| 1. |
- Barg, Sebastian, et al.
(författare)
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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
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Ingår i: Diabetes. - 1939-327X .- 0012-1797. ; 51 Suppl 1, s. 74-82
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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.
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| 2. |
- Barg, Sebastian, et al.
(författare)
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Tight coupling between electrical activity and exocytosis in mouse glucagon-secreting alpha-cells
- 2000
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Ingår i: Diabetes. - 1939-327X .- 0012-1797. ; 49:9, s. 1500-1510
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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.
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| 3. |
- Cederberg, Jonas, et al.
(författare)
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Increased mRNA levels of Mn-SOD and catalase in embryos of diabetic rats from a malformation-resistant strain
- 2000
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Ingår i: Diabetes. - : American Diabetes Association. - 0012-1797 .- 1939-327X. ; 49, s. 101-107
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Tidskriftsartikel (refereegranskat)abstract
- Previous studies have suggested that reactive oxygen species (ROS) are mediators in the teratogenic process of diabetic pregnancy. In an animal model for diabetic pregnancy, offspring of the H rat strain show minor dysmorphogenesis when the mother is diabetic, whereas the offspring of diabetic rats of a sister strain, U, display major morphologic malformations. Earlier studies have shown that embryonic catalase activity is higher in the H than in the U strain, and maternal diabetes increases this difference in activity. The aim of this study was to characterize the influence of genetic predisposition on diabetic embryopathy by comparing the mRNA levels of ROS-metabolizing enzymes in the two strains. We determined the mRNA levels of catalase, glutathione peroxidase, gamma-glutamylcystein-synthetase, glutathione reductase, and superoxide dismutase (CuZn-SOD and Mn-SOD) in day 11 embryos of normal and diabetic H and U rats using semiquantitative reverse transcription-polymerase chain reaction. The mRNA levels of catalase and Mn-SOD were increased in H embryos as a response to maternal diabetes, and no differences were found for the other genes. Sequence analysis of the catalase promoter indicated that the difference in mRNA levels may result from different regulation of transcription. Sequence analysis of the catalase cDNA revealed no differences between the two strains in the translated region, suggesting that the previously observed difference in the electrophoretic mobility in zymograms is due to posttranslational modifications. An impaired expression of scavenging enzymes in response to ROS excess can thus be an integral part of a genetic predisposition to embryonic dysmorphogenesis.
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| 4. |
- Islam, M. Shahidul
(författare)
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The ryanodine receptor calcium channel of beta-cells : molecular regulation and physiological significance
- 2002
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Ingår i: Diabetes. - : American Diabetes Association. - 0012-1797 .- 1939-327X. ; 51:5, s. 1299-1309
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Tidskriftsartikel (refereegranskat)abstract
- The list of Ca(2+) channels involved in stimulus-secretion coupling in beta-cells is increasing. In this respect the roles of the voltage-gated Ca(2+) channels and IP(3) receptors are well accepted. There is a lack of consensus about the significance of a third group of Ca(2+) channels called ryanodine (RY) receptors. These are large conduits located on Ca(2+) storage organelle. Ca(2+) gates these channels in a concentration- and time-dependent manner. Activation of these channels by Ca(2+) leads to fast release of Ca(2+) from the stores, a process called Ca(2+)-induced Ca(2+) release (CICR). A substantial body of evidence confirms that beta-cells have RY receptors. CICR by RY receptors amplifies Ca(2+) signals. Some properties of RY receptors ensure that this amplification process is engaged in a context-dependent manner. Several endogenous molecules and processes that modulate RY receptors determine the appropriate context. Among these are several glycolytic intermediates, long-chain acyl CoA, ATP, cAMP, cADPR, NO, and high luminal Ca(2+) concentration, and all of these have been shown to sensitize RY receptors to the trigger action of Ca(2+). RY receptors, thus, detect co-incident signals and integrate them. These Ca(2+) channels are targets for the action of cAMP-linked incretin hormones that stimulate glucose-dependent insulin secretion. In beta-cells some RY receptors are located on the secretory vesicles. Thus, despite their low abundance, RY receptors are emerging as distinct players in beta-cell function by virtue of their large conductance, strategic locations, and their ability to amplify Ca(2+) signals in a context-dependent manner.
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| 5. |
- Krook, Henrik, et al.
(författare)
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A distinct Th1 immune response precedes the described Th2 response in Islet xenograft rejection
- 2002
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Ingår i: Diabetes. - : American Diabetes Association. - 0012-1797 .- 1939-327X. ; 51:1, s. 79-86
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Tidskriftsartikel (refereegranskat)abstract
- Previous studies using semiquantitative reverse transcription-polymerase chain reaction (RT-PCR) have demonstrated that islet xenograft rejection in mice is dominated by Th2-associated cytokines, i.e., interleukin (IL)-4 and IL-10. However, immunohistochemical stainings show that the morphological pattern in this model is more reminiscent of a delayed-type hypersensitivity (DTH) reaction, which is associated with a Th1 response. This study was designed to resolve the mechanisms of acute cellular xenograft rejection in rats transplanted with fetal porcine islet-like cell clusters (ICCs). Real-time quantitative RT-PCR was used to quantify the mRNA expression of cytokines in the grafts and lymph nodes, and the findings were related to the immunopathology of the rejecting grafts. By day 1, mRNA expression levels of IL-1 beta, IL-2, IL-12p40, interferon-gamma, and tumor necrosis factor-alpha were already induced in the lymph nodes. From days 3 to 12, an increasing amount of activated macrophages was seen in the grafts, whereas T- and NK-cells were fewer and mainly accumulated in the periphery of the grafts. Most of the ICCs were rejected by day 5. Transcripts of Th1-associated cytokines were dominant in both regional lymph nodes and in the grafts, with peak levels on days 3 and 5, respectively. The mRNA expression of IL-4 was increased on day 12, and it correlated with the infiltration of eosinophils and an increased level of xenoreactive IgG. The data presented indicate that an islet xenograft triggers a sequential activation of 1) a Th1-associated response characterized by graft destruction in a DTH-like reaction and then 2) a subsequent Th2-associated response characterized by increased levels of xenoreactive antibodies.
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| 6. |
- Mattsson, Göran, et al.
(författare)
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Evidence of Functional Impairment of Syngeneically Transplanted Mouse Pancreatic Islets Retrieved from the Liver
- 2004
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Ingår i: Diabetes. - : American Diabetes Association. - 0012-1797 .- 1939-327X. ; 53:4, s. 948-954
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Tidskriftsartikel (refereegranskat)abstract
- A drawback in pancreatic islet transplantation is the large number of islets needed to obtain insulin independence in patients with diabetes. This most likely reflects extensive posttransplantation islet cell death and functional impairment of the remaining endocrine cells. We aimed to develop an experimental method to retrieve transplanted islets from the mouse liver, which would enable comparisons of transplanted and endogenous islets and provide valuable information on functional changes induced by intraportal transplantation. Transplanted islets were obtained by retrograde perfusion of the liver with collagenase. The identity of retrieved tissue as transplanted islets was confirmed by intravital staining, immunohistochemistry, and electron microscopy. The retrieved islets, irrespective of whether they had resided in diabetic or nondiabetic recipients, had a markedly lower insulin content and glucose-stimulated insulin release when compared with isolated endogenous islets. The glucose oxidation rate was also markedly lower in the retrieved islets, suggesting mitochondrial dysfunction. These disturbances in insulin content, insulin release, and glucose oxidation rate were not reversed by a few days of culture after retrieval. The results implicate changes in islet function after intraportal transplantation. Such dysfunction may contribute to the high number of islets needed for successful transplantation in diabetic individuals.
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| 7. |
- Nielsen, Jens H, et al.
(författare)
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Regulation of the beta cell mass by hormones and growth factors
- 2001
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Ingår i: Diabetes. - 0012-1797 .- 1939-327X. ; 50:Suppl . 1, s. 25-29
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Tidskriftsartikel (refereegranskat)abstract
- Substantial new information has accumulated on molecular mechanisms of pancreas development, regulation of beta-cell gene expression, and the role of growth factors in the differentiation, growth, and regeneration of beta-cells. The present review focuses on some recent studies on the mechanism of action of cytokines such as growth hormone (GH) and prolactin (PRL) in beta-cell proliferation and gene expression-in particular, the role of signal transducers and activators of transcription (STAT) proteins. The implication of the discovery of suppressors of cytokine signaling (SOCS) proteins for the interaction between stimulatory and inhibitory cytokines, including GH, PRL, leptin, and the proinflammatory cytokines interleukin-1 and interferon-gamma, in beta-cell survival is not yet clear. Recent studies indicate a role of cell adhesion molecules and the delta-like protein preadipocyte factor 1/fetal antigen 1 (Pref-1/FA-1) in cytokine-induced beta-cell growth and development. Surprisingly, glucagon-like peptide-1 (GLP-1) was recently found to stimulate not only insulin secretion but also beta-cell replication and differentiation, which may present a new perspective in treatment of type 2 diabetes. Together with the intriguing reports on positive effects of insulin on both beta-cell growth and function, a picture is emerging of an integrated network of signaling events acting in concert to control beta-cell mass adaptation to insulin demand.
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| 8. |
- Renström, Erik, et al.
(författare)
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Sulfonylurea-Mediated Stimulation of Insulin Exocytosis via an ATP-Sensitive K(+) Channel--Independent Action.
- 2002
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Ingår i: Diabetes. - 1939-327X .- 0012-1797. ; 51:Suppl 1, s. 33-36
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Tidskriftsartikel (refereegranskat)abstract
- Several reports indicate that hypoglycemic sulfonylureas augment Ca(2+)-dependent insulin secretion via mechanisms other than inhibition of the ATP-sensitive K(+) channel. The effect involves a 65-kd protein in the granule membrane and culminates in intragranular acidification. Lowering of granule pH is necessary for the insulin granule to gain release competence. Proton pumping into the granule is driven by a v-type H(+)-ATPase, but requires simultaneous Cl(-) uptake into the granule via metabolically regulated ClC-3 Cl(-) channels to maintain electroneutrality. Here we discuss the possibility that modulation of granule ClC-3 channels represents the mechanism whereby sulfonylureas directly potentiate the beta-cell exocytotic machinery.
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| 9. |
- Sandqvist, M M, et al.
(författare)
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Increased lactate release per fat cell in normoglycemic first-degree relatives of individuals with type 2 diabetes.
- 2001
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Ingår i: Diabetes. - 0012-1797 .- 1939-327X. ; 50:10, s. 2344-8
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Tidskriftsartikel (refereegranskat)abstract
- The aim of this study was to examine subcutaneous lactate production in the relatives of individuals with type 2 diabetes. Therefore, we recruited seven healthy first-degree relatives of type 2 diabetic patients and seven pairwise, matched, healthy control subjects without any heredity for diabetes. All subjects were studied with a euglycemic insulin clamp at approximately 600 pmol/l, abdominal subcutaneous microdialysis, and (133)Xe clearance. Furthermore, a subcutaneous needle biopsy was performed to determine fat cell size. In the fasting state, interstitial lactate was 40% higher in relatives than in control subjects (P = 0.043), but net lactate production was similar in both groups. However, during the insulin clamp, interstitial lactate (2.50 +/- 0.29 vs. 1.98 +/- 0.26 mmol/l, P = 0.018), interstitial-arterial lactate concentration difference (1.08 +/- 0.30 vs. 0.53 +/- 0.24 mmol/l, P = 0.028), and net lactate release per fat cell (10.9 +/- 3.7 vs. 2.8 +/- 1.3 fmol. cell(-1). min(-1), P = 0.018) were increased in the relatives. We conclude that first-degree relatives of type 2 diabetic patients may have an enhanced net lactate release per fat cell in abdominal subcutaneous tissue. This could suggest a pathological regulation in adipose tissue that is of importance for the metabolic defects known in type 2 diabetic relatives.
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| 10. |
- Schmidt, Peter, et al.
(författare)
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A new murine model of islet xenograft rejection : Graft destruction is dependent on a major histocompatibility-specific interaction between T-cells and macrophages
- 2003
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Ingår i: Diabetes. - : American Diabetes Association. - 0012-1797 .- 1939-327X. ; 52:5, s. 1111-1118
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Tidskriftsartikel (refereegranskat)abstract
- A new murine model of porcine islet-like cell cluster (ICC) xenograft rejection, avoiding interference of unspecific inflammation, was introduced and used to investigate rejection mechanisms. Athymic (nu/nu) mice were transplanted with syngeneic, allogeneic, or xenogeneic islets under the kidney capsule. After the original transplantation, immune cells in porcine ICC xenografts undergoing rejection in native immunocompetent mice were transferred to the peritoneal cavity of the athymic mice. At defined time points after transfer, the primary grafts were evaluated by immunohistochemistry and real-time quantitative RT-PCR to estimate cytokine and chemokine mRNA expression. Transfer of immunocompetent cells enabled athymic (nu/nu) mice to reject a previously tolerated ICC xenograft only when donor and recipient were matched for major histocompatibility complex (MHC). In contrast, allogeneic and syngeneic islets were not rejected. The ICC xenograft rejection was mediated by transferred T-cells. The main effector cells, macrophages, were shown to be part of a specific immune response. By day 4 after transplantation, there was an upreglation of both Th1- and Th2-associated cytokine transcripts. The transferred T-cells were xenospecific and required MHC compatibility to induce rejection. Interaction between the TCR of transferred T-cells and MHC on host endothelial cells and/or macrophages seems necessary for inducing ICC xenograft rejection.
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