probably mediated by the increased MAFA), and increased insulin secretion at 16.8 mM glucose. Conclusion: T3 is a useful in vitro stimulus to promote human β-cell maturation as shown in both human fetal ICCs and differentiated hESCs. The degree of maturation induced varied in the two models, possibly due to the different developmental status at the beginning of the study. PMID:26207953

  16. Peribiliary Glands as a Niche of Extrapancreatic Precursors Yielding Insulin-Producing Cells in Experimental and Human Diabetes.

    PubMed

    Carpino, Guido; Puca, Rosa; Cardinale, Vincenzo; Renzi, Anastasia; Scafetta, Gaia; Nevi, Lorenzo; Rossi, Massimo; Berloco, Pasquale B; Ginanni Corradini, Stefano; Reid, Lola M; Maroder, Marella; Gaudio, Eugenio; Alvaro, Domenico

    2016-05-01

    Peribiliary glands (PBGs) are niches in the biliary tree and containing heterogeneous endodermal stem/progenitors cells that can differentiate, in vitro and in vivo, toward pancreatic islets. The aim of this study was to evaluate, in experimental and human diabetes, proliferation of cells in PBGs and differentiation of the biliary tree stem/progenitor cells (BTSCs) toward insulin-producing cells. Diabetes was generated in mice by intraperitoneal injection of a single dose of 200 mg/kg (N = 12) or 120 mg/kg (N = 12) of streptozotocin. Liver, pancreas, and extrahepatic biliary trees were en bloc dissected and examined. Cells in PBGs proliferated in experimental diabetes, and their proliferation was greatest in the PBGs of the hepatopancreatic ampulla, and inversely correlated with the pancreatic islet area. In rodents, the cell proliferation in PBGs was characterized by the expansion of Sox9-positive stem/progenitor cells that gave rise to insulin-producing cells. Insulin-producing cells were located mostly in PBGs in the portion of the biliary tree closest to the duodenum, and their appearance was associated with upregulation of MafA and Gli1 gene expression. In patients with type 2 diabetes, PBGs at the level of the hepatopancreatic ampulla contained cells showing signs of proliferation and pancreatic fate commitment. In vitro, high glucose concentrations induced the differentiation of human BTSCs cultures toward pancreatic beta cell fates. The cells in PBGs respond to diabetes with proliferation and differentiation towards insulin-producing cells indicating that PBG niches may rescue pancreatic islet impairment in diabetes. These findings offer important implications for the pathophysiology and complications of this disease. Stem Cells 2016;34:1332-1342. PMID:26850087

  17. Acute regulation by insulin of phosphatidylinositol-3-kinase, Rad, Glut 4, and lipoprotein lipase mRNA levels in human muscle.

    PubMed

    Laville, M; Auboeuf, D; Khalfallah, Y; Vega, N; Riou, J P; Vidal, H

    1996-07-01

    We have investigated the acute regulation by insulin of the mRNA levels of nine genes involved in insulin action, in muscle biopsies obtained before and at the end of a 3-h euglycemic hyperinsulinemic clamp. Using reverse transcription-competitive PCR, we have measured the mRNAs encoding the two insulin receptor variants, the insulin receptor substrate-1, the p85alpha subunit of phosphatidylinositol-3-kinase, Ras associated to diabetes (Rad), the glucose transporter Glut 4, glycogen synthase, 6-phosphofructo-l-kinase, lipoprotein lipase, and the hormone-sensitive lipase. Insulin infusion induced a significant increase in the mRNA level of Glut 4 (+56 +/- 13%), Rad (+96 +/- 25%), the p85alpha subunit of phosphatidylinositol-3-kinase (+92 +/- 18%) and a decrease in the lipoprotein lipase mRNA level (-49 +/- 5%), while the abundance of the other mRNAs was unaffected. The relative expression of the two insulin receptor variants was not modified. These results demonstrate an acute coordinated regulation by insulin of the expression of genes coding key proteins involved in its action in human skeletal muscle and suggest that Rad and the p85alpha regulatory subunit of phosphatidylinositol-3-kinase can be added to the list of the genes controlled by insulin. PMID:8690802

  18. The human insulin mRNA is partly translated via a cap- and eIF4A-independent mechanism

    SciTech Connect

    Fred, Rikard G.; Sandberg, Monica; Pelletier, Jerry; Welsh, Nils

    2011-09-09

    Highlights: {yields} The polypyrimidine tract binding protein binds to the 5'-UTR of the insulin mRNA. {yields} Insulin mRNA can be translated via a cap-independent mechanism. {yields} The fraction cap-independent insulin synthesis increases during conditions of stress. {yields} The {beta}-cell is able to uphold basal insulin biosynthesis under conditions of stress. -- Abstract: The aim of this study was to investigate whether cap-independent insulin mRNA translation occurs in human pancreatic islets at basal conditions, during stimulation at a high glucose concentration and at conditions of nitrosative stress. We also aimed at correlating cap-independent insulin mRNA translation with binding of the IRES trans-acting factor polypyrimidine tract binding protein (PTB) to the 5'-UTR of insulin mRNA. For this purpose, human islets were incubated for 2 h in the presence of low (1.67 mM) or high glucose (16.7 mM). Nitrosative stress was induced by addition of 1 mM DETA/NO and cap-dependent mRNA translation was inhibited with hippuristanol. Insulin biosynthesis rates were determined by radioactive labeling and immunoprecipitation. PTB affinity to insulin mRNA 5'-UTR was assessed by a magnetic micro bead pull-down procedure. We observed that in the presence of 1.67 mM glucose, approximately 70% of the insulin mRNA translation was inhibited by hippuristanol. Corresponding value from islets incubated at 16.7 mM glucose was 93%. DETA/NO treatment significantly decreased the translation of insulin by 85% in high glucose incubated islets, and by 50% at a low glucose concentration. The lowered insulin biosynthesis rates of DETA/NO-exposed islets were further suppressed by hippuristanol with 55% at 16.7 mM glucose but not at 1.67 mM glucose. Thus, hippuristanol-induced inhibition of insulin biosynthesis was less pronounced in DETA/NO-treated islets as compared to control islets. We observed also that PTB bound specifically to the insulin mRNA 5'-UTR in vitro, and that this

  19. Dynamics and structure of turbulent premixed flames

    NASA Technical Reports Server (NTRS)

    Bilger, R. W.; Swaminathan, N.; Ruetsch, G. R.; Smith, N. S. A.

    1995-01-01

    In earlier work (Mantel & Bilger, 1994) the structure of the turbulent premixed flame was investigated using statistics based on conditional averaging with the reaction progress variable as the conditioning variable. The DNS data base of Trouve and Poinsot (1994) was used in this investigation. Attention was focused on the conditional dissipation and conditional axial velocity in the flame with a view to modeling these quantities for use in the conditional moment closure (CMC) approach to analysis of kinetics in premixed flames (Bilger, 1993). Two remarkable findings were made: there was almost no acceleration of the axial velocity in the flame front itself; and the conditional scalar dissipation remained as high, or higher, than that found in laminar premixed flames. The first finding was surprising since in laminar flames all the fluid acceleration occurs through the flame front, and this could be expected also for turbulent premixed flames at the flamelet limit. The finding gave hope of inventing a new approach to the dynamics of turbulent premixed flames through use of rapid distortion theory or an unsteady Bernoulli equation. This could lead to a new second order closure for turbulent premixed flames. The second finding was contrary to our measurements with laser diagnostics in lean hydrocarbon flames where it is found that conditional scalar dissipation drops dramatically below that for laminar flamelets when the turbulence intensity becomes high. Such behavior was not explainable with a one-step kinetic model, even at non-unity Lewis number. It could be due to depletion of H2 from the reaction zone by preferential diffusion. The capacity of the flame to generate radicals is critically dependent on the levels of H2 present (Bilger, et al., 1991). It seemed that a DNS computation with a multistep reduced mechanism would be worthwhile if a way could be found to make this feasible. Truly innovative approaches to complex problems often come only when there is the

  20. Albert Renold Memorial Lecture: Molecular Background of Nutritionally Induced Insulin Resistance Leading to Type 2 Diabetes – From Animal Models to Humans

    PubMed Central

    Shafrir, Eleazar

    2001-01-01

    Albert Renold strived to gain insight into the abnormalities of human diabetes by defining the pathophysiology of the disease peculiar to a given animal. He investigated the Israeli desert-derived spiny mice (Acomys cahirinus), which became obese on fat-rich seed diet. After a few months hyperplasia and hypertrophy of β-cells occurred leading to a sudden rupture, insulin loss and ketosis. Spiny mice were low insulin responders, which is probably a characteristic of certain desert animals, protecting against insulin oversecretion when placed on an abundant diet. We have compared the response to overstimulation of several mutant diabetic species and nutritionally induced nonmutant animals when placed on affluent diet. Some endowed with resilient β-cells sustain long-lasting oversecretion, compensating for the insulin resistance, without lapsing into overt diabetes. Some with labile beta cells exhibit apoptosis and lose their capacity of coping with insulin resistance after a relatively short period. The wide spectrum of response to insulin resistance among different diabetes prone species seems to represent the varying response of human beta cells among the populations. In search for the molecular background of insulin resistance resulting from overnutrition we have studied the Israeli desert gerbil Psammomys obesus (sand rat), which progresses through hyperinsulinemia, followed by hyperglycemia and irreversible beta cell loss. Insulin resistance was found to be the outcome of reduced activation of muscle insulin receptor tyrosine kinase by insulin, in association with diminished GLUT4 protein and DNA content and overexpression of PKC isoenzymes, notably of PKCε. This overexpression and translocation to the membrane was discernible even prior to hyperinsulinemia and may reflect the propensity to diabetes in nondiabetic species and represent a marker for preventive action. By promoting the phosphorylation of serine/threonine residues on certain proteins of the

  1. Differentiating of banked human umbilical cord blood-derived mesenchymal stem cells into insulin-secreting cells.

    PubMed

    Phuc, Pham Van; Nhung, Truong Hai; Loan, Dang Thi Tung; Chung, Doan Chinh; Ngoc, Phan Kim

    2011-01-01

    Umbilical cord blood (UCB)-derived mesenchymal stem cells (MSCs) are multipotent cells. They are able to differentiate into functional cells from not only mesoderm but also endoderm. Many researches showed that cells derived from fresh human UCB could transdifferentiate into insulin-secreting cells. In this study, transdifferentiating potential of cryopreserved human UCB-derived MSCs into insulin-secreting cell was investigated. Fresh human UCB was enriched the mononuclear cells by Ficoll-Paque centrifugation. The mononuclear cell population was cryopreserved in cryo-medium containing Iscove's modified Dulbecco's media (IMDM) with 10% DMSO at -196°C for 1 yr. After thawing, mononuclear cells were cultured to isolate MSCs in medium IMDM with 20% FBS supplemented with growth factors. At the fifth passages, MSCs were confirmed by flow cytometry about expression of CD13, CD14, CD34, CD45, CD166, and HLA-DR markers; after that, they were induced to differentiate into adipocytes and osteoblasts. After inducing with specific medium for islet differentiation, there were many clusters of cell like islet at day 14-28. Using real-time reverse transcription polymerase chain reaction (RT-PCR) to analyze the expression of functional genes, the result showed that Nestin, Pdx-1, Ngn3, Ils-1, Pax6, Pax4, Nkx2.2, Nkx6.1, Glut-2, Insulin genes expressed. The results showed that MSCs derived from banked cord blood can differentiate into functional pancreatic islet-like cells in vitro. If human MSCs, especially MSCs from banked cord blood of diabetes patients themselves can be isolated, proliferated, differentiated into functional pancreatic islet-like cells, and transplanted back into them (autologous transplantation), their high-proliferation potency and rejection avoidance will provide one promising therapy for diabetes.

  2. Relative contribution of PDX-1, MafA and E47/β2 to the regulation of the human insulin promoter

    PubMed Central

    2005-01-01

    The insulin promoter binds a number of tissue-specific and ubiquitous transcription factors. Of these, the homoeodomain protein PDX-1 (pancreatic duodenal homeobox factor-1), the basic leucine zipper protein MafA and the basic helix–loop–helix heterodimer E47/BETA2 (β-cell E box transactivator 2; referred to here as β2) bind to important regulatory sites. Previous studies have shown that PDX-1 can interact synergistically with E47 and β2 to activate the rat insulin 1 promoter. The aim of the present study was to determine the relative contribution of PDX-1, MafA and E47/β2 in regulating the human insulin promoter, and whether these factors could interact synergistically in the context of the human promoter. Mutagenesis of the PDX-1, MafA and E47/β2 binding sites reduced promoter activity by 60, 74 and 94% respectively, in INS-1 β-cells. In the islet glucagonoma cell line αTC1.6, overexpression of PDX-1 and MafA separately increased promoter activity approx. 2.5–3-fold, and in combination approx. 6-fold, indicating that their overall effect was additive. Overexpression of E47 and β2 had no effect. In HeLa cells, PDX-1 stimulated the basal promoter by approx. 40-fold, whereas MafA, E47 and β2 each increased activity by less than 2-fold. There was no indication of any synergistic effects on the human insulin promoter. On the other hand, the rat insulin 1 promoter and a mutated version of the human insulin promoter, in which the relevant regulatory elements were separated by the same distances as in the rat insulin 1 promoter, did exhibit synergy. PDX-1 was shown further to activate the endogenous insulin 1 gene in αTC1.6 cells, whereas MafA activated the insulin 2 gene. In combination, PDX-1 and MafA activated both insulin genes. Chromatin immunoprecipitation assays confirmed that PDX-1 increased the association of acetylated histones H3 and H4 with the insulin 1 gene and MafA increased the association of acetylated histone H3 with the insulin 2 gene

  3. Vasodilator effects of L-arginine are stereospecific and augmented by insulin in humans.

    PubMed

    Dallinger, Susanne; Sieder, Anna; Strametz, Jeanette; Bayerle-Eder, Michaela; Wolzt, Michael; Schmetterer, Leopold

    2003-06-01

    The amino acid l-arginine, the precursor of nitric oxide (NO) synthesis, induces vasodilation in vivo, but the mechanism behind this effect is unclear. There is, however, some evidence to assume that the l-arginine membrane transport capacity is dependent on insulin plasma levels. We hypothesized that vasodilator effects of l-arginine may be dependent on insulin plasma levels. Accordingly, we performed two randomized, double-blind crossover studies in healthy male subjects. In protocol 1 (n = 15), subjects received an infusion of insulin (6 mU x kg(-1) x min(-1) for 120 min) or placebo and, during the last 30 min, l-arginine or d-arginine (1 g/min for 30 min) x In protocol 2 (n = 8), subjects received l-arginine in stepwise increasing doses in the presence (1.5 mU x kg(-1) x min(-1)) or absence of insulin. Renal plasma flow and glomerular filtration rate were assessed by the para-aminohippurate and inulin plasma clearance methods, respectively. Pulsatile choroidal blood flow was assessed with laser interferometric measurement of fundus pulsation, and mean flow velocity in the ophthalmic artery was measured with Doppler sonography. l-arginine, but not d-arginine, significantly increased renal and ocular hemodynamic parameters. Coinfusion of l-arginine with insulin caused a dose-dependent leftward shift of the vasodilator effect of l-arginine. This stereospecific renal and ocular vasodilator potency of l-arginine is enhanced by insulin, which may result from facilitated l-arginine membrane transport, enhanced intracellular NO formation, or increased NO bioavailability.

  4. Insulin resistance after a 72-h fast is associated with impaired AS160 phosphorylation and accumulation of lipid and glycogen in human skeletal muscle

    PubMed Central

    Vendelbo, M. H.; Clasen, B. F. F.; Treebak, J. T.; Møller, L.; Krusenstjerna-Hafstrøm, T.; Madsen, M.; Nielsen, T. S.; Stødkilde-Jørgensen, H.; Pedersen, S. B.; Jørgensen, J. O. L.; Goodyear, L. J.; Wojtaszewski, J. F. P.; Møller, N.

    2012-01-01

    During fasting, human skeletal muscle depends on lipid oxidation for its energy substrate metabolism. This is associated with the development of insulin resistance and a subsequent reduction of insulin-stimulated glucose uptake. The underlying mechanisms controlling insulin action on skeletal muscle under these conditions are unresolved. In a randomized design, we investigated eight healthy subjects after a 72-h fast compared with a 10-h overnight fast. Insulin action on skeletal muscle was assessed by a hyperinsulinemic euglycemic clamp and by determining insulin signaling to glucose transport. In addition, substrate oxidation, skeletal muscle lipid content, regulation of glycogen synthesis, and AMPK signaling were assessed. Skeletal muscle insulin sensitivity was reduced profoundly in response to a 72-h fast and substrate oxidation shifted to predominantly lipid oxidation. This was associated with accumulation of both lipid and glycogen in skeletal muscle. Intracellular insulin signaling to glucose transport was impaired by regulation of phosphorylation at specific sites on AS160 but not TBC1D1, both key regulators of glucose uptake. In contrast, fasting did not impact phosphorylation of AMPK or insulin regulation of Akt, both of which are established upstream kinases of AS160. These findings show that insulin resistance in muscles from healthy individuals is associated with suppression of site-specific phosphorylation of AS160, without Akt or AMPK being affected. This impairment of AS160 phosphorylation, in combination with glycogen accumulation and increased intramuscular lipid content, may provide the underlying mechanisms for resistance to insulin in skeletal muscle after a prolonged fast. PMID:22028408

  5. Reduced Insulin Sensitivity Is Related to Less Endogenous Dopamine at D2/3 Receptors in the Ventral Striatum of Healthy Nonobese Humans

    PubMed Central

    Caravaggio, Fernando; Borlido, Carol; Hahn, Margaret; Feng, Zhe; Fervaha, Gagan; Gerretsen, Philip; Nakajima, Shinichiro; Plitman, Eric; Chung, Jun Ku; Iwata, Yusuke; Wilson, Alan; Remington, Gary

    2015-01-01

    Background: Food addiction is a debated topic in neuroscience. Evidence suggests diabetes is related to reduced basal dopamine levels in the nucleus accumbens, similar to persons with drug addiction. It is unknown whether insulin sensitivity is related to endogenous dopamine levels in the ventral striatum of humans. We examined this using the agonist dopamine D2/3 receptor radiotracer [11C]-(+)-PHNO and an acute dopamine depletion challenge. In a separate sample of healthy persons, we examined whether dopamine depletion could alter insulin sensitivity. Methods: Insulin sensitivity was estimated for each subject from fasting plasma glucose and insulin using the Homeostasis Model Assessment II. Eleven healthy nonobese and nondiabetic persons (3 female) provided a baseline [11C]-(+)-PHNO scan, 9 of which provided a scan under dopamine depletion, allowing estimates of endogenous dopamine at dopamine D2/3 receptor. Dopamine depletion was achieved via alpha-methyl-para-tyrosine (64mg/kg, P.O.). In 25 healthy persons (9 female), fasting plasma and glucose was acquired before and after dopamine depletion. Results: Endogenous dopamine at ventral striatum dopamine D2/3 receptor was positively correlated with insulin sensitivity (r(7)=.84, P=.005) and negatively correlated with insulin levels (r(7)=-.85, P=.004). Glucose levels were not correlated with endogenous dopamine at ventral striatum dopamine D2/3 receptor (r(7)=-.49, P=.18). Consistently, acute dopamine depletion in healthy persons significantly decreased insulin sensitivity (t(24)=2.82, P=.01), increased insulin levels (t(24)=-2.62, P=.01), and did not change glucose levels (t(24)=-0.93, P=.36). Conclusion: In healthy individuals, diminished insulin sensitivity is related to less endogenous dopamine at dopamine D2/3 receptor in the ventral striatum. Moreover, acute dopamine depletion reduces insulin sensitivity. These findings may have important implications for neuropsychiatric populations with metabolic

  6. Studies of Premixed Laminar and Turbulent Flames at Microgravity

    NASA Technical Reports Server (NTRS)

    Kwon, O. C.; Abid, M.; Porres, J.; Liu, J. B.; Ronney, P. D.; Struk, P. M.; Weiland, K. J.

    2003-01-01

    Several topics relating to premixed flame behavior at reduced gravity have been studied. These topics include: (1) flame balls; (2) flame structure and stability at low Lewis number; (3) experimental simulation of buoyancy effects in premixed flames using aqueous autocatalytic reactions; and (4) premixed flame propagation in Hele-Shaw cells. Because of space limitations, only topic (1) is discussed here, emphasizing results from experiments on the recent STS-107 Space Shuttle mission, along with numerical modeling efforts.

  7. Insulin-like growth factor-I receptor blockade by a specific tyrosine kinase inhibitor for human gastrointestinal carcinomas.

    PubMed

    Piao, Wenhua; Wang, Yu; Adachi, Yasushi; Yamamoto, Hiroyuki; Li, Rong; Imsumran, Arisa; Li, Hua; Maehata, Tadateru; Ii, Masanori; Arimura, Yoshiaki; Lee, Choon-Taek; Shinomura, Yasuhisa; Carbone, David P; Imai, Kohzoh

    2008-06-01

    Insulin-like growth factor-I receptor (IGF-IR) signaling is required for carcinogenicity and proliferation of gastrointestinal (GI) cancers. In this study, we sought to evaluate the effect of a new tyrosine kinase inhibitor of IGF-IR, NVP-AEW541, on the signal transduction and the progression of GI carcinomas. We assessed the effect of NVP-AEW541 on signal transduction, proliferation, survival, and migration in four GI cancer cells: colorectal adenocarcinoma HT29, pancreatic adenocarcinoma BxPC3, esophageal squamous cell carcinoma TE1, and hepatoma PLC/PRF/5. The effects of NVP-AEW541 alone and with chemotherapy were studied in vitro and in nude mouse xenografts. We also analyzed the effects of NVP-AEW541 on insulin signals and hybrid receptor formation between IGF-IR and insulin receptor. NVP-AEW541 blocked autophosphorylation of IGF-IR and both Akt and extracellular signal-regulated kinase activation by IGF but not by insulin. NVP-AEW541 suppressed proliferation and tumorigenicity in vitro in a dose-dependent manner in all cell lines. The drug inhibited tumor as a single agent and, when combined with stressors, up-regulated apoptosis in a dose-dependent fashion and inhibited mobility. NVP-AEW541 augmented the effects of chemotherapy on in vitro growth and induction of apoptosis. Moreover, the combination of NVP-AEW541 and chemotherapy was highly effective against tumors in mice. This compound did not influence hybrid receptor formation. Thus, NVP-AEW541 may have significant therapeutic utility in human GI carcinomas both alone and in combination with chemotherapy.

  8. Flame front configuration of turbulent premixed flames

    SciTech Connect

    Furukawa, Junichi; Maruta, Kaoru; Hirano, Toshisuke

    1998-02-01

    The present study is performed to explore dependence of the wrinkle scale of propane-air turbulent premixed flames on the characteristics of turbulence in the nonreacting flow, burner size, and mixture ratio. The wrinkle scales are examined and expressed in the frequency distribution of the radii of flame front curvatures. The average wrinkle scale depends not only on the characteristics of turbulence in the nonreacting flow but also on burner diameter and mixture ratio. The average wrinkle scale of a lean propane-air flame is larger than those of the near stoichiometric and rich flames. The smallest wrinkle scale of turbulent premixed flame is in the range of 0.75--1.0 mm, which is much larger than the Kolmogorov scale of turbulence in the nonreacting flow.

  9. Lifted Partially Premixed Flames in Microgravity

    NASA Technical Reports Server (NTRS)

    Lock, Andrew J.; Ganguly, Ranjan; Puri, Ishwar K.; Aggarwal, Suesh K.; Hegde, Uday

    2004-01-01

    Lifted Double and Triple flames are established in the UIC-NASA Partially Premixed microgravity rig. The flames examined in this paper are established above a coannular burner because its axisymmetric geometry allows for future implementation of other non-intrusive optical diagnostic techniques easily. Both burner-attached stable flames and lifted flames are established at normal and microgravity conditions in the drop tower facility.

  10. Gravity Effects Observed In Partially Premixed Flames

    NASA Technical Reports Server (NTRS)

    Puri, Ishwar K.; Aggarwal, Suresh K.; Lock, Andrew J.; Gauguly, Ranjan; Hegde, Uday

    2003-01-01

    Partially premixed flames (PPFs) contain a rich premixed fuel air mixture in a pocket or stream, and, for complete combustion to occur, they require the transport of oxidizer from an appropriately oxidizer-rich (or fuel-lean) mixture that is present in another pocket or stream. Partial oxidation reactions occur in fuel-rich portions of the mixture and any remaining unburned fuel and/or intermediate species are consumed in the oxidizer-rich portions. Partial premixing, therefore, represents that condition when the equivalence ratio (phi) in one portion of the flowfield is greater than unity, and in another section its value is less than unity. In general, for combustion to occur efficiently, the global equivalence ratio is in the range fuel-lean to stoichiometric. These flames can be established by design by placing a fuel-rich mixture in contact with a fuel-lean mixture, but they also occur otherwise in many practical systems, which include nonpremixed lifted flames, turbulent nonpremixed combustion, spray flames, and unwanted fires. Other practical applications of PPFs are reported elsewhere. Although extensive experimental studies have been conducted on premixed and nonpremixed flames under microgravity, there is a absence of previous experimental work on burner stabilized PPFs in this regard. Previous numerical studies by our group employing a detailed numerical model showed gravity effects to be significant on the PPF structure. We report on the results of microgravity experiments conducted on two-dimensional (established on a Wolfhard-Parker slot burner) and axisymmetric flames (on a coannular burner) that were investigated in a self-contained multipurpose rig. Thermocouple and radiometer data were also used to characterize the thermal transport in the flame.

  11. Displacement speeds in turbulent premixed flame simulations

    SciTech Connect

    Day, Marcus S.; Shepherd, Ian G.; Bell, J.; Grcar, Joseph F.; Lijewski, Michael J.

    2007-07-01

    The theory of turbulent premixed flames is based on acharacterization of the flame as a discontinuous surface propagatingthrough the fluid. The displacement speed, defined as the local speed ofthe flame front normal to itself, relative to the unburned fluid,provides one characterization of the burning velocity. In this paper, weintroduce a geometric approach to computing displacement speed anddiscuss the efficacy of the displacement speed for characterizing aturbulent flame.

  12. The role of dietary acid load and mild metabolic acidosis in insulin resistance in humans.

    PubMed

    Williams, Rebecca S; Kozan, Pinar; Samocha-Bonet, Dorit

    2016-05-01

    Type 2 diabetes is increasingly being recognised as a global health crisis (World Health Organisation). Insulin resistance is closely associated with obesity and precedes the development of type 2 diabetes. However, there is now increasing evidence to suggest that diet itself may independently be associated with type 2 diabetes risk. A diet with a high acid load (or high potential renal net acid load, PRAL) can result in a decrease in pH towards the lower end of the normal physiological range, which may in turn lead to the development of insulin resistance. Conversely, reducing dietary acid load (the so called 'alkaline diet') may be protective and prevent the onset of type 2 diabetes. Here, we explore the influence of dietary acid load on the development of mild metabolic acidosis and induction of insulin resistance. Whilst large prospective cohort studies link high dietary acid load or low serum bicarbonate with the development of type 2 diabetes, the effect of a diet with a low acid (or high alkaline) load remains unclear. Further interventional studies are required to investigate the influence of dietary composition on the body's acid/base balance, insulin resistance and incidence of type 2 diabetes. PMID:26363101

  13. Immunohistochemical localization of glucose transporters and insulin receptors in human fetal membranes at term.

    PubMed

    Wolf, H J; Desoye, G

    1993-11-01

    The localization has been investigated of the isoforms GLUT1, GLUT3 and GLUT4 of glucose transporter proteins as well as of insulin receptors. Fetal membranes (n = 10) were examined by immunohistochemical methods at the light and electron microscopic levels using mono- and polyclonal antibodies. In all amnion epithelial cells, GLUT1 and GLUT3 antibodies were bound to the apical membrane. Very rarely the GLUT1 antibody also immunostained the basolateral membrane and reacted weakly with the endomembrane system and membranes of the lateral cell protrusions. Fibroblasts reacted with the antibodies against GLUT1, GLUT4 and insulin receptor, whereas they were labelled only in one case with GLUT3 antibody. Cytotrophoblast cells were only stained with antibodies against GLUT1 and GLUT3. Antibodies against GLUT4 only reacted with fibroblasts in the membranes. On amnion epithelial cells, weak immunoreactivity with insulin receptor antibodies was detected only at the electron microscopic level. The data indicate: (1) GLUT1 is located on all cells of the amnion, whereas GLUT3 is present in detectable amounts only on amnion epithelial cells and cytotrophoblast; (2) GLUT1 and GLUT3 on amnion epithelial cells are predominantly located on the apical surface; (3) GLUT4 and insulin receptors are not regularly expressed. We suggest that amnion epithelial cells cover their basal glucose requirements from the amniotic fluid and not from the maternal circulation.

  14. Insulin, catecholamines, glucose and antioxidant enzymes in oxidative damage during different loads in healthy humans.

    PubMed

    Koska, J; Blazícek, P; Marko, M; Grna, J D; Kvetnanský, R; Vigas, M

    2000-01-01

    Exercise, insulin-induced hypoglycemia and oral glucose loads (50 g and 100 g) were used to compare the production of malondialdehyde and the activity of antioxidant enzymes in healthy subjects. Twenty male volunteers participated in the study. Exercise consisted of three consecutive work loads on a bicycle ergometer of graded intensity (1.5, 2.0, and 2.5 W/kg, 6 min each). Hypoglycemia was induced by insulin (Actrapid MC Novo, 0.1 IU/kg, i.v.). Oral administration of 50 g and 100 g of glucose was given to elevate plasma glucose. The activity of superoxide dismutase (SOD) was determined in red blood cells, whereas glutathione peroxidase (GSH-Px) activity was measured in whole blood. The concentration of malondialdehyde (MDA) was determined by HPLC, catecholamines were assessed radioenzymatically and glucose was measured by the glucose-oxidase method. Exercise increased MDA concentrations, GSH-Px and SOD activities as well as plasma noradrenaline and adrenaline levels. Insulin hypoglycemia increased plasma adrenaline levels, but the concentrations of MDA and the activities of GSH-Px and SOD were decreased. Hyperglycemia increased plasma MDA concentrations, but the activities of GSH-Px and SOD were significantly higher after a larger dose of glucose only. Plasma catecholamines were unchanged. These results indicate that the transient increase of plasma catecholamine and insulin concentrations did not induce oxidative damage, while glucose already in the low dose was an important triggering factor for oxidative stress. PMID:10984077

  15. Simulation of lean premixed turbulent combustion

    SciTech Connect

    Bell, John B.; Day, Marcus S.; Almgren, Ann S.; Lijewski, MichaelJ.; Rendleman, Charles A.; Cheng, Robert K.; Shepherd, Ian G.

    2006-06-25

    There is considerable technological interest in developingnew fuel-flexible combustion systems that can burn fuels such ashydrogenor syngas. Lean premixed systems have the potential to burn thesetypes of fuels with high efficiency and low NOx emissions due to reducedburnt gas temperatures. Although traditional scientific approaches basedon theory and laboratory experiment have played essential roles indeveloping our current understanding of premixed combustion, they areunable to meet the challenges of designing fuel-flexible lean premixedcombustion devices. Computation, with itsability to deal with complexityand its unlimited access to data, hasthe potential for addressing thesechallenges. Realizing this potential requires the ability to perform highfidelity simulations of turbulent lean premixed flames under realisticconditions. In this paper, we examine the specialized mathematicalstructure of these combustion problems and discuss simulation approachesthat exploit this structure. Using these ideas we can dramatically reducecomputational cost, making it possible to perform high-fidelitysimulations of realistic flames. We illustrate this methodology byconsidering ultra-lean hydrogen flames and discuss how this type ofsimulation is changing the way researchers study combustion.

  16. Premixed Turbulent Flame Propagation in Microgravity

    NASA Technical Reports Server (NTRS)

    Menon, Suresh

    1999-01-01

    A combined numerical-experimental study has been carried out to investigate the structure and propagation characteristics of turbulent premixed flames with and without the influence of buoyancy. Experimentally, the premixed flame characteristics are studied in the wrinkled regime using a Couette flow facility and an isotropic flow facility in order to resolve the scale of flame wrinkling. Both facilities were chosen for their ability to achieve sustained turbulence at low Reynolds number. This implies that conventional diagnostics can be employed to resolve the smallest scales of wrinkling. The Couette facility was also built keeping in mind the constraints imposed by the drop tower requirements. Results showed that the flow in this Couette flow facility achieves full-developed turbulence at low Re and all turbulence statistics are in good agreement with past measurements on large-scale facilities. Premixed flame propagation studies were then carried out both using the isotropic box and the Couette facility. Flame imaging showed that fine scales of wrinkling occurs during flame propagation. Both cases in Ig showed significant buoyancy effect. To demonstrate that micro-g can remove this buoyancy effect, a small drop tower was built and drop experiments were conducted using the isotropic box. Results using the Couette facility confirmed the ability to carry out these unique reacting flow experiments at least in 1g. Drop experiments at NASA GRC were planned but were not completed due to termination of this project.

  17. Insulin responsiveness of protein metabolism in vivo following bedrest in humans

    SciTech Connect

    Shangraw, R.E.; Stuart, C.A.; Prince, M.J.; Peters, E.J.; Wolfe, R.R. )

    1988-10-01

    To test the influence of bedrest on insulin regulation of leucine metabolism, six normal young men were subjected to a five-step hyperinsulinemic euglycemic clamp before and after 7 days of strict bedrest. A primed-constant infusion of (1-{sup 13}C)leucine was used. Before bedrest, the basal rate of appearance (R{sub a}) of intracellular leucine and leucine oxidation were 2.79{plus minus}0.17 and 0.613{plus minus}0.070 {mu}mol{center dot}kg{sup {minus}1}{center dot}min{sup {minus}1}, respectively. Insulin caused a dose-dependent reduction of the intracellular leucine R{sub a} and leucine oxidation to a minimum of 1.64{plus minus}0.08 and 0.322{plus minus}0.039 {mu}mol{center dot}kg{sup {minus}1}{center dot}min{sup {minus}1}, respectively, in nonbedrested subjects. Insulin also caused a dose-dependent reduction of plasma leucine concentration. After bedrest, subjects exhibited decreased glucose tolerance and increased endogenous insulin secretion, but basal and insulin-suppressed intracellular leucine R{sub a} and leucine oxidation rates were not different from control. Magnetic resonance imaging of the back and lower extremities revealed a 1-4% decrease in muscle volume and a 2-5% increase in fat volume secondary to bedrest. Bedrest also resulted in a negative nitrogen balance as compared with the control period. Thus because negative nitrogen balance and skeletal muscle atrophy occurred in six rested subjects in the absence of changes in the two indices of protein breakdown used in this study, it seems likely that muscle protein synthesis was inhibited.

  18. Overview of Clinical Trial Program and Applicability of Insulin Degludec/Insulin Aspart in Diabetes Management.

    PubMed

    Bantwal, Ganapathi; Wangnoo, Subhash K; Shunmugavelu, M; Nallaperumal, S; Harsha, K P; Bhattacharyya, Arpandev

    2015-05-01

    Insulin degludec/insulin aspart (IDegAsp) is the first soluble coformulation combining a long-acting insulin degludec (IDeg) and rapid-acting insulin aspart (IAsp). In patients with uncontrolled type 2 diabetes (T2DM) previously treated with insulins, IDegAsp twice daily effectively improves glycated haemoglobin (HbA1c) and fasting plasma glucose (FPG) levels with fewer hypoglycaemic episodes versus premix insulins. Further, insulin initiation with IDegAsp once daily provides superior long-term glycaemic control compared to insulin glargine with similar FPG and insulin doses, and numerically lower rates of overall and nocturnal hypoglycaemia. In patients with type 1 diabetes mellitus (T1DM), IDegAsp once daily and IAsp at remaining meals provides more convenient three injection regimen per day over conventional 4-5 injections based basal-bolus therapy. IDegAsp is an appropriate and reasonable option for intensifying insulin therapy in patients with T2DM and a relatively less complex treatment option for the management of T1DM. PMID:26548031

  19. Zinc-α2-Glycoprotein Modulates AKT-Dependent Insulin Signaling in Human Adipocytes by Activation of the PP2A Phosphatase

    PubMed Central

    Duran, Xavier; Pachón, Gisela; Vázquez-Carballo, Ana; Roche, Kelly; Núñez-Roa, Catalina; Garrido-Sánchez, Lourdes; Tinahones, Francisco J.; Vendrell, Joan; Fernández-Veledo, Sonia

    2015-01-01

    Objective Evidence from mouse models suggests that zinc-α2-glycoprotein (ZAG) is a novel anti-obesity adipokine. In humans, however, data are controversial and its physiological role in adipose tissue (AT) remains unknown. Here we explored the molecular mechanisms by which ZAG regulates carbohydrate metabolism in human adipocytes. Methods ZAG action on glucose uptake and insulin action was analyzed. β1 and β2-adrenoreceptor (AR) antagonists and siRNA targeting PP2A phosphatase were used to examine the mechanisms by which ZAG modulates insulin sensitivity. Plasma levels of ZAG were measured in a lean patient cohort stratified for HOMA-IR. Results ZAG treatment increased basal glucose uptake, correlating with an increase in GLUT expression, but induced insulin resistance in adipocytes. Pretreatment of adipocytes with propranolol and a specific β1-AR antagonist demonstrated that ZAG effects on basal glucose uptake and GLUT4 expression are mediated via β1-AR, whereas inhibition of insulin action is dependent on β2-AR activation. ZAG treatment correlated with an increase in PP2A activity. Silencing of the PP2A catalytic subunit abrogated the negative effect of ZAG on insulin-stimulated AKT phosphorylation and glucose uptake but not on GLUT4 expression and basal glucose uptake. ZAG circulating levels were unchanged in a lean patient cohort stratified for HOMA-IR. Neither glucose nor insulin was associated with plasma ZAG. Conclusions ZAG inhibits insulin-induced glucose uptake in human adipocytes by impairing insulin signaling at the level of AKT in a β2-AR- and PP2A-dependent manner. PMID:26068931

  20. The MAPK pathway and HIF-1 are involved in the induction of the human PAI-1 gene expression by insulin in the human hepatoma cell line HepG2.

    PubMed

    Dimova, Elitsa Y; Kietzmann, Thomas

    2006-12-01

    Enhanced levels of plasminogen activator inhibitor-1 (PAI-1) are considered to be a risk factor for pathological conditions associated with hypoxia or hyperinsulinemia. The expression of the PAI-1 gene is increased by insulin in different cells, although, the molecular mechanisms behind insulin-induced PAI-1 expression are not fully known yet. Here, we show that insulin upregulates human PAI-1 gene expression and promoter activity in HepG2 cells and that mutation of the hypoxia-responsive element (HRE)-binding hypoxia-inducible factor-1 (HIF-1) abolished the insulin effects. Mutation of E-boxes E4 and E5 abolished the insulin-dependent activation of the PAI-1 promoter only under normoxia, but did not affect it under hypoxia. Furthermore, the insulin effect was associated with activation of HIF-1alpha via mitogen-activated protein kinases (MAPKs) but not PDK1 and PKB in HepG2 cells. Furthermore, mutation of a putative FoxO1 binding site which was supposed to be involved in insulin-dependent PAI-1 gene expression influenced the insulin-dependent activation only under normoxia. Thus, insulin-dependent PAI-1 gene expression might be regulated by the action of both HIF-1 and FoxO1 transcription factors.

  1. Insulin pump therapy in pregnancy.

    PubMed

    Kesavadev, Jothydev

    2016-09-01

    Control of blood glucose during pregnancy is difficult because of wide variations, ongoing hormonal changes and mood swings. The need for multiple injections, pain at the injection site, regular monitoring and skillful handling of the syringes/pen further makes insulin therapy inconvenient. Insulin pump is gaining popularity in pregnancy because it mimics the insulin delivery of a healthy human pancreas. Multiple guidelines have also recommended the use of insulin pump in pregnancy to maintain the glycaemic control. The pump can release small doses of insulin continuously (basal), or a bolus dose close to mealtime to control the spike in blood glucose after a meal and the newer devices can shut down insulin delivery before the occurrence of hypoglycaemia. Pump insulin of choice is rapid acting analogue insulin. This review underscores the role of insulin pump in pregnancy, their usage, advantages and disadvantages in the light of existing literature and clinic experience. PMID:27582150

  2. Genetic insulin resistance is a potent regulator of gene expression and proliferation in human iPS cells.

    PubMed

    Iovino, Salvatore; Burkart, Alison M; Kriauciunas, Kristina; Warren, Laura; Hughes, Katelyn J; Molla, Michael; Lee, Youn-Kyoung; Patti, Mary-Elizabeth; Kahn, C Ronald

    2014-12-01

    Insulin resistance is central to diabetes and metabolic syndrome. To define the consequences of genetic insulin resistance distinct from those secondary to cellular differentiation or in vivo regulation, we generated induced pluripotent stem cells (iPSCs) from individuals with insulin receptor mutations and age-appropriate control subjects and studied insulin signaling and gene expression compared with the fibroblasts from which they were derived. iPSCs from patients with genetic insulin resistance exhibited altered insulin signaling, paralleling that seen in the original fibroblasts. Insulin-stimulated expression of immediate early genes and proliferation were also potently reduced in insulin resistant iPSCs. Global gene expression analysis revealed marked differences in both insulin-resistant iPSCs and corresponding fibroblasts compared with control iPSCs and fibroblasts. Patterns of gene expression in patients with genetic insulin resistance were particularly distinct in the two cell types, indicating dependence on not only receptor activity but also the cellular context of the mutant insulin receptor. Thus, iPSCs provide a novel approach to define effects of genetically determined insulin resistance. This study demonstrates that effects of insulin resistance on gene expression are modified by cellular context and differentiation state. Moreover, altered insulin receptor signaling and insulin resistance can modify proliferation and function of pluripotent stem cell populations. PMID:25059784

  3. Recombinant human insulin-like growth factor I induces its own specific carrier protein in hypophysectomized and diabetic rats.

    PubMed Central

    Zapf, J; Hauri, C; Waldvogel, M; Futo, E; Häsler, H; Binz, K; Guler, H P; Schmid, C; Froesch, E R

    1989-01-01

    The physiology of the specific serum binding proteins which constitute the main storage pool for insulin-like growth factors (IGFs) in mammals is still incompletely understood. We have, therefore, investigated the regulation of these proteins in (i) hypophysectomized (hypox) rats infused with recombinant human growth hormone (rhGH) or recombinant human IGF 1 (rhIGF I) and (ii) streptozotocin-diabetic rats infused with insulin or rhIGF I. The main carrier protein, a GH-dependent complex of apparent molecular mass 200 kDa, contains N-glycosylated IGF-binding subunits (42, 45, and 49 kDa) that differ in their glycosyl but not in their protein moiety. These subunits are lacking in hypox and diabetic rats. They are induced by GH and insulin, respectively, and appear in the 200-kDa complex. Infusion of rhIGF I induces the subunits in both states; however, only in diabetic, not in hypox, rats do they form the 200-kDa complex. Glycosylated carrier protein subunits do not appear before 8 hr of rhIGF I infusion. During that period, hypox rats may become severely hypoglycemic. After 16 hr, glycosylated subunits are clearly induced, and blood sugar values are normal. We conclude: (i) The N-glycosylated subunits of the 200-kDa complex reflect the IGF I status. (ii) IGF I may mediate the induction of these subunits by GH. (iii) Significant association to the 200-kDa complex occurs only in the presence of GH. It is likely that GH, but not IGF I, induces a component, which itself does not bind IGF, but associates with the glycosylated IGF-binding subunits. (iv) The glycosylated subunits protect against IGF-induced hypoglycemia and may be involved in tissue-specific targeting of IGFs. Images PMID:2471192

  4. Effect of pulmonary surfactant and phospholipid hexadecanol tyloxapol on recombinant human-insulin absorption from intratracheally administered dry powders in diabetic rats.

    PubMed

    Zheng, Jianheng; Zhang, Ge; Lu, Yang; Fang, Fang; He, Jiake; Li, Ning; Talbi, Amer; Zhang, Ying; Tang, Yue; Zhu, Jiabi; Chen, Xijing

    2010-12-01

    The purpose of the present study was to evaluate the enhancement effect of the natural pulmonary surfactant (PS) or its artificial substitute, phospholipid hexadecanol tyloxapol (PHT) on the bioavailability and hypoglycemic activity of recombinant human insulin (rh-insulin) in a pulmonary delivery system. PS- or PHT-loaded insulin formulation was administered to streptozotocin induced diabetic rats, at doses of 5 U/kg, 10 U/kg and 20 U/kg insulin, respectively. The hypoglycemic effect caused by PS or PHT containing rh-insulin was analyzed and the area above the curves (AAC) of serum glucose levels versus time, the minimum glucose concentration (C(min)), the time to C(min) (T(min)) and the pharmacological availability (PA%) were derived from the serum glucose profiles. Results showed that PS and PHT caused significantly decrease in serum glucose levels. The decrease in plasma glucose levels continued for about 5 h after the nadir. The highest AAC value was obtained when 20 U/kg rh-insulin with PS or PHT as absorption enhancer was administered to rats. AAC(0-360 min) of PS- or PHT-loaded rh-insulin was 2-3 times as much as that without PS or PHT and PA% increased by 1.3-2 fold. Thus, the extent of oral absorption of insulin from PS- or PHT-loaded particles was significantly greater when compared with that without them. In addition, PHT as well as PS did not change the lactate dehydrogenase (LDH) activity, alkaline phosphatase (AKP) activity and N-acetyl-β-D-glucoaminidase (NAG) activity in bronch fluid which are sensitive indicators of acute toxicity to lung cells in bronchoalveolar lavage (BAL). It is concluded that PS and PHT is a promising absorption enhancer for pulmonary delivery systems of large molecule drugs as rh-insulin.

  5. Granulocyte colony-stimulating factor (G-CSF): A saturated fatty acid-induced myokine with insulin-desensitizing properties in humans

    PubMed Central

    Ordelheide, Anna-Maria; Gommer, Nadja; Böhm, Anja; Hermann, Carina; Thielker, Inga; Machicao, Fausto; Fritsche, Andreas; Stefan, Norbert; Häring, Hans-Ulrich; Staiger, Harald

    2016-01-01

    Objective Circulating long-chain free fatty acids (FFAs) are important metabolic signals that acutely enhance fatty acid oxidation, thermogenesis, energy expenditure, and insulin secretion. However, if chronically elevated, they provoke inflammation, insulin resistance, and β-cell failure. Moreover, FFAs act via multiple signaling pathways as very potent regulators of gene expression. In human skeletal muscle cells differentiated in vitro (myotubes), we have shown in previous studies that the expression of CSF3, the gene encoding granulocyte colony-stimulating factor (G-CSF), is markedly induced upon FFA treatment and exercise. Methods and results We now report that CSF3 is induced in human myotubes by saturated, but not unsaturated, FFAs via Toll-like receptor 4-dependent and -independent pathways including activation of Rel-A, AP-1, C/EBPα, Src, and stress kinases. Furthermore, we show that human adipocytes and myotubes treated with G-CSF become insulin-resistant. In line with this, a functional polymorphism in the CSF3 gene affects adipose tissue- and whole-body insulin sensitivity and glucose tolerance in human subjects with elevated plasma FFA concentrations. Conclusion G-CSF emerges as a new player in FFA-induced insulin resistance and thus may be of interest as a target for prevention and treatment of type 2 diabetes. PMID:27069870

  6. IL-6 induction of TLR-4 gene expression via STAT3 has an effect on insulin resistance in human skeletal muscle.

    PubMed

    Kim, Tae Ho; Choi, Sung E; Ha, Eun Suk; Jung, Jong Gab; Han, Seung Jin; Kim, Hae Jin; Kim, Dae Jung; Kang, Yup; Lee, Kwan Woo

    2013-04-01

    We investigated the cytokines and mechanisms involved in the induction of insulin resistance in human skeletal muscle. Ten subjects with impaired glucose tolerance (IGT) and 10 control subjects were recruited. We performed biopsies on the vastus lateralis muscle and used immunoblotting to determine levels of inflammatory cytokines, Toll-like receptor (TLR) gene expression, and insulin signaling. We also used a human myotube culture system to examine the mechanisms underlying TLR-4 gene expression. To identify inflammatory cytokines associated with insulin resistance, we measured the levels of IL-6, TNF-α, TLR-2, and TLR-4 in skeletal muscle from non-obese patients with IGT and control subjects. Levels of IL-6, TNF-α, and TLR-4, but not TLR-2, were significantly increased in the IGT group. Insulin resistance decreased significantly in HSMMs following long-term IL-6 treatment. TLR-4 gene expression was significantly increased in human skeletal muscle myoblasts (HSMMs) treated with IL-6. To determine the main signaling pathway for IL-6-induced TLR-4 gene expression, we examined several signaling factors associated with IL-6 signaling pathways. We found that the active form of "signal transducer and activator of transcription 3" (STAT3) was increased. "Stattic" (a STAT3 inhibitor) markedly inhibited TLR-4 gene expression. IL-6 induction of TLR-4 gene expression via STAT3 is one of the main mechanisms underlying insulin resistance in human skeletal muscle.

  7. Insulin-like growth factor 1 receptors in human breast tumour: localisation and quantification by histo-autoradiographic analysis.

    PubMed Central

    Jammes, H.; Peyrat, J. P.; Ban, E.; Vilain, M. O.; Haour, F.; Djiane, J.; Bonneterre, J.

    1992-01-01

    To assess the precise role of IGF1 in benign and malignant breast diseases, we analysed the tissular localisation, characterised, and quantified specific insulin-like growth factor 1 (IGF1) binding sites in these heterogenous tissues, using histo-autoradiographic analysis (HAA). The 125I-IGF1 binding was performed on frozen tissue sections and analysed using 3H Ultrofilm autoradiography coupled to computerised image analysis. Competitive binding experiments using unlabelled IGF1, IGF2 and insulin showed that the tissues exhibited typical type I IGF binding sites. This specificity was confirmed by the use of alpha IR-3 monoclonal antibody, as inhibitor of 125I-IGF1 binding. IGF1 binding sites were detected in 18 human primary breast cancers, 12 benign breast tumours and two normal breast tissues. Using HAA we found that the human breast carcinomas studied exhibit a specific and high binding capacity for 125I-IGF1 exclusively localised on the proliferative epithelial component. The 125I-IGF1 binding activity of benign breast tumours or normal breast tissue was significantly lower than in cancerous tissues. There was a significant correlation between IGF1-R concentrations detected with HAA and those detected with a classical biochemical method. Moreover, HAA could be useful in further detailing whether a tumour is IGF1-R positive or negative HAA appears to be a useful method for the detection of growth factor receptors, specially in small biopsy specimens. Images Figure 2 Figure 3 PMID:1323990

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

    PubMed

    Lizunov, Vladimir A; Stenkula, Karin G; Blank, Paul S; Troy, Aaron; Lee, Jo-Ping; Skarulis, Monica C; Cushman, Samuel W; Zimmerberg, Joshua

    2015-01-01

    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. PMID:25768970

  9. Adipose tissue macrophages, low grade inflammation and insulin resistance in human obesity.

    PubMed

    Heilbronn, Leonie K; Campbell, Lesley V

    2008-01-01

    Obesity was first described as a low-grade inflammatory condition more than a decade ago. However, it is only relatively recently that obese individuals have been described with increased macrophage infiltration of adipose tissue, as well as an increase in the number of "M1" or "classically activated" macrophages. Furthermore, macrophages have been identified as the primary source of many of the circulating inflammatory molecules that are detected in the obese state and are postulated to be causal both in the development of insulin resistance and in the progression to type 2 diabetes. There is also novel evidence to suggest that macrophages inhibit adipocyte differentiation, potentially leading to adipocyte hypertrophy, altered secretion of adipokines and ectopic storage of lipid within liver, muscle and other non-adipose tissues. Currently, it is not clear what causes increased macrophage infiltration of adipose tissue in obese individuals. Theories include altered signalling by adipocytes, nutritional induction of metabolic endotoxemia or reduced angiogenesis and local adipose cell hypoxia. Importantly, PPAR-gamma agonists have been shown to alter macrophage phenotype to "M2" or an "alternatively activated" anti-inflammatory phenotype and may induce macrophage specific cell death. Consequently, excitement surrounds the potential for specific inhibition of macrophage infiltration of adipose tissue via pharmacotherapy for obese patients and more particularly as adjunct therapy to improve insulin sensitivity in obese individuals with insulin resistance and overt type 2 diabetes.

  10. Glucose metabolism and the response to insulin by human adipose tissue in spontaneous and experimental obesity. Effects of dietary composition and adipose cell size.

    PubMed

    Salans, L B; Bray, G A; Cushman, S W; Danforth, E; Glennon, J A; Horton, E S; Sims, E A

    1974-03-01

    [1-(14)C]glucose oxidation to CO(2) and conversion into glyceride by adipose tissue from nonobese and obese subjects has been studied in vitro in the presence of varying medium glucose and insulin concentrations as functions of adipose cell size, the composition of the diet, and antecedent weight gain or loss. Increasing medium glucose concentrations enhance the incorporation of glucose carbons by human adipose tissue into CO(2) and glyceride-glycerol. Insulin further stimulates the conversion of glucose carbons into CO(2), but not into glyceride-glycerol. Incorporation of [1-(14)C]glucose into glyceride-fatty acids by these tissues could not be demonstrated under any of the conditions tested. Both adipose cell size and dietary composition influence the in vitro metabolism of glucose in, and the response to insulin by, human adipose tissue. During periods of ingestion of weight-maintenance isocaloric diets of similar carbohydrate, fat, and protein composition, increasing adipose cell size is associated with (a) unchanging rates of glucose oxidation and increasing rates of glucose carbon incorporation into glyceride-glycerol in the absence of insulin, but (b) decreasing stimulation of glucose oxidation by insulin. On the other hand, when cell size is kept constant, increasing dietary carbohydrate intake is associated with an increased basal rate of glucose metabolism and response to insulin by both small and large adipose cells. Thus, the rate of glucose oxidation and the magnitude of the insulin response of large adipose cells from individuals ingesting a high carbohydrate diet may be similar to or greater than that in smaller cells from individuals ingesting an isocaloric lower carbohydrate diet.The alterations in basal glucose metabolism and insulin response observed in adipose tissue from patients with spontaneous obesity are reproduced by weight gain induced experimentally in nonobese volunteers; these metabolic changes are reversible with weight loss. The

  11. Diabetes-associated mutations in human insulin: crystal structure and photo-cross-linking studies of a-chain variant insulin Wakayama.

    PubMed

    Wan, Zhu-li; Huang, Kun; Xu, Bin; Hu, Shi-Quan; Wang, Shuhua; Chu, Ying-Chi; Katsoyannis, Panayotis G; Weiss, Michael A

    2005-04-01

    Naturally occurring mutations in insulin associated with diabetes mellitus identify critical determinants of its biological activity. Here, we describe the crystal structure of insulin Wakayama, a clinical variant in which a conserved valine in the A chain (residue A3) is substituted by leucine. The substitution occurs within a crevice adjoining the classical receptor-binding surface and impairs receptor binding by 500-fold, an unusually severe decrement among mutant insulins. To resolve whether such decreased activity is directly or indirectly mediated by the variant side chain, we have determined the crystal structure of Leu(A3)-insulin and investigated the photo-cross-linking properties of an A3 analogue containing p-azidophenylalanine. The structure, characterized in a novel crystal form as an R(6) zinc hexamer at 2.3 A resolution, is essentially identical to that of the wild-type R(6) hexamer. The variant side chain remains buried in a nativelike crevice with small adjustments in surrounding side chains. The corresponding photoactivatable analogue, although of low affinity, exhibits efficient cross-linking to the insulin receptor. The site of photo-cross-linking lies within a 14 kDa C-terminal domain of the alpha-subunit. This domain, unrelated in sequence to the major insulin-binding region in the N-terminal L1 beta-helix, is also contacted by photoactivatable probes at positions A8 and B25. Packing of Val(A3) at this interface may require a conformational change in the B chain to expose the A3-related crevice. The structure of insulin Wakayama thus evokes the reasoning of Sherlock Holmes in "the curious incident of the dog in the night": the apparent absence of structural perturbations (like the dog that did not bark) provides a critical clue to the function of a hidden receptor-binding surface.

  12. Microencapsulation of human insulin DEAE-dextran complex and the complex in liposomes by the emulsion non-solvent addition method.

    PubMed

    Manosroi, A; Manosroi, J

    1997-01-01

    Human insulin-DEAE (diethyl amino ethyl) dextran complex and human insulin DEAE-dextran complex in liposomes were encapsulated in cellulose acetate butyrate (CAB) microcapsules by the emulsion non-solvent addition method. The ratio of core-to-coat used was 1:1. The average diameters of the complex microcapsules and the complex liposome microcapsules were 239.5 +/- 77.5 and 182.9 +/- 52.2 microns respectively. In vitro dissolution studies of both types of microcapsules in simulated intestinal fluid at pH 7.2 showed a sustained release of the complex and the complex liposome microcapsules with t50 = 1.5 h and 4 h respectively. This study can be applied to the further development of oral formulations of human insulin liposomes for diabetic treatment.

  13. Cell motility in models of wounded human skin is improved by Gap27 despite raised glucose, insulin and IGFBP-5

    SciTech Connect

    Wright, Catherine S.; Berends, Rebecca F.; Flint, David J.; Martin, Patricia E.M.

    2013-02-15

    Reducing Cx43 expression stimulates skin wound healing. This is mimicked in models when Cx43 function is blocked by the connexin mimetic peptide Gap27. IGF-I also stimulates wound healing with IGFBP-5 attenuating its actions. Further, the IGF-I to IGFBP-5 ratio is altered in diabetic skin, where wound closure is impaired. We investigated whether Gap27 remains effective in augmenting scrape-wound closure in human skin wound models simulating diabetes-induced changes, using culture conditions with raised glucose, insulin and IGFBP-5. Gap27 increased scrape-wound closure in normal glucose and insulin (NGI) and to a lesser extent in high glucose and insulin (HGI). IGF-I enhanced scrape-wound closure in keratinocytes whereas IGFBP-5 inhibited this response. Gap27 overcame the inhibitory effects of IGFBP-5 on IGF-I activity. Connexin-mediated communication (CMC) was reduced in HGI, despite raised Cx43, and Gap27 significantly decreased CMC in NGI and HGI. IGF-I and IGFBP-5 did not affect CMC. IGF-I increased keratinocyte proliferation in NGI, and Gap27 increased proliferation in NGI to a greater extent than in HGI. We conclude that IGF-I and Gap27 stimulate scrape-wound closure by independent mechanisms with Gap27 inhibiting Cx43 function. Gap27 can enhance wound closure in diabetic conditions, irrespective of the IGF-I:IGFBP-5 balance. - Highlights: ► Human organotypic and keratinocyte ‘diabetic’ skin models were used to demonstrate the ability of Gap27 to improve scrape-wound closure. ► Gap27 enhanced scrape-wound closure by reducing Cx43-mediated communication, whereas IGFBP-5 retarded cell migration. ► IGF-I and IGFBP-5 did not affect connexin-mediated pathways. ► Gap27 can override altered glucose, insulin, IGF-I, and IGFBP-5 in ‘diabetic’ skin models and thus has therapeutic potential.

  14. Action of a semi-synthetic human insulin preparation (30/70 NPH insulin Organon): comparison with another 30/70 NPH (Actraphane Novo).

    PubMed

    Gin, H; Nivet, A M; Morlat, P; Aubertin, J

    1989-01-01

    The action of a new semi-synthetic insulin preparation (30% soluble, 70% NPH insulin from Organon Laboratories Eragny sur Epte France) was studied in 6 healthy male volunteers using the euglycemic clamp technique (Biostator GCIIS) and compared with another 30/70 NPH (Actraphane Novo). Insulin levels, inhibition of C peptide secretion and glucose consumption were determined. There was a time lag between the maximum glucose need (167 +/- 18 mg/Kg/15 min at the 195th minute after the injection) and the peak plasma insulin level (98.3 +/- 8.5 uu/ml at the 105th minute following the injection). The maximum glucose need was followed by a slow fall in insulin levels with a duration of action of 17 hours. The total glucose need was the same as for Actraphane, although Actraphane had a slower action with a lower peak glucose need (144 +/- 18 mg/Kg/15 min at the 280th minute after injection). The two preparations had the same duration of action.

  15. Adipose triglyceride lipase expression in human adipose tissue and muscle. Role in insulin resistance and response to training and pioglitazone

    PubMed Central

    Yao-Borengasser, Aiwei; Varma, Vijayalakshmi; Coker, Robert H.; Ranganathan, Gouri; Phanavanh, Bounleut; Rasouli, Neda; Kern, Philip A.

    2010-01-01

    Objective Adipose triglyceride lipase (ATGL) catalyzes the first step in adipocyte and muscle triglyceride hydrolysis, and Comparative Gene Identification-58 (CGI-58) is an essential cofactor. We studied the expression of ATGL and CGI-58 in human adipose and muscle, and examined correlations with markers of muscle fatty acid oxidation. Materials/Methods Non diabetic volunteers were studied. Subjects with impaired glucose tolerance were treated with pioglitazone or metformin for 10 weeks. Normal glucose tolerant subjects underwent a 12 week training program. We examined changes in ATGL and CGI-58 with obesity and insulin resistance, and effects of exercise and pioglitazone. Results ATGL mRNA expression showed no correlation with either body mass index (BMI) or insulin sensitivity (SI) in either adipose or muscle. However, adipose ATGL protein levels were inversely correlated with BMI (r=−0.64, p<0.02), and positively correlated with SI (r=0.67, p<0.02). In muscle, ATGL mRNA demonstrated a strong positive relationship with carnitine palmitoyltransferase I mRNA (r=0.82, p<0.0001), and the adiponectin receptors AdipoR1 mRNA (r=0.71, p<0.0001), and AdipoR2 mRNA (r=0.74, p<0.0001). Muscle CGI-58 mRNA was inversely correlated with intramyocellular triglyceride in both type 1 (r=−0.35, p<0.05) and type 2 (r=−0.40, p<0.05) fibers. Exercise training resulted in increased muscle ATGL and pioglitazone increased adipose ATGL by 31% (p<0.05). Pioglitazone also increased ATGL in adipocytes. Conclusions Adipose ATGL protein is decreased with insulin resistance and obesity, and muscle ATGL mRNA is associated with markers of fatty acid oxidation in muscle, as is CGI-58. The regulation of ATGL and CGI-58 have important implications for the control of lipotoxicity. PMID:21129760

  16. Portal Vein Glucose Sensors Do Not Play a Major Role in Modulating Physiological Responses to Insulin-Induced Hypoglycemia in Humans

    PubMed Central

    Rossetti, Paolo; Porcellati, Francesca; Lucidi, Paola; Busciantella Ricci, Natalia; Candeloro, Paola; Cioli, Patrizia; Santeusanio, Fausto; Bolli, Geremia B.; Fanelli, Carmine G.

    2009-01-01

    OBJECTIVE—Experimental data from animal studies indicate that portal vein glucose sensors play a key role in the responses to slow-fall hypoglycemia. However, their role in modulating these responses in humans is not well understood. The aim of the present study was to examine in humans the potential role of portal vein glucose sensors in physiological responses to insulin-induced hypoglycemia mimicking the slow fall of insulin-treated diabetic subjects. RESEARCH DESIGN AND METHODS—Ten nondiabetic subjects were studied on two different occasions during intravenous insulin (2 mU · kg−1 · min−1) plus variable glucose for 160 minutes. In both studies, after 60 min of normal plasma glucose concentrations, hypoglycemia (47 mg/dl) was induced slowly (60 min) and maintained for 60 min. Hypoglycemia was preceded by the ingestion of either oral placebo or glucose (28 g) given at 30 min. RESULTS—Plasma glucose and insulin were not different with either placebo or glucose (P > 0.2). Similarly, counterregulatory hormones, substrates, and symptoms were not different with either placebo or glucose. The Stroop color and colored words subtest of the Stroop test deteriorated less (P < 0.05) with glucose than placebo. CONCLUSIONS—In contrast to animals, in humans, prevention of portal hypoglycemia with oral glucose from the beginning of insulin-induced slow-fall hypoglycemia has no effect on sympathoadrenal and symptomatic responses to hypoglycemia. PMID:18852332

  17. Quantitative measurement of full-length and C-terminal proteolyzed RBP4 in serum of normal and insulin-resistant humans using a novel mass spectrometry immunoassay.

    PubMed

    Yang, Qin; Eskurza, Iratxe; Kiernan, Urban A; Phillips, David A; Blüher, Matthias; Graham, Timothy E; Kahn, Barbara B

    2012-03-01

    Serum retinol-binding protein 4 (RBP4) levels are increased in insulin-resistant humans and correlate with severity of insulin resistance in metabolic syndrome. Quantitative Western blotting (qWestern) has been the most accurate method for serum RBP4 measurements, but qWestern is technically complex and labor intensive. The lack of a reliable, high-throughput method for RBP4 measurements has resulted in variability in findings in insulin-resistant humans. Many commonly used ELISAs have limited dynamic range. Neither the current ELISAs nor qWestern distinguish among full-length and carboxyl terminus proteolyzed forms of circulating RBP4 that are altered in different medical conditions. Here, we report the development of a novel quantitative mass spectrometry immunoaffinity assay (qMSIA) to measure full-length and proteolyzed forms of RBP4. qMSIA and qWestern of RBP4 were performed in identical serum aliquots from insulin-sensitive/normoglycemic or insulin-resistant humans with impaired glucose tolerance or type 2 diabetes. Total RBP4 qMSIA measurements were highly similar to qWestern and correlated equally well with clinical severity of insulin resistance (assessed by clamp glucose disposal rate, r = -0.74), hemoglobin A1c (r = 0.63), triglyceride/high-density lipoprotein (r = 0.55), waist/hip (r = 0.61), and systolic blood pressure (r = 0.53, all P < 0.001). Proteolyzed forms of RBP4 accounted for up to 50% of total RBP4 in insulin-resistant subjects, and des(Leu)-RBP4 (cleavage of last leucine) correlated highly with insulin resistance (assessed by glucose disposal rate, r = -0.69). In multiple regression analysis, insulin resistance but not glomerular filtration rate was the strongest, independent predictor of serum RBP4 levels. Thus, qMSIA provides a novel tool for accurately measuring serum RBP4 levels as a biomarker for severity of insulin resistance and risk for type 2 diabetes and metabolic syndrome. PMID:22253430

  18. Can we characterize turbulence in premixed flames?

    SciTech Connect

    Lipatnikov, A.N.

    2009-06-15

    Modeling of premixed turbulent combustion involves averaging reaction rates in turbulent flows. The focus of most approaches to resolving this problem has been placed on determining the dependence of the mean rate w of product creation on the laminar flame speed S{sub L}, the rms turbulence velocity u', etc. The goal of the present work is to draw attention to another issue: May the input quantity u{sup '} for a model of w= w(u'/S{sub L},..) be considered to be known? The point is that heat release substantially affects turbulence and, hence, turbulence characteristics in premixed flames should be modeled. However, standard moment methods for numerically simulating turbulent flows do not allow us to evaluate the true turbulence characteristics in a flame. For instance, the Reynolds stresses in premixed flames are affected not only by turbulence itself, but also by velocity jump across flamelets. A common way to resolving this problem consists of considering the Reynolds stresses conditioned on unburned (or burned) mixture to be the true turbulence characteristics. In the present paper, this widely accepted but never proved hypothesis is put into question, first, by considering simple model constant-density problems (flame motion in an oscillating one-dimensional laminar flow; flame stabilized in a periodic shear, one-dimensional, laminar flow; turbulent mixing). In all the cases, the magnitude of velocity fluctuations, calculated using the conditioned Reynolds stresses, is affected by the intermittency of reactants and products and, hence, is not the true rms velocity. Second, the above claim is further supported by comparing balance equations for the mean and conditioned Reynolds stresses. The conditioned Reynolds stresses do not characterize the true turbulence in flames, because conditional averaging cuts off flow regions characterized by either high or low velocities. (author)

  19. Premixed rapid-setting calcium phosphate composites for bone repair.

    PubMed

    Carey, Lisa E; Xu, Hockin H K; Simon, Carl G; Takagi, Shozo; Chow, Laurence C

    2005-08-01

    Although calcium phosphate cement (CPC) is promising for bone repair, its clinical use requires on site powder-liquid mixing. To shorten surgical time and improve graft properties, it is desirable to develop premixed CPC in which the paste remains stable during storage and hardens only after placement into the defect. The objective of this study was to develop premixed CPC with rapid setting when immersed in a physiological solution. Premixed CPCs were formulated using the following approach: Premixed CPC = CPC powder + nonaqueous liquid + gelling agent + hardening accelerator. Three premixed CPCs were developed: CPC-monocalcium phosphate monohydrate (MCPM), CPC-chitosan, and CPC-tartaric. Setting time for these new premixed CPCs ranged from 5.3 to 7.9 min, significantly faster than 61.7 min for a premixed control CPC reported previously (p < 0.05). SEM revealed the formation of nano-sized needle-like hydroxyapatite crystals after 1 d immersion and crystal growth after 7 d. Diametral tensile strength for premixed CPCs at 7 d ranged from 2.8 to 6.4 MPa, comparable to reported strengths for cancellous bone and sintered porous hydroxyapatite implants. Osteoblast cells attained a normal polygonal morphology on CPC-MCPM and CPC-chitosan with cytoplasmic extensions adhering to the nano-hydroxyapatite crystals. In summary, fast-setting premixed CPCs were developed to avoid the powder-liquid mixing in surgery. The pastes hardened rapidly once immersed in physiological solution and formed hydroxyapatite. The cements had strengths matching those of cancellous bone and sintered porous hydroxyapatite and non-cytotoxicity similar to conventional non-premixed CPC.

  20. Premixed rapid-setting calcium phosphate composites for bone repair✩

    PubMed Central

    Carey, Lisa E.; Xu, Hockin H.K.; Simon, Carl G.; Takagi, Shozo; Chow, Laurence C.

    2009-01-01

    Although calcium phosphate cement (CPC) is promising for bone repair, its clinical use requires on site powder–liquid mixing. To shorten surgical time and improve graft properties, it is desirable to develop premixed CPC in which the paste remains stable during storage and hardens only after placement into the defect. The objective of this study was to develop premixed CPC with rapid setting when immersed in a physiological solution. Premixed CPCs were formulated using the following approach: Premixed CPC = CPC powder+nonaqueous liquid+gelling agent+hardening accelerator. Three premixed CPCs were developed: CPC–monocalcium phosphate monohydrate (MCPM), CPC–chitosan, and CPC–tartaric. Setting time for these new premixed CPCs ranged from 5.3 to 7.9 min, significantly faster than 61.7 min for a premixed control CPC reported previously (p<05). SEM revealed the formation of nano-sized needle-like hydroxyapatite crystals after 1 d immersion and crystal growth after 7 d. Diametral tensile strength for premixed CPCs at 7 d ranged from 2.8 to 6.4 MPa, comparable to reported strengths for cancellous bone and sintered porous hydroxyapatite implants. Osteoblast cells attained a normal polygonal morphology on CPC–MCPM and CPC–chitosan with cytoplasmic extensions adhering to the nano-hydroxyapatite crystals. In summary, fast-setting premixed CPCs were developed to avoid the powder–liquid mixing in surgery. The pastes hardened rapidly once immersed in physiological solution and formed hydroxyapatite. The cements had strengths matching those of cancellous bone and sintered porous hydroxyapatite and non-cytotoxicity similar to conventional non-premixed CPC. PMID:15769536

  1. Premixed Combustion Model for Boron Clouds

    NASA Astrophysics Data System (ADS)

    Wang, Mengze; Han, Wang; Chen, Zheng

    2015-11-01

    Boron particle is an ideal additive in solid propellants and fuels due to its very high volumetric heat release. In this study, a premixed combustion model for boron clouds is developed based on a previous combustion model for single boron particle. The flame structure is assumed to be composed of three zones: the preheat zone, the ignition zone, and the reaction zone, and analytical solutions are derived from the governing equations. Consequently the influence of the boron clouds' physical properties on the flame propagation process is investigated.

  2. Polypeptides with nonsuppressible insulin-like and cell-growth promoting activities in human serum: isolation, chemical characterization, and some biological properties of forms I and II.

    PubMed

    Rinderknecht, E; Humbel, R E

    1976-07-01

    Serum contains a polypeptide with insulin-like activity not suppressible by insulin antibodies (NSILA). A large-scale isolation procedure for NSILA is described, starting from an acid ethanol extract of a Cohn fraction (precipitate B) obtained from human plasma. Two homogenous polypeptides with insulin-like and cell-growth promoting activities could be isolated by gel filtration, ion exchange chromatography, and preparative polyacrylamide gel electrophoresis. Both components are slightly basic polypeptides with a minimal molecular weight of 5800 +/- 400. Both are single-chain molecules with two intrachain disulfide bridges each and no free sulfhydryl groups. NSILA I and II differ, however, in their amino acid compositions. The N-terminal amino acid sequences are Gly-Pro-Glu- in NSILA I, and Ala-Tyr-Arg- and Tyr-Arg- in NSILA II. Both NSILA I and II enhance net gas exchange in adipose tissue with a specific activity 60 times lower than that of insulin. In the range of 1-50 ng/ml, both substances stimulate [3H]thymidine incorporation into DNA of chick embryo fibroblasts. The same effect can be obtained with insulin but only at concentrations 50-100 times higher than those of NSILA. These results suggest that NSILA I and II are two forms of an insulin-like hormone with predominating effects on cell and tissue growth parameters.

  3. Insulin Signaling And Insulin Resistance

    PubMed Central

    Beale, Elmus G.

    2013-01-01

    Insulin resistance or its sequelae may be the common etiology of maladies associated with metabolic syndrome (e.g., hypertension, type 2 diabetes, atherosclerosis, heart attack, stroke and kidney failure). It is thus important to understand those factors that affect insulin sensitivity. This review stems from the surprising discovery that interference with angiotensin signaling improves insulin sensitivity and it provides a general overview of insulin action and factors that control insulin sensitivity. PMID:23111650

  4. Analysis of the add-on effect of α-glucosidase inhibitor, acarbose in insulin therapy: A pilot study

    PubMed Central

    Li, Feng-Fei; Fu, Li-Yuan; Xu, Xiao-Hua; Su, Xiao-Fei; Wu, Jin-Dan; Ye, Lei; Ma, Jian-Hua

    2016-01-01

    The aim of the present study was to evaluate the add-on effect of acarbose therapy in oxidative stress, and the lipid and inflammatory profiles of patients with type 2 diabetes mellitus (T2DM) treated with insulin. This was an open and unblended study. Patients (n=134) with T2DM (haemoglobin A1c range, 9.0–12.0%) were recruited. After continuous subcutaneous insulin infusion for 7 days for initial rapid correction of hyperglycaemia, a premixed insulin titration period (duration, 4–6 days) subsequently followed. Patients were then randomized (1:1) into two groups as follows: An acarbose plus pre-mixed 30/70 insulin group or a pre-mixed 30/70 insulin only group; each group received treatment for 2 weeks. Plasma high-sensitivity C-reactive protein (Hs-CRP), 8-iso-prostaglandin F2α (8-iso PGF2α), tumor necrosis factor-α (TNF-α), interleukin (IL)-1β, and IL-6 levels were measured before and after therapy. Patients that received acarbose plus insulin demonstrated greater reduction in 8-iso PGF2α, Hs-CRP, TNF-α, IL-1β and IL-6 levels when compared with the insulin only patients. Thus, acarbose add-on insulin therapy was identified to be associated with greater improvements in oxidative stress and inflammation in patients with T2DM when compared with those that received insulin only therapy.

  5. Induction of human umbilical cord blood-derived stem cells with embryonic stem cell phenotypes into insulin producing islet-like structure.

    PubMed

    Sun, Bo; Roh, Kyung-Hwan; Lee, Sae-Rom; Lee, Yong-Soon; Kang, Kyung-Sun

    2007-03-23

    Success in islet-transplantation-based therapies for type I diabetes, coupled with a worldwide shortage of transplant-ready islets, has motivated efforts to develop renewable sources of islet-replacement tissue. Embryonic stem cells (ESCs) have been successfully induced into insulin producing islet-like structure in several studies. However, the source of the ESCs has presented ethical and technical concerns. Here, we isolated a population of stem cells from human cord blood (UCB), which expressed embryo stage specific maker, SSEA-4, and the multi-potential stem cell marker, Oct4. Subsequently, we successfully induced them into insulin-producing islet-like structures, which co-express insulin and C-peptide. These findings might have a significant potential to advance human UCB derived stem-cell-based therapeutics for diabetes.

  6. Molecular bases for the recognition of short peptide substrates and cysteine-directed modifications of human insulin-degrading enzyme

    PubMed Central

    Malito, Enrico; Ralat, Luis A.; Manolopoulou, Marika; Tsay, Julie L.; Wadlington, Natasha L.; Tang, Wei-Jen

    2009-01-01

    Insulin degrading enzyme (IDE) utilizes a large catalytic chamber to selectively bind and degrade peptide substrates such as insulin and amyloid β (Aβ). Tight interactions with substrates occur at an exosite located ~30Å away from the catalytic center that anchors the N-terminus of substrates to facilitate binding and subsequent cleavages at the catalytic site. However, IDE also degrades peptide substrates that are too short to occupy both the catalytic site and the exosite simultaneously. Here, we use kinins as a model system to address the kinetics and regulation of human IDE with short peptides. IDE specifically degrades bradykinin and kallidin at the Pro/Phe site. A 1.9Å crystal structure of bradykinin-bound IDE reveals the binding of bradykinin to the exosite, and not to the catalytic site. In agreement with observed high Km values, this suggests low affinity of bradykinin for IDE. This structure also provides the molecular basis on how the binding of short peptides at the exosite could regulate substrate recognition. We also found that human IDE is potently inhibited by physiologically relevant concentrations of S-nitrosylation and oxidation agents. Cysteine-directed modifications play a key role, since an IDE mutant devoid of all thirteen cysteines is insensitive to the inhibition by S-nitroso-glutathione, hydrogen peroxide, or N-ethylmaleimide. Specifically, cysteine 819 of human IDE is located inside the catalytic chamber pointing towards an extended hydrophobic pocket and is critical for the inactivation. Thiol-directed modification of this residue likely causes local structural perturbation to reduce substrate binding and catalysis. PMID:18986166

  7. Molecular bases for the recognition of short peptide substrates and cysteine-directed modifications of human insulin-degrading enzyme.

    PubMed

    Malito, Enrico; Ralat, Luis A; Manolopoulou, Marika; Tsay, Julie L; Wadlington, Natasha L; Tang, Wei-Jen

    2008-12-01

    Insulin degrading enzyme (IDE) utilizes a large catalytic chamber to selectively bind and degrade peptide substrates such as insulin and amyloid beta (Abeta). Tight interactions with substrates occur at an exosite located approximately 30 A away from the catalytic center that anchors the N-terminus of substrates to facilitate binding and subsequent cleavages at the catalytic site. However, IDE also degrades peptide substrates that are too short to occupy both the catalytic site and the exosite simultaneously. Here, we use kinins as a model system to address the kinetics and regulation of human IDE with short peptides. IDE specifically degrades bradykinin and kallidin at the Pro/Phe site. A 1.9 A crystal structure of bradykinin-bound IDE reveals the binding of bradykinin to the exosite and not to the catalytic site. In agreement with observed high K(m) values, this suggests low affinity of bradykinin for IDE. This structure also provides the molecular basis on how the binding of short peptides at the exosite could regulate substrate recognition. We also found that human IDE is potently inhibited by physiologically relevant concentrations of S-nitrosylation and oxidation agents. Cysteine-directed modifications play a key role, since an IDE mutant devoid of all 13 cysteines is insensitive to the inhibition by S-nitrosoglutathione, hydrogen peroxide, or N-ethylmaleimide. Specifically, cysteine 819 of human IDE is located inside the catalytic chamber pointing toward an extended hydrophobic pocket and is critical for the inactivation. Thiol-directed modification of this residue likely causes local structural perturbation to reduce substrate binding and catalysis.

  8. Molecular Bases for the Recognition of Short Peptide Substrates and Cysteine-Directed Modifications of Human Insulin-Degrading Enzyme

    SciTech Connect

    Malito, Enrico; Ralat, Luis A.; Manolopoulou, Marika; Tsay, Julie L.; Wadlington, Natasha L.; Tang, Wei-Jen

    2009-12-01

    Insulin degrading enzyme (IDE) utilizes a large catalytic chamber to selectively bind and degrade peptide substrates such as insulin and amyloid {beta} (A{beta}). Tight interactions with substrates occur at an exosite located 30 {angstrom} away from the catalytic center that anchors the N-terminus of substrates to facilitate binding and subsequent cleavages at the catalytic site. However, IDE also degrades peptide substrates that are too short to occupy both the catalytic site and the exosite simultaneously. Here, we use kinins as a model system to address the kinetics and regulation of human IDE with short peptides. IDE specifically degrades bradykinin and kallidin at the Pro/Phe site. A 1.9 {angstrom} crystal structure of bradykinin-bound IDE reveals the binding of bradykinin to the exosite and not to the catalytic site. In agreement with observed high K{sub m} values, this suggests low affinity of bradykinin for IDE. This structure also provides the molecular basis on how the binding of short peptides at the exosite could regulate substrate recognition. We also found that human IDE is potently inhibited by physiologically relevant concentrations of S-nitrosylation and oxidation agents. Cysteine-directed modifications play a key role, since an IDE mutant devoid of all 13 cysteines is insensitive to the inhibition by S-nitrosoglutathione, hydrogen peroxide, or N-ethylmaleimide. Specifically, cysteine 819 of human IDE is located inside the catalytic chamber pointing toward an extended hydrophobic pocket and is critical for the inactivation. Thiol-directed modification of this residue likely causes local structural perturbation to reduce substrate binding and catalysis.

  9. Effect of N-acetylcysteine infusion on exercise-induced modulation of insulin sensitivity and signaling pathways in human skeletal muscle.

    PubMed

    Trewin, Adam J; Lundell, Leonidas S; Perry, Ben D; Patil, Kim Vikhe; Chibalin, Alexander V; Levinger, Itamar; McQuade, Leon R; Stepto, Nigel K

    2015-08-15

    -Reactive oxygen species (ROS) produced in skeletal muscle may play a role in potentiating the beneficial responses to exercise; however, the effects of exercise-induced ROS on insulin action and protein signaling in humans has not been fully elucidated. Seven healthy, recreationally active participants volunteered for this double-blind, randomized, repeated-measures crossover study. Exercise was undertaken with infusion of saline (CON) or the antioxidant N-acetylcysteine (NAC) to attenuate ROS. Participants performed two 1-h cycling exercise sessions 7-14 days apart, 55 min at 65% V̇o2peak plus 5 min at 85%V̇o2peak, followed 3 h later by a 2-h hyperinsulinemic euglycemic clamp (40 mIU·min(-1)·m(2)) to determine insulin sensitivity. Four muscle biopsies were taken on each trial day, at baseline before NAC infusion (BASE), after exercise (EX), after 3-h recovery (REC), and post-insulin clamp (PI). Exercise, ROS, and insulin action on protein phosphorylation were evaluated with immunoblotting. NAC tended to decrease postexercise markers of the ROS/protein carbonylation ratio by -13.5% (P = 0.08) and increase the GSH/GSSG ratio twofold vs. CON (P < 0.05). Insulin sensitivity was reduced (-5.9%, P < 0.05) by NAC compared with CON without decreased phosphorylation of Akt or AS160. Whereas p-mTOR was not significantly decreased by NAC after EX or REC, phosphorylation of the downstream protein synthesis target kinase p70S6K was blunted by 48% at PI with NAC compared with CON (P < 0.05). We conclude that NAC infusion attenuated muscle ROS and postexercise insulin sensitivity independent of Akt signaling. ROS also played a role in normal p70S6K phosphorylation in response to insulin stimulation in human skeletal muscle.

  10. Coordinated Defects in Hepatic Long Chain Fatty Acid Metabolism and Triglyceride Accumulation Contribute to Insulin Resistance in Non-Human Primates

    PubMed Central

    Gastaldelli, Amalia; Casiraghi, Francesca; Halff, Glenn A.; Abrahamian, Gregory A.; Davalli, Alberto M.; Bastarrachea, Raul A.; Comuzzie, Anthony G.; Guardado-Mendoza, Rodolfo; Jimenez-Ceja, Lilia M.; Mattern, Vicki; Paez, Ana Maria; Ricotti, Andrea; Tejero, Mary E.; Higgins, Paul B.; Rodriguez-Sanchez, Iram Pablo; Tripathy, Devjit; DeFronzo, Ralph A.; Dick, Edward J.; Cline, Gary W.; Folli, Franco

    2011-01-01

    Non-Alcoholic fatty liver disease (NAFLD) is characterized by accumulation of triglycerides (TG) in hepatocytes, which may also trigger cirrhosis. The mechanisms of NAFLD are not fully understood, but insulin resistance has been proposed as a key determinant. Aims To determine the TG content and long chain fatty acyl CoA composition profile in liver from obese non-diabetic insulin resistant (IR) and lean insulin sensitive (IS) baboons in relation with hepatic and peripheral insulin sensitivity. Methods Twenty baboons with varying grades of adiposity were studied. Hepatic (liver) and peripheral (mainly muscle) insulin sensitivity was measured with a euglycemic clamp and QUICKI. Liver biopsies were performed at baseline for TG content and LCFA profile by mass spectrometry, and histological analysis. Findings were correlated with clinical and biochemical markers of adiposity and insulin resistance. Results Obese IR baboons had elevated liver TG content compared to IS. Furthermore, the concentration of unsaturated (LC-UFA) was greater than saturated (LC-SFA) fatty acyl CoA in the liver. Interestingly, LC-FA UFA and SFA correlated with waist, BMI, insulin, NEFA, TG, QUICKI, but not M/I. Histological findings of NAFLD ranging from focal to diffuse hepatic steatosis were found in obese IR baboons. Conclusion Liver TG content is closely related with both hepatic and peripheral IR, whereas liver LC-UFA and LC-SFA are closely related only with hepatic IR in non-human primates. Mechanisms leading to the accumulation of TG, LC-UFA and an altered UFA: LC-SFA ratio may play an important role in the pathophysiology of fatty liver disease in humans. PMID:22125617

  11. Insulin degludec and insulin degludec/insulin aspart in Ramadan: A single center experience

    PubMed Central

    Kalra, Sanjay

    2016-01-01

    This study aimed to document the utility and safety of insulin degludec (IDeg) and insulin degludec aspart (IDegAsp) in persons with type 2 diabetes, observing the Ramadan fast. An observational study was conducted at a single center, in the real world setting, on six persons who either switched to IDeg or IDegAsp a month before Ramadan or changed time of administration of IDegAsp at the onset of Ramadan, to keep the fast in a safe manner. Subjects were kept under regular monitoring and surveillance before, during, and after Ramadan, and counseled in an opposite manner. Four persons, who shifted from premixed insulin to IDegAsp, experienced a 12–18% dose reduction after 14 days. At the onset of Ramadan, the Suhur dose was reduced by 30%, and this remained unchanged during the fasting month. The Iftar dose had to be increased by 4 units. One person who shifted from neutral protamine hagedorn to IDeg demonstrated a 25% dose reduction at 20 days, without any further change in insulin requirement during Ramadan. One person who changed time of injection of IDegAsp from morning to night reported no change in dosage. No episode of major hypoglycemia was reported. IDeg and IDegAsp are effective, safe, and well-tolerated means of achieving glycemic control in persons with type 2 diabetes who wish to fast. PMID:27366727

  12. High oxygen condition facilitates the differentiation of mouse and human pluripotent stem cells into pancreatic progenitors and insulin-producing cells.

    PubMed

    Hakim, Farzana; Kaitsuka, Taku; Raeed, Jamiruddin Mohd; Wei, Fan-Yan; Shiraki, Nobuaki; Akagi, Tadayuki; Yokota, Takashi; Kume, Shoen; Tomizawa, Kazuhito

    2014-04-01

    Pluripotent stem cells have potential applications in regenerative medicine for diabetes. Differentiation of stem cells into insulin-producing cells has been achieved using various protocols. However, both the efficiency of the method and potency of differentiated cells are insufficient. Oxygen tension, the partial pressure of oxygen, has been shown to regulate the embryonic development of several organs, including pancreatic β-cells. In this study, we tried to establish an effective method for the differentiation of induced pluripotent stem cells (iPSCs) into insulin-producing cells by culturing under high oxygen (O2) conditions. Treatment with a high O2 condition in the early stage of differentiation increased insulin-positive cells at the terminus of differentiation. We found that a high O2 condition repressed Notch-dependent gene Hes1 expression and increased Ngn3 expression at the stage of pancreatic progenitors. This effect was caused by inhibition of hypoxia-inducible factor-1α protein level. Moreover, a high O2 condition activated Wnt signaling. Optimal stage-specific treatment with a high O2 condition resulted in a significant increase in insulin production in both mouse embryonic stem cells and human iPSCs and yielded populations containing up to 10% C-peptide-positive cells in human iPSCs. These results suggest that culturing in a high O2 condition at a specific stage is useful for the efficient generation of insulin-producing cells.

  13. The effect of the putative endogenous imidazoline receptor ligand, clonidine-displacing substance, on insulin secretion from rat and human islets of Langerhans

    PubMed Central

    Chan, Susan L F; Atlas, Daphne; James, Roger F L; Morgan, Noel G

    1997-01-01

    The effects of a rat brain extract containing clonidine-displacing substance (CDS), a putative endogenous imidazoline receptor ligand, on insulin release from rat and human isolated islets of Langerhans were investigated.CDS was able to potentiate the insulin secretory response of rat islets incubated at 6 mM glucose, in a dose-dependent manner. The magnitude of this effect was similar to that in response to the well-characterized imidazoline secretagogue, efaroxan.CDS, like other imidazoline secretagogues, was also able to reverse the inhibitory action of diazoxide on glucose-induced insulin release, in both rat and human islets.These effects of CDS on secretion were reversed by the imidazoline secretagogue antagonists, RX801080 and the newly defined KU14R, providing the first evidence that imidazoline-mediated actions of CDS can be blocked by specific imidazoline antagonists.The effects of CDS on insulin secretion were unaffected when the method of preparation involved centri-filtration through a 3,000 Da cut-off membrane or when the extract was treated with protease. These results confirm that the active principle is of low molecular weight and is not a peptide.Overall, the data suggest that CDS behaves as a potent endogenous insulin secretagogue acting at the islet imidazoline receptor. PMID:9138700

  14. A model for premixed combustion oscillations

    SciTech Connect

    Janus, M.C.; Richards, G.A.

    1996-03-01

    Combustion oscillations are receiving renewed research interest due to increasing application of lean premix (LPM) combustion to gas turbines. A simple, nonlinear model for premixed combustion is described; it was developed to explain experimental results and to provide guidance for developing active control schemes based on nonlinear concepts. The model can be used to quickly examine instability trends associated with changes in equivalence ratio, mass flow rate, geometry, ambient conditions, etc. The model represents the relevant processes occurring in a fuel nozzle and combustor analogous to current LPM turbine combustors. Conservation equations for the nozzle and combustor are developed from simple control volume analysis, providing ordinary differential equations that can be solved on a PC. Combustion is modeled as a stirred reactor, with bimolecular reaction between fuel and air. Although focus is on the model, it and experimental results are compared to understand effects of inlet air temperature and open loop control schemes. The model shows that both are related to changes in transport time.

  15. Active control for turbulent premixed flame simulations

    SciTech Connect

    Bell, John B.; Day, Marcus S.; Grcar, Joseph F.; Lijewski, Michael J.

    2004-03-26

    Many turbulent premixed flames of practical interest are statistically stationary. They occur in combustors that have anchoring mechanisms to prevent blow-off and flashback. The stabilization devices often introduce a level of geometric complexity that is prohibitive for detailed computational studies of turbulent flame dynamics. As a result, typical detailed simulations are performed in simplified model configurations such as decaying isotropic turbulence or inflowing turbulence. In these configurations, the turbulence seen by the flame either decays or, in the latter case, increases as the flame accelerates toward the turbulent inflow. This limits the duration of the eddy evolutions experienced by the flame at a given level of turbulent intensity, so that statistically valid observations cannot be made. In this paper, we apply a feedback control to computationally stabilize an otherwise unstable turbulent premixed flame in two dimensions. For the simulations, we specify turbulent in flow conditions and dynamically adjust the integrated fueling rate to control the mean location of the flame in the domain. We outline the numerical procedure, and illustrate the behavior of the control algorithm. We use the simulations to study the propagation and the local chemical variability of turbulent flame chemistry.

  16. Quantitative variation in obesity-related traits and insulin precursors linked to the OB gene region on human chromosome 7

    SciTech Connect

    Duggirala, R.; Stern, M.P.; Reinhart, L.J.

    1996-09-01

    Despite the evidence that human obesity has strong genetic determinants, efforts at identifying specific genes that influence human obesity have largely been unsuccessful. Using the sibship data obtained from 32 low-income Mexican American pedigrees ascertained on a type II diabetic proband and a multipoint variance-components method, we tested for linkage between various obesity-related traits plus associated metabolic traits and 15 markers on human chromosome 7. We found evidence for linkage between markers in the OB gene region and various traits, as follows: D7S514 and extremity skinfolds (LOD = 3.1), human carboxypeptidase A1 (HCPA1) and 32,33-split proinsulin level (LOD = 4.2), and HCPA1 and proinsulin level (LOD = 3.2). A putative susceptibility locus linked to the marker D7S514 explained 56% of the total phenotypic variation in extremity skinfolds. Variation at the HCPA1 locus explained 64% of phenotypic variation in proinsulin level and {approximately}73% of phenotypic variation in split proinsulin concentration, respectively. Weaker evidence for linkage to several other obesity-related traits (e.g., waist circumference, body-mass index, fat mass by bioimpedance, etc.) was observed for a genetic location, which is {approximately}15 cM telomeric to OB. In conclusion, our study reveals that the OB region plays a significant role in determining the phenotypic variation of both insulin precursors and obesity-related traits, at least in Mexican Americans. 66 refs., 3 figs., 4 tabs.

  17. Inhibition of insulin amyloid fibril formation by cyclodextrins.

    PubMed

    Kitagawa, Keisuke; Misumi, Yohei; Ueda, Mitsuharu; Hayashi, Yuya; Tasaki, Masayoshi; Obayashi, Konen; Yamashita, Taro; Jono, Hirofumi; Arima, Hidetoshi; Ando, Yukio

    2015-01-01

    Localized insulin-derived amyloid masses occasionally form at the site of repeated insulin injections in patients with insulin-dependent diabetes and cause subcutaneous insulin resistance. Various kinds of insulin including porcine insulin, human insulin, and insulin analogues reportedly formed amyloid fibrils in vitro and in vivo, but the impact of the amino acid replacement in insulin molecules on amyloidogenicity is largely unknown. In the present study, we demonstrated the difference in amyloid fibril formation kinetics of human insulin and insulin analogues, which suggests an important role of the C-terminal domain of the insulin B chain in nuclear formation of amyloid fibrils. Furthermore, we determined that cyclodextrins, which are widely used as drug carriers in the pharmaceutical field, had an inhibitory effect on the nuclear formation of insulin amyloid fibrils. These findings have significant implications for the mechanism underlying insulin amyloid fibril formation and for developing optimal additives to prevent this subcutaneous adverse effect.

  18. Human insulin analogues modified at the B26 site reveal a hormone conformation that is undetected in the receptor complex

    SciTech Connect

    Žáková, Lenka; Kletvíková, Emília; Lepšík, Martin; Collinsová, Michaela; Watson, Christopher J.; Turkenburg, Johan P.; Jiráček, Jiří; Brzozowski, Andrzej M.

    2014-10-01

    [AsnB26]- and [GlyB26]-insulin mutants attain a B26-turn like fold without assistance of chemical modifications. Their structures match the insulin receptor interface and expand the spectrum of insulin conformations. The structural characterization of the insulin–insulin receptor (IR) interaction still lacks the conformation of the crucial B21–B30 insulin region, which must be different from that in its storage forms to ensure effective receptor binding. Here, it is shown that insulin analogues modified by natural amino acids at the TyrB26 site can represent an active form of this hormone. In particular, [AsnB26]-insulin and [GlyB26]-insulin attain a B26-turn-like conformation that differs from that in all known structures of the native hormone. It also matches the receptor interface, avoiding substantial steric clashes. This indicates that insulin may attain a B26-turn-like conformation upon IR binding. Moreover, there is an unexpected, but significant, binding specificity of the AsnB26 mutant for predominantly the metabolic B isoform of the receptor. As it is correlated with the B26 bend of the B-chain of the hormone, the structures of AsnB26 analogues may provide the first structural insight into the structural origins of differential insulin signalling through insulin receptor A and B isoforms.

  19. Structural and functional characterization of the human T lymphocyte receptor for insulin-like growth factor I in vitro.

    PubMed Central

    Tapson, V F; Boni-Schnetzler, M; Pilch, P F; Center, D M; Berman, J S

    1988-01-01

    Growth factor receptors for T lymphocytes, such as interleukin 2 and insulin, are present on activated but not resting T lymphocytes. We sought to determine if insulin-like growth factor I (IGF-I) could act as a growth factor for human T cells and to characterize its receptor on resting and activated cells. Recombinant IGF-I induced two separate functions. It was chemotactic for and increased incorporation of tritiated thymidine into both unactivated (resting) and mitogen-activated T cells. High-affinity 125I-IGF-I binding to human T cells was saturable with an apparent Kd of 1.2 +/- .6 X 10(-10) M for binding to activated T cells and 1.2 +/- .9 X 10(-10) for unactivated T cells. The calculated binding for activated cells was 330 +/- 90 and for resting cells 45 +/- 9 high-affinity receptor sites per cell. Affinity cross-linking of 125I-IGF-I to resting or activated T cells revealed a radioligand-receptor complex of 360,000 mol wt when analyzed by SDS-PAGE without reduction and complexes of 270,000 and 135,000 mol wt upon reduction; prior incubation with excess unlabeled IGF-I prevented formation of the 125I-IGF-I receptor complex. Our data suggest that both resting and activated T lymphocytes bear functional IGF-I receptors similar to those found in other tissues. These receptors may mediate T cell growth and chemotaxis. Images PMID:3262126

  20. Protein kinase B modulates the sensitivity of human neuroblastoma cells to insulin-like growth factor receptor inhibition.

    PubMed

    Guerreiro, Ana S; Boller, Danielle; Shalaby, Tarek; Grotzer, Michael A; Arcaro, Alexandre

    2006-12-01

    The potential of the novel insulin-like growth factor receptor (IGF-IR) inhibitor NVP-AEW541 as an antiproliferative agent in human neuroblastoma was investigated. Proliferation of a panel of neuroblastoma cell lines was inhibited by NVP-AEW541 with IC(50) values ranging from 0.15 to 5 microM. Experiments using an IGF-IR neutralizing antibody confirmed that the IGF-IR was essential to support growth of neuroblastoma cell lines. The expression levels of the IGF-IR in individual neuroblastoma cell lines did not correlate with the sensitivities to NVP-AEW541, while coexpression of the IGF-IR and the insulin receptor (IR) correlated with lower sensitivity to the inhibitor in some cell lines. Intriguingly, high levels of activation of Akt/protein kinase B (PKB) and phosphorylation of the ribosomal S6 protein were observed in neuroblastoma cell lines with decreased sensitivities to NVP-AEW541. Inhibition of Akt/PKB activity restored the sensitivity of neuroblastoma cells to the IGF-IR inhibitor. Transfection of neuroblastoma cells with activated Akt or ribosomal protein S6 kinase (S6K) decreased the sensitivity of the cells to NVP-AEW541. IGF-I-stimulated proliferation of neuroblastoma cell lines was completely blocked by NVP-AEW541, or by a combination of an inhibitor of phosphoinositide 3-kinase and rapamycin. In addition to its antiproliferative effects, NVP-AEW541 sensitized neuroblastoma cells to cisplatin-induced apoptosis. Together, our data demonstrate that NVP-AEW541 in combination with Akt/PKB inhibitors or chemotherapeutic agents may represent a novel approach to target human neuroblastoma cell proliferation.

  1. Functional properties of an isolated. cap alpha beta. heterodimeric human placenta insulin-like growth factor 1 receptor complex

    SciTech Connect

    Feltz, S.M.; Swanson, M.L.; Wemmie, J.A.; Pessin, J.E.

    1988-05-03

    Treatment of human placenta membranes at pH 8.5 in the presence of 2.0 mM dithiothreitol (DTT) for 5 min, followed by the simultaneous removal of the DTT and pH adjustment of pH 7.6, resulted in the formation of a functional ..cap alpha beta.. heterodimeric insulin-like growth factor 1 (IGF-1) receptor complex from the native ..cap alpha../sub 2/..beta../sub 2/ heterotetrameric disulfide-linked state. The membrane-bound ..cap alpha beta.. heterodimeric complex displayed similar curvilinear /sup 125/I-IGF-1 equilibrium binding compared to the ..cap alpha../sub 2/..beta../sub 2/ heterotetrameric complex. /sup 125/I-IGF-1 binding to both the isolated ..cap alpha../sub 2/..beta../sub 2/ heterotetrameric and ..cap alpha beta.. heterodimeric complexes demonstrated a marked straightening of the Scatchard plots, compared to the placenta membrane-bound IGF-1 receptors, with a 2-fold increase in the high-affinity binding component. IGF-1 stimulation of IGF-1 receptor autophosphorylation indicated that the ligand-dependent activation of ..cap alpha beta.. heterodimeric protein kinase activity occurred concomitant with the reassociation into a covalent ..cap alpha../sub 2/..beta../sub 2/ heterotetrameric state. These data demonstrate that (i) a combination of alkaline pH and DTT treatment of human placenta membranes results in the formation of an ..cap alpha beta.. heterodimeric IGF-1 receptor complex, (ii) unlike the insulin receptor, high-affinity homogeneous IGF-1 binding occurs in both the ..cap alpha../sub 2/..beta../sub 2/ heterotetrameric and ..cap alpha beta.. heterodimeric complexes, and (iii) IGF-1-dependent autophosphorylation of the ..cap alpha beta.. heterodimeric IGF-1 receptor complex correlates wit an IGF-1 dependent covalent reassociation into an ..cap alpha../sub 2/..beta../sub 2/ heterotetrameric disulfide-linked state.

  2. Conformational states and recognition of amyloidogenic peptides of human insulin-degrading enzyme.

    PubMed

    McCord, Lauren A; Liang, Wenguang G; Dowdell, Evan; Kalas, Vasilios; Hoey, Robert J; Koide, Akiko; Koide, Shohei; Tang, Wei-Jen

    2013-08-20

    Insulin-degrading enzyme (IDE) selectively degrades the monomer of amyloidogenic peptides and contributes to clearance of amyloid β (Aβ). Thus, IDE retards the progression of Alzheimer's disease. IDE possesses an enclosed catalytic chamber that engulfs and degrades its peptide substrates; however, the molecular mechanism of IDE function, including substrate access to the chamber and recognition, remains elusive. Here, we captured a unique IDE conformation by using a synthetic antibody fragment as a crystallization chaperone. An unexpected displacement of a door subdomain creates an ~18-Å opening to the chamber. This swinging-door mechanism permits the entry of short peptides into the catalytic chamber and disrupts the catalytic site within IDE door subdomain. Given the propensity of amyloidogenic peptides to convert into β-strands for their polymerization into amyloid fibrils, they also use such β-strands to stabilize the disrupted catalytic site resided at IDE door subdomain for their degradation by IDE. Thus, action of the swinging door allows IDE to recognize amyloidogenicity by substrate-induced stabilization of the IDE catalytic cleft. Small angle X-ray scattering (SAXS) analysis revealed that IDE exists as a mixture of closed and open states. These open states, which are distinct from the swinging door state, permit entry of larger substrates (e.g., Aβ, insulin) to the chamber and are preferred in solution. Mutational studies confirmed the critical roles of the door subdomain and hinge loop joining the N- and C-terminal halves of IDE for catalysis. Together, our data provide insights into the conformational changes of IDE that govern the selective destruction of amyloidogenic peptides.

  3. Ubiquitin is a Novel Substrate for Human Insulin-Degrading Enzyme

    PubMed Central

    Ralat, Luis A.; Kalas, Vasilios; Zheng, Zhongzhou; Goldman, Robert D.; Sosnick, Tobin R.; Tang, Wei-Jen

    2011-01-01

    Insulin-degrading enzyme (IDE) can degrade insulin and amyloid-β (Aβ), peptides involved in diabetes and Alzheimer's disease, respectively. IDE selects its substrates based on size, charge, and flexibility. From these criteria, we predict that IDE can cleave and inactivate ubiquitin (Ub). Here, we show that IDE cleaves Ub in a biphasic manner, first, by rapidly removing the two C-terminal glycines (kcat = 2 sec-1) followed by a slow cleavage between residues 72-73 (kcat = 0.07 sec-1), thereby producing the inactive Ub1-74 and Ub1-72. IDE is a ubiquitously expressed cytosolic protein, where monomeric Ub is also present. Thus, Ub degradation by IDE should be regulated. IDE is known to bind the cytoplasmic intermediate filament protein nestin with high affinity. We found that nestin potently inhibits the cleavage of Ub by IDE. In addition, Ub1-72 has a markedly increased affinity for IDE (∼90 fold). Thus, the association of IDE with cellular regulators and product inhibition by Ub1-72 can prevent inadvertent proteolysis of cellular Ub by IDE. Ub is a highly stable protein. However, IDE instead prefers to degrade peptides with high intrinsic flexibility. Indeed, we demonstrate that IDE is exquisitely sensitive to Ub stability. Mutations that only mildly destabilize Ub (ΔΔG ‹ 0.6 kcal/mol) render IDE hypersensitive to Ub with rate enhancements greater than 12-fold. The Ub-bound IDE structure and IDE mutants reveal that interaction of the exosite with the N-terminus of Ub guides the unfolding of Ub, allowing its sequential cleavages. Together, our studies link the control of Ub clearance with IDE. PMID:21185309

  4. Studies on the autophosphorylation of the insulin receptor from human placenta. Analysis of the sites phosphorylated by two-dimensional peptide mapping.

    PubMed Central

    Tavaré, J M; Denton, R M

    1988-01-01

    1. A partially purified preparation of human placental insulin receptors was incubated with [gamma-32P]ATP in the presence or absence of insulin. The 32P-labelled insulin-receptor beta-subunits were then isolated, cleaved with trypsin followed by protease V8 and the [32P]phosphopeptides generated were analysed by thin layer electrophoresis and chromatography. This approach revealed that insulin stimulates autophosphorylation of the insulin-receptor beta-subunit in vitro on at least seven tyrosine residues distributed among three distinct domains. 2. One domain (domain 2), containing tyrosine residues 1146, 1150 and 1151 was the most rapidly phosphorylated and could be recovered as mono-, di- and triphosphorylated peptides cleaved by trypsin at Arg-1143 and either Lys-1153 or Lys-1156. Multiple phosphorylation of this domain appears to partially inhibit the cleavage at Lys-1153 by trypsin. 3. In a second domain (domain 3) containing two phosphorylated tyrosine residues at positions 1316 and 1322 the tyrosines were phosphorylated more slowly than those in domain 2. This domain is close to the C-terminus of the beta-subunit polypeptide chain. 4. At least two further tyrosine residues appeared to be phosphorylated after those in domains 2 and 3. These residues probably residue within a domain lying in close proximity to the inner face of the plasma membrane containing tyrosines 953, 960 and 972, but conclusive evidence is still required. 5. The two-dimensional thin-layer analysis employed in this study to investigate insulin-receptor phosphorylation has several advantages over previous methods based on reverse-phase chromatography. It allows greater resolution of 32P-labelled tryptic peptides and, when coupled to radioautography, is considerably more sensitive. The approach can be readily adapted to study phosphorylation of the insulin receptor within intact cells. Images Fig. 1. Fig. 2. Fig. 3. Fig. 4. PMID:3166375

  5. Enhancement of human granulopoiesis in vitro by biosynthetic insulin-like growth factor I/somatomedin C and human growth hormone.

    PubMed Central

    Merchav, S; Tatarsky, I; Hochberg, Z

    1988-01-01

    The effect of biosynthetic recombinant insulin-like growth factor I/somatomedin C (IGF-I/Sm-C) and human growth hormone (hGH) on the in vitro growth and maturation of human marrow myeloid progenitors was investigated. Myeloid colony formation was maximally enhanced by 60 ng/ml IGF-I/Sm-C and by 250 ng/ml hGH, resulting in an increase in colony numbers of 41 +/- 7 and 38 +/- 4%, respectively (P less than 0.001). Both peptides induced a 1.5-2.5-fold increase in the frequency of colonies composed of granulocytes alone, but did not alter the numbers of monocyte/macrophage or mixed granulocyte/macrophage colonies. IGF-I/Sm-C and hGH were also found to enhance myeloid maturation towards mature granulocytes in suspension cultures of human marrow cells. The effect of both peptides on human marrow granulopoiesis was similarly demonstrable in serum-free cultures stimulated with human recombinant granulocyte/macrophage colony-stimulating factor. Enhancement of human marrow granulopoiesis in vitro by hGH required the presence of marrow adherent cells and was abrogated by specific monoclonal antibodies directed against IGF-I/Sm-C receptors. The effect of hGH on marrow myeloid progenitors thus appears to be mediated by paracrine IGF-I/Sm-C. PMID:2963830

  6. Insulin Resistance in Alzheimer's Disease

    PubMed Central

    Dineley, Kelly T; Jahrling, Jordan B; Denner, Larry

    2014-01-01

    Insulin is a key hormone regulating metabolism. Insulin binding to cell surface insulin receptors engages many signaling intermediates operating in parallel and in series to control glucose, energy, and lipids while also regulating mitogenesis and development. Perturbations in the function of any of these intermediates, which occur in a variety of diseases, cause reduced sensitivity to insulin and insulin resistance with consequent metabolic dysfunction. Chronic inflammation ensues which exacerbates compromised metabolic homeostasis. Since insulin has a key role in learning and memory as well as directly regulating ERK, a kinase required for the type of learning and memory compromised in early Alzheimer's disease (AD), insulin resistance has been identified as a major risk factor for the onset of AD. Animal models of AD or insulin resistance or both demonstrate that AD pathology and impaired insulin signaling form a reciprocal relationship. Of note are human and animal model studies geared toward improving insulin resistance that have led to the identification of the nuclear receptor and transcription factor, peroxisome proliferator-activated receptor gamma (PPARγ) as an intervention tool for early AD. Strategic targeting of alternate nodes within the insulin signaling network has revealed disease-stage therapeutic windows in animal models that coalesce with previous and ongoing clinical trial approaches. Thus, exploiting the connection between insulin resistance and AD provides powerful opportunities to delineate therapeutic interventions that slow or block the pathogenesis of AD. PMID:25237037

  7. Deletion of Asn{sup 281} in the {alpha}-subunit of the human insulin receptor causes constitutive activation of the receptor and insulin desensitization

    SciTech Connect

    Desbois-Mouthon, C.; Sert-Langeron, C.; Magre, J.; Blivet, M.J.

    1996-02-01

    We studied the structure and function of the insulin receptor (IR) in two sisters with leprechaunism. The patients had inherited alterations in the IR gene and were compound heterozygotes. Their paternal IR allele carried a major deletion, including exons 10-13, which shifted the reading frame and introduced a premature chain termination codon in the IR sequence. This allele was expressed at a very low level in cultured fibroblasts (<10% of total IR messenger ribonucleic acid content) and encoded a truncated protein lacking transmembrane and tyrosine kinase domains. The maternal IR allele was deleted of 3 bp in exon 3, causing the loss of Asn{sup 281} in the {alpha}-subunit. This allele generated levels of IR messenger ribonucleic acid and cell surface receptors similar to those seen in control fibroblasts. However, IRs from patients` cells had impaired insulin binding and exhibited in vivo and in vitro constitutive activation of autophosphorylation and tyrosine kinase activity. As a result of this IR-preactivated state, the cells were desensitized to insulin stimulation of glycogen and DNA syntheses. These findings strongly suggest that Asn{sup 281} of the IR {alpha}-subunit plays a critical role in the inhibitory constraint exerted by the extracellular {alpha}-subunit over the intracellular kinase activity. 59 refs., 6 figs.

  8. Effects of Type 1 Insulin-Like Growth Factor Receptor Silencing in a Human Adrenocortical Cell Line.

    PubMed

    Ribeiro, T C; Jorge, A A; Montenegro, L R; Almeida, M Q; Ferraz-de-Souza, B; Nishi, M Y; Mendonca, B B; Latronico, A C

    2016-07-01

    Type 1 insulin-like growth factor receptor (IGF-1R) is overexpressed in a variety of human cancers, including adrenocortical tumors. The aim of the work was to investigate the effects of IGF-1R downregulation in a human adrenocortical cell line by small interfering RNA (siRNA). The human adrenocortical tumor cell line NCI H295R was transfected with 2 specific IGF1R siRNAs (# 1 and # 2) and compared with untreated cells and a negative control siRNA. IGF1R expression was determined by quantitative reverse-transcription PCR (qRTPCR) and Western blot. The effects of IGF-1R downregulation on cell proliferation and apoptosis were assessed. IGF-1R levels were significantly decreased in cells treated with IGF-1R siRNA # 1 or # 2. Relative expression of IGF1R mRNA decreased approximately 50% and Western blot analysis revealed a 30% of reduction in IGF-1R protein. Downregulation of this gene resulted in 40% reduction in cell growth in vitro and 45% increase in apoptosis using siRNA # 2. These findings demonstrate that decreasing IGF-1R mRNA and protein expression in NCI H295R cells can partially inhibit adrenal tumor cell growth in vitro. Targeting IGF1R is a promising therapy for pediatric malignant adrenocortical tumor and can still be an option for adult adrenocortical cancer based on personalized genomic tumor profiling. PMID:27246621

  9. Orthologs of Human Disease Associated Genes and RNAi Analysis of Silencing Insulin Receptor Gene in Bombyx mori

    PubMed Central

    Zhang, Zan; Teng, Xiaolu; Chen, Maohua; Li, Fei

    2014-01-01

    The silkworm, Bombyx mori L., is an important economic insect that has been domesticated for thousands of years to produce silk. It is our great interest to investigate the possibility of developing the B. mori as human disease model. We searched the orthologs of human disease associated genes in the B. mori by bi-directional best hits of BLAST and confirmed by searching the OrthoDB. In total, 5006 genes corresponding to 1612 kinds of human diseases had orthologs in the B. mori, among which, there are 25 genes associated with diabetes mellitus. Of these, we selected the insulin receptor gene of the B. mori (Bm-INSR) to study its expression in different tissues and at different developmental stages and tissues. Quantitative PCR showed that Bm-INSR was highly expressed in the Malpighian tubules but expressed at low levels in the testis. It was highly expressed in the 3rd and 4th instar larvae, and adult. We knocked down Bm-INSR expression using RNA interference. The abundance of Bm-INSR transcripts were dramatically reduced to ~4% of the control level at 6 days after dsRNA injection and the RNAi-treated B. mori individuals showed apparent growth inhibition and malformation such as abnormal body color in black, which is the typical symptom of diabetic patients. Our results demonstrate that B. mori has potential use as an animal model for diabetic mellitus research. PMID:25302617

  10. Expression of a synthetic gene encoding human insulin-like growth factor I in cultured mouse fibroblasts

    SciTech Connect

    Bayne, M.L.; Cascieri, M.A.; Kelder, B.; Applebaum, J.; Chicchi, G.; Shapiro, J.A.; Pasleau, F.; Kopchick, J.J.

    1987-05-01

    A synthetic gene encoding human insulin-like growth factor I (hIGF-I) was assembled and inserted into an expression vector containing the cytomegalovirus immediate early (CMV-IE) transcriptional regulatory region and portions of the bovine growth hormone gene. The recombinant plasmid encodes a 97 amino acid fusion protein containing the first 27 amino acids of the bovine growth hormone precursor and the 70 amino acids of hIGF-I. This plasmid, when transiently introduced into cultured mouse fibroblasts, directs synthesis of the fusion protein, subsequent proteolytic removal of the bovine growth hormone signal peptide, and secretion of hIGF-I into the culture medium. Conditioned medium from transfected cells inhibits binding of /sup 125/I-labeled IGF-I to type I IGF receptors on human placental membranes and to acid-stable human serum carrier proteins. The recombinant hIGF-I produced is biologically active, as monitored by the stimulation of DNA synthesis in vascular smooth muscle cells.

  11. Pdx1 and controlled culture conditions induced differentiation of human amniotic fluid-derived stem cells to insulin-producing clusters.

    PubMed

    Chun, So Young; Mack, David L; Moorefield, Emily; Oh, Se Heang; Kwon, Tae Gyun; Pettenati, Mark J; Yoo, James J; Coppi, Paolo De; Atala, Anthony; Soker, Shay

    2015-05-01

    This study investigated the differentiation of human amniotic fluid-derived stem cells (hAFSCs) into insulin-producing clusters in vitro. Adenovirally-delivered mouse Pdx1 (Ad-Pdx1) induced human Pdx1 expression in hAFSCs and enhanced the coordinated expression of downstream β-cell markers. When Ad-Pdx1-transduced hAFSCs were sequentially treated with activin A, bFGF and nicotinamide and the culture plate surface coated with poly-l-ornithine, the expression of islet-associated human mRNAs for Pdx1, Pax6, Ngn3 and insulin was increased. C-peptide ELISA confirmed that Ad-Pdx1-transduced hAFSCs processed and secreted insulin in a manner consistent with that pathway in pancreatic β-cells. To sustain the β-cell-like phenotype and investigate the effect of three-dimensional (3D) conformation on the differentiation of hAFSCs, Pdx1-transduced cells were encapsulated in alginate and cultured long-term under serum-free conditions. Over 2 weeks, partially differentiated hAFSC clusters increased in size and increased insulin secretion. Taken together, these data demonstrate that ectopic Pdx1 expression initiates pancreatic differentiation in hAFSCs and that a β-cell-like phenotype can be augmented by culture conditions that mimic the stromal components and 3D geometry associated with pancreatic islets.

  12. [Novel insulins].

    PubMed

    Eriksson, Johan G; Laine, Merja K

    2016-01-01

    Novel insulins have entered the market during recent years. The ultra-long acting insulins, insulin degludek and insulin glargine, the latter having a strength of 300 U/ml, exhibit a steady and predictable action curve. Studies have indicated that significantly fewer hypoglycemiae occur when using degludek in patients with either type 1 or type 2 diabetes, whereas similar evidence about glargine (300 U/mI) has been obtained in the treatment of type 2 diabetes. The long duration of action of both insulins brings long-needed flexibility to.their dosing. PMID:27089618

  13. Identification of novel genes for glucose metabolism based upon expression pattern in human islets and effect on insulin secretion and glycemia.

    PubMed

    Taneera, Jalal; Fadista, Joao; Ahlqvist, Emma; Atac, David; Ottosson-Laakso, Emilia; Wollheim, Claes B; Groop, Leif

    2015-04-01

    Normal glucose homeostasis is characterized by appropriate insulin secretion and low HbA1c. Gene expression signatures associated with these two phenotypes could be essential for islet function and pathophysiology of type 2 diabetes (T2D). Herein, we employed a novel approach to identify candidate genes involved in T2D by correlating islet microarray gene expression data (78 donors) with insulin secretion and HbA1c level. The expression of 649 genes (P < 0.05) was correlated with insulin secretion and HbA1c. Of them, five genes (GLR1A, PPP1R1A, PLCDXD3, FAM105A and ENO2) correlated positively with insulin secretion/negatively with HbA1c and one gene (GNG5) correlated negatively with insulin secretion/positively with HbA1c were followed up. The five positively correlated genes have lower expression levels in diabetic islets, whereas GNG5 expression is higher. Exposure of human islets to high glucose for 24 h resulted in up-regulation of GNG5 and PPP1R1A expression, whereas the expression of ENO2 and GLRA1 was down-regulated. No effect was seen on the expression of FAM105A and PLCXD3. siRNA silencing in INS-1 832/13 cells showed reduction in insulin secretion for PPP1R1A, PLXCD3, ENO2, FAM105A and GNG5 but not GLRA1. Although no SNP in these gene loci passed the genome-wide significance for association with T2D in DIAGRAM+ database, four SNPs influenced gene expression in cis in human islets. In conclusion, we identified and confirmed PPP1R1A, FAM105A, ENO2, PLCDX3 and GNG5 as potential regulators of islet function. We provide a list of candidate genes as a resource for exploring their role in the pathogenesis of T2D.

  14. Protective role of Cys-178 against the inactivation and oligomerization of human insulin-degrading enzyme by oxidation and nitrosylation.

    PubMed

    Ralat, Luis A; Ren, Min; Schilling, Alexander B; Tang, Wei-Jen

    2009-12-01

    Insulin-degrading enzyme (IDE), a 110-kDa metalloendopeptidase, hydrolyzes several physiologically relevant peptides, including insulin and amyloid-beta (Abeta). Human IDE has 13 cysteines and is inhibited by hydrogen peroxide and S-nitrosoglutathione (GSNO), donors of reactive oxygen and nitrogen species, respectively. Here, we report that the oxidative burst of BV-2 microglial cells leads to oxidation or nitrosylation of secreted IDE, leading to the reduced activity. Hydrogen peroxide and GSNO treatment of IDE reduces the V(max) for Abeta degradation, increases IDE oligomerization, and decreases IDE thermostability. Additionally, this inhibitory response of IDE is substrate-dependent, biphasic for Abeta degradation but monophasic for a shorter bradykinin-mimetic substrate. Our mutational analysis of IDE and peptide mass fingerprinting of GSNO-treated IDE using Fourier transform-ion cyclotron resonance mass spectrometer reveal a surprising interplay of Cys-178 with Cys-110 and Cys-819 for catalytic activity and with Cys-789 and Cys-966 for oligomerization. Cys-110 is near the zinc-binding catalytic center and is normally buried. The oxidation and nitrosylation of Cys-819 allow Cys-110 to be oxidized or nitrosylated, leading to complete inactivation of IDE. Cys-789 is spatially adjacent to Cys-966, and their nitrosylation and oxidation together trigger the oligomerization and inhibition of IDE. Interestingly, the Cys-178 modification buffers the inhibition caused by Cys-819 modification and prevents the oxidation or nitrosylation of Cys-110. The Cys-178 modification can also prevent the oligomerization-mediated inhibition. Thus, IDE can be intricately regulated by reactive oxygen or nitrogen species. The structure of IDE reveals the molecular basis for the long distance interactions of these cysteines and how they regulate IDE function.

  15. Overexpression of human insulin receptor substrate 1 induces cellular transformation with activation of mitogen-activated protein kinases.

    PubMed Central

    Ito, T; Sasaki, Y; Wands, J R

    1996-01-01

    The receptor insulin substrate 1 protein (IRS-1) is a specific substrate for insulin receptor tyrosine kinase. Expression and tyrosyl phosphorylation of IRS-1 play an important role during normal hepatocyte growth, and the gene is overexpressed in hepatocellular carcinoma tissue. We determined if IRS-1 overexpression directly contributes to cellular transformation. The human IRS-1 gene was subcloned into a mammalian expression vector driven by the cytomegalovirus early promoter. NIH 3T3 cells transiently transfected with this vector subsequently developed transformed foci. Several stably transfected cell lines were established, and they grew efficiently under low-serum conditions and formed colonies when plated in soft agar. Cell lines overexpressing IRS-1 displayed increased tyrosyl phosphorylation of IRS-1 and association with Grb2 but not with the p85 subunit of phosphatidylinositol 3' kinase. Since Grb2 is a component of the son-of-sevenless-Ras pathway and upstream in the mitogen-activated protein kinase (MAPK) cascade, enzymatic activities of the major components of this cascade, such as MAPK kinase and MAPK were evaluated and found to be substantially increased in three independent cell lines with IRS-1 protein overexpression. Such cells, when injected into nude mice, were highly tumorigenic, and there may be a correlation between the degree of MAPK activation and tumor growth rate. This report describes the generation of a transformed phenotype by overexpression of a molecule without a catalytic domain far upstream in the signal transduction cascade and suggests that prolonged activation of MAPKs by this mechanism may be one of the molecular events related to hepatocellular transformation. PMID:8622697

  16. Structural and functional characterization of pathogenic non- synonymous genetic mutations of human insulin-degrading enzyme by in silico methods.

    PubMed

    Shaik, Noor A; Kaleemuddin, Mohammed; Banaganapalli, Babajan; Khan, Fazal; Shaik, Nazia S; Ajabnoor, Ghada; Al-Harthi, Sameer E; Bondagji, Nabeel; Al-Aama, Jumana Y; Elango, Ramu

    2014-04-01

    Insulin-degrading enzyme (IDE) is a key protease involved in degrading insulin and amyloid peptides in human body. Several non-synonymous genetic mutations of IDE gene have been recently associated with susceptibility to both diabetes and Alzheimer's diseases. However, the consequence of these mutations on the structure of IDE protein and its substrate binding characteristics is not well elucidated. The computational investigation of genetic mutation consequences on structural level of protein is recently found to be an effective alternate to traditional in vivo and in vitro approaches. Hence, by using a combination of empirical rule and support vector machine based in silico algorithms, this study was able to identify that the pathogenic nonsynonymous genetic mutations corresponding to p.I54F, p.P122T, p.T533R, p.P581A and p.Y609A have more potential role in structural and functional deviations of IDE activity. Moreover, molecular modeling and secondary structure analysis have also confirmed their impact on the stability and secondary properties of IDE protein. The molecular docking analysis of IDE with combinational substrates has revealed that peptide inhibitors compared to small non-peptide inhibitor molecules possess good inhibitory activity towards mutant IDE. This finding may pave a way to design novel potential small peptide inhibitors for mutant IDE. Additionally by un-translated region (UTR) scanning analysis, two regulatory pathogenic genetic mutations i.e., rs5786997 (3' UTR) and rs4646954 (5' UTR), which can influence the translation pattern of IDE gene through sequence alteration of upstream-Open Reading Frame and Internal Ribosome Entry Site elements were identified. Our findings are expected to help in narrowing down the number of IDE genetic variants to be screened for disease association studies and also to select better competitive inhibitors for IDE related diseases.

  17. NOx Formation in a Premixed Syngas Flame

    SciTech Connect

    Yilmaz, S.L.; Givi, P.; Strakey, P.; Casleton, K.

    2006-11-01

    Reduction of NOx is a subject of significant current interest in stationary gas turbines. The objective of this study is to examine the effects of turbulence on non-thermal NOx formation in a syngas flame. This is archived by a detailed parametric study via PDF simulations of a partially stirred reactor and a dumped axisymmetric premixed flame. Several different detailed and reduced kinetics schemes are considered. The simulated results demonstrate the strong dependence of combustion process on turbulence. It is shown that the amount of NOx formation is significantly influenced by the inlet conditions. That is, the turbulence intensity can be tweaked to attain optimal ultra-low NOx emissions at a given temperature.

  18. Lagrangian formulation of turbulent premixed combustion.

    PubMed

    Pagnini, Gianni; Bonomi, Ernesto

    2011-07-22

    The lagrangian point of view is adopted to study turbulent premixed combustion. The evolution of the volume fraction of combustion products is established by the Reynolds transport theorem. It emerges that the burned-mass fraction is led by the turbulent particle motion, by the flame front velocity, and by the mean curvature of the flame front. A physical requirement connecting particle turbulent dispersion and flame front velocity is obtained from equating the expansion rates of the flame front progression and of the unburned particles spread. The resulting description compares favorably with experimental data. In the case of a zero-curvature flame, with a non-markovian parabolic model for turbulent dispersion, the formulation yields the Zimont equation extended to all elapsed times and fully determined by turbulence characteristics. The exact solution of the extended Zimont equation is calculated and analyzed to bring out different regimes.

  19. Premixed flames in closed cylindrical tubes

    NASA Astrophysics Data System (ADS)

    Metzener, Philippe; Matalon, Moshe

    2001-09-01

    We consider the propagation of a premixed flame, as a two-dimensional sheet separating unburned gas from burned products, in a closed cylindrical tube. A nonlinear evolution equation, that describes the motion of the flame front as a function of its mean position, is derived. The equation contains a destabilizing term that results from the gas motion induced by thermal expansion and has a memory term associated with vorticity generation. Numerical solutions of this equation indicate that, when diffusion is stabilizing, the flame evolves into a non-planar form whose shape, and its associated symmetry properties, are determined by the Markstein parameter, and by the initial data. In particular, we observe the development of convex axisymmetric or non-axisymmetric flames, tulip flames and cellular flames.

  20. Human Monoclonal Islet Cell Antibodies From a Patient with Insulin- Dependent Diabetes Mellitus Reveal Glutamate Decarboxylase as the Target Antigen

    NASA Astrophysics Data System (ADS)

    Richter, Wiltrud; Endl, Josef; Eiermann, Thomas H.; Brandt, Michael; Kientsch-Engel, Rosemarie; Thivolet, Charles; Jungfer, Herbert; Scherbaum, Werner A.

    1992-09-01

    The autoimmune phenomena associated with destruction of the β cell in pancreatic islets and development of type 1 (insulin-dependent) diabetes mellitus (IDDM) include circulating islet cell antibodies. We have immortalized peripheral blood lymphocytes from prediabetic individuals and patients with newly diagnosed IDDM by Epstein-Barr virus transformation. IgG-positive cells were selected by anti-human IgG-coupled magnetic beads and expanded in cell culture. Supernatants were screened for cytoplasmic islet cell antibodies using the conventional indirect immunofluorescence test on cryostat sections of human pancreas. Six islet cell-specific B-cell lines, originating from a patient with newly diagnosed IDDM, could be stabilized on a monoclonal level. All six monoclonal islet cell antibodies (MICA 1-6) were of the IgG class. None of the MICA reacted with human thyroid, adrenal gland, anterior pituitary, liver, lung, stomach, and intestine tissues but all six reacted with pancreatic islets of different mammalian species and, in addition, with neurons of rat cerebellar cortex. MICA 1-6 were shown to recognize four distinct antigenic epitopes in islets. Islet cell antibody-positive diabetic sera but not normal human sera blocked the binding of the monoclonal antibodies to their target epitopes. Immunoprecipitation of 35S-labeled human islet cell extracts revealed that a protein of identical size to the enzyme glutamate decarboxylase (EC 4.1.1.15) was a target of all MICA. Furthermore, antigen immunotrapped by the MICA from brain homogenates showed glutamate decarboxylase enzyme activity. MICA 1-6 therefore reveal glutamate decarboxylase as the predominant target antigen of cytoplasmic islet cell autoantibodies in a patient with newly diagnosed IDDM.

  1. Implementation of Premixed Equilibrium Chemistry Capability in OVERFLOW

    NASA Technical Reports Server (NTRS)

    Olsen, Mike E.; Liu, Yen; Vinokur, M.; Olsen, Tom

    2004-01-01

    An implementation of premixed equilibrium chemistry has been completed for the OVERFLOW code, a chimera capable, complex geometry flow code widely used to predict transonic flowfields. The implementation builds on the computational efficiency and geometric generality of the solver.

  2. Studies of premixed laminar and turbulent flames at microgravity

    NASA Technical Reports Server (NTRS)

    Ronney, Paul D.

    1993-01-01

    A two and one-half year experimental and theoretical research program on the properties of laminar and turbulent premixed gas flames at microgravity was conducted. Progress during this program is identified and avenues for future studies are discussed.

  3. Mechanisms of combustion limits in premixed gas flames at microgravity

    NASA Technical Reports Server (NTRS)

    Ronney, Paul D.

    1991-01-01

    A three-year experimental and theoretical research program on the mechanisms of combustion limits of premixed gasflames at microgravity was conducted. Progress during this program is identified and avenues for future studies are discussed.

  4. Implementation of Premixed Equilibrium Chemistry Capability in OVERFLOW

    NASA Technical Reports Server (NTRS)

    Olsen, M. E.; Liu, Y.; Vinokur, M.; Olsen, T.

    2003-01-01

    An implementation of premixed equilibrium chemistry has been completed for the OVERFLOW code, a chimera capable, complex geometry flow code widely used to predict transonic flowfields. The implementation builds on the computational efficiency and geometric generality of the solver.

  5. Intensifying Insulin Therapy in Type 2 Diabetes: Choices & Challenges.

    PubMed

    Kumar, Ajay; Kesavadev, Jothydev; Sethi, Bipin; Jain, Sunil M; Guruprasad, C S; Shah, Siddharth N

    2015-05-01

    Insulin therapy remains the cornerstone of effective diabetes management. Timely intensification of insulin therapy reduces the progression of diabetes and the development of diabetes-related complications. Given that overall hyperglycaemia is a relative contribution of both fasting and postprandial hyperglycaemia, use of basal insulin alone may not achieve optimal glucose control due to its inability to cover postprandial glucose excursions. Intensifying therapy with addition of bolus insulin or switching to premixed insulin is a viable option in patients failing on basal alone therapy. Although the benefits of early insulin treatment are well established, a considerable delay in intensifying insulin therapy in patients with sub-optimal glycaemic control is still observed. Most of the patients and physicians are reluctant to intensify therapy due to the fear of hypoglycaemia, regimen complexity, and increased burden of multiple daily injections. In this context, there is a need for a flexible, alternative intensification option taking into account individual patient considerations to achieve or maintain individual glycaemic targets. An ideal insulin regimen should mimic physiological insulin release while providing optimal glycaemic control with low risk of hypoglycaemia, weight gain and fewer daily injections. The current paper reviews the challenges of insulin intensification in patients with type 2 diabetes mellitus poorly controlled on current treatment regimens. PMID:26548029

  6. Hypomorphism in human NSMCE2 linked to primordial dwarfism and insulin resistance

    PubMed Central

    Payne, Felicity; Colnaghi, Rita; Rocha, Nuno; Seth, Asha; Harris, Julie; Carpenter, Gillian; Bottomley, William E.; Wheeler, Eleanor; Wong, Stephen; Saudek, Vladimir; Savage, David; O’Rahilly, Stephen; Carel, Jean-Claude; Barroso, Inês; O’Driscoll, Mark; Semple, Robert

    2014-01-01

    Structural maintenance of chromosomes (SMC) complexes are essential for maintaining chromatin structure and regulating gene expression. Two the three known SMC complexes, cohesin and condensin, are important for sister chromatid cohesion and condensation, respectively; however, the function of the third complex, SMC5–6, which includes the E3 SUMO-ligase NSMCE2 (also widely known as MMS21) is less clear. Here, we characterized 2 patients with primordial dwarfism, extreme insulin resistance, and gonadal failure and identified compound heterozygous frameshift mutations in NSMCE2. Both mutations reduced NSMCE2 expression in patient cells. Primary cells from one patient showed increased micronucleus and nucleoplasmic bridge formation, delayed recovery of DNA synthesis, and reduced formation of foci containing Bloom syndrome helicase (BLM) after hydroxyurea-induced replication fork stalling. These nuclear abnormalities in patient dermal fibroblast were restored by expression of WT NSMCE2, but not a mutant form lacking SUMO-ligase activity. Furthermore, in zebrafish, knockdown of the NSMCE2 ortholog produced dwarfism, which was ameliorated by reexpression of WT, but not SUMO-ligase–deficient NSMCE. Collectively, these findings support a role for NSMCE2 in recovery from DNA damage and raise the possibility that loss of its function produces dwarfism through reduced tolerance of replicative stress. PMID:25105364

  7. Human conditions of insulin-like growth factor-I (IGF-I) deficiency

    PubMed Central

    2012-01-01

    Insulin-like growth factor I (IGF-I) is a polypeptide hormone produced mainly by the liver in response to the endocrine GH stimulus, but it is also secreted by multiple tissues for autocrine/paracrine purposes. IGF-I is partly responsible for systemic GH activities although it possesses a wide number of own properties (anabolic, antioxidant, anti-inflammatory and cytoprotective actions). IGF-I is a closely regulated hormone. Consequently, its logical therapeutical applications seems to be limited to restore physiological circulating levels in order to recover the clinical consequences of IGF-I deficiency, conditions where, despite continuous discrepancies, IGF-I treatment has never been related to oncogenesis. Currently the best characterized conditions of IGF-I deficiency are Laron Syndrome, in children; liver cirrhosis, in adults; aging including age-related-cardiovascular and neurological diseases; and more recently, intrauterine growth restriction. The aim of this review is to summarize the increasing list of roles of IGF-I, both in physiological and pathological conditions, underlying that its potential therapeutical options seem to be limited to those proven states of local or systemic IGF-I deficiency as a replacement treatment, rather than increasing its level upper the normal range. PMID:23148873

  8. Site-specific covalent modifications of human insulin by catechol estrogens: Reactivity and induced structural and functional changes

    NASA Astrophysics Data System (ADS)

    Ku, Ming-Chun; Fang, Chieh-Ming; Cheng, Juei-Tang; Liang, Huei-Chen; Wang, Tzu-Fan; Wu, Chih-Hsing; Chen, Chiao-Chen; Tai, Jung-Hsiang; Chen, Shu-Hui

    2016-06-01

    Proteins, covalently modified by catechol estrogens (CEs), were identified recently from the blood serum of diabetic patients and referred to as estrogenized proteins. Estrogenization of circulating insulin may occur and affect its molecular functioning. Here, the chemical reactivity of CEs towards specific amino acid residues of proteins and the structural and functional changes induced by the estrogenization of insulin were studied using cyclic voltammetry, liquid chromatography-mass spectrometry, circular dichroism spectroscopy, molecular modeling, and bioassays. Our results indicate that CEs, namely, 2- and 4-hydroxyl estrogens, were thermodynamically and kinetically more reactive than the catechol moiety. Upon co-incubation, intact insulin formed a substantial number of adducts with one or multiple CEs via covalent conjugation at its Cys 7 in the A or B chain, as well as at His10 or Lys29 in the B chain. Such conjugation was coupled with the cleavage of inter-chain disulfide linkages. Estrogenization on these sites may block the receptor-binding pockets of insulin. Insulin signaling and glucose uptake levels were lower in MCF-7 cells treated with modified insulin than in cells treated with native insulin. Taken together, our findings demonstrate that insulin molecules are susceptible to active estrogenization, and that such modification may alter the action of insulin.

  9. Site-specific covalent modifications of human insulin by catechol estrogens: Reactivity and induced structural and functional changes

    PubMed Central

    Ku, Ming-Chun; Fang, Chieh-Ming; Cheng, Juei-Tang; Liang, Huei-Chen; Wang, Tzu-Fan; Wu, Chih-Hsing; Chen, Chiao-Chen; Tai, Jung-Hsiang; Chen, Shu-Hui

    2016-01-01

    Proteins, covalently modified by catechol estrogens (CEs), were identified recently from the blood serum of diabetic patients and referred to as estrogenized proteins. Estrogenization of circulating insulin may occur and affect its molecular functioning. Here, the chemical reactivity of CEs towards specific amino acid residues of proteins and the structural and functional changes induced by the estrogenization of insulin were studied using cyclic voltammetry, liquid chromatography-mass spectrometry, circular dichroism spectroscopy, molecular modeling, and bioassays. Our results indicate that CEs, namely, 2- and 4-hydroxyl estrogens, were thermodynamically and kinetically more reactive than the catechol moiety. Upon co-incubation, intact insulin formed a substantial number of adducts with one or multiple CEs via covalent conjugation at its Cys 7 in the A or B chain, as well as at His10 or Lys29 in the B chain. Such conjugation was coupled with the cleavage of inter-chain disulfide linkages. Estrogenization on these sites may block the receptor-binding pockets of insulin. Insulin signaling and glucose uptake levels were lower in MCF-7 cells treated with modified insulin than in cells treated with native insulin. Taken together, our findings demonstrate that insulin molecules are susceptible to active estrogenization, and that such modification may alter the action of insulin. PMID:27353345

  10. 21 CFR 522.1160 - Insulin.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS...) of insulin. (2) Each mL of protamine zinc recombinant human insulin suspension contains 40 IU of... or on the order of a licensed veterinarian. (2) Cats—(i) Amount—(A) Porcine insulin zinc....

  11. Studies of Premixed Laminar and Turbulent Flames at Microgravity

    NASA Technical Reports Server (NTRS)

    Ronney, Paul D.

    1993-01-01

    The work of the Principal Investigator (PI) has encompassed four topics related to the experimental and theoretical study of combustion limits in premixed flames at microgravity, as discussed in the following sections. These topics include: (1) radiation effects on premixed gas flames; (2) flame structure and stability at low Lewis number; (3) flame propagation and extinction is cylindrical tubes; and (4) experimental simulation of combustion processes using autocatalytic chemical reactions.

  12. Fully Premixed Low Emission, High Pressure Multi-Fuel Burner

    NASA Technical Reports Server (NTRS)

    Nguyen, Quang-Viet (Inventor)

    2012-01-01

    A low-emissions high-pressure multi-fuel burner includes a fuel inlet, for receiving a fuel, an oxidizer inlet, for receiving an oxidizer gas, an injector plate, having a plurality of nozzles that are aligned with premix face of the injector plate, the plurality of nozzles in communication with the fuel and oxidizer inlets and each nozzle providing flow for one of the fuel and the oxidizer gas and an impingement-cooled face, parallel to the premix face of the injector plate and forming a micro-premix chamber between the impingement-cooled face and the in injector face. The fuel and the oxidizer gas are mixed in the micro-premix chamber through impingement-enhanced mixing of flows of the fuel and the oxidizer gas. The burner can be used for low-emissions fuel-lean fully-premixed, or fuel-rich fully-premixed hydrogen-air combustion, or for combustion with other gases such as methane or other hydrocarbons, or even liquid fuels.

  13. Molecular genetics of human growth hormone, insulin-like growth factors and their pathways in common disease.

    PubMed

    Rodriguez, Santiago; Gaunt, Tom R; Day, Ian N M

    2007-08-01

    The human growth hormone gene (GH1) and the insulin-like growth factor 1 and 2 genes (IGF1 and IGF2) encode the central elements of a key pathway influencing growth in humans. This "growth pathway" also includes transcription factors, agonists, antagonists, receptors, binding proteins, and endocrine factors that constitute an intrincate network of feedback loops. GH1 is evolutionarily coupled with other genes in linkage disequilibrium in 17q24.2, and the same applies to IGF2 in 11p15.5. In contrast, IGF1 in 12q22-24.1 is not in strong linkage disequilibrium with neighbouring genes. Knowledge of the functional architecture of these regions is important for the understanding of the combined evolution and function of GH1, IGF2 and IGF1 in relation to complex diseases. A number of mutations accounting for rare Mendelian disorders have been described in GH-IGF elements. The constellation of genes in this key pathway contains potential candidates in a number of complex diseases, including growth disorders, metabolic syndrome, diabetes (notably IGF2BP2) cardiovascular disease, and central nervous system diseases, and in longevity, aging and cancer. We review these genes and their associations with disease phenotypes, with special attention to metabolic risk traits. PMID:17534663

  14. Recombinant human insulin-like growth factor I stimulates growth and has distinct effects on organ size in hypophysectomized rats.

    PubMed Central

    Guler, H P; Zapf, J; Scheiwiller, E; Froesch, E R

    1988-01-01

    Recombinant human insulin-like growth factor I (rhIGF-I) was infused subcutaneously into hypophysectomized rats for as long as 18 days. Three hundred micrograms (39 nmol) of rhIGF-I per day and 200 milliunits (4.5 nmol) of human growth hormone (hGH) per day increased body weight, tibial epiphyseal width, longitudinal bone growth, and trabecular bone formation similarly. Weight gains of the kidneys and spleen, however, were greater with rhIGF-I than with hGH, whereas the weight of the epididymal fat pads was reduced with rhIGF-I. The weight of the thymus was increased by rhIGF-I treatment. Thus, IGF-I administered over a prolonged period of time mimics GH effects in hypophysectomized rats. Quantitative differences between rhIGF-I and hGH treatment with respect to organ weights may be related to different forms of circulating IGF-I or may be due to independent effects of GH and IGF-I. The results support the somatomedin hypothesis, but they also stress the role of GH as a modulator of IGF-I action. Images PMID:3387445

  15. Interaction between Fibrinogen and Insulin-Like Growth Factor-Binding Protein-1 in Human Plasma under Physiological Conditions.

    PubMed

    Gligorijević, N; Nedić, O

    2016-02-01

    Fibrinogen is a plasma glycoprotein and one of the principle participants in blood coagulation. It interacts with many proteins during formation of a blood clot, including insulin-like growth factors (IGFs) and their binding proteins (IGFBP). Fibrinogen complexes were found as minor fractions in fibrinogen preparations independently of the coagulation process, and their presence influences the kinetics of polymerization. The idea of this work was to investigate whether fibrinogen in human plasma interacts with IGFBPs independently of the tissue injury or coagulation process. The results have shown that fibrinogen forms complexes with IGFBP-1 under physiological conditions. Several experimental approaches have confirmed that complexes are co-isolated with fibrinogen from plasma, they are relatively stable, and they appear as a general feature of human plasma. Several other experiments excluded the possibility that alpha-2 macroglobulin/IGFBP-1 complexes or IGFBP-1 oligomers contributed to IGFBP-1 immunoreactivity. The role of fibrinogen/IGFBP-1 complexes is still unknown. Further investigation in individuals expressing both impaired glucose control and coagulopathy could contribute to identification and understanding of their possible physiological role. PMID:27260393

  16. [Insulin and physical exercise].

    PubMed

    Louis-Sylvestre, J

    1987-04-01

    Secretion of some pituitary hormones and sympatho-adrenal activity increase very early during exercise. Sympathetic activation is of major importance in cardiovascular adaptation, thermoregulation, etc. Furthermore among the hormonal consequences of such activation those related to insulin are capital. In animal and human subjects basal insulin level decrease during prolonged and progressive exercise. With habitual exercise, both basal and stimulated insulin levels are reduced. It seems that the reduced basal level could be due to alpha-adrenergic inhibition of the islets of Langerhans, while the reduced stimulated response could be the consequence of increased clearance. In trained subjects, in spite of reduced insulin secretion tolerance to glucose is normal due to increased sensitivity to insulin. Sensitivity to insulin is particularly enhanced at the muscular tissue level; it is accompanied by increased hexokinase and glycogen synthetase activity. As a consequence glucose uptake remains optimal at the muscular level. In the liver, both insulin sensitivity and glucokinase activity are reduced, so that glucose is spared and the muscular glycogen store can be restored. At the adipocyte level, metabolic adaptations are such that triglyceride turnover is greatly increased, favouring fuel supply and resaturation of stores.

  17. Differential half-maximal effects of human insulin and its analogs for in situ glucose transport and protein synthesis in rat soleus muscle

    NASA Technical Reports Server (NTRS)

    Weinstein, Randi B.; Eleid, Noura; LeCesne, Catherine; Durando, Bianca; Crawford, Julie T.; Heffner, Michelle; Layton, Christle; O'Keefe, Matthew; Robinson, Jennifer; Rudinsky, Suzy; Henriksen, Erik J.; Tischler, Marc E.

    2002-01-01

    Analogs of human insulin have been used to discriminate between responses of metabolic and mitogenic (growth-related) pathways. This study compared the stimulatory effects of human insulin (HI) and 2 analogs (X2, B-Asp(9), B-Glu(27) and H2, A-His(8),B-His(4),B-Glu(10), B-His(27)) on glucose uptake and protein synthesis in rat soleus muscle in situ. Glucose uptake, estimated by intramuscular (IM) injection of 2-deoxy[1,2-3H]glucose with or without insulin, was maximally increased at 10(-6) mol/L for HI and X2 and 10(-7) mol/L for H2. HI had a larger effect (318%) than either X2 (156%) or H2 (124%). The half-maximal effect (ED(50)) values for HI, X2, and H2 were 3.3 x10(-8) mol/L, 1.7 x 10(-7) mol/L, and 1.6 x 10(-9) mol/L, respectively. Protein synthesis, estimated by protein incorporation of [(3)H]phenylalanine injected into muscles with or without insulin, was maximally increased at 10(-5) mol/L for HI and 10(-6) for X2 and H2. HI had a larger effect in stimulating protein synthesis (34%) than either X2 (25%) or H2 (19.8%). The ED(50) values for HI, X2, and H2 were 3.0 x 10(-7) mol/L, 3.2 x 10(-7) mol/L, and 1.0 x 10(-9) mol/L, respectively. The biological potency of each analog (ED(50)insulin/ED(50)analog) showed X2 to be less potent than HI for both glucose uptake (0.2) and protein synthesis (0.9), whereas H2 is more potent than HI with ratios of 20 and 300, respectively. These data suggest that this approach for studying insulin responsiveness in a single muscle in situ may be a useful tool for investigating insulin signaling in muscle in vivo. Copyright 2002, Elsevier Science (USA). All rights reserved.

  18. Differentiation of Human Bone Marrow-Derived Mesenchymal Stem Cells into Insulin-Producing Cells: Evidence for Further Maturation In Vivo

    PubMed Central

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

    2015-01-01

    The aim of this study was to provide evidence for further in vivo maturation of insulin-producing cells (IPCs) derived from human bone marrow-derived mesenchymal stem cells (HBM-MSCs). HBM-MSCs were obtained from three insulin-dependent type 2 diabetic volunteers. Following expansion, cells were differentiated according to a trichostatin-A/GLP protocol. One million cells were transplanted under the renal capsule of 29 diabetic nude mice. Blood glucose, serum human insulin and c-peptide levels, and glucose tolerance curves were determined. Mice were euthanized 1, 2, 4, or 12 weeks after transplantation. IPC-bearing kidneys were immunolabeled, number of IPCs was counted, and expression of relevant genes was determined. At the end of in vitro differentiation, all pancreatic endocrine genes were expressed, albeit at very low values. The percentage of IPCs among transplanted cells was small (≤3%). Diabetic animals became euglycemic 8 ± 3 days after transplantation. Thereafter, the percentage of IPCs reached a mean of ~18% at 4 weeks. Relative gene expression of insulin, glucagon, and somatostatin showed a parallel increase. The ability of the transplanted cells to induce euglycemia was due to their further maturation in the favorable in vivo microenvironment. Elucidation of the exact mechanism(s) involved requires further investigation. PMID:26064925

  19. Differentiation of Human Bone Marrow-Derived Mesenchymal Stem Cells into Insulin-Producing Cells: Evidence for Further Maturation In Vivo.

    PubMed

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

    2015-01-01

    The aim of this study was to provide evidence for further in vivo maturation of insulin-producing cells (IPCs) derived from human bone marrow-derived mesenchymal stem cells (HBM-MSCs). HBM-MSCs were obtained from three insulin-dependent type 2 diabetic volunteers. Following expansion, cells were differentiated according to a trichostatin-A/GLP protocol. One million cells were transplanted under the renal capsule of 29 diabetic nude mice. Blood glucose, serum human insulin and c-peptide levels, and glucose tolerance curves were determined. Mice were euthanized 1, 2, 4, or 12 weeks after transplantation. IPC-bearing kidneys were immunolabeled, number of IPCs was counted, and expression of relevant genes was determined. At the end of in vitro differentiation, all pancreatic endocrine genes were expressed, albeit at very low values. The percentage of IPCs among transplanted cells was small (≤3%). Diabetic animals became euglycemic 8 ± 3 days after transplantation. Thereafter, the percentage of IPCs reached a mean of ~18% at 4 weeks. Relative gene expression of insulin, glucagon, and somatostatin showed a parallel increase. The ability of the transplanted cells to induce euglycemia was due to their further maturation in the favorable in vivo microenvironment. Elucidation of the exact mechanism(s) involved requires further investigation.

  20. Activation of L-arginine transport (system y+) and nitric oxide synthase by elevated glucose and insulin in human endothelial cells.

    PubMed Central

    Sobrevia, L; Nadal, A; Yudilevich, D L; Mann, G E

    1996-01-01

    1. Modulation of L-arginine transport (system y+) and release of nitric oxide (NO) and prostacyclin (PGI2) by elevated glucose and insulin were investigated in human cultured umbilical vein endothelial cells. 2. Elevated glucose induced a time- (6-12 h) and concentration-dependent stimulation of L-arginine transport, which was reversible and associated with a 3-fold increase in intracellular cGMP accumulation (index of NO synthesis) and 75% decrease in PGI2 production. 3. Elevated glucose had no effect on the initial transport rates for L-serine, L-citrulline, L-leucine, L-cystine or 2-deoxyglucose. 4. Resting membrane potential was unaffected by elevated glucose whereas basal intracellular [Ca2+] increased from 65 +/- 5 nM to 136 +/- 16 nM. 5. Insulin induced a protein synthesis-dependent stimulation of L-arginine transport and increased NO and PGI2 production in cells exposed to 5 mM glucose. 6. In cells exposed to high glucose, insulin downregulated elevated rates of L-arginine transport and cGMP accumulation but had no effect on the depressed PGI2 production. 7. Our findings suggest that insulin's normal stimulatory action on human endothelial cell vasodilator pathways may be impaired under conditions of sustained hyperglycaemia. PMID:8683475

  1. [Insulinization in type 2 diabetes mellitus. Intensification options].

    PubMed

    Fuente, Graciela V; Sinay, Isaac; Costa Gil, José E; Puchulu, Félix; Dieuzeide, Guillermo; Rodríguez, Martín; Faingold, María C; Litwak, León E

    2016-01-01

    Diabetes mellitus is associated with vascular complications and high rates of morbidity and mortality. Timely insulin therapy, intensified when necessary, represent appropriate measures to prevent or delay the onset of complications. However, the incidence of hypoglycemia and difficulties in treatment adherence represent barriers to achieve therapeutic success. Premixes analogs and, specially, combinations of insulin analogues are associated with pharmacokinetic and pharmacodynamic advantages, that translate into clinical benefits such as improved metabolic control, decreased hypoglycemic events and, for their simplicity, potentially greater adherence. PMID:27295707

  2. Recombinant human insulin-like growth factor binding proteins 4, 5, and 6: biological and physiochemical characterization.

    PubMed

    Kiefer, M C; Schmid, C; Waldvogel, M; Schläpfer, I; Futo, E; Masiarz, F R; Green, K; Barr, P J; Zapf, J

    1993-03-01

    We have recently cloned cDNAs encoding human insulin-like growth factor binding proteins (IGFBP)-4, -5 and -6 and have now expressed these BPs in yeast as ubiquitin (Ub)-IGFBP fusion proteins. Western ligand blotting with 125I-IGF II under nonreducing conditions of recombinant human (rh) IGFBP-containing yeast lysates revealed specific binding bands for IGFBP-4, -5, and -6 at apparent molecular masses of 24-26, 30-32, and 24-26 kDa, respectively, indicating expression and processing of the fusion proteins. HPLC purified rhIGFBPs had virtually the same amino acid composition, amino acid number, and NH2-terminal sequences as the native BPs. Rabbit antiserum directed against each rhIGFBP-4, -5 and -6 reacted specifically with the respective rhIGFBP as well as with the native human counterpart and displayed very low cross-reactivity with other IGFBPs. Except for the affinity of rhIGFBP-6 for IGF I (Ka = 8.5 x 10(8) M-1), the affinity constants of the three IGFBPs for IGF I and II lie between 1.7 and 3.3 x 10(10) M-1. When present in excess, rhIGFBP-4, -5, and -6 inhibited IGF I- and II-stimulated DNA and glycogen synthesis in human osteoblastic cells, although rh-IGFBP-6 had only a weak inhibitory effect on IGF I in agreement with its relatively lower IGF I affinity constant. PMID:7683532

  3. Biosimilar Insulins

    PubMed Central

    Hompesch, Marcus

    2014-01-01

    Until now most of the insulin used in developed countries has been manufactured and distributed by a small number of multinational companies. Beyond the established insulin manufacturers, a number of new players have developed insulin manufacturing capacities based on modern biotechnological methods. Because the patents for many of the approved insulin formulations have expired or are going to expire soon, these not yet established companies are increasingly interested in seeking market approval for their insulin products as biosimilar insulins (BI) in highly regulated markets like the EU and the United States. Differences in the manufacturing process (none of the insulin manufacturing procedures are 100% identical) can lead to insulins that to some extent may differ from the originator insulin. The key questions are if subtle differences in the structure of the insulins, purity, and so on are clinically relevant and may result in different biological effects. The aim of this article is to introduce and discuss basic aspects that may be of relevance with regard to BI. PMID:24876530

  4. Stimulation of glucose uptake by insulin-like growth factor II in human muscle is not mediated by the insulin-like growth factor II/mannose 6-phosphate receptor.

    PubMed Central

    Burguera, B; Elton, C W; Caro, J F; Tapscott, E B; Pories, W J; Dimarchi, R; Sakano, K; Dohm, G L

    1994-01-01

    Although the growth-promoting effects of insulin-like growth factor II (IGF-II) have been intensively studied, the acute actions of this hormone on glucose metabolism have been less well evaluated, especially in skeletal muscle of humans. We and other groups have shown that IGFs reduce glycaemic levels in humans and stimulate glucose uptake in rat muscle. The purpose of the present study was to evaluate the effect of IGF-II on glucose transport in muscle of normal and obese patients with and without non-insulin-dependent diabetes mellitus (NIDDM), as well as to identify the receptor responsible for this action. 2-Deoxyglucose transport was determined in vitro using a muscle-fibre strip preparation. IGF-II were investigated in biopsy material of rectus abdominus muscle taken from lean and obese patients and obese patients with NIDDM at the time of surgery. In the lean group, IGF-II (100 nM) stimulated glucose transport 2.1-fold, which was slightly less than stimulation by insulin (2.8-fold) at the same concentration. Binding of IGF-II was approx. 25% of that of insulin at 1 nM concentrations of both hormones. Obesity with or without NIDDM significantly reduced IGF-II-stimulated glucose uptake compared with the lean group. In order to explore which receptor mediated the IGF-II effect, we compared glucose uptake induced by IGF-II and two IGF-II analogues: [Leu27]IGF-II, with high affinity for the IGF-II/Man 6-P receptor but markedly reduced affinity for the IGF-I and insulin receptors, and [Arg54,Arg55]IGF-II was similar to that of IGF-II, whereas [Leu27]IGF-II had a very diminished effect. Results show that IGF-II is capable of stimulating muscle glucose uptake in lean but not in obese subjects and this effect seems not to be mediated via an IGF-II/Man 6-P receptor. Images Figure 2 PMID:8010960

  5. Identification and characterization of a functional retinoic acid/thyroid hormone-response element upstream of the human insulin gene enhancer.

    PubMed Central

    Clark, A R; Wilson, M E; London, N J; James, R F; Docherty, K

    1995-01-01

    A deletion analysis of the human insulin gene extending to 2 kb upstream of the transcription start site provided evidence of regulatory sequences located upstream of the insulin-linked polymorphic region (ILPR). Within this ILPR-distal region is a sequence (Ink, for insulin kilobase upstream) which contains three potential nuclear hormone-receptor half-sites, closely matching the consensus sequence AGGTCA. These sequences are arranged as a palindromic element with zero spacing over-lapping a direct repeat with 2 bp spacing. The Ink sequence was used in electrophoretic mobility-shift assays within nuclear extracts from COS-7 cells overexpressing the vitamin D, thyroid hormone or retinoic acid receptors, or from an insulin-expressing hamster cell line, HIT-T15. These studies suggest that the insulin-expressing cell line contains thyroid hormone and retinoic acid receptors at least, and that these receptors are able to recognize the Ink sequence. Three copies of the Ink sequence were placed upstream of the thymidine kinase promoter and firefly luciferase reporter gene. In COS-7 cells expressing the appropriate nuclear hormone receptor, this construct was responsive to both thyroid hormone (18-fold) and all-trans-retinoic acid (31-fold). In HIT-T15 cells the same construct responded to all-trans-retinoic acid, but not to thyroid hormone. Within the context of a 2 kb insulin gene fragment, the Ink sequence was shown to be activated by retinoic acid and by the retinoic acid receptor, but acted as a negative element in the presence of both retinoic acid and the retinoic acid receptor. Mutagenesis studies demonstrated that the palindromic sequence was important for the retinoic acid response, and for binding of complexes containing retinoic acid receptor. In human islets of Langerhans, retinoic acid was shown to stimulate insulin mRNA levels. These results demonstrate that a functional nuclear hormone-receptor-response element is located upstream of the human ILPR. As

  6. Insulin Delivery System

    NASA Technical Reports Server (NTRS)

    1988-01-01

    When Programmable Implantable Medication System (PIMS) is implanted in human body, it delivers precise programmed amounts of insulin over long periods of time. Mini-Med Technologies has been refining the Technologies since initial development at APL. The size of a hockey puck, and encased in titanium shell, PIMS holds about 2 1/2 teaspoons of insulin at a programmed basal rate. If a change in measured blood sugar level dictates a different dose, the patient can vary the amount of insulin delivered by holding a small radio transceiver over the implanted system and dialing in a specific program held in the PIMS computer memory. Insulin refills are accomplished approximately 4 times a year by hypodermic needle.

  7. Premixer Design for High Hydrogen Fuels

    SciTech Connect

    Benjamin P. Lacy; Keith R. McManus; Balachandar Varatharajan; Biswadip Shome

    2005-12-16

    This 21-month project translated DLN technology to the unique properties of high hydrogen content IGCC fuels, and yielded designs in preparation for a future testing and validation phase. Fundamental flame characterization, mixing, and flame property measurement experiments were conducted to tailor computational design tools and criteria to create a framework for predicting nozzle operability (e.g., flame stabilization, emissions, resistance to flashback/flame-holding and auto-ignition). This framework was then used to establish, rank, and evaluate potential solutions to the operability challenges of IGCC combustion. The leading contenders were studied and developed with the most promising concepts evaluated via computational fluid dynamics (CFD) modeling and using the design rules generated by the fundamental experiments, as well as using GE's combustion design tools and practices. Finally, the project scoped the necessary steps required to carry the design through mechanical and durability review, testing, and validation, towards full demonstration of this revolutionary technology. This project was carried out in three linked tasks with the following results. (1) Develop conceptual designs of premixer and down-select the promising options. This task defined the ''gap'' between existing design capabilities and the targeted range of IGCC fuel compositions and evaluated the current capability of DLN pre-mixer designs when operated at similar conditions. Two concepts (1) swirl based and (2) multiple point lean direct injection based premixers were selected via a QFD from 13 potential design concepts. (2) Carry out CFD on chosen options (1 or 2) to evaluate operability risks. This task developed the leading options down-selected in Task 1. Both a GE15 swozzle based premixer and a lean direct injection concept were examined by performing a detailed CFD study wherein the aerodynamics of the design, together with the chemical kinetics of the combustion process, were

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

    PubMed

    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.

  9. The melanocortin system and insulin resistance in humans: insights from a patient with complete POMC deficiency and type 1 diabetes mellitus.

    PubMed

    Aslan, I R; Ranadive, S A; Valle, I; Kollipara, S; Noble, J A; Vaisse, C

    2014-01-01

    The central melanocortin system is essential for the regulation of long-term energy homeostasis in humans. Rodent experiments suggest that this system also affects glucose metabolism, in particular by modulating peripheral insulin sensitivity independently of its effect on adiposity. Rare patients with complete genetic defects in the central melanocortin system can provide insight into the role of this system in glucose homeostasis in humans. We here describe the eighth individual with complete proopiomelanocortin (POMC) deficiency and the first with coincidental concomitant type 1 diabetes, which provides a unique opportunity to determine the role of melanocortins in glucose homeostasis in human. Direct sequencing of the POMC gene in this severely obese patient with isolated adrenocorticotropic hormone deficiency identified a homozygous 5' untranslated region mutation -11C>A, which we find to abolish normal POMC protein synthesis, as assessed in vitro. The patient's insulin requirements were as expected for his age and pubertal development. This unique patient suggests that in humans the central melanocortin system does not seem to affect peripheral insulin sensitivity, independently of its effect on adiposity. PMID:23649472

  10. Autocrine insulin-like growth factor-I signaling promotes growth and survival of human acute myeloid leukemia cells via the phosphoinositide 3-kinase/Akt pathway.

    PubMed

    Doepfner, K T; Spertini, O; Arcaro, A

    2007-09-01

    Insulin-like growth factor (IGF) signaling plays an important role in various human cancers. Therefore, the role of insulin-like growth factor I (IGF-I) signaling in growth and survival of acute myeloid leukemia (AML) cells was investigated. Expression of the IGF-I receptor (IGF-IR) and its ligand IGF-I were detected in a panel of human AML blasts and cell lines. IGF-I and insulin promoted the growth of human AML blasts in vitro and activated the phosphoinositide 3-kinase (PI3K)/Akt and the extracellular signal-regulated kinase (Erk) pathways. IGF-I-stimulated growth of AML blasts was blocked by an inhibitor of the PI3K/Akt pathway. Moreover, downregulation of the class Ia PI3K isoforms p110beta and p110delta by RNA interference impaired IGF-I-stimulated Akt activation, cell growth and survival in AML cells. Proliferation of a panel of AML cell lines and blasts isolated from patients with AML was inhibited by the IGF-IR kinase inhibitor NVP-AEW541 or by an IGF-IR neutralizing antibody. In addition to its antiproliferative effects, NVP-AEW541 sensitized primary AML blasts and cell lines to etoposide-induced apoptosis. Together, our data describe a novel role for autocrine IGF-I signaling in the growth and survival of primary AML cells. IGF-IR inhibitors in combination with chemotherapeutic agents may represent a novel approach to target human AML.

  11. Insulin oedema.

    PubMed Central

    Evans, D. J.; Pritchard-Jones, K.; Trotman-Dickenson, B.

    1986-01-01

    A 35 year old markedly underweight woman presented with uncontrolled diabetes. Following insulin therapy she developed gross fluid retention with extensive peripheral oedema, bilateral pleural effusions and weight gain of 18.8 kg in 22 days, accompanied by a fall in plasma albumin. She responded well to treatment with diuretics and salt-poor albumin, losing 10.3 kg in 6 days without recurrence of oedema. Severe insulin oedema is an uncommon complication of insulin therapy and may be due to effects of insulin on both vascular permeability and the renal tubule. Images Figure 2 PMID:3529068

  12. Therapeutic potential of the dual peroxisome proliferator activated receptor (PPAR)α/γ agonist aleglitazar in attenuating TNF-α-mediated inflammation and insulin resistance in human adipocytes.

    PubMed

    Massaro, Marika; Scoditti, Egeria; Pellegrino, Mariangela; Carluccio, Maria Annunziata; Calabriso, Nadia; Wabitsch, Martin; Storelli, Carlo; Wright, Matthew; De Caterina, Raffaele

    2016-05-01

    Adipose tissue inflammation is a mechanistic link between obesity and its related sequelae, including insulin resistance and type 2 diabetes. Dual ligands of peroxisome proliferator activated receptor (PPAR)α and γ, combining in a single molecule the metabolic and inflammatory-regulatory properties of α and γ agonists, have been proposed as a promising therapeutic strategy to antagonize adipose tissue inflammation. Here we investigated the effects of the dual PPARα/γ agonist aleglitazar on human adipocytes challenged with inflammatory stimuli. Human Simpson-Golabi-Behmel syndrome (SGBS) adipocytes were treated with aleglitazar or - for comparison - the selective agonists for PPARα or γ fenofibrate or rosiglitazone, respectively, for 24h before stimulation with TNF-α. Aleglitazar, at concentrations as low as 10nmol/L, providing the half-maximal transcriptional activation of both PPARα and PPARγ, reduced the stimulated expression of several pro-inflammatory mediators including interleukin (IL)-6, the chemokine CXC-L10, and monocyte chemoattractant protein (MCP)-1. Correspondingly, media from adipocytes treated with aleglitazar reduced monocyte migration, consistent with suppression of MCP-1 secretion. Under the same conditions, aleglitazar also reversed the TNF-α-mediated suppression of insulin-stimulated ser473 Akt phosphorylation and decreased the TNF-α-induced ser312 IRS1 phosphorylation, two major switches in insulin-mediated metabolic activities, restoring glucose uptake in insulin-resistant adipocytes. Such effects were similar to those obtainable with a combination of single PPARα and γ agonists. In conclusion, aleglitazar reduces inflammatory activation and dysfunction in insulin signaling in activated adipocytes, properties that may benefit diabetic and obese patients. The effect of aleglitazar was consistent with dual PPARα and γ agonism, but with no evidence of synergism. PMID:26976796

  13. Insulin Resistance and Body Weight: Recent Human Studies Documenting the Benefits of Supplemental Chromium

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The essentiality of chromium for humans was confirmed more than three decades ago with the studies showing that patients on total parenteral nutritrion (TPN) developed severe diabetic-like symptoms that could be reversed by the addition of chromium to their parenteral nutrition solutions. Since the...

  14. The challenge of improved secretory production of active pharmaceutical ingredients in Saccharomyces cerevisiae: a case study on human insulin analogs.

    PubMed

    Kazemi Seresht, Ali; Palmqvist, Eva A; Schluckebier, Gerd; Pettersson, Ingrid; Olsson, Lisbeth

    2013-10-01

    The yeast Saccharomyces cerevisiae has widely been used as a host for the production of heterologous proteins. Great attention has been put on improved secretory production of active pharmaceutical ingredients, and the secretory pathway of this eukaryotic host has been the playground of diverse strain engineering studies, aiming at enhanced cellular capacities for folding and trafficking of the target proteins. However, the cellular quality assessment for secretory proteins remains mostly unpredictable, and different target proteins often do not picture similar secretion yields, underlining the dependency of efficient secretion on the physicochemical properties of the protein of interest. In this study, two human insulin analog precursors (IAPs) with minor differences in their amino acid sequences were used as model secretory proteins. No differences between cells expressing these two proteins were found in the IAP transcript levels, gene copy numbers, or intra-cellularly accumulated proteins, yet a more than sevenfold difference in their secretion yields was found. Physiological characterization of cells expressing these proteins in batch processes revealed no significant difference in their specific growth rate, but an altered overflow metabolism. Global transcriptome analysis carried out in chemostat experiments pinpointed distinct steps during the protein maturation pathway to be differentially regulated and indicated an increased degradation of the IAP with the low secretion yield. In silico protein structure modeling of the IAPs suggested a difference in conformational stability, induced by the amino acid substitution, which most likely resulted in disparity in trafficking through the secretory pathway and thus a large difference in secretion yields. PMID:23592021

  15. Insulin-like growth factor-1 delays Fas-mediated apoptosis in human neutrophils through the phosphatidylinositol-3 kinase pathway.

    PubMed

    Himpe, Eddy; Degaillier, Céline; Coppens, Astrid; Kooijman, Ron

    2008-10-01

    Apoptosis of human neutrophils is a crucial mechanism for the resolution of inflammation. We previously showed that insulin-like growth factor-1 (IGF1) delays spontaneous neutrophil apoptosis without influencing the secretion of cytokines by these cells. In the present study, we further addressed the role of IGF1 in regulating neutrophil survival in the presence of other factors present during inflammation, and the mechanism involved in delaying apoptosis. We show that IGF1 delays neutrophil apoptosis triggered by the agonistic anti-Fas antibody CH11 and that the effect of IGF1 is comparable in magnitude to that of the acknowledged anti-apoptotic cytokines interferon-gamma (IFNG) and granulocyte-macrophage colony-stimulating factor (GM-CSF; now known as CSF2). Furthermore, IGF1 exerted additional effects on cell survival in the presence of these cytokines. IGF1 did not affect Fas expression or activation by anti-Fas of caspase-8, but inhibited the depolarization of the mitochondrial membrane. Inhibitor studies indicate that the phosphatidylinositol-3 kinase (PI3K) pathway, but not the MEK-ERK pathway, mediates the effects of IGF1. However, in contrast to CSF2, IGF1 did not induce phosphorylation and translocation to the membrane of AKT, the canonical downstream target of PI3K. We therefore speculate that other downstream targets of PI3K are involved in the delay of neutrophil apoptosis by IGF1, possibly through stabilization of the mitochondrial membrane.

  16. HNF4α Antagonists Discovered by a High-Throughput Screen for Modulators of the Human Insulin Promoter

    PubMed Central

    Kiselyuk, Alice; Lee, Seung-Hee; Farber-Katz, Suzette; Zhang, Mingjun; Athavankar, Sonalee; Cohen, Tom; Pinkerton, Anthony B.; Ye, Mao; Bushway, Paul; Richardson, Adam D.; Hostetler, Heather A.; Rodriguez-Lee, Mariam; Huang, Li; Spangler, Benjamin; Smith, Layton; Higginbotham, Jennifer; Cashman, John; Freeze, Hudson; Itkin-Ansari, Pamela; Dawson, Marcia I.; Schroeder, Friedhelm; Cang, Yong; Mercola, Mark; Levine, Fred

    2012-01-01

    SUMMARY Hepatocyte Nuclear Factor (HNF)4α is a central regulator of gene expression in cell types that play a critical role in metabolic homeostasis, including hepatocytes, enterocytes, and pancreatic β-cells. Although fatty acids were found to occupy the HNF4α ligand-binding pocket and proposed to act as ligands, there is controversy about both the nature of HNF4α ligands as well as the physiological role of the binding. Here, we report the discovery of potent synthetic HNF4α antagonists through a high-throughput screen for effectors of the human insulin promoter. These molecules bound to HNF4α with high affinity and modulated the expression of known HNF4α target genes. Notably, they were found to be selectively cytotoxic to cancer cell lines in vitro and in vivo, although in vivo potency was limited by suboptimal pharmacokinetic properties. The discovery of bioactive modulators for HNF4α raises the possibility that diseases involving HNF4α, such as diabetes and cancer, might be amenable to pharmacologic intervention by modulation of HNF4α activity. PMID:22840769

  17. A case of hypersensitivity to soluble and isophane insulins but not to insulin glargine

    PubMed Central

    Belhekar, Mahesh N.; Pai, Sarayu; Tayade, Parimal; Dalwadi, Pradip; Munshi, Renuka; Varthakavi, Prema

    2015-01-01

    Insulin is an important agent for the treatment of diabetes mellitus (DM). Allergic reactions to insulin therapy, although rare, have been evident since animal insulin became available for the treatment of DM in 1922. Hypersensitivity to insulin has considerably been reduced with the introduction of human insulin produced by recombinant deoxyribonucleic acid technology. Here, we present a case of Type 2 DM who demonstrated immediate (Type 1) hypersensitivity reaction on the sites of subcutaneous injection of soluble and isophane insulin but insulin glargine was tolerated well and provided good glycemic control. PMID:25878390

  18. A case of hypersensitivity to soluble and isophane insulins but not to insulin glargine.

    PubMed

    Belhekar, Mahesh N; Pai, Sarayu; Tayade, Parimal; Dalwadi, Pradip; Munshi, Renuka; Varthakavi, Prema

    2015-01-01

    Insulin is an important agent for the treatment of diabetes mellitus (DM). Allergic reactions to insulin therapy, although rare, have been evident since animal insulin became available for the treatment of DM in 1922. Hypersensitivity to insulin has considerably been reduced with the introduction of human insulin produced by recombinant deoxyribonucleic acid technology. Here, we present a case of Type 2 DM who demonstrated immediate (Type 1) hypersensitivity reaction on the sites of subcutaneous injection of soluble and isophane insulin but insulin glargine was tolerated well and provided good glycemic control. PMID:25878390

  19. Consuming fructose-sweetened, not glucose-sweetened, beverages increases visceral adiposity and lipids and decreases insulin sensitivity in overweight/obese humans

    PubMed Central

    Stanhope, Kimber L.; Schwarz, Jean Marc; Keim, Nancy L.; Griffen, Steven C.; Bremer, Andrew A.; Graham, James L.; Hatcher, Bonnie; Cox, Chad L.; Dyachenko, Artem; Zhang, Wei; McGahan, John P.; Seibert, Anthony; Krauss, Ronald M.; Chiu, Sally; Schaefer, Ernst J.; Ai, Masumi; Otokozawa, Seiko; Nakajima, Katsuyuki; Nakano, Takamitsu; Beysen, Carine; Hellerstein, Marc K.; Berglund, Lars; Havel, Peter J.

    2009-01-01

    Studies in animals have documented that, compared with glucose, dietary fructose induces dyslipidemia and insulin resistance. To assess the relative effects of these dietary sugars during sustained consumption in humans, overweight and obese subjects consumed glucose- or fructose-sweetened beverages providing 25% of energy requirements for 10 weeks. Although both groups exhibited similar weight gain during the intervention, visceral adipose volume was significantly increased only in subjects consuming fructose. Fasting plasma triglyceride concentrations increased by approximately 10% during 10 weeks of glucose consumption but not after fructose consumption. In contrast, hepatic de novo lipogenesis (DNL) and the 23-hour postprandial triglyceride AUC were increased specifically during fructose consumption. Similarly, markers of altered lipid metabolism and lipoprotein remodeling, including fasting apoB, LDL, small dense LDL, oxidized LDL, and postprandial concentrations of remnant-like particle–triglyceride and –cholesterol significantly increased during fructose but not glucose consumption. In addition, fasting plasma glucose and insulin levels increased and insulin sensitivity decreased in subjects consuming fructose but not in those consuming glucose. These data suggest that dietary fructose specifically increases DNL, promotes dyslipidemia, decreases insulin sensitivity, and increases visceral adiposity in overweight/obese adults. PMID:19381015

  20. Effects of Common Genetic Variants Associated With Type 2 Diabetes and Glycemic Traits on α- and β-Cell Function and Insulin Action in Humans

    PubMed Central

    Jonsson, Anna; Ladenvall, Claes; Ahluwalia, Tarunveer Singh; Kravic, Jasmina; Krus, Ulrika; Taneera, Jalal; Isomaa, Bo; Tuomi, Tiinamaija; Renström, Erik; Groop, Leif; Lyssenko, Valeriya

    2013-01-01

    Although meta-analyses of genome-wide association studies have identified >60 single nucleotide polymorphisms (SNPs) associated with type 2 diabetes and/or glycemic traits, there is little information on whether these variants also affect α-cell function. The aim of the current study was to evaluate the effects of glycemia-associated genetic loci on islet function in vivo and in vitro. We studied 43 SNPs in 4,654 normoglycemic participants from the Finnish population-based Prevalence, Prediction, and Prevention of Diabetes-Botnia (PPP-Botnia) Study. Islet function was assessed, in vivo, by measuring insulin and glucagon concentrations during oral glucose tolerance test, and, in vitro, by measuring glucose-stimulated insulin and glucagon secretion from human pancreatic islets. Carriers of risk variants in BCL11A, HHEX, ZBED3, HNF1A, IGF1, and NOTCH2 showed elevated whereas those in CRY2, IGF2BP2, TSPAN8, and KCNJ11 showed decreased fasting and/or 2-h glucagon concentrations in vivo. Variants in BCL11A, TSPAN8, and NOTCH2 affected glucagon secretion both in vivo and in vitro. The MTNR1B variant was a clear outlier in the relationship analysis between insulin secretion and action, as well as between insulin, glucose, and glucagon. Many of the genetic variants shown to be associated with type 2 diabetes or glycemic traits also exert pleiotropic in vivo and in vitro effects on islet function. PMID:23557703

  1. Obesity–insulin targeted genes in the 3p26-25 region in human studies and LG/J and SM/J mice

    PubMed Central

    Kraja, Aldi T.; Lawson, Heather A.; Arnett, Donna K.; Borecki, Ingrid B.; Broeckel, Ulrich; de las Fuentes, Lisa; Hunt, Steven C.; Province, Michael A.; Cheverud, James; Rao, D.C.

    2012-01-01

    Identifying metabolic syndrome (MetS) genes is important for novel drug development and health care. This study extends the findings on human chromosome 3p26-25 for an identified obesity–insulin factor QTL, with an LOD score above 3. A focused association analysis comprising up to 9578 African American and Caucasian subjects from the HyperGEN Network (908 African Americans and 1025 whites), the Family Heart Study (3035 whites in time 1 and 1943 in time 2), and the Framingham Heart Study (1317 in Offspring and 1320 in Generation 3) was performed. The homologous mouse region was explored in an F16 generation of an advanced intercross between the LG/J and SM/J inbred strains, in an experiment where 1002 animals were fed low-fat (247 males; 254 females) or high-fat (253 males; 248 females) diets. Association results in humans indicate pleiotropic effects for SNPs within or surrounding CNTN4 on obesity, lipids and blood pressure traits and for SNPs near IL5RA, TRNT1, CRBN, and LRRN1 on central obesity and blood pressure. Linkage analyses of this region in LG/J × SM/J mice identify a highly significant pleiotropic QTL peak for insulin and glucose levels, as well as response to glucose challenge. The mouse results show that insulin and glucose levels interact with high and low fat diets and differential gene expression was identified for Crbn and Arl8b. In humans, ARL8B resides ~137 kbps away from BHLHE40, expression of which shows up-regulation in response to insulin treatment. This focused human genetic analysis, incorporating mouse research evidenced that 3p26-25 has important genetic contributions to MetS components. Several of the candidate genes have functions in the brain. Their interaction with MetS and the brain warrants further investigation. PMID:22386932

  2. Obesity-insulin targeted genes in the 3p26-25 region in human studies and LG/J and SM/J mice.

    PubMed

    Kraja, Aldi T; Lawson, Heather A; Arnett, Donna K; Borecki, Ingrid B; Broeckel, Ulrich; de las Fuentes, Lisa; Hunt, Steven C; Province, Michael A; Cheverud, James; Rao, D C

    2012-08-01

    Identifying metabolic syndrome (MetS) genes is important for novel drug development and health care. This study extends the findings on human chromosome 3p26-25 for an identified obesity-insulin factor QTL, with an LOD score above 3. A focused association analysis comprising up to 9578 African American and Caucasian subjects from the HyperGEN Network (908 African Americans and 1025 whites), the Family Heart Study (3035 whites in time 1 and 1943 in time 2), and the Framingham Heart Study (1317 in Offspring and 1320 in Generation 3) was performed. The homologous mouse region was explored in an F(16) generation of an advanced intercross between the LG/J and SM/J inbred strains, in an experiment where 1002 animals were fed low-fat (247 males; 254 females) or high-fat (253 males; 248 females) diets. Association results in humans indicate pleiotropic effects for SNPs within or surrounding CNTN4 on obesity, lipids and blood pressure traits and for SNPs near IL5RA, TRNT1, CRBN, and LRRN1 on central obesity and blood pressure. Linkage analyses of this region in LG/J×SM/J mice identify a highly significant pleiotropic QTL peak for insulin and glucose levels, as well as response to glucose challenge. The mouse results show that insulin and glucose levels interact with high and low fat diets and differential gene expression was identified for Crbn and Arl8b. In humans, ARL8B resides ~137kbps away from BHLHE40, expression of which shows up-regulation in response to insulin treatment. This focused human genetic analysis, incorporating mouse research evidenced that 3p26-25 has important genetic contributions to MetS components. Several of the candidate genes have functions in the brain. Their interaction with MetS and the brain warrants further investigation.

  3. Insulin has a biphasic effect on the ability of human chorionic gonadotropin to induce ovarian cysts in the rat.

    PubMed

    Bogovich, K; Clemons, J; Poretsky, L

    1999-08-01

    Hyperinsulinemia enhances the ability of subovulatory doses of human chorionic gonadotropin (hCG) to induce ovarian follicular cysts in the rat. To determine the relative contribution of these hormones to the development of ovarian cysts, adult female rats were treated with either (1) vehicle alone (controls), (2) a high-fat diet (HFD) to control for the effects of weight gain, (3) 1.5 to 6 IU hCG twice daily plus 6 U insulin (Ins)/d, or (4) 1.5 to 9 U Ins/d plus 3 IU hCG twice daily. On day 23 of the in vivo treatments, all groups that received at least 6 U Ins/d displayed increased body weight compared with control and HFD rats (P < or = .05). No control rats and only one HFD rat displayed ovarian cysts on this day. Plasma estrone (E1) and androstenedione (A4) were elevated in HFD rats with noncystic follicles compared with control rats (P < or = .05). Between 64% and 80% of rats on 6 U Ins/d plus twice-daily injections of 1.5 to 6 IU hCG displayed ovarian cysts on day 23. Plasma estradiol (E2) concentrations for these treatment groups were similar to those of control rats. Of the hormonally treated animals, only those that had ovarian cysts in response to twice-daily injections of 4.5 or 6 IU hCG plus 6 U Ins/d displayed elevated plasma A4 and/or testosterone compared with controls. In contrast, plasma E1 concentrations were elevated on day 23 for animals bearing ovarian cysts in response to increasing doses of hCG plus the fixed dose of 6 U Ins/d. Between 70% and 80% of rats treated twice daily with 3 IU hCG plus a daily dose of 1.5 to 6 U Ins displayed ovarian cysts on day 23. In marked contrast, only 25% of rats treated with this dose of hCG plus 9 U Ins/d developed cystic follicles. Of the plasma steroids tested, only E1 and A4 were elevated in these treatment groups compared with controls. However, these increases in plasma steroid concentrations did not correlate with the dose of insulin. We conclude from these data that, although the mechanisms remain to

  4. Studies in premixed combustion. Progress report, November 1, 1990--October 31, 1992

    SciTech Connect

    Sivashinsky, G.I.

    1992-08-01

    This report discusses the following topics on premixed combustion: theory of turbulent flame propagation; pattern formation in premixed flames and related problems; and pattern formation in extended systems. (LSP)

  5. A review of piscine islet xenotransplantation using wild-type tilapia donors and the production of transgenic tilapia expressing a "humanized" tilapia insulin.

    PubMed

    Wright, James R; Yang, Hua; Hyrtsenko, Olga; Xu, Bao-You; Yu, Weiming; Pohajdak, Bill

    2014-01-01

    Most islet xenotransplantation laboratories have focused on porcine islets, which are both costly and difficult to isolate. Teleost (bony) fish, such as tilapia, possess macroscopically visible distinct islet organs called Brockmann bodies which can be inexpensively harvested. When transplanted into diabetic nude mice, tilapia islets maintain long-term normoglycemia and provide human-like glucose tolerance profiles. Like porcine islets, when transplanted into euthymic mice, they are rejected in a CD4 T-cell-dependent manner. However, unlike pigs, tilapia are so phylogenetically primitive that their cells do not express α(1,3)Gal and, because tilapia are highly evolved to live in warm stagnant waters nearly devoid of dissolved oxygen, their islet cells are exceedingly resistant to hypoxia, making them ideal for transplantation within encapsulation devices. Encapsulation, especially when combined with co-stimulatory blockade, markedly prolongs tilapia islet xenograft survival in small animal recipients, and a collaborator has shown function in diabetic cynomolgus monkeys. In anticipation of preclinical xenotransplantation studies, we have extensively characterized tilapia islets (morphology, embryologic development, cell biology, peptides, etc.) and their regulation of glucose homeostasis. Because tilapia insulin differs structurally from human insulin by 17 amino acids, we have produced transgenic tilapia whose islets stably express physiological levels of humanized insulin and have now bred these to homozygosity. These transgenic fish can serve as a platform for further development into a cell therapy product for diabetes.

  6. Induction of steroid sulfatase expression in PC-3 human prostate cancer cells by insulin-like growth factor II.

    PubMed

    Sung, Chul-Hoon; Im, Hee-Jung; Park, Nahee; Kwon, Yeojung; Shin, Sangyun; Ye, Dong-Jin; Cho, Nam-Hyeon; Park, Young-Shin; Choi, Hyung-Kyoon; Kim, Donghak; Chun, Young-Jin

    2013-11-25

    Human steroid sulfatase (STS) plays an important role in regulating the formation of biologically active estrogens and may be a promising target for treating estrogen-mediated carcinogenesis. The molecular mechanism of STS gene expression, however, is still not clear. Growth factors are known to increase STS activity but the changes in STS expression have not been completely understood. To determine whether insulin-like growth factor (IGF)-II can induce STS gene expression, the effects of IGF-II on STS expression were studied in PC-3 human prostate cancer cells. RT-PCR and Western blot analysis showed that IGF-II treatment significantly increased the expression of STS mRNA and protein in concentration- and time-dependent manners. To understand the signaling pathway by which IGF-II induces STS gene expression, the effects of specific PI3-kinase/Akt and NF-κB inhibitors were determined. When the cells were treated with IGF-II and PI3-kinase/Akt inhibitors, such as LY294002, wortmannin, or Akt inhibitor IV, STS expression induced by IGF-II was significantly blocked. Moreover, we found that NF-κB inhibitors, such as MG-132, bortezomib, Bay 11-7082 or Nemo binding domain (NBD) binding peptide, also strongly prevented IGF-II from inducing STS gene expression. We assessed whether IGF-II activates STS promoter activity using transient transfection with a luciferase reporter. IGF-II significantly stimulated STS reporter activity. Furthermore, IGF-II induced expression of 17β-hydroxysteroid dehydrogenase (HSD) 1 and 3, whereas it reduced estrone sulfotransferase (EST) gene expression, causing enhanced estrone and β-estradiol production. Taken together, these results strongly suggest that IGF-II induces STS expression via a PI3-kinase/Akt-NF-κB signaling pathway in PC-3 cells and may induce estrogen production and estrogen-mediated carcinogenesis. PMID:24055520

  7. Induction of steroid sulfatase expression in PC-3 human prostate cancer cells by insulin-like growth factor II.

    PubMed

    Sung, Chul-Hoon; Im, Hee-Jung; Park, Nahee; Kwon, Yeojung; Shin, Sangyun; Ye, Dong-Jin; Cho, Nam-Hyeon; Park, Young-Shin; Choi, Hyung-Kyoon; Kim, Donghak; Chun, Young-Jin

    2013-11-25

    Human steroid sulfatase (STS) plays an important role in regulating the formation of biologically active estrogens and may be a promising target for treating estrogen-mediated carcinogenesis. The molecular mechanism of STS gene expression, however, is still not clear. Growth factors are known to increase STS activity but the changes in STS expression have not been completely understood. To determine whether insulin-like growth factor (IGF)-II can induce STS gene expression, the effects of IGF-II on STS expression were studied in PC-3 human prostate cancer cells. RT-PCR and Western blot analysis showed that IGF-II treatment significantly increased the expression of STS mRNA and protein in concentration- and time-dependent manners. To understand the signaling pathway by which IGF-II induces STS gene expression, the effects of specific PI3-kinase/Akt and NF-κB inhibitors were determined. When the cells were treated with IGF-II and PI3-kinase/Akt inhibitors, such as LY294002, wortmannin, or Akt inhibitor IV, STS expression induced by IGF-II was significantly blocked. Moreover, we found that NF-κB inhibitors, such as MG-132, bortezomib, Bay 11-7082 or Nemo binding domain (NBD) binding peptide, also strongly prevented IGF-II from inducing STS gene expression. We assessed whether IGF-II activates STS promoter activity using transient transfection with a luciferase reporter. IGF-II significantly stimulated STS reporter activity. Furthermore, IGF-II induced expression of 17β-hydroxysteroid dehydrogenase (HSD) 1 and 3, whereas it reduced estrone sulfotransferase (EST) gene expression, causing enhanced estrone and β-estradiol production. Taken together, these results strongly suggest that IGF-II induces STS expression via a PI3-kinase/Akt-NF-κB signaling pathway in PC-3 cells and may induce estrogen production and estrogen-mediated carcinogenesis.

  8. [Local lipohypertrophy in insulin treatment].

    PubMed

    Herold, D A; Albrecht, G

    1993-01-01

    Local lipoatrophy and lipohypertrophy at injection sites are well known side effects of treatment with insulin. Conditions favouring these local complications are created when repeated or continuous injections are given into the same areas. We report on a 27-year-old female patient who suffered from persistent local swellings after use of an external pump which continuously injected human insulin via indwelling cannulas.

  9. Flame front geometry in premixed turbulent flames

    SciTech Connect

    Shepherd, I.G. ); Ashurst, W.T. )

    1991-12-01

    Experimental and numerical determinations of flame front curvature and orientation in premixed turbulent flames are presented. The experimental data is obtained from planar, cross sectional images of stagnation point flames at high Damkoehler number. A direct numerical simulation of a constant energy flow is combined with a zero-thickness, constant density flame model to provide the numerical results. The computational domain is a 32{sup 3} cube with periodic boundary conditions. The two-dimensional curvature distributions of the experiments and numerical simulations compare well at similar q{prime}/S{sub L} values with means close to zero and marked negative skewness. At higher turbulence levels the simulations show that the distributions become symmetric about zero. These features are also found in the three dimensional distributions of curvature. The simulations support assumptions which make it possible to determine the mean direction cosines from the experimental data. This leads to a reduction of 12% in the estimated flame surface area density in the middle of the flame brush. 18 refs.

  10. Numerical simulation of premixed turbulent methane combustion

    SciTech Connect

    Bell, John B.; Day, Marcus S.; Grcar, Joseph F.

    2001-12-14

    In this paper we study the behavior of a premixed turbulent methane flame in three dimensions using numerical simulation. The simulations are performed using an adaptive time-dependent low Mach number combustion algorithm based on a second-order projection formulation that conserves both species mass and total enthalpy. The species and enthalpy equations are treated using an operator-split approach that incorporates stiff integration techniques for modeling detailed chemical kinetics. The methodology also incorporates a mixture model for differential diffusion. For the simulations presented here, methane chemistry and transport are modeled using the DRM-19 (19-species, 84-reaction) mechanism derived from the GRIMech-1.2 mechanism along with its associated thermodynamics and transport databases. We consider a lean flame with equivalence ratio 0.8 for two different levels of turbulent intensity. For each case we examine the basic structure of the flame including turbulent flame speed and flame surface area. The results indicate that flame wrinkling is the dominant factor leading to the increased turbulent flame speed. Joint probability distributions are computed to establish a correlation between heat release and curvature. We also investigate the effect of turbulent flame interaction on the flame chemistry. We identify specific flame intermediates that are sensitive to turbulence and explore various correlations between these species and local flame curvature. We identify different mechanisms by which turbulence modulates the chemistry of the flame.

  11. Premixed flame propagation in vertical tubes

    NASA Astrophysics Data System (ADS)

    Kazakov, Kirill A.

    2016-04-01

    Analytical treatment of the premixed flame propagation in vertical tubes with smooth walls is given. Using the on-shell flame description, equations for a quasi-steady flame with a small but finite front thickness are obtained and solved numerically. It is found that near the limits of inflammability, solutions describing upward flame propagation come in pairs having close propagation speeds and that the effect of gravity is to reverse the burnt gas velocity profile generated by the flame. On the basis of these results, a theory of partial flame propagation driven by a strong gravitational field is developed. A complete explanation is given of the intricate observed behavior of limit flames, including dependence of the inflammability range on the size of the combustion domain, the large distances of partial flame propagation, and the progression of flame extinction. The role of the finite front-thickness effects is discussed in detail. Also, various mechanisms governing flame acceleration in smooth tubes are identified. Acceleration of methane-air flames in open tubes is shown to be a combined effect of the hydrostatic pressure difference produced by the ambient cold air and the difference of dynamic gas pressure at the tube ends. On the other hand, a strong spontaneous acceleration of the fast methane-oxygen flames at the initial stage of their evolution in open-closed tubes is conditioned by metastability of the quasi-steady propagation regimes. An extensive comparison of the obtained results with the experimental data is made.

  12. Premixed Gas Combustion: An Excitable System

    NASA Technical Reports Server (NTRS)

    Pearlman, Howard

    1997-01-01

    Rotating spiral and target patterns have been observed experimentally on freely-propagating premixed gas flames in large diameter tubes at normal gravity (1-g). These modes of propagation occur in near-limit mixtures which have a Lewis number (Le, defined as the ratio of the thermal diffusivity of the cold mixture to the mass diffusivity of the scarce component into the mixture) sufficiently greater than one. However, at 1-g, buoyant flows strongly distort the flame curvature, hydrodynamics (thus stretch) and convective transport of species and heat. In turn, these alter the critical Le required for onset of instability. To isolate and better understand the mechanisms which drive the observed patterns and their dynamics, 1-g and microgravity (micro-g) experiments are being conducted to determine: (1) the structure and dynamics of the patterns, (2) a map of the critical Le and heat loss for their occurrence, (3) the relative significance of the chemical kinetics, and (4) the effect of curvature (local wave and global flame front) on wave propagation. With this in hand, we will be better prepared to discuss an additional mode, a state of 'chemical turbulence,' which seems to be the ultimate fate of many of these near-limit flames prior to extinction.

  13. Premixed turbulent combustion to opposed streams

    SciTech Connect

    Kostiuk, L.W.; Cheng, R.K.

    1992-03-01

    Premixed turbulent combustion in opposed streams has been studied experimentally by the use of two component laser doppler aneomometry. This flow geometry is part of a class of stagnating flows used to study turbulent combustion in recent years. It does not involve any surface near the flames because of the flow symmetry thus circumventing many of the effects of flame surface interaction. The mean non-reacting flow is found to be self-similar for all the conditions studied in this and the stagnation plate configuration. A homogeneous region of plane straining is produced in the vicinity of the stagnation and there is a strong interaction between the turbulence in the flow and the mean straining which can increase the rms velocity as the flow stagnates. The reacting flow fields are found to be symmetric about the free stagnation point. The traverses of mean axial velocity in the stagnation streamlines for reaction flows are not dramatically different from the non-reaction flows. These results differ from turbulent combustion experiments where the flow is stagnated by a flat plate. The extinction limits was studied for propane:air mixtures. 11 refs.

  14. Transplacental passage of insulin complexed to antibody.

    PubMed Central

    Bauman, W A; Yalow, R S

    1981-01-01

    The passage of plasma proteins across the placental barrier in humans is known to be highly selective. Thus, free maternal insulin has been reported not to cross the normal maternofetal barrier, although insulin-binding antibodies have been detected in newborn infants whose diabetic mothers received insulin therapy. In this report we demonstrate, with the use of a human antiserum that permits distinction between human and animal insulins, that insulin in the cord blood of each of two neonates of insulin-treated diabetic mothers was, in part, animal insulin. The higher the antibody titer of the mother the greater was the total insulin in the cord plasma and the greater was the fraction that was animal insulin. In case 1 cord plasma insulin was 0.7 unit/liter, of which 10% was animal insulin; in case 2 cord plasma insulin was 3.5 units/liter, of which 25% was animal insulin. The demonstration that antigen restricted from transplacental passage can be transferred while complexed to antibody raises the question whether such fetal exposure would induce partial or total immunologic unresponsiveness subsequently if the fetus were rechallenged with the same antigen. PMID:7027265

  15. Soot Formation in Freely-Propagating Laminar Premixed Flames

    NASA Technical Reports Server (NTRS)

    Lin, K.-C.; Hassan, M. I.; Faeth, G. M.

    1997-01-01

    Soot formation within hydrocarbon-fueled flames is an important unresolved problem of combustion science. Thus, the present study is considering soot formation in freely-propagating laminar premixed flames, exploiting the microgravity environment to simplify measurements at the high-pressure conditions of interest for many practical applications. The findings of the investigation are relevant to reducing emissions of soot and continuum radiation from combustion processes, to improving terrestrial and spacecraft fire safety, and to developing methods of computational combustion, among others. Laminar premixed flames are attractive for studying soot formation because they are simple one-dimensional flows that are computationally tractable for detailed numerical simulations. Nevertheless, studying soot-containing burner-stabilized laminar premixed flames is problematical: spatial resolution and residence times are limited at the pressures of interest for practical applications, flame structure is sensitive to minor burner construction details so that experimental reproducibility is not very good, consistent burner behavior over the lengthy test programs needed to measure soot formation properties is hard to achieve, and burners have poor durability. Fortunately, many of these problems are mitigated for soot-containing, freely-propagating laminar premixed flames. The present investigation seeks to extend work in this laboratory for various soot processes in flames by observing soot formation in freely-propagating laminar premixed flames. Measurements are being made at both Normal Gravity (NG) and MicroGravity (MG), using a short-drop free-fall facility to provide MG conditions.

  16. Insulin signaling and addiction

    PubMed Central

    Daws, Lynette C.; Avison, Malcolm J.; Robertson, Sabrina D.; Niswender, Kevin D.; Galli, Aurelio; Saunders, Christine

    2012-01-01

    Across species, the brain evolved to respond to natural rewards such as food and sex. These physiological responses are important for survival, reproduction and evolutionary processes. It is no surprise, therefore, that many of the neural circuits and signaling pathways supporting reward processes are conserved from Caenorhabditis elegans to Drosophilae, to rats, monkeys and humans. The central role of dopamine (DA) in encoding reward and in attaching salience to external environmental cues is well recognized. Less widely recognized is the role of reporters of the “internal environment”, particularly insulin, in the modulation of reward. Insulin has traditionally been considered an important signaling molecule in regulating energy homeostasis and feeding behavior rather than a major component of neural reward circuits. However, research over recent decades has revealed that DA and insulin systems do not operate in isolation from each other, but instead, work together to orchestrate both the motivation to engage in consummatory behavior and to calibrate the associated level of reward. Insulin signaling has been found to regulate DA neurotransmission and to affect the ability of drugs that target the DA system to exert their neurochemical and behavioral effects. Given that many abused drugs target the DA system, the elucidation of how dopaminergic, as well as other brain reward systems, are regulated by insulin will create opportunities to develop therapies for drug and potentially food addiction. Moreover, a more complete understanding of the relationship between DA neurotransmission and insulin may help to uncover etiological bases for “food addiction” and the growing epidemic of obesity. This review focuses on the role of insulin signaling in regulating DA homeostasis and DA signaling, and the potential impact of impaired insulin signaling in obesity and psychostimulant abuse. PMID:21420985

  17. Mitogenic properties of insulin-like growth factors I and II, insulin-like growth factor binding protein-3 and epidermal growth factor on human breast stromal cells in primary culture.

    PubMed

    Strange, Karen S; Wilkinson, Darcy; Edin, Glenn; Emerman, Joanne T

    2004-03-01

    Insulin-like growth factors I and II (IGF-I and IGF-II) are growth factors implicated in both normal mammary gland development and breast cancer. We have previously reported on the effects of components of the IGF system on breast epithelial cells. Since data suggests that stromal-epithelial interactions play a crucial role in breast cancer, we have now investigated the mitogenic properties of IGF-I, IGF-II, insulin-like growth factor binding protein-3 (IGFBP-3) and epidermal growth factor (EGF) on human breast stromal cells in primary culture. We show that, under serum-free conditions, stromal cells are stimulated to grow in response to IGF-I and IGF-II in a dose-dependent manner. IGF-I and EGF, a potent stimulator of human breast epithelial cell growth in primary culture and also associated with breast cancer, appear to stimulate stromal cell growth in a synergistic manner. IGFBP-3 does not inhibit the stimulation of growth by IGF-I, or IGF-I plus EGF. However, IGFBP-3 does inhibit the stimulation of growth by IGF-II. In contrast to our previous results with human breast epithelial cells, IGFBP-3 does not have an IGF-independent inhibitory effect on stromal cell growth. This study is the first to address the effects of IGF-I, IGF-II and IGFBP-3 alone and in combination with EGF on human breast stromal cell growth in primary culture. Characterizing the role of the IGF system in both normal breast epithelial cells and stromal cells will aid in our understanding of the mechanisms behind the role of the IGF system in breast cancer.

  18. Carob pulp preparation rich in insoluble dietary fibre and polyphenols increases plasma glucose and serum insulin responses in combination with a glucose load in humans.

    PubMed

    Gruendel, Sindy; Otto, Baerbel; Garcia, Ada L; Wagner, Karen; Mueller, Corinna; Weickert, Martin O; Heldwein, Walter; Koebnick, Corinna

    2007-07-01

    Dietary fibre consumption is associated with improved glucose homeostasis. In contrast, dietary polyphenols have been suggested to exert both beneficial and detrimental effects on glucose and insulin metabolism. Recently, we reported that a polyphenol-rich insoluble dietary fibre preparation from carob pulp (carob fibre) resulted in lower postprandial acylated ghrelin levels after a liquid meal challenge test compared with a control meal without supplementation. The effects may, however, differ when a different food matrix is used. Thus, we investigated the effects of carob fibre on glucose, insulin and ghrelin responses in healthy humans in combination with a glucose load. In a randomized single-blind cross-over study involving twenty healthy subjects (aged 22-62 years), plasma glucose, total and acylated ghrelin, and serum insulin were repeatedly assessed before and after the ingestion of 200 ml water with 50 g glucose and 0, 5, 10 or 20 g carob fibre over a period of 180 min. The intake of 5 and 10 g carob fibre increased the plasma glucose by 47 % and 64 % (P < 0.001), and serum insulin by 19.9 and 24.8 % (P < 0.001), compared with the control. Plasma acylated ghrelin concentrations did not change significantly after the consumption of carob-enriched glucose solution. Total ghrelin decreased only after 10 g carob fibre (P < 0.001) compared with control. In conclusion, we showed that polyphenol-rich carob fibre, administered within a water-glucose solution, increases postprandial glucose and insulin responses, suggesting a deterioration in glycaemic control.

  19. 21 CFR 170.60 - Nitrites and/or nitrates in curing premixes.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 3 2014-04-01 2014-04-01 false Nitrites and/or nitrates in curing premixes. 170... nitrates in curing premixes. (a) Nitrites and/or nitrates are food additives when combined in curing... nitrates, when packaged separately from flavoring and seasoning in curing premixes, may continue to be...

  20. The Generation of Insulin Producing Cells from Human Mesenchymal Stem Cells by MiR-375 and Anti-MiR-9

    PubMed Central

    Jafarian, Arefeh; Taghikani, Mohammad; Abroun, Saeid; Allahverdi, Amir; Lamei, Maryam; Lakpour, Niknam; Soleimani, Masoud

    2015-01-01

    Background MicroRNAs (miRNAs) are a group of endogenous small non-coding RNAs that regulate gene expression at the post-transcriptional level. A number of studies have led to the notion that some miRNAs have key roles in control of pancreatic islet development and insulin secretion. Based on some studies on miRNAs pattern, the researchers in this paper investigated the pancreatic differentiation of human bone marrow mesenchymal stem cells (hBM-MSCs) by up-regulation of miR-375 and down-regulation of miR-9 by lentiviruses containing miR-375 and anti-miR-9. Methodology After 21 days of induction, islet-like clusters containing insulin producing cells (IPCs) were confirmed by dithizone (DTZ) staining. The IPCs and β cell specific related genes and proteins were detected using qRT-PCR and immunofluorescence on days 7, 14 and 21 of differentiation. Glucose challenge test was performed at different concentrations of glucose so extracellular and intracellular insulin and C-peptide were assayed using ELISA kit. Although derived IPCs by miR-375 alone were capable to express insulin and other endocrine specific transcription factors, the cells lacked the machinery to respond to glucose. Conclusion It was found that over-expression of miR-375 led to a reduction in levels of Mtpn protein in derived IPCs, while treatment with anti-miR-9 following miR-375 over-expression had synergistic effects on MSCs differentiation and insulin secretion in a glucose-regulated manner. The researchers reported that silencing of miR-9 increased OC-2 protein in IPCs that may contribute to the observed glucose-regulated insulin secretion. Although the roles of miR-375 and miR-9 are well known in pancreatic development and insulin secretion, the use of these miRNAs in transdifferentiation was never demonstrated. These findings highlight miRNAs functions in stem cells differentiation and suggest that they could be used as therapeutic tools for gene-based therapy in diabetes mellitus. PMID:26047014

  1. The Role of Human Factors in the Design and Development of an Insulin Pump

    PubMed Central

    Schaeffer, Noel E.

    2012-01-01

    This article discusses human factors (HF) processes and how they are applied during the development of a medical device to minimize the risk that the user interface design could lead to patient errors, adverse events, and product recalls. This process is best defined as “prevention through design.” The HF design process is exemplified by three distinct phases: (1) preliminary analysis, (2) formative design evaluation and modification, and (3) design validation. Additional benefits of employing HF principles during medical device development are briefly reviewed, including reduced patient risk by eliminating design flaws, increased patient adherence through the reduction in the complexity of therapeutic regimes, and reduced likelihood for product recalls. PMID:22538134

  2. Quantification of adipose tissue insulin sensitivity.

    PubMed

    Søndergaard, Esben; Jensen, Michael D

    2016-06-01

    In metabolically healthy humans, adipose tissue is exquisitely sensitive to insulin. Similar to muscle and liver, adipose tissue lipolysis is insulin resistant in adults with central obesity and type 2 diabetes. Perhaps uniquely, however, insulin resistance in adipose tissue may directly contribute to development of insulin resistance in muscle and liver because of the increased delivery of free fatty acids to those tissues. It has been hypothesized that insulin adipose tissue resistance may precede other metabolic defects in obesity and type 2 diabetes. Therefore, precise and reproducible quantification of adipose tissue insulin sensitivity, in vivo, in humans, is an important measure. Unfortunately, no consensus exists on how to determine adipose tissue insulin sensitivity. We review the methods available to quantitate adipose tissue insulin sensitivity and will discuss their strengths and weaknesses.

  3. Two Cases of Allergy to Insulin in Gestational Diabetes

    PubMed Central

    Kim, Gi Jun; Kim, Shin Bum; Jo, Seong Il; Shin, Jin Kyeong; Kwon, Hee Sun; Jeong, Heekyung; Son, Jang Won; Lee, Seong Su; Kim, Sung Rae; Kim, Byung Kee

    2015-01-01

    Allergic reaction to insulin is uncommon since the introduction of human recombinant insulin preparations and is more rare in pregnant than non-pregnant females due to altered immune reaction during pregnancy. Herein, we report two cases of allergic reaction to insulin in gestational diabetes that were successfully managed. One case was a 33-year-old female using isophane-neutral protamine Hagedorn human insulin and insulin lispro. She experienced dyspnea, cough, urticaria and itching sensation at the sites of insulin injection immediately after insulin administration. We discontinued insulin therapy and started oral hypoglycemic agents with metformin and glibenclamide. The other case was a 32-year-old female using insulin lispro and insulin detemer. She experienced pruritus and burning sensation and multiple nodules at the sites of insulin injection. We changed the insulin from insulin lispro to insulin aspart. Assessments including immunoglobulin E (IgE), IgG, eosinophil, insulin antibody level and skin biopsy were performed. In the two cases, the symptoms were resolved after changing the insulin to oral agents or other insulin preparations. We report two cases of allergic reaction to human insulin in gestational diabetes due to its rarity. PMID:26435137

  4. Two Cases of Allergy to Insulin in Gestational Diabetes.

    PubMed

    Kim, Gi Jun; Kim, Shin Bum; Jo, Seong Il; Shin, Jin Kyeong; Kwon, Hee Sun; Jeong, Heekyung; Son, Jang Won; Lee, Seong Su; Kim, Sung Rae; Kim, Byung Kee; Yoo, Soon Jib

    2015-09-01

    Allergic reaction to insulin is uncommon since the introduction of human recombinant insulin preparations and is more rare in pregnant than non-pregnant females due to altered immune reaction during pregnancy. Herein, we report two cases of allergic reaction to insulin in gestational diabetes that were successfully managed. One case was a 33-year-old female using isophane-neutral protamine Hagedorn human insulin and insulin lispro. She experienced dyspnea, cough, urticaria and itching sensation at the sites of insulin injection immediately after insulin administration. We discontinued insulin therapy and started oral hypoglycemic agents with metformin and glibenclamide. The other case was a 32-year-old female using insulin lispro and insulin detemer. She experienced pruritus and burning sensation and multiple nodules at the sites of insulin injection. We changed the insulin from insulin lispro to insulin aspart. Assessments including immunoglobulin E (IgE), IgG, eosinophil, insulin antibody level and skin biopsy were performed. In the two cases, the symptoms were resolved after changing the insulin to oral agents or other insulin preparations. We report two cases of allergic reaction to human insulin in gestational diabetes due to its rarity. PMID:26435137

  5. A filtered tabulated chemistry model for LES of premixed combustion

    SciTech Connect

    Fiorina, B.; Auzillon, P.; Darabiha, N.; Gicquel, O.; Veynante, D.; Vicquelin, R.

    2010-03-15

    A new modeling strategy called F-TACLES (Filtered Tabulated Chemistry for Large Eddy Simulation) is developed to introduce tabulated chemistry methods in Large Eddy Simulation (LES) of turbulent premixed combustion. The objective is to recover the correct laminar flame propagation speed of the filtered flame front when subgrid scale turbulence vanishes as LES should tend toward Direct Numerical Simulation (DNS). The filtered flame structure is mapped using 1-D filtered laminar premixed flames. Closure of the filtered progress variable and the energy balance equations are carefully addressed in a fully compressible formulation. The methodology is first applied to 1-D filtered laminar flames, showing the ability of the model to recover the laminar flame speed and the correct chemical structure when the flame wrinkling is completely resolved. The model is then extended to turbulent combustion regimes by including subgrid scale wrinkling effects in the flame front propagation. Finally, preliminary tests of LES in a 3-D turbulent premixed flame are performed. (author)

  6. Annular fuel and air co-flow premixer

    DOEpatents

    Stevenson, Christian Xavier; Melton, Patrick Benedict; York, William David

    2013-10-15

    Disclosed is a premixer for a combustor including an annular outer shell and an annular inner shell. The inner shell defines an inner flow channel inside of the inner shell and is located to define an outer flow channel between the outer shell and the inner shell. A fuel discharge annulus is located between the outer flow channel and the inner flow channel and is configured to inject a fuel flow into a mixing area in a direction substantially parallel to an outer airflow through the outer flow channel and an inner flow through the inner flow channel. Further disclosed are a combustor including a plurality of premixers and a method of premixing air and fuel in a combustor.

  7. Laminar Premixed and Diffusion Flames (Ground-Based Study)

    NASA Technical Reports Server (NTRS)

    Dai, Z.; El-Leathy, A. M.; Lin, K.-C.; Sunderland, P. B.; Xu, F.; Faeth, G. M.; Urban, D. L. (Technical Monitor); Yuan, Z.-G. (Technical Monitor)

    2000-01-01

    Ground-based studies of soot processes in laminar flames proceeded in two phases, considering laminar premixed flames and laminar diffusion flames, in turn. The test arrangement for laminar premixed flames involved round flat flame burners directed vertically upward at atmospheric pressure. The test arrangement for laminar jet diffusion flames involved a round fuel port directed vertically upward with various hydrocarbon fuels burning at atmospheric pressure in air. In both cases, coflow was used to prevent flame oscillations and measurements were limited to the flame axes. The measurements were sufficient to resolve soot nucleation, growth and oxidation rates, as well as the properties of the environment needed to evaluate mechanisms of these processes. The experimental methods used were also designed to maintain capabilities for experimental methods used in corresponding space-based experiments. This section of the report will be limited to consideration of flame structure for both premixed and diffusion flames.

  8. Quantitative Phosphoproteomics Analysis Reveals a Key Role of Insulin Growth Factor 1 Receptor (IGF1R) Tyrosine Kinase in Human Sperm Capacitation*

    PubMed Central

    Wang, Jing; Qi, Lin; Huang, Shaoping; Zhou, Tao; Guo, Yueshuai; Wang, Gaigai; Guo, Xuejiang; Zhou, Zuomin; Sha, Jiahao

    2015-01-01

    One of the most important changes during sperm capacitation is the enhancement of tyrosine phosphorylation. However, the mechanisms of protein tyrosine phosphorylation during sperm capacitation are not well studied. We used label-free quantitative phosphoproteomics to investigate the overall phosphorylation events during sperm capacitation in humans and identified 231 sites with increased phosphorylation levels. Motif analysis using the NetworKIN algorithm revealed that the activity of tyrosine phosphorylation kinases insulin growth factor 1 receptor (IGF1R)/insulin receptor is significantly enriched among the up-regulated phosphorylation substrates during capacitation. Western blotting further confirmed inhibition of IGF1R with inhibitors GSK1904529A and NVP-AEW541, which inhibited the increase in tyrosine phosphorylation levels during sperm capacitation. Additionally, sperm hyperactivated motility was also inhibited by GSK1904529A and NVP-AEW541 but could be up-regulated by insulin growth factor 1, the ligand of IGF1R. Thus, the IGF1R-mediated tyrosine phosphorylation pathway may play important roles in the regulation of sperm capacitation in humans and could be a target for improvement in sperm functions in infertile men. PMID:25693802

  9. Quantitative phosphoproteomics analysis reveals a key role of insulin growth factor 1 receptor (IGF1R) tyrosine kinase in human sperm capacitation.

    PubMed

    Wang, Jing; Qi, Lin; Huang, Shaoping; Zhou, Tao; Guo, Yueshuai; Wang, Gaigai; Guo, Xuejiang; Zhou, Zuomin; Sha, Jiahao

    2015-04-01

    One of the most important changes during sperm capacitation is the enhancement of tyrosine phosphorylation. However, the mechanisms of protein tyrosine phosphorylation during sperm capacitation are not well studied. We used label-free quantitative phosphoproteomics to investigate the overall phosphorylation events during sperm capacitation in humans and identified 231 sites with increased phosphorylation levels. Motif analysis using the NetworKIN algorithm revealed that the activity of tyrosine phosphorylation kinases insulin growth factor 1 receptor (IGF1R)/insulin receptor is significantly enriched among the up-regulated phosphorylation substrates during capacitation. Western blotting further confirmed inhibition of IGF1R with inhibitors GSK1904529A and NVP-AEW541, which inhibited the increase in tyrosine phosphorylation levels during sperm capacitation. Additionally, sperm hyperactivated motility was also inhibited by GSK1904529A and NVP-AEW541 but could be up-regulated by insulin growth factor 1, the ligand of IGF1R. Thus, the IGF1R-mediated tyrosine phosphorylation pathway may play important roles in the regulation of sperm capacitation in humans and could be a target for improvement in sperm functions in infertile men.

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

    PubMed

    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. PMID:25059983

  11. Identification of novel genes for glucose metabolism based upon expression pattern in human islets and effect on insulin secretion and glycemia.

    PubMed

    Taneera, Jalal; Fadista, Joao; Ahlqvist, Emma; Atac, David; Ottosson-Laakso, Emilia; Wollheim, Claes B; Groop, Leif

    2015-04-01

    Normal glucose homeostasis is characterized by appropriate insulin secretion and low HbA1c. Gene expression signatures associated with these two phenotypes could be essential for islet function and pathophysiology of type 2 diabetes (T2D). Herein, we employed a novel approach to identify candidate genes involved in T2D by correlating islet microarray gene expression data (78 donors) with insulin secretion and HbA1c level. The expression of 649 genes (P < 0.05) was correlated with insulin secretion and HbA1c. Of them, five genes (GLR1A, PPP1R1A, PLCDXD3, FAM105A and ENO2) correlated positively with insulin secretion/negatively with HbA1c and one gene (GNG5) correlated negatively with insulin secretion/positively with HbA1c were followed up. The five positively correlated genes have lower expression levels in diabetic islets, whereas GNG5 expression is higher. Exposure of human islets to high glucose for 24 h resulted in up-regulation of GNG5 and PPP1R1A expression, whereas the expression of ENO2 and GLRA1 was down-regulated. No effect was seen on the expression of FAM105A and PLCXD3. siRNA silencing in INS-1 832/13 cells showed reduction in insulin secretion for PPP1R1A, PLXCD3, ENO2, FAM105A and GNG5 but not GLRA1. Although no SNP in these gene loci passed the genome-wide significance for association with T2D in DIAGRAM+ database, four SNPs influenced gene expression in cis in human islets. In conclusion, we identified and confirmed PPP1R1A, FAM105A, ENO2, PLCDX3 and GNG5 as potential regulators of islet function. We provide a list of candidate genes as a resource for exploring their role in the pathogenesis of T2D. PMID:25489054

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

    PubMed

    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.

  13. Insulin action on the liver in vivo.

    PubMed

    Cherrington, A D; Moore, M C; Sindelar, D K; Edgerton, D S

    2007-11-01

    Insulin has a potent inhibitory effect on hepatic glucose production by direct action at hepatic receptors. The hormone also inhibits glucose production by suppressing both lipolysis in the fat cell and secretion of glucagon by the alpha-cell. Neural sensing of insulin levels appears to participate in control of hepatic glucose production in rodents, but a role for brain insulin sensing has not been documented in dogs or humans. The primary effect of insulin on the liver is its direct action.

  14. Failure of human and mouse leptin to affect insulin, glucagon and somatostatin secretion by the perfused rat pancreas at physiological glucose concentration.

    PubMed

    Leclercq-Meyer, V; Malaisse, W J

    1998-06-25

    In isolated perfused pancreas from normal rats, a rise in d-glucose concentration from 3.3 to 8.3 mM provoked a rapid phasic stimulation of both insulin and somatostatin secretion and rapid fall in glucagon output, these changes being reversed when the concentration of the hexose was brought back to its initial low level. In the presence of 8.3 mM d-glucose, the administration of either human or mouse leptin (10 nM in both cases) for 15 min failed to affect significantly the perfusion pressure and release of the three hormones. It is concluded that leptin does not exert any major immediate and direct effect upon pancreatic insulin, glucagon and somatostatin secretion, at least at the physiological concentration of d-glucose normally found in the plasma of fed rats. PMID:9723892

  15. Pulp-capping with recombinant human insulin-like growth factor I (rhIGF-I) in rat molars.

    PubMed

    Lovschall, H; Fejerskov, O; Flyvbjerg, A

    2001-08-01

    The aim of this study was to explore pulp healing and reparative dentinogenesis following pulp-capping by using recombinant human insulin-like growth factor I (rhIGF-I). Exposures were made through the mesial pulp horn in first upper molars in two-month-old Wistar rats. The pulp was covered with one dose of sterile 4% methylcellulose gel containing either 400 ng rhIGF-I or saline in contralateral controls. The exposure site was closed with sterile Teflon membrane, and the cavity was filled with IRM cement. Additional molars were capped with Dycal as controls. After 3, 7, or 28 days, animals were anesthetized and fixed by intravascular glutaraldehyde perfusion. Molars were decalcified and processed for histological analysis and cut with membrane and residual methacrylate from IRM in situ. Only specimens with acceptable pulp sealing according to blinded microscopy control were included. On day 3, identical inflammatory responses in the upper pulp were observed in molars with rhIGF-I gel or control gel. On day 7, granulation tissue ingrowth had partly replaced inflammatory infiltration in both groups. After 28 days, complete dentin bridging and tubular dentin formation were observed more frequently and closer to the test substance containing rhIGF-I. The reparative dentin response to capping with rhIGF-I was similar to that after the use of Dycal. In conclusion, microscopic control of membrane sealing in situ gives valid information on the more subtle pulp effects of growth factors. The observations suggest that pulp-capping of rat molars by means of rhIGF-I enhances reparative dentinogenesis in comparison with vehicle controls. PMID:12640754

  16. Human Cortical Neural Stem Cells Expressing Insulin-Like Growth Factor-I: A Novel Cellular Therapy for Alzheimer's Disease.

    PubMed

    McGinley, Lisa M; Sims, Erika; Lunn, J Simon; Kashlan, Osama N; Chen, Kevin S; Bruno, Elizabeth S; Pacut, Crystal M; Hazel, Tom; Johe, Karl; Sakowski, Stacey A; Feldman, Eva L

    2016-03-01

    Alzheimer's disease (AD) is the most prevalent age-related neurodegenerative disorder and a leading cause of dementia. Current treatment fails to modify underlying disease pathologies and very little progress has been made to develop effective drug treatments. Cellular therapies impact disease by multiple mechanisms, providing increased efficacy compared with traditional single-target approaches. In amyotrophic lateral sclerosis, we have shown that transplanted spinal neural stem cells (NSCs) integrate into the spinal cord, form synapses with the host, improve inflammation, and reduce disease-associated pathologies. Our current goal is to develop a similar "best in class" cellular therapy for AD. Here, we characterize a novel human cortex-derived NSC line modified to express insulin-like growth factor-I (IGF-I), HK532-IGF-I. Because IGF-I promotes neurogenesis and synaptogenesis in vivo, this enhanced NSC line offers additional environmental enrichment, enhanced neuroprotection, and a multifaceted approach to treating complex AD pathologies. We show that autocrine IGF-I production does not impact the cell secretome or normal cellular functions, including proliferation, migration, or maintenance of progenitor status. However, HK532-IGF-I cells preferentially differentiate into gamma-aminobutyric acid-ergic neurons, a subtype dysregulated in AD; produce increased vascular endothelial growth factor levels; and display an increased neuroprotective capacity in vitro. We also demonstrate that HK532-IGF-I cells survive peri-hippocampal transplantation in a murine AD model and exhibit long-term persistence in targeted brain areas. In conclusion, we believe that harnessing the benefits of cellular and IGF-I therapies together will provide the optimal therapeutic benefit to patients, and our findings support further preclinical development of HK532-IGF-I cells into a disease-modifying intervention for AD.

  17. Acoustic radiation from weakly wrinkled premixed flames

    SciTech Connect

    Lieuwen, Tim; Mohan, Sripathi; Rajaram, Rajesh; Preetham,

    2006-01-01

    This paper describes a theoretical analysis of acoustic radiation from weakly wrinkled (i.e., u'/S{sub L}<1) premixed flames. Specifically, it determines the transfer function relating the spectrum of the acoustic pressure oscillations, P'({omega}), to that of the turbulent velocity fluctuations in the approach flow, U'({omega}). In the weakly wrinkled limit, this transfer function is local in frequency space; i.e., velocity fluctuations at a frequency {omega} distort the flame and generate sound at the same frequency. This transfer function primarily depends upon the flame Strouhal number St (based on mean flow velocity and flame length) and the correlation length, {lambda}, of the flow fluctuations. For cases where the ratio of the correlation length and duct radius {lambda}/a>>1, the acoustic pressure and turbulent velocity power spectra are related by P'({omega})-{omega}{sup 2}U'({omega}) and P'({omega})-U'({omega}) for St<<1 and St>>1, respectively. For cases where {lambda}/a<<1, the transfer functions take the form P'({omega})-{omega}{sup 2}({lambda}/a){sup 2}U'({omega}) and P'({omega})-{omega}{sup 2}({lambda}/a){sup 2}({psi}-{delta}ln({lambda}/a))U'({omega}) for St<<1 and St>>1, respectively, where (PS) and {delta} are constants. The latter result demonstrates that this transfer function does not exhibit a simple power law relationship in the high frequency region of the spectra. The simultaneous dependence of this pressure-velocity transfer function upon the Strouhal number and correlation length suggests a mechanism for the experimentally observed maximum in acoustic spectra and provides some insight into the controversy in the literature over how this peak should scale with the flame Strouhal number.

  18. The lipogenic transcription factor ChREBP dissociates hepatic steatosis from insulin resistance in mice and humans

    PubMed Central

    Benhamed, Fadila; Denechaud, Pierre-Damien; Lemoine, Maud; Robichon, Céline; Moldes, Marthe; Bertrand-Michel, Justine; Ratziu, Vlad; Serfaty, Lawrence; Housset, Chantal; Capeau, Jacqueline; Girard, Jean; Guillou, Hervé; Postic, Catherine

    2012-01-01

    Nonalcoholic fatty liver disease (NAFLD) is associated with all features of the metabolic syndrome. Although deposition of excess triglycerides within liver cells, a hallmark of NAFLD, is associated with a loss of insulin sensitivity, it is not clear which cellular abnormality arises first. We have explored this in mice overexpressing carbohydrate responsive element–binding protein (ChREBP). On a standard diet, mice overexpressing ChREBP remained insulin sensitive, despite increased expression of genes involved in lipogenesis/fatty acid esterification and resultant hepatic steatosis (simple fatty liver). Lipidomic analysis revealed that the steatosis was associated with increased accumulation of monounsaturated fatty acids (MUFAs). In primary cultures of mouse hepatocytes, ChREBP overexpression induced expression of stearoyl-CoA desaturase 1 (Scd1), the enzyme responsible for the conversion of saturated fatty acids (SFAs) into MUFAs. SFA impairment of insulin-responsive Akt phosphorylation was therefore rescued by the elevation of Scd1 levels upon ChREBP overexpression, whereas pharmacological or shRNA-mediated reduction of Scd1 activity decreased the beneficial effect of ChREBP on Akt phosphorylation. Importantly, ChREBP-overexpressing mice fed a high-fat diet showed normal insulin levels and improved insulin signaling and glucose tolerance compared with controls, despite having greater hepatic steatosis. Finally, ChREBP expression in liver biopsies from patients with nonalcoholic steatohepatitis was increased when steatosis was greater than 50% and decreased in the presence of severe insulin resistance. Together, these results demonstrate that increased ChREBP can dissociate hepatic steatosis from insulin resistance, with beneficial effects on both glucose and lipid metabolism. PMID:22546860

  19. Combustion-acoustic stability analysis for premixed gas turbine combustors

    NASA Technical Reports Server (NTRS)

    Darling, Douglas; Radhakrishnan, Krishnan; Oyediran, Ayo; Cowan, Lizabeth

    1995-01-01

    Lean, prevaporized, premixed combustors are susceptible to combustion-acoustic instabilities. A model was developed to predict eigenvalues of axial modes for combustion-acoustic interactions in a premixed combustor. This work extends previous work by including variable area and detailed chemical kinetics mechanisms, using the code LSENS. Thus the acoustic equations could be integrated through the flame zone. Linear perturbations were made of the continuity, momentum, energy, chemical species, and state equations. The qualitative accuracy of our approach was checked by examining its predictions for various unsteady heat release rate models. Perturbations in fuel flow rate are currently being added to the model.

  20. The Placental Variant of Human Growth Hormone Reduces Maternal Insulin Sensitivity in a Dose-Dependent Manner in C57BL/6J Mice.

    PubMed

    Liao, Shutan; Vickers, Mark H; Stanley, Joanna L; Ponnampalam, Anna P; Baker, Philip N; Perry, Jo K

    2016-03-01

    The human placental GH variant (GH-V) is secreted continuously from the syncytiotrophoblast layer of the placenta during pregnancy and is thought to play a key role in the maternal adaptation to pregnancy. Maternal GH-V concentrations are closely related to fetal growth in humans. GH-V has also been proposed as a potential candidate to mediate insulin resistance observed later in pregnancy. To determine the effect of maternal GH-V administration on maternal and fetal growth and metabolic outcomes during pregnancy, we examined the dose-response relationship for GH-V administration in a mouse model of normal pregnancy. Pregnant C57BL/6J mice were randomized to receive vehicle or GH-V (0.25, 1, 2, or 5 mg/kg · d) by osmotic pump from gestational days 12.5 to 18.5. Fetal linear growth was slightly reduced in the 5 mg/kg dose compared with vehicle and the 0.25 mg/kg groups, respectively, whereas placental weight was not affected. GH-V treatment did not affect maternal body weights or food intake. However, treatment with 5 mg/kg · d significantly increased maternal fasting plasma insulin concentrations with impaired insulin sensitivity observed at day 18.5 as assessed by homeostasis model assessment. At 5 mg/kg · d, there was also an increase in maternal hepatic GH receptor/binding protein (Ghr/Ghbp) and IGF binding protein 3 (Igfbp3) mRNA levels, but GH-V did not alter maternal plasma IGF-1 concentrations or hepatic Igf-1 mRNA expression. Our findings suggest that at higher doses, GH-V treatment can cause hyperinsulinemia and is a likely mediator of the insulin resistance associated with late pregnancy.

  1. Insulin Test

    MedlinePlus

    ... people with type 2 diabetes , polycystic ovarian syndrome (PCOS) , prediabetes or heart disease , or metabolic syndrome . A ... resistance), especially in obese individuals and those with PCOS . This test involves an IV-infusion of insulin, ...

  2. Insulin-like growth factors and their binding proteins in human colonocytes: preferential degradation of insulin-like growth factor binding protein 2 in colonic cancers.

    PubMed Central

    Michell, N. P.; Langman, M. J.; Eggo, M. C.

    1997-01-01

    We have compared the expression of insulin-like growth factors (IGFs) and IGF binding proteins (IGFBPs) in ten paired samples of normal and tumour colonic tissue with regard to both mRNA and protein. We have compared sensitivity of these tissues to IGF-I using primary cultures of epithelial cells of colonic mucosa, and we have examined the production of IGFs and IGFBPs by these cells. In the tissues, IGFBP-2 mRNA was expressed in all normal and cancer samples but other IGFBPs showed variable expression. mRNAs for IGF-I were expressed in all normal and cancer tissues but IGF-II mRNA was only detected in cancer tissue (3 out of 10). Immunostaining of sections of normal and cancer tissue was negative for IGF-I and IGF-II; IGFBP-2 was positive in 2 out of 10 cancer tissues and 7 out of 10 normal tissues; IGFBP-3 was positive in 7 out of 10 cancer tissues and 7 out of 10 normal tissues; and IGFBP-4 was positive in 5 out of 10 cancer tissues and 6 out of 10 normal tissues. In the cells in culture, cancer cells showed increased incorporation of [35S]methionine into protein and [3H]thymidine into DNA (P < 0.02) when treated with IGF-I. Western blotting of serum-free conditioned media from cells in culture showed that 8 out of 10 normal and 3 out of 10 cancer cultures produced a 32-kDa immunoreactive IGFBP-2. No IGFBP-3 was secreted by any culture but 24-kDa IGFBP-4 was found in 3 out of 10 normal and 5 out of 10 cancer tissues. Because of the discrepancy between mRNA and protein expression for IGFBP-2, degradation of native IGFBPs was assessed using tissue extracts. Colon cancer extracts were able to degrade exogenous IGFBP-2, IGFBP-3 and IGFBP-4, whereas normal tissue extracts were without effect on IGFBP-2. We conclude that IGFBPs are synthesized and secreted by cells of the colonic mucosa but that proteolysis of secreted IGFBP-2 occurs in colon cancer tissue. This selective degradation may confer a growth advantage. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5

  3. The Transcriptional Response in Human Umbilical Vein Endothelial Cells Exposed to Insulin: A Dynamic Gene Expression Approach

    PubMed Central

    Di Camillo, Barbara; Sanavia, Tiziana; Iori, Elisabetta; Bronte, Vincenzo; Roncaglia, Enrica; Maran, Alberto; Avogaro, Angelo; Toffolo, Gianna; Cobelli, Claudio

    2010-01-01

    Background In diabetes chronic hyperinsulinemia contributes to the instability of the atherosclerotic plaque and stimulates cellular proliferation through the activation of the MAP kinases, which in turn regulate cellular proliferation. However, it is not known whether insulin itself could increase the transcription of specific genes for cellular proliferation in the endothelium. Hence, the characterization of transcriptional modifications in endothelium is an important step for a better understanding of the mechanism of insulin action and the relationship between endothelial cell dysfunction and insulin resistance. Methodology and principal findings The transcriptional response of endothelial cells in the 440 minutes following insulin stimulation was monitored using microarrays and compared to a control condition. About 1700 genes were selected as differentially expressed based on their treated minus control profile, thus allowing the detection of even small but systematic changes in gene expression. Genes were clustered in 7 groups according to their time expression profile and classified into 15 functional categories that can support the biological effects of insulin, based on Gene Ontology enrichment analysis. In terms of endothelial function, the most prominent processes affected were NADH dehydrogenase activity, N-terminal myristoylation domain binding, nitric-oxide synthase regulator activity and growth factor binding. Pathway-based enrichment analysis revealed “Electron Transport Chain” significantly enriched. Results were validated on genes belonging to “Electron Transport Chain” pathway, using quantitative RT-PCR. Conclusions As far as we know, this is the first systematic study in the literature monitoring transcriptional response to insulin in endothelial cells, in a time series microarray experiment. Since chronic hyperinsulinemia contributes to the instability of the atherosclerotic plaque and stimulates cellular proliferation, some of the

  4. An Open-Label Trial of Recombinant Human Insulin-Like Growth Factor-I/Recombinant Human Insulin-Like Growth Factor Binding Protein-3 (rhIGF-1/rhIGFBP-3) in Myotonic Dystrophy Type 1

    PubMed Central

    Heatwole, Chad R.; Eichinger, Katy J.; Friedman, Deborah I.; Hilbert, James E.; Jackson, Carlayne E.; Logigian, Eric L.; Martens, William B.; McDermott, Michael P.; Pandya, Shree K.; Quinn, Christine; Smirnow, Alexis M.; Thornton, Charles A.; Moxley, Richard T.

    2012-01-01

    Objective To evaluate the safety and tolerability of recombinant human insulin-like growth factor-1 (rhIGF-1) complexed with IGF binding protein-3 (rhIGF-1/rhIGFBP-3) in patients with myotonic dystrophy type 1 (DM1). Design Open-label dose-escalation clinical trial. Setting University medical center. Participants Fifteen moderately affected ambulatory participants with genetically-proven DM1. Intervention Participants received escalating dosages of subcutaneous rhIGF-1/rhIGFBP-3 over 24 weeks followed by a 16 week washout period. Outcome Measures Serial assessments of safety, muscle mass, muscle function, and metabolic state were performed. The primary outcome variable was the ability of participants to complete 24 weeks on rhIGF-1/rhIGFBP-3 treatment. Results All participants tolerated rhIGF-1/rhIGFBP-3. There were no significant changes in muscle strength or functional outcomes measures. Lean body muscle mass measured by dual energy x-ray absorptiometry increased by 1.95 kg (p=0.0007) after treatment. Participants also experienced a mean reduction in triglyceride levels of 47 mg/dL (p=0.002), a mean increase in HDL levels of 5.0 mg/dL (p=0.03), a mean reduction in HbA1c of 0.15% (p=0.03), and a mean increase in testosterone level (in men) of 203 ng/dL (p=0.002) while on rhIGF-1/rhIGFBP-3. Mild reactions at the injection site occurred (n=9 participants), as did mild transient hypoglycemia (n=3), lightheadedness (n=2), and transient papilledema (n=1). Conclusions rhIGF-1/rhIGFBP-3 treatment was generally well tolerated in DM1. rhIGF-1/rhIGFBP-3 was associated with increased lean body mass and improvements in metabolism, but not with increased muscle strength or function. Larger randomized controlled trials would be needed to further evaluate the efficacy and safety of this medication in patients with neuromuscular disease. PMID:20837825

  5. Effects of Probiotics and Synbiotics on Obesity, Insulin Resistance Syndrome, Type 2 Diabetes and Non-Alcoholic Fatty Liver Disease: A Review of Human Clinical Trials.

    PubMed

    Sáez-Lara, Maria Jose; Robles-Sanchez, Candido; Ruiz-Ojeda, Francisco Javier; Plaza-Diaz, Julio; Gil, Angel

    2016-01-01

    The use of probiotics and synbiotics in the prevention and treatment of different disorders has dramatically increased over the last decade. Both probiotics and synbiotics are well known ingredients of functional foods and nutraceuticals and may provide beneficial health effects because they can influence the intestinal microbial ecology and immunity. The present study reviews the effects of probiotics and synbiotics on obesity, insulin resistance syndrome (IRS), type 2 diabetes (T2D) and non-alcoholic fatty liver disease (NAFLD) in human randomized clinical trials. Select probiotics and synbiotics provided beneficial effects in patients with obesity, mainly affecting the body mass index and fat mass. Some probiotics had beneficial effects on IRS, decreasing the cell adhesion molecule-1 levels, and the synbiotics decreased the insulin resistance and plasma lipid levels. Moreover, select probiotics improved the carbohydrate metabolism, fasting blood glucose, insulin sensitivity and antioxidant status and also reduced metabolic stress in subjects with T2D. Some probiotics and synbiotics improved the liver and metabolic parameters in patients with NAFLD. The oral intake of probiotics and synbiotics as co-adjuvants for the prevention and treatment of obesity, IRS, T2D and NAFLD is partially supported by the data shown in the present review. However, further studies are required to understand the precise mechanism of how probiotics and synbiotics affect these metabolic disorders. PMID:27304953

  6. Preservation of Cellular Glutathione Status and Mitochondrial Membrane Potential by N-Acetylcysteine and Insulin Sensitizers Prevent Carbonyl Stress-Induced Human Brain Endothelial Cell Apoptosis

    PubMed Central

    Okouchi, Masahiro; Okayama, Naotsuka; Aw, Tak Yee

    2011-01-01

    Oxidative stress-induced cerebral endothelial cell dysfunction is associated with cerebral microvascular complication of primary diabetic encephaolopathy, a neurodegenerative disorder of long-standing diabetes, but the injury mechanisms are poorly understood. This study sought to determine the contribution of carbonyl (methylglyoxal, MG) stress to human brain endothelial cell (IHEC) apoptosis, the relationship to cellular redox status and mitochondrial membrane potential, and the protection by thiol antioxidant and insulin sensitizers. MG exposure induced IHEC apoptosis in association with perturbed cellular glutathione (GSH) redox status, decreased mitochondrial membrane potential (Δψm), activation of caspase-9 and -3, and cleavage of polyADP-ribose polymerase. Insulin sensitizers such as biguanides or AMP-activated protein kinase activator, but not glitazones, afforded cytoprotection through preventing Δψm collapse and activation of caspase-9 that was independent of cellular GSH. Similarly, cyclosporine A prevented Δψm collapse, while N-acetylcysteine (NAC) mediated the recovery of cellular GSH redox balance that secondarily preserved Δψm. Collectively, these results provide mechanistic insights into the role of GSH redox status and mitochondrial potential in carbonyl stress-induced apoptosis of brain endothelial cells, with implications for cerebral microvascular complications associated with primary diabetic encephalopathy. The findings that thiol antioxidant and insulin sensitizers afforded cytoprotection suggest potential therapeutic approaches. PMID:19807652

  7. Sustained Brown Fat Stimulation and Insulin Sensitization by a Humanized Bispecific Antibody Agonist for Fibroblast Growth Factor Receptor 1/βKlotho Complex

    PubMed Central

    Kolumam, Ganesh; Chen, Mark Z.; Tong, Raymond; Zavala-Solorio, Jose; Kates, Lance; van Bruggen, Nicholas; Ross, Jed; Wyatt, Shelby K.; Gandham, Vineela D.; Carano, Richard A.D.; Dunshee, Diana Ronai; Wu, Ai-Luen; Haley, Benjamin; Anderson, Keith; Warming, Søren; Rairdan, Xin Y.; Lewin-Koh, Nicholas; Zhang, Yingnan; Gutierrez, Johnny; Baruch, Amos; Gelzleichter, Thomas R.; Stevens, Dale; Rajan, Sharmila; Bainbridge, Travis W.; Vernes, Jean-Michel; Meng, Y. Gloria; Ziai, James; Soriano, Robert H.; Brauer, Matthew J.; Chen, Yongmei; Stawicki, Scott; Kim, Hok Seon; Comps-Agrar, Laëtitia; Luis, Elizabeth; Spiess, Christoph; Wu, Yan; Ernst, James A.; McGuinness, Owen P.; Peterson, Andrew S.; Sonoda, Junichiro

    2015-01-01

    Dissipating excess calories as heat through therapeutic stimulation of brown adipose tissues (BAT) has been proposed as a potential treatment for obesity-linked disorders. Here, we describe the generation of a humanized effector-less bispecific antibody that activates fibroblast growth factor receptor (FGFR) 1/βKlotho complex, a common receptor for FGF21 and FGF19. Using this molecule, we show that antibody-mediated activation of FGFR1/βKlotho complex in mice induces sustained energy expenditure in BAT, browning of white adipose tissue, weight loss, and improvements in obesity-associated metabolic derangements including insulin resistance, hyperglycemia, dyslipidemia and hepatosteatosis. In mice and cynomolgus monkeys, FGFR1/βKlotho activation increased serum high-molecular-weight adiponectin, which appears to contribute over time by enhancing the amplitude of the metabolic benefits. At the same time, insulin sensitization by FGFR1/βKlotho activation occurs even before the onset of weight loss in a manner that is independent of adiponectin. Together, selective activation of FGFR1/βKlotho complex with a long acting therapeutic antibody represents an attractive approach for the treatment of type 2 diabetes and other obesity-linked disorders through enhanced energy expenditure, insulin sensitization and induction of high-molecular-weight adiponectin. PMID:26288846

  8. Effects of Probiotics and Synbiotics on Obesity, Insulin Resistance Syndrome, Type 2 Diabetes and Non-Alcoholic Fatty Liver Disease: A Review of Human Clinical Trials

    PubMed Central

    Sáez-Lara, Maria Jose; Robles-Sanchez, Candido; Ruiz-Ojeda, Francisco Javier; Plaza-Diaz, Julio; Gil, Angel

    2016-01-01

    The use of probiotics and synbiotics in the prevention and treatment of different disorders has dramatically increased over the last decade. Both probiotics and synbiotics are well known ingredients of functional foods and nutraceuticals and may provide beneficial health effects because they can influence the intestinal microbial ecology and immunity. The present study reviews the effects of probiotics and synbiotics on obesity, insulin resistance syndrome (IRS), type 2 diabetes (T2D) and non-alcoholic fatty liver disease (NAFLD) in human randomized clinical trials. Select probiotics and synbiotics provided beneficial effects in patients with obesity, mainly affecting the body mass index and fat mass. Some probiotics had beneficial effects on IRS, decreasing the cell adhesion molecule-1 levels, and the synbiotics decreased the insulin resistance and plasma lipid levels. Moreover, select probiotics improved the carbohydrate metabolism, fasting blood glucose, insulin sensitivity and antioxidant status and also reduced metabolic stress in subjects with T2D. Some probiotics and synbiotics improved the liver and metabolic parameters in patients with NAFLD. The oral intake of probiotics and synbiotics as co-adjuvants for the prevention and treatment of obesity, IRS, T2D and NAFLD is partially supported by the data shown in the present review. However, further studies are required to understand the precise mechanism of how probiotics and synbiotics affect these metabolic disorders. PMID:27304953

  9. Vorticity transformation in high Karlovitz number premixed flames

    NASA Astrophysics Data System (ADS)

    Bobbitt, Brock; Lapointe, Simon; Blanquart, Guillaume

    2016-01-01

    To better understand the two-way coupling between turbulence and chemistry, the changes in turbulence characteristics through a premixed flame are investigated. Specifically, this study focuses on vorticity, ω, which is characteristic of the smallest length and time scales of turbulence, analyzing its behavior within and across high Karlovitz number (Ka) premixed flames. This is accomplished through a series of direct numerical simulations (DNS) of premixed n-heptane/air flames, modeled with a 35-species finite-rate chemical mechanism, whose conditions span a wide range of unburnt Karlovitz numbers and flame density ratios. The behavior of the terms in the enstrophy, ω2 = ω ṡ ω, transport equation is analyzed, and a scaling is proposed for each term. The resulting normalized enstrophy transport equation involves only a small set of parameters. Specifically, the theoretical analysis and DNS results support that, at high Karlovitz number, enstrophy transport obtains a balance of the viscous dissipation and production/vortex stretching terms. It is shown that, as a result, vorticity scales in the same manner as in homogeneous, isotropic turbulence within and across the flame, namely, scaling with the inverse of the Kolmogorov time scale, τη. As τη is a function only of the viscosity and dissipation rate, this work supports the validity of Kolmogorov's first similarity hypothesis in premixed turbulent flames for sufficiently high Ka numbers. Results are unaffected by the transport model, chemical model, turbulent Reynolds number, and finally the physical configuration.

  10. Effect of increasing doses of recombinant human insulin-like growth factor-I on glucose, lipid, and leucine metabolism in man.

    PubMed

    Turkalj, I; Keller, U; Ninnis, R; Vosmeer, S; Stauffacher, W

    1992-11-01

    The metabolic effects of recombinant human insulin-like growth factor-I (IGF-I) were assessed in five groups of normal male overnight-fasted volunteers receiving infusions of either 0, 5, 7.5, 15, or 30 micrograms/kg.h IGF-I during 8 h, resulting in total plasma IGF-I concentrations 127 +/- 7, 247 +/- 30, 389 +/- 39, 573 +/- 62, 620 +/- 105 ng/ml, respectively. Glucose consumption (euglycemic glucose clamp) increased dose dependently during IGF-I infusion (P < 0.001) up to 6.7 +/- 1.3 mg/kg. min in the 30 micrograms/kg.h group. Plasma triglyceride concentrations decreased with increasing doses of IGF-I (P < 0.03); the fall was 43% in the 30 micrograms/kg.h group. Plasma free fatty acid concentrations decreased during 7.5, 15, and 30 micrograms/kg.h IGF-I by 23%, 34%, and 48%, respectively. IGF-I lowered plasma beta-hydroxybutyrate concentrations in a dose-dependent manner (P < 0.025). Plasma concentrations of leucine and alpha-ketoisocaproate decreased dose dependently (P < 0.001 and P < 0.015). Whole body leucine flux (1-13C-leucine infusion technique) decreased with increasing doses of IGF-I by 41% during 30 micrograms/kg.h, indicating decreased whole body protein breakdown. Leucine oxidation into 13CO2 decreased with increasing doses of IGF-I (P < 0.045) by 57% in the 30 micrograms/kg.h group, suggesting inhibition of irreversible loss of leucine. Plasma C-peptide and insulin concentrations decreased dose dependently (P < 0.005 and P < 0.02), indicating diminished insulin secretion. Thus, acute elevation of plasma IGF-I concentrations in man results in metabolic effects which are qualitatively similar to those described previously of insulin. PMID:1430077

  11. Kinetic modeling of human hepatic glucose metabolism in type 2 diabetes mellitus predicts higher risk of hypoglycemic events in rigorous insulin therapy.

    PubMed

    König, Matthias; Holzhütter, Hermann-Georg

    2012-10-26

    A major problem in the insulin therapy of patients with diabetes type 2 (T2DM) is the increased occurrence of hypoglycemic events which, if left untreated, may cause confusion or fainting and in severe cases seizures, coma, and even death. To elucidate the potential contribution of the liver to hypoglycemia in T2DM we applied a detailed kinetic model of human hepatic glucose metabolism to simulate changes in glycolysis, gluconeogenesis, and glycogen metabolism induced by deviations of the hormones insulin, glucagon, and epinephrine from their normal plasma profiles. Our simulations reveal in line with experimental and clinical data from a multitude of studies in T2DM, (i) significant changes in the relative contribution of glycolysis, gluconeogenesis, and glycogen metabolism to hepatic glucose production and hepatic glucose utilization; (ii) decreased postprandial glycogen storage as well as increased glycogen depletion in overnight fasting and short term fasting; and (iii) a shift of the set point defining the switch between hepatic glucose production and hepatic glucose utilization to elevated plasma glucose levels, respectively, in T2DM relative to normal, healthy subjects. Intriguingly, our model simulations predict a restricted gluconeogenic response of the liver under impaired hormonal signals observed in T2DM, resulting in an increased risk of hypoglycemia. The inability of hepatic glucose metabolism to effectively counterbalance a decline of the blood glucose level becomes even more pronounced in case of tightly controlled insulin treatment. Given this Janus face mode of action of insulin, our model simulations underline the great potential that normalization of the plasma glucagon profile may have for the treatment of T2DM.

  12. Structural organization of the porcine and human genes coding for a leydig cell-specific insulin-like peptide (LEY I-L) and chromosomal localization of the human gene (INSL3)

    SciTech Connect

    Burkhardt E.; Adham, I.M.; Brosig, B.; Gastmann, A.; Engel, W. ); Mattei, M.G. )

    1994-03-01

    Leydig insulin-like protein (LEY I-L) is a member of the insulin-like hormone superfamily. The LEY I-L gene (designated INSL3) is expressed exclusively in prenatal and postnatal Leydig cells. The authors report here the cloning and nucleotide sequence of porcine and human LEY I-L genes including the 5[prime] regions. Both genes consist of two exons and one intron. The organization of the LEY I-L gene is similar to that of insulin and relaxin. The transcription start site in the porcine and human LEY I-L gene is localized 13 and 14 bp upstream of the translation start site, respectively. Alignment of the 5[prime] flanking regions of both genes reveals that the first 107 nucleotides upstream of the transcription start site exhibit an overall sequence similarity of 80%. This conserved region contains a consensus TATAA box, a CAAT-like element (GAAT), and a consensus SP1 sequence (GGGCGG) at equivalent positions in both genes and therefore may play a role in regulation of expression of the LEY I-L gene. The porcine and human genome contains a single copy of the LEY I-L gene. By in situ hybridization, the human gene was assigned to bands p13.2-p12 of the short arm of chromosome 19. 25 refs., 6 figs.

  13. Molecular Basis for the Recognition and Cleavages of IGF-II, TGF-[alpha], and Amylin by Human Insulin-Degrading Enzyme

    SciTech Connect

    Guo, Qing; Manolopoulou, Marika; Bian, Yao; Schilling, Alexander B.; Tang, Wei-Jen

    2010-02-11

    Insulin-degrading enzyme (IDE) is involved in the clearance of many bioactive peptide substrates, including insulin and amyloid-{beta}, peptides vital to the development of diabetes and Alzheimer's disease, respectively. IDE can also rapidly degrade hormones that are held together by intramolecular disulfide bond(s) without their reduction. Furthermore, IDE exhibits a remarkable ability to preferentially degrade structurally similar peptides such as the selective degradation of insulin-like growth factor (IGF)-II and transforming growth factor-{alpha} (TGF-{alpha}) over IGF-I and epidermal growth factor, respectively. Here, we used high-accuracy mass spectrometry to identify the cleavage sites of human IGF-II, TGF-{alpha}, amylin, reduced amylin, and amyloid-{beta} by human IDE. We also determined the structures of human IDE-IGF-II and IDE-TGF-{alpha} at 2.3 {angstrom} and IDE-amylin at 2.9 {angstrom}. We found that IDE cleaves its substrates at multiple sites in a biased stochastic manner. Furthermore, the presence of a disulfide bond in amylin allows IDE to cut at an additional site in the middle of the peptide (amino acids 18-19). Our amylin-bound IDE structure offers insight into how the structural constraint from a disulfide bond in amylin can alter IDE cleavage sites. Together with NMR structures of amylin and the IGF and epidermal growth factor families, our work also reveals the structural basis of how the high dipole moment of substrates complements the charge distribution of the IDE catalytic chamber for the substrate selectivity. In addition, we show how the ability of substrates to properly anchor their N-terminus to the exosite of IDE and undergo a conformational switch upon binding to the catalytic chamber of IDE can also contribute to the selective degradation of structurally related growth factors.

  14. Molecular basis for the recognition and cleavages of IGF-II, TGF-α, and amylin by human insulin degrading enzyme

    PubMed Central

    Guo, Qing; Manolopoulou, Marika; Bian, Yao; Schilling, Alexander B.; Tang, Wei-Jen

    2009-01-01

    Insulin-degrading enzyme (IDE) is involved in the clearance of many bioactive peptide substrates, including insulin and amyloid-β (Aβ), peptides vital to the development of diabetes and Alzheimer's disease, respectively. IDE can also rapidly degrade hormones that are held together by intra-molecular disulfide bond(s) without their reduction. Furthermore, IDE exhibits a remarkable ability to preferentially degrade structurally similar peptides such as the selective degradation of insulin-growth factor-II (IGF-II) and transforming growth factor-α (TGF-α) over IGF-I and epidermal growth factor (EGF), respectively. Here, we used high accuracy mass spectrometry to identify the cleavage sites of human IGF-II, TGF-α, amylin, reduced amylin, and Aβ by human IDE. We also determined the structures of human IDE-IGF-II and IDE-TGF-α at 2.3 Å and IDE-amylin at 2.9 Å. We found that IDE cleaves its substrates at multiple sites in a biased stochastic manner. Furthermore, the presence of a disulfide bond in amylin allows IDE to cut at an additional site in the middle of the peptide (aa 18-19). Our amylin-bound IDE structure offers insight into how the structural constraint from a disulfide bond in amylin can alter IDE cleavage sites. Together with NMR structures of amylin and the IGF and EGF families, our work also reveals the structural basis of how the high dipole moment of substrates complements the charge distribution of the IDE catalytic chamber for the substrate selectivity. In addition, we show how the ability of substrates to properly anchor their N-terminus to the exosite of IDE and undergo a conformational switch upon binding to the catalytic chamber of IDE can also contribute to the selective degradation of structurally related growth factors. PMID:19896952

  15. EJE PRIZE 2015: How does insulin resistance arise, and how does it cause disease? Human genetic lessons.

    PubMed

    Semple, R K

    2016-05-01

    Insulin orchestrates physiological responses to ingested nutrients; however, although it elicits widely ramifying metabolic and trophic responses from diverse tissues, 'insulin resistance (IR)', a pandemic metabolic derangement commonly associated with obesity, is usually defined solely by blunting of insulin's hypoglycaemic effect. Recent study of monogenic forms of IR has established that biochemical subphenotypes of IR exist, clustering into those caused by primary disorders of adipose tissue and those caused by primary defects in proximal insulin signalling. IR is often first recognised by virtue of its associated disorders including type 2 diabetes, dyslipidaemia (DL), fatty liver and polycystic ovary syndrome (PCOS). Although these clinically observed associations are confirmed by cross-sectional and longitudinal population-based studies, causal relationships among these phenomena have been more difficult to establish. Single gene IR is important to recognise in order to optimise clinical management and also permits testing of causal relationships among components of the IR syndrome using the principle of Mendelian randomisation. Thus, where a precisely defined genetic defect is identified that directly produces one component of the syndrome, then phenomena that are causally linked to that component should be seen. Where this is not the case, then a simple causal link is refuted. This article summarises known forms of monogenic severe IR and considers the lessons to be learned about the pathogenic mechanisms both upstream from common IR and those downstream linking it to disorders such as DL, fatty liver, PCOS and cancer. PMID:26865583

  16. A Comparison of the Anorexic Effects of Chicken, Porcine, Human and Bovine Insulin on the Central Nervous System of Chicks

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The aim of the present study was to determine if some naturally-occurring substitutions of amino acid residues of insulin could act differentially within the central nervous system (CNS) of neonatal chicks to control ingestive behavior. Intracerebroventricular (ICV) administration of chicken insuli...

  17. EJE PRIZE 2015: How does insulin resistance arise, and how does it cause disease? Human genetic lessons.

    PubMed

    Semple, R K

    2016-05-01

    Insulin orchestrates physiological responses to ingested nutrients; however, although it elicits widely ramifying metabolic and trophic responses from diverse tissues, 'insulin resistance (IR)', a pandemic metabolic derangement commonly associated with obesity, is usually defined solely by blunting of insulin's hypoglycaemic effect. Recent study of monogenic forms of IR has established that biochemical subphenotypes of IR exist, clustering into those caused by primary disorders of adipose tissue and those caused by primary defects in proximal insulin signalling. IR is often first recognised by virtue of its associated disorders including type 2 diabetes, dyslipidaemia (DL), fatty liver and polycystic ovary syndrome (PCOS). Although these clinically observed associations are confirmed by cross-sectional and longitudinal population-based studies, causal relationships among these phenomena have been more difficult to establish. Single gene IR is important to recognise in order to optimise clinical management and also permits testing of causal relationships among components of the IR syndrome using the principle of Mendelian randomisation. Thus, where a precisely defined genetic defect is identified that directly produces one component of the syndrome, then phenomena that are causally linked to that component should be seen. Where this is not the case, then a simple causal link is refuted. This article summarises known forms of monogenic severe IR and considers the lessons to be learned about the pathogenic mechanisms both upstream from common IR and those downstream linking it to disorders such as DL, fatty liver, PCOS and cancer.

  18. Insulin-responsiveness of tumor growth.

    PubMed

    Chantelau, Ernst

    2009-05-01

    In October 2008, the 2nd International Insulin & Cancer Workshop convened roughly 30 researchers from eight countries in Düsseldorf/Germany. At this meeting, which was industry-independent like the preceding one in 2007, the following issues were discussed a) association between certain cancers and endogenous insulin production in humans, b) growth-promoting effects of insulin in animal experiments, c) mitogenic and anti-apoptotic activity of pharmaceutic insulin and insulin analogues in in vitro experiments