Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0011849 (diabetes)
 277,896 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Glucose-6-phosphatase (Glu-6-Pase) catalyzes the terminal step of gluconeogenesis, the conversion of glucose 6-phosphate (Glu-6-P) to free glucose. This enzyme activity is thought to be conferred by a complex of proteins residing in the endoplasmic reticulum (ER), including a Glu-6-P translocase that transports Glu-6-P into the lumen of the ER, a phosphohydrolase catalytic subunit residing in the lumen, and putative glucose and inorganic phosphate transporters that allow exit of the products of the reaction. In this study, we have investigated the effect of adenovirus-mediated overexpression of the Glu-6-Pase catalytic subunit on glucose metabolism and insulin secretion, using a well differentiated insulinoma cell line, INS-1. We found that the overexpressed Glu-6-Pase catalytic subunit was normally glycosylated, correctly sorted to the ER, and caused a 10-fold increase in Glu-6-Pase enzymatic activity in in vitro assays. Consistent with these findings, a 4.2-fold increase in 3H2O incorporation into glucose was observed in INS-1 cells treated with the recombinant adenovirus containing the Glu-6-Pase catalytic subunit cDNA (AdCMV-Glu-6-Pase). 3-[3H]Glucose usage was decreased by 32% in AdCMV-Glu-6-Pase-treated cells relative to controls, resulting in a proportional 30% decrease in glucose-stimulated insulin secretion. Our findings indicate that overexpression of the Glu-6-Pase catalytic subunit significantly impacts glucose metabolism and insulin secretion in islet beta-cells. However, INS-1 cells treated with AdCMV-Glu-6-Pase do not exhibit the severe alterations of beta-cell function and metabolism associated with islets from rodent models of obesity and non-insulin-dependent diabetes mellitus, suggesting the involvement of genes in addition to the catalytic subunit of Glu-6-Pase in the etiology of such beta-cell dysfunction.
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PMID:Adenovirus-mediated expression of the catalytic subunit of glucose-6-phosphatase in INS-1 cells. Effects on glucose cycling, glucose usage, and insulin secretion. 931 82

Interferon-gamma is among the cytokines which have been implicated as effector molecules of beta-cell destruction in autoimmune diabetes. Its mechanism of action is, however, largely unknown. In the present study rat pancreatic beta-cells, INS-1, were incubated with rat interferon-gamma (rIRN-gamma) for 24 h. rIFN-gamma at 1-1000 U/ml caused a dose-dependent inhibition of insulin release and cell metabolism with maximal inhibition being observed at 100 U/ml (insulin release: 51.2%, cell metabolism: 43.3% of control, respectively). In addition, 100 U/ml rIFN-gamma induced a 4- and 8.3-fold increase in apoptotic cell death after 24 and 48 h of incubation, respectively. These effects were not mediated by nitric oxide (NO), since IFN-gamma failed to induce nitric oxide synthase and NO production. Similarly, beta-cell dysfunction and death were not prevented by coincubation of the INS-1 cells with the poly(ADP-ribose) polymerase inhibitors benzamide, 3-aminobenzamide, and 4-aminobenzamide, the oxygen free radical scavenger Trolox, and the antioxidant N-acetylcysteine, indicating that NO, poly(ADP-ribose) polymerase, and oxygen free radicals are not involved in IFN-gamma induced beta-cell dysfunction and death.
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PMID:Interferon-gamma inhibits insulin release and induces cell death in the pancreatic beta-cell line INS-1 independently of nitric oxide production. 941 85

Mitochondrial dysfunction due to alterations in the mitochondrial genome (mtDNA) has recently attracted much attention, with the finding that mutations in the mitochondrially encoded proteins perturb cell function. Several disorders have been linked to such genetic changes, including a specific diabetic phenotype. Using ethidium bromide (EtBr) that intercalates into mtDNA, we have effectively eliminated functions under the control of mtDNA from the highly differentiated INS-1 insulin-secreting cell line. We have investigated the consequences on insulin secretion, mitochondrial enzyme activity, organelle structure, and membrane polarization in such cells (INS-1 rho0). Under these conditions, the mitochondrial membrane potential fails to hyperpolarize in response to either glucose or methylsuccinate. In agreement with this finding, the morphology of the mitochondria is altered in the presence of EtBr, sharing similarities with mitochondria in which the membrane potential has been collapsed with the protonophore carbonyl cyanide p-trifluoromethoxyphenylhydrazone (FCCP). In addition, there is no effect of either nutrient secretagogue at the level of the plasma membrane potential, although the effect of the depolarizing agent KCl on membrane depolarization is completely preserved. Similarly, glucose and methylsuccinate fail to increase insulin secretion, whereas KCl is still effective. To test further the effects of mtDNA depletion on exocytosis, we permeabilized INS-1 cells with Staphylococcus aureus alpha-toxin, which forms small holes in the plasma membrane. In contrast to control cells, mitochondrial substrates were incapable of stimulating insulin secretion in mtDNA-deficient cells, emphasizing that the defect in secretion lies at the level of mitochondrial function rather than in the exocytotic process. The results indicate the paramount importance of the mitochondria in the downstream effects elicited by exposure to elevated concentrations of nutrient secretagogue.
Diabetes 1998 Mar
PMID:Effects of depletion of mitochondrial DNA in metabolism secretion coupling in INS-1 cells. 951 42

The fact that insulin-producing islet beta-cells are susceptible to the cytotoxic effects of inflammatory cytokines represents a potential hinderance to the use of such cells for transplantation therapy of insulin-dependent diabetes mellitus (IDDM). In the current study, we show that IL-1beta induces destruction of INS-1 insulinoma cells, while having no effect on a second insulinoma cell line RIN1046-38 and its engineered derivatives, and that this difference is correlated with a higher level of expression of manganese superoxide dismutase (MnSOD) in the latter cells. Stable overexpression of MnSOD in INS-1 cells provides complete protection against IL-1beta-mediated cytotoxicity, and also results in markedly reduced killing when such cells are exposed to conditioned media from activated human or rat PBMC. Further, overexpression of MnSOD in either RIN- or INS-1-derived lines results in a sharp reduction in IL-1beta-induced nitric oxide (NO) production, a finding that correlates with reduced levels of the inducible form of nitric oxide synthase (iNOS). Treatment of INS-1 cells with L-NMMA, an inhibitor of iNOS, provides the same degree of protection against IL-1beta or supernatants from LPS-activated rat PBMC as MnSOD overexpression, supporting the idea that MnSOD protects INS-1 cells by interfering with the normal IL-1beta-mediated increase in iNOS. Because NO and its derivatives have been implicated as critical mediators of beta-cell destruction in IDDM, we conclude that well regulated insulinoma cell lines engineered for MnSOD overexpression may be an attractive alternative to isolated islets as vehicles for insulin replacement in autoimmune diabetes.
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PMID:Stable expression of manganese superoxide dismutase (MnSOD) in insulinoma cells prevents IL-1beta- induced cytotoxicity and reduces nitric oxide production. 957 43

Chronic exposure of pancreatic beta-cells to high glucose has pleiotropic action on beta-cell function. In particular, it induces key glycolytic genes, promotes glycogen deposition, and causes beta-cell proliferation and altered insulin secretion characterized by sensitization to low glucose. Postglycolytic events, in particular, anaplerosis and lipid signaling, are thought to be implicated in beta-cell activation by glucose. To understand the biochemical nature of the beta-cell adaptive process to hyperglycemia, we studied the regulation by glucose of lipogenic genes in the beta-cell line INS-1. A 3-day exposure of cells to elevated glucose (5-25 mmol/l) increased the enzymatic activities of fatty acid synthase 3-fold, acetyl-CoA carboxylase 30-fold, and malic enzyme 1.3-fold. Pyruvate carboxylase and citrate lyase expression remained constant. Similar observations were made at the protein and mRNA levels except for malic enzyme mRNA, which did not vary. Metabolic gene expression changes were associated with chronically elevated levels of citrate, malate, malonyl-CoA, and conversion of glucose carbon into lipids, even in cells that were subsequently exposed to low glucose. Similarly, fatty acid oxidation was suppressed and phospholipid and triglyceride synthesis was enhanced independently of the external glucose concentration in cells preexposed to high glucose. The results suggest that a coordinated induction of glycolytic and lipogenic genes in conjunction with glycogen and triglyceride deposition, as well as increased anaplerosis and altered lipid partitioning, contribute to the adaptive process to hyperglycemia and glucose sensitization of the beta-cell.
Diabetes 1998 Jul
PMID:Long-term exposure of beta-INS cells to high glucose concentrations increases anaplerosis, lipogenesis, and lipogenic gene expression. 964 32

Hepatocyte nuclear factors (HNFs) are a heterogeneous class of evolutionarily conserved transcription factors that are required for cellular differentiation and metabolism. Mutations in HNF-1alphaand HNF-4alpha genes impair insulin secretion and cause type 2 diabetes. Regulation of HNF-4/HNF-1 expression by HNF-3alpha and HNF-3beta was studied in embryoid bodies in which one or both HNF-3alpha or HNF-3beta alleles were inactivated. HNF-3beta positively regulated the expression of HNF-4alpha/HNF-1alpha and their downstream targets, implicating a role in diabetes. HNF-3beta was also necessary for expression of HNF-3alpha. In contrast, HNF-3alpha acts as a negative regulator of HNF-4alpha/HNF-1alpha demonstrating that HNF-3alpha and HNF-3beta have antagonistic transcriptional regulatory functions in vivo. HNF-3alpha does not appear to act as a classic biochemical repressor but rather exerts its negative effect by competing for HNF-3 binding sites with the more efficient activator HNF-3beta. In addition, the HNF-3alpha/HNF-3beta ratio is modulated by the presence of insulin, providing evidence that the HNF network may have important roles in mediating the action of insulin.
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PMID:Regulation of a transcription factor network required for differentiation and metabolism. 968 61

Tyrosine kinases are involved in various intracellular signalling cascades of different cells: Genistein has been shown to inhibit tyrosine kinase in INS-1 cells, an insulin-secreting cell line (Verspohl et al., 1995). It is, however, not established how specific and selective the tyrosine kinase inhibitors and their controls are. The tyrosine kinase inhibitors genistein and tyrphostin 25 increased insulin release, but not their negative controls with isoflavonoid structure (daidzein and genistin). In addition to this short-term effect a long-term effect was investigated. Genistein (100 microM) time-dependently increased insulin mRNA levels in INS-1 cells. On the other hand the tyrosine kinase inhibitors tyrphostin 25 and lavendustin A (both at 100 microM), which are structurally different from genistein, failed to increase the insulin mRNA whereas daidzein and genistin, normally used as negative controls, increased insulin mRNA as potently as genistein did. However, an examination of the incubation medium revealed that genistin was degraded to genistein by about 50% probably by nonspecific glucosidases first seen after 2 hours of incubation; genistin, therefore, does not appear to be a proper control though often used in this way. In conclusion, the suitability of the compounds used in recent studies is doubtful since other effects than the inhibition of tyrosine kinases are possible. Whereas the involvement of tyrosine kinase in a short-term effect (insulin release) is obvious and clearly substantiated by using the established pharmacological tools (negative controls), the involvement of tyrosine kinases in long-term effects is not that clear; only compounds with isoflavonoid structure are effective independent whether they normally are thought to be inhibitors or negative controls. One has to be cautious in using the above-mentioned compounds in an uncritical way.
Exp Clin Endocrinol Diabetes 1998
PMID:The specificity of tyrosine kinase inhibitors: their effect on insulin release (short-term effect) and insulin mRNA (long-term effect) in an insulin-secreting cell line (INS-1). 979 61

A role of diadenosine polyphosphates as second messengers was suggested for insulin-secreting cells. It has not yet been investigated whether specific receptors for these compounds exist and how these extracellular compounds and their degradation products may contribute to insulin release. Specific saturable binding sites for diadenosine polyphosphates exist in INS-1 cells and rat pancreatic islets. In INS-1 cells, the rank order of diadenosine polyphosphates displacing [3H]Ap4A from binding sites was Ap4A = Ap5A >Ap3A = Ap6A. Binding was specific, since suramin was not able to displace the binding; adenosine, ATP, UTP, alpha,beta-methylene ATP, beta,gamma-methylene ATP, ADP-betaS, 2-methylthio ATP, and pyridoxalphosphate-6-azophenyl-2',4'-disulfonic acid (PPADS) were able to displace [3H]Ap4A from its binding sites. Insulin release was investigated in INS-1 cells. Perifusion experiments showed an increase in insulin release stimulated by the diadenosine polyphosphates in the presence of 8.3 mmol/l glucose; in static incubations (90 min), however, insulin release was inhibited dose dependently by the four diadenosine polyphosphates. This discrepancy might be due to the instability of the compounds. [3H]Ap4A was degraded in the extracellular medium to mainly adenosine and low concentrations of ATP, ADP, AMP, and inosine (half-maximal degradation after 25 min). The insulin stimulatory effect is due to the original compounds (acute perifusion experiments), and the insulin inhibitory effect (static incubation experiments) is due to the production of inhibitory compounds, such as adenosine, in the medium. Small amounts of intact [3H]Ap4A, but mainly [3H]ATP, accumulated in the cells within 20 min. The uptake of labeled compounds is dependent on an intact metabolism and intact receptor internalization. This data indicates that 1) specific bindings sites for diadenosine polyphosphates exist in INS-1 cells and rat pancreatic islets mediating insulin release; 2) the receptors involved in INS-1 cells may be diadenosine polyphosphate receptors, albeit others, such as P2X-receptors, cannot be ruled out; and 3) diadenosine polyphosphates, and mainly their degradation products in the extracellular space, are to a high degree accumulated within cells with unknown function. Thus, diadenosine polyphosphates are worth being investigated more closely in physiological and pathophysiological terms.
Diabetes 1998 Nov
PMID:Diadenosine polyphosphates in insulin-secreting cells: interaction with specific receptors and degradation. 979 42

Mutations in the hepatocyte nuclear factor-1alpha (HNF-1alpha) have been linked to subtype 3 of maturity-onset diabetes of the young (MODY3), which is characterized by a primary defect in insulin secretion. The role of HNF-1alpha in the regulation of pancreatic beta-cell function was investigated. Gene manipulation allowed graded overexpression of HNF-1alpha and controlled dominant-negative suppression of HNF-1alpha function in insulinoma INS-1 cells. We show that HNF-1alpha is essential for insulin gene transcription, as demonstrated by a pronounced decrease in insulin mRNA expression and in insulin promoter activity under dominant-negative conditions. The expression of genes involved in glucose transport and metabolism including glucose transporter-2 and L-type pyruvate kinase is also regulated by HNF-1alpha. Loss of HNF-1alpha function leads to severe defects in insulin secretory responses to glucose and leucine, resulting from impaired glucose utilization and mitochondrial oxidation. The nutrient-evoked ATP production and subsequent changes in plasma membrane potential and intracellular Ca2+ were diminished by suppression of HNF-1alpha function. These results suggest that HNF-1alpha function is essential for maintaining insulin storage and nutrient-evoked release. The defective mitochondrial oxidation of metabolic substrates causes impaired insulin secretion, indicating a molecular basis for the diabetic phenotype of MODY3 patients.
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PMID:Dominant-negative suppression of HNF-1alpha function results in defective insulin gene transcription and impaired metabolism-secretion coupling in a pancreatic beta-cell line. 982 13

Triglycerides in the beta-cell may be important for stimulus-secretion coupling, through provision of a lipid-derived signal, and for pathogenetic events in NIDDM, where lipids may adversely affect beta-cell function. In adipose tissues, hormone-sensitive lipase (HSL) is rate-limiting in triglyceride hydrolysis. Here, we investigated whether this enzyme is also expressed and active in beta-cells. Northern blot analysis and reverse transcription-polymerase chain reaction demonstrated that HSL is expressed in rat islets and in the clonal beta-cell lines INS-1, RINm5F, and HIT-T15. Western blot analysis identified HSL in mouse and rat islets and the clonal beta-cells. In mouse and rat, immunocytochemistry showed a predominant occurrence of HSL in beta-cells, with a presumed cytoplasmic localization. Lipase activity in homogenates of the rodent islets and clonal beta-cells constituted 2.1 +/- 0.6% of that in adipocytes; this activity was immunoinhibited by use of antibodies to HSL. The established HSL expression and activity in beta-cells offer a mechanism whereby lipids are mobilized from intracellular stores. Because HSL in adipocytes is activated by cAMP-dependent protein kinase (PKA), PKA-regulated triglyceride hydrolysis in beta-cells may participate in the regulation of insulin secretion, possibly by providing a lipid-derived signal, e.g., long-chain acyl-CoA and diacylglycerol.
Diabetes 1999 Jan
PMID:Hormone-sensitive lipase, the rate-limiting enzyme in triglyceride hydrolysis, is expressed and active in beta-cells. 989 50


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