| 31. |
- Banfi, C, et al.
(författare)
-
Transcriptional regulation of plasminogen activator inhibitor type 1 gene by insulin: insights into the signaling pathway
- 2001
-
Ingår i: Diabetes. - : American Diabetes Association. - 0012-1797 .- 1939-327X. ; 50:7, s. 1522-1530
-
Tidskriftsartikel (refereegranskat)abstract
- Impairment of the fibrinolytic system, caused primarily by increases in the plasma levels of plasminogen activator inhibitor (PAI) type 1, are frequently found in diabetes and the insulin-resistance syndrome. Among the factors responsible for the increases of PAI-1, insulin has recently attracted attention. In this study, we analyzed the effects of insulin on PAI-1 biosynthesis in HepG2 cells, paying particular attention to the signaling network evoked by this hormone. Experiments performed in CHO cells overexpressing the insulin receptor indicate that insulin increases PAI-1 gene transcription through interaction with its receptor. By using inhibitors of the different signaling pathways evoked by insulin-receptor binding, it has been shown that the biosynthesis of PAI-1 is due to phosphatidylinositol (PI) 3-kinase activation, followed by protein kinase C and ultimately by mitogen-activated protein (MAP) kinase activation and extracellular signal–regulated kinase 2 phosphorylation. We also showed that this pathway is Ras-independent. Transfection of HepG2 cells with several truncations of the PAI-1 promoter coupled to a CAT gene allowed us to recognize two major response elements located in the regions between −804 and −708 and between −211 and −54. Electrophoretic mobility shift assay identified three binding sites for insulin-induced factors, all colocalized with putative Sp1 binding sites. Using supershifting antibodies, the binding of Sp1 could only be confirmed at the binding site located just upstream from the transcription start site of the PAI-1 promoter. A construct comprising four tandem repeat copies of the −93/−62 region of the PAI-1 promoter linked to CAT was transcriptionally activated in HepG2 cells by insulin. These results outline the central role of MAP kinase activation in the regulation of PAI-1 induced by insulin.
|
|
| 32. |
- Barbarroja, Nuria, et al.
(författare)
-
Increased dihydroceramide/ceramide ratio mediated by defective expression of degs1 impairs adipocyte differentiation and function
- 2015
-
Ingår i: Diabetes. - : American Diabetes Association. - 0012-1797 .- 1939-327X. ; 64:4, s. 1180-1192
-
Tidskriftsartikel (refereegranskat)abstract
- Adipose tissue dysfunction is an important determinant of obesity-associated, lipid-induced metabolic complications. Ceramides are well-known mediators of lipid-induced insulin resistance in peripheral organs such as muscle. DEGS1 is the desaturase catalyzing the last step in the main ceramide biosynthetic pathway. Functional suppression of DEGS1 activity results in substantial changes in ceramide species likely to affect fundamental biological functions such as oxidative stress, cell survival, and proliferation. Here, we show that degs1 expression is specifically decreased in the adipose tissue of obese patients and murine models of genetic and nutritional obesity. Moreover, loss-of-function experiments using pharmacological or genetic ablation of DEGS1 in preadipocytes prevented adipogenesis and decreased lipid accumulation. This was associated with elevated oxidative stress, cellular death, and blockage of the cell cycle. These effects were coupled with increased dihydroceramide content. Finally, we validated in vivo that pharmacological inhibition of DEGS1 impairs adipocyte differentiation. These data identify DEGS1 as a new potential target to restore adipose tissue function and prevent obesity-associated metabolic disturbances.
|
|
| 33. |
- Barg, Sebastian, et al.
(författare)
-
A Subset of 50 Secretory Granules in Close Contact With L-Type Ca(2+) Channels Accounts for First-Phase Insulin Secretion in Mouse beta-Cells.
- 2002
-
Ingår i: Diabetes. - 1939-327X .- 0012-1797. ; 51 Suppl 1, s. 74-82
-
Tidskriftsartikel (refereegranskat)abstract
- Capacitance measurements were applied to mouse pancreatic beta-cells to elucidate the cellular mechanisms underlying biphasic insulin secretion. We report here that only <50 of the beta-cell's >10,000 granules are immediately available for release. The releasable granules tightly associate with the voltage-gated alpha(1C) Ca(2+) channels, and it is proposed that the release of these granules accounts for first-phase insulin secretion. Subsequent replenishment of the releasable pool by priming of previously nonreleasable granules is required for second-phase insulin secretion. The latter reaction depends on intragranular acidification due to the concerted action of granular bafilomycin-sensitive v-type H(+)-ATPase and 4,4-diisothiocyanostilbene-2,2-disulfonate--blockable ClC-3 Cl(-) channels. Lowering the cytoplasmic ATP/ADP ratio prevents granule acidification, granule priming, and refilling of the releasable pool. The latter finding provides an explanation to the transient nature of insulin secretion elicited by, for example, high extracellular K(+) in the absence of metabolizable fuels.
|
|
| 34. |
|
|
| 35. |
- Barg, Sebastian, et al.
(författare)
-
Tight coupling between electrical activity and exocytosis in mouse glucagon-secreting alpha-cells
- 2000
-
Ingår i: Diabetes. - 1939-327X .- 0012-1797. ; 49:9, s. 1500-1510
-
Tidskriftsartikel (refereegranskat)abstract
- alpha-Cells were identified in preparations of dispersed mouse islets by immunofluorescence microscopy. A high fraction of alpha-cells correlated with a small cell size measured as the average cell diameter (10 microm) and whole-cell capacitance (<4 pF). The alpha-cells generated action potentials at a low frequency (1 Hz) in the absence of glucose. These action potentials were reversibly inhibited by elevation of the glucose concentration to 20 mmol/l. The action potentials originated from a membrane potential more negative than -50 mV, had a maximal upstroke velocity of 5 V/s, and peaked at +1 mV. Voltage-clamp experiments revealed the ionic conductances underlying the generation of action potentials. alpha-Cells are equipped with a delayed tetraethyl-ammonium-blockable outward current (activating at voltages above -20 mV), a large tetrodotoxin-sensitive Na+ current (above -30 mV; peak current 200 pA at +10 mV), and a small Ca2+ current (above -50 mV; peak current 30 pA at +10 mV). The latter flowed through omega-conotoxin GVIA (25%)- and nifedipine-sensitive (50%) Ca(2+)-channels. Mouse alpha-cells contained, on average, 7,300 granules, which undergo Ca(2+)-induced exocytosis when the alpha-cell is depolarized. Three functional subsets of granules were identified, and the size of the immediately releasable pool was estimated as 80 granules, or 1% of the total granule number. The maximal rate of exocytosis (1.5 pF/s) was observed 21 ms after the onset of the voltage-clamp depolarization, which is precisely the duration of Ca(2+)-influx during an action potential. Our results suggest that the secretory machinery of the alpha-cell is optimized for maximal efficiency in the use of Ca2+ for exocytosis.
|
|
| 36. |
- Barker, Adam, et al.
(författare)
-
Association of genetic loci with glucose levels in childhood and adolescence a meta-analysis of over 6,000 children
- 2011
-
Ingår i: Diabetes. - : American Diabetes Association. - 0012-1797 .- 1939-327X. ; 60:6, s. 1805-1812
-
Tidskriftsartikel (refereegranskat)abstract
- OBJECTIVE-To investigate whether associations of common genetic variants recently identified for fasting glucose or insulin levels in nondiabetic adults are detectable in healthy children and adolescents. RESEARCH DESIGN AND METHODS-A total of 16 single nucleotide polymorphisms (SNPs) associated with fasting glucose were genotyped in six studies of children and adolescents of European origin, including over 6,000 boys and girls aged 9-16 years. We performed meta-analyses to test associations of individual SNPs and a weighted risk score of the 16 loci with fasting glucose. RESULTS-Nine loci were associated with glucose levels in healthy children and adolescents, with four of these associations reported in previous studies and five reported here for the first time (GLIS3, PROX1, SLC2A2, ADCY5, and CRY2). Effect sizes were similar to those in adults, suggesting age-independent effects of these fasting glucose loci. Children and adolescents carrying glucose-raising alleles of G6PC2, MTNR1B, GCK, and GLIS3 also showed reduced p-cell function, as indicated by homeostasis model assessment of beta-cell function. Analysis using a weighted risk score showed an increase [beta (95% CI)] in fasting glucose level of 0.026 mrnol/L (0.021-0.031) for each unit increase in the score. CONCLUSIONS-Novel fasting glucose loci identified in genome-wide association studies of adults are associated with altered fasting glucose levels in healthy children and adolescents with effect sizes comparable to adults. In nondiabetic adults, fasting glucose changes little over time, and our results suggest that age-independent effects of fasting glucose loci contribute to long-term interindividual differences in glucose levels from childhood onwards. Diabetes 60:1805-1812, 2011
|
|
| 37. |
- Barker, CJ, et al.
(författare)
-
XPR1 Mediates the Pancreatic β-Cell Phosphate Flush
- 2021
-
Ingår i: Diabetes. - : American Diabetes Association. - 1939-327X .- 0012-1797. ; 70:1, s. 111-118
-
Tidskriftsartikel (refereegranskat)abstract
- Glucose-stimulated insulin secretion is the hallmark of the pancreatic β-cell, a critical player in the regulation of blood glucose concentration. In 1974, the remarkable observation was made that an efflux of intracellular inorganic phosphate (Pi) accompanied the events of stimulated insulin secretion. The mechanism behind this “phosphate flush,” its association with insulin secretion, and its regulation have since then remained a mystery. We recapitulated the phosphate flush in the MIN6m9 β-cell line and pseudoislets. We demonstrated that knockdown of XPR1, a phosphate transporter present in MIN6m9 cells and pancreatic islets, prevented this flush. Concomitantly, XPR1 silencing led to intracellular Pi accumulation and a potential impact on Ca2+ signaling. XPR1 knockdown slightly blunted first-phase glucose-stimulated insulin secretion in MIN6m9 cells, but had no significant impact on pseudoislet secretion. In keeping with other cell types, basal Pi efflux was stimulated by inositol pyrophosphates, and basal intracellular Pi accumulated following knockdown of inositol hexakisphosphate kinases. However, the glucose-driven phosphate flush occurred despite inositol pyrophosphate depletion. Finally, while it is unlikely that XPR1 directly affects exocytosis, it may protect Ca2+ signaling. Thus, we have revealed XPR1 as the missing mediator of the phosphate flush, shedding light on a 45-year-old mystery.
|
|
| 38. |
- Barnes, BR, et al.
(författare)
-
Changes in exercise-induced gene expression in 5'-AMP-activated protein kinase gamma3-null and gamma3 R225Q transgenic mice
- 2005
-
Ingår i: Diabetes. - : American Diabetes Association. - 0012-1797 .- 1939-327X. ; 54:12, s. 3484-3489
-
Tidskriftsartikel (refereegranskat)abstract
- 5′-AMP–activated protein kinase (AMPK) is important for metabolic sensing. We used AMPKγ3 mutant–overexpressing Tg-Prkag3225Q and AMPKγ3-knockout Prkag3−/− mice to determine the role of the AMPKγ3 isoform in exercise-induced metabolic and gene regulatory responses in skeletal muscle. Mice were studied after 2 h swimming or 2.5 h recovery. Exercise increased basal and insulin-stimulated glucose transport, with similar responses among genotypes. In Tg-Prkag3225Q mice, acetyl-CoA carboxylase (ACC) phosphorylation was increased and triglyceride content was reduced after exercise, suggesting that this mutation promotes greater reliance on lipid oxidation. In contrast, ACC phosphorylation and triglyceride content was similar between wild-type and Prkag3−/− mice. Expression of genes involved in lipid and glucose metabolism was altered by genetic modification of AMPKγ3. Expression of lipoprotein lipase 1, carnitine palmitoyl transferase 1b, and 3-hydroxyacyl–CoA dehydrogenase was increased in Tg-Prkag3225Q mice, with opposing effects in Prkag3−/− mice after exercise. GLUT4, hexokinase II (HKII), and glycogen synthase mRNA expression was increased in Tg-Prkag3225Q mice after exercise. GLUT4 and HKII mRNA expression was increased in wild-type mice and blunted in Prkag3−/− mice after recovery. In conclusion, the Prkag3225Q mutation, rather than presence of a functional AMPKγ3 isoform, directly promotes metabolic and gene regulatory responses along lipid oxidative pathways in skeletal muscle after endurance exercise.
|
|
| 39. |
- Barnes, BR, et al.
(författare)
-
Isoform-specific regulation of 5' AMP-activated protein kinase in skeletal muscle from obese Zucker (fa/fa) rats in response to contraction
- 2002
-
Ingår i: Diabetes. - : American Diabetes Association. - 0012-1797 .- 1939-327X. ; 51:9, s. 2703-2708
-
Tidskriftsartikel (refereegranskat)abstract
- Glucose transport can be activated in skeletal muscle in response to insulin via activation of phosphoinositide (PI) 3-kinase and in response to contractions or hypoxia, presumably via activation of 5′ AMP-activated protein kinase (AMPK). We determined the effects of insulin and muscle contraction/hypoxia on PI 3-kinase, AMPK, and glucose transport activity in epitrochlearis skeletal muscle from insulin-resistant Zucker (fa/ fa) rats. Insulin-stimulated glucose transport in isolated skeletal muscle was reduced 47% in obese versus lean rats, with a parallel 42% reduction in tyrosine-associated PI 3-kinase activity. Contraction and hypoxia elicited normal responses for glucose transport in skeletal muscle from insulin-resistant obese rats. Isoform-specific AMPK activity was measured in skeletal muscle in response to insulin, contraction, or hypoxia. Contraction increased AMPKα1 activity 2.3-fold in lean rats, whereas no effect was noted in obese rats. Hypoxia increased AMPKα1 activity to a similar extent (more than sixfold) in lean and obese rats. Regardless of genotype, contraction, and hypoxia, each increased AMPKα2 activity more than fivefold, whereas insulin did not alter either AMPKα1 or -α2 activity in skeletal muscle. In conclusion, obesity-related insulin resistance is associated with an isoform-specific impairment in AMPKα1 in response to contraction. However, this impairment does not appear to affect contraction-stimulated glucose transport. Activation of AMPKα2 in response to muscle contraction/ exercise is associated with a parallel and normal increase in glucose transport in insulin-resistant skeletal muscle.
|
|
| 40. |
- Baumeier, Christian, et al.
(författare)
-
Hepatic DPP4 DNA methylation associates with fatty liver
- 2017
-
Ingår i: Diabetes. - : American Diabetes Association. - 0012-1797 .- 1939-327X. ; 66:1, s. 25-35
-
Tidskriftsartikel (refereegranskat)abstract
- Hepatic DPP4 expression is elevated in subjects with ectopic fat accumulation in the liver. However, whether increased dipeptidyl peptidase 4 (DPP4) is involved in the pathogenesis or is rather a consequence ofmetabolic disease is not known. We therefore studied the transcriptional regulation of hepatic Dpp4 in young mice prone to diet-induced obesity. Already at 6 weeks of age, expression of hepatic Dpp4 was increased in mice with high weight gain, independent of liver fat content. In the same animals, methylation of four intronic CpG sites was decreased, amplifying glucose-induced transcription of hepatic Dpp4. In older mice, hepatic triglyceride content was increased only in animals with elevated Dpp4 expression. Expression and release of DPP4 were markedly higher in the liver compared with adipose depots. Analysis of human liver biopsy specimens revealed a correlation of DPP4 expression and DNA methylation to stages of hepatosteatosis and nonalcoholic steatohepatitis. In summary, our results indicate a crucial role of the liver in participation to systemic DPP4 levels. Furthermore, the data show that glucoseinduced expression of Dpp4 in the liver is facilitated by demethylation of the Dpp4 gene early in life. This might contribute to early deteriorations in hepatic function, which in turn result in metabolic disease such as hepatosteatosis later in life.
|
|
