| 1. |
- Abels, Mia, et al.
(författare)
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CART is overexpressed in human type 2 diabetic islets and inhibits glucagon secretion and increases insulin secretion
- 2016
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Ingår i: Diabetologia. - : Springer Science and Business Media LLC. - 0012-186X .- 1432-0428. ; 59:9, s. 1928-1937
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Tidskriftsartikel (refereegranskat)abstract
- Aims/hypothesis Insufficient insulin release and hyperglucagonaemia are culprits in type 2 diabetes. Cocaine- and amphetamine-regulated transcript (CART, encoded by Cartpt) affects islet hormone secretion and beta cell survival in vitro in rats, and Cart(-/-) mice have diminished insulin secretion. We aimed to test if CART is differentially regulated in human type 2 diabetic islets and if CART affects insulin and glucagon secretion in vitro in humans and in vivo in mice. Methods CART expression was assessed in human type 2 diabetic and non-diabetic control pancreases and rodent models of diabetes. Insulin and glucagon secretion was examined in isolated islets and in vivo in mice. Ca2+ oscillation patterns and exocytosis were studied in mouse islets. Results We report an important role of CART in human islet function and glucose homeostasis in mice. CART was found to be expressed in human alpha and beta cells and in a subpopulation of mouse beta cells. Notably, CART expression was several fold higher in islets of type 2 diabetic humans and rodents. CART increased insulin secretion in vivo in mice and in human and mouse islets. Furthermore, CART increased beta cell exocytosis, altered the glucose-induced Ca2+ signalling pattern in mouse islets from fast to slow oscillations and improved synchronisation of the oscillations between different islet regions. Finally, CART reduced glucagon secretion in human and mouse islets, as well as in vivo in mice via diminished alpha cell exocytosis. Conclusions/interpretation We conclude that CART is a regulator of glucose homeostasis and could play an important role in the pathophysiology of type 2 diabetes. Based on the ability of CART to increase insulin secretion and reduce glucagon secretion, CART-based agents could be a therapeutic modality in type 2 diabetes.
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| 2. |
- Buda, Pawel, et al.
(författare)
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Eukaryotic translation initiation factor 3 subunit e controls intracellular calcium homeostasis by regulation of cav1.2 surface expression.
- 2013
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Ingår i: PLoS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 8:5
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Tidskriftsartikel (refereegranskat)abstract
- Inappropriate surface expression of voltage-gated Ca(2+)channels (CaV) in pancreatic ß-cells may contribute to the development of type 2 diabetes. First, failure to increase intracellular Ca(2+) concentrations at the sites of exocytosis impedes insulin release. Furthermore, excessive Ca(2+) influx may trigger cytotoxic effects. The regulation of surface expression of CaV channels in the pancreatic β-cells remains unknown. Here, we used real-time 3D confocal and TIRFM imaging, immunocytochemistry, cellular fractionation, immunoprecipitation and electrophysiology to study trafficking of L-type CaV1.2 channels upon β-cell stimulation. We found decreased surface expression of CaV1.2 and a corresponding reduction in L-type whole-cell Ca(2+) currents in insulin-secreting INS-1 832/13 cells upon protracted (15-30 min) stimulation. This internalization occurs by clathrin-dependent endocytosis and could be prevented by microtubule or dynamin inhibitors. eIF3e (Eukaryotic translation initiation factor 3 subunit E) is part of the protein translation initiation complex, but its effect on translation are modest and effects in ion channel trafficking have been suggested. The factor interacted with CaV1.2 and regulated CaV1.2 traffic bidirectionally. eIF3e silencing impaired CaV1.2 internalization, which resulted in an increased intracellular Ca(2+) load upon stimulation. These findings provide a mechanism for regulation of L-type CaV channel surface expression with consequences for β-cell calcium homeostasis, which will affect pancreatic β-cell function and insulin production.
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| 3. |
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| 4. |
- Krus, Ulrika, et al.
(författare)
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The Complement Inhibitor CD59 Regulates Insulin Secretion by Modulating Exocytotic Events.
- 2014
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Ingår i: Cell Metabolism. - : Elsevier BV. - 1550-4131 .- 1932-7420. ; 19:5, s. 883-890
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Tidskriftsartikel (refereegranskat)abstract
- Type 2 diabetes is triggered by reduced insulin production, caused by genetic and environmental factors such as inflammation originating from the innate immune system. Complement proteins are a component of innate immunity and kill non-self cells by perforating the plasma membrane, a reaction prevented by CD59. Human pancreatic islets express CD59 at very high levels. CD59 is primarily known as a plasma membrane protein in membrane rafts, but most CD59 protein in pancreatic β cells is intracellular. Removing extracellular CD59 disrupts membrane rafts and moderately stimulates insulin secretion, whereas silencing intracellular CD59 markedly suppresses regulated secretion by exocytosis, as demonstrated by TIRF imaging. CD59 interacts with the exocytotic proteins VAMP2 and Syntaxin-1. CD59 expression is reduced by glucose and in rodent diabetes models but upregulated in human diabetic islets, potentially reflecting compensatory reactions. This unconventional action of CD59 broadens the established view of innate immunity in type 2 diabetes.
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| 5. |
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| 6. |
- Nagaraj, Vini, et al.
(författare)
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Complement inhibitor CD55 governs the integrity of membrane rafts in pancreatic beta cells, but plays no role in insulin secretion.
- 2015
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Ingår i: Biochemical and Biophysical Research Communications. - : Elsevier BV. - 1090-2104 .- 0006-291X. ; 460:3, s. 518-524
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Tidskriftsartikel (refereegranskat)abstract
- CD55 is a glycosylphosphatidylinositol-anchored protein, which inhibits complement activation by acting on the complement C3 convertases. CD55 is widely localized in the cholesterol rich regions of the cell plasma membrane termed membrane rafts. CD55 is attached to these specialized regions via a GPI link on the outer leaflet of the plasma membrane. Membrane rafts anchor many important signaling proteins, which control several cellular functions within the cell. For example, we recently demonstrated that the membrane raft protein and complement inhibitor CD59 also controls insulin secretion by an intracellular mechanism. Therefore, we have in this study aimed at addressing the expression and function of CD55 in pancreatic beta cells. To this end, we observe that CD55 is highly expressed in INS1 832/13 beta cells as well as human pancreatic islets. Diabetic human islets show a tendency for increased expression of CD55 when compared to the healthy controls. Importantly, silencing of CD55 in INS1 832/13 cells does not affect their insulin secretory capacity. On the other hand, silencing of CD55 diminished the intensity of membrane rafts as determined by Atto-SM staining. We hence conclude that CD55 expression is affected by glycemic status in human islets and plays a critical role in maintaining the conserved structure of rafts in pancreatic islets, which is similar to that of the related complement inhibitor CD59. However CD55 does not interfere with insulin secretion in beta cells, which is in sharp contrast to the action of the complement inhibitor CD59.
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| 7. |
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| 8. |
- Nagaraj, Vini, et al.
(författare)
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Elevated basal insulin secretion in type 2 diabetes caused by reduced plasma membrane cholesterol
- 2016
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Ingår i: Molecular Endocrinology. - : Endocrine Society. - 0888-8809 .- 1944-9917. ; 30:10, s. 1059-1069
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Tidskriftsartikel (refereegranskat)abstract
- Elevated basal insulin secretion under fasting conditions together with insufficient stimulated insulin release is an important hallmark of type 2 diabetes, but the mechanisms controlling basal insulin secretion remain unclear. Membrane rafts exist in pancreatic islet cells and spatially organize membrane ion channels and proteins controlling exocytosis, which may contribute to the regulation of insulin secretion. Membrane rafts (cholesterol and sphingolipid containing microdomains) were dramatically reduced in human type 2 diabetic and diabetic Goto-Kakizaki (GK) rat islets when compared with healthy islets. Oxidation of membrane cholesterol markedly reduced microdomain staining intensity in healthy human islets, but was without effect in type 2 diabetic islets. Intriguingly, oxidation of cholesterol affected glucose-stimulated insulin secretion only modestly, whereas basal insulin release was elevated. This was accompanied by increased intracellular Ca2+ spike frequency and Ca2+ influx and explained by enhanced single Ca2+ channel activity. These results suggest that the reduced presence of membrane rafts could contribute to the elevated basal insulin secretion seen in type 2 diabetes.
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| 9. |
- Nagaraj, Vini
(författare)
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Membrane rafts and its GPI anchored complement inhibitors regulate insulin secretion
- 2015
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Type 2 diabetic patients usually exhibit an abnormal lipid profile. One such lipid that is constantly elevated in T2D is cholesterol. At the cellular level, cholesterol aids in tight packing of sphingolipids in certain regions of the plasma membrane and these specialized regions are termed ‘membrane rafts’, which act as signalling hubs. In pancreatic beta cells, these structures have been found to anchor important exocytotic proteins like VAMP2, syntaxin1A, SNAP25, Ca2+ channels and K+ channels and there are increasing number of evidences for its role in insulin secretory process. This thesis has aimed at unravelling the mechanisms by which membrane rafts regulate insulin exocytosis. Pancreatic islets from type 2 diabetic human donors show markedly decreased membrane raft staining intensity when compared to islets from healthy donors. Oxidation of membrane raft cholesterol using cholesterol oxidase (CO) significantly reduced membrane rafts, thereby disrupting its native structure and thus mimicking the diabetic condition. We have here shown that membrane raft disruption causes increased secretion of insulin under non-stimulatory conditions, depicting the loss of regulated insulin secretion. The increased basal insulin levels are accounted for by increased voltage-gated Ca2+ channel activity, which is further translated into increased influx of Ca2+ under basal conditions. Membrane rafts contain two important GPI anchored proteins, CD59 and CD55 which are crucial inhibitors of the complement system. Our results suggest that CD59 is the most highly expressed complement gene in human and rat pancreatic islets, the expression of which is downregulated in diabetic rodents. Intracellular CD59 is found colocalized with insulin, VAMP2 and syntaxin1A and this association is increased by high glucose incubations. Cells lacking CD59 exhibit decreased insulin exocytosis, which is caused by the lost CD59-mediated pairing of the SNARE proteins VAMP2 and syntaxin1A. CD59 thus plays an important role in insulin secretion. The extracellular CD59 is further observed to be important for maintaining the structural integrity of membrane rafts. CD55 is highly expressed in pancreatic islets and beta cells. Interestingly, in chronic hyperglycemia (diabetes), the expression of CD55 is further increased and is positively correlated with HbA1c. In fact, this is probably directly related to the hyperglycemic condition as glucose incubations regulate CD55 expression in human islets. In contrast to CD59, CD55 neither interacts with, nor regulates the insulin secretion machinery. However, CD55 shares the important capacity to regulate membrane rafts, which are disintegrated by silencing of CD55. Taken together, this thesis has shown that membrane rafts regulate insulin secretion by controlling the activity of voltage-gated Ca2+ channels. Furthermore, membrane raft marker protein and complement inhibitor CD59 is involved in the exocytosis process. This action is mediated by intracellular assistance in pairing of exocytosis-regulating SNARE proteins.
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| 10. |
- Rosengren, Anders, et al.
(författare)
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Overexpression of alpha2A-adrenergic receptors contributes to type 2 diabetes
- 2010
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Ingår i: Science. - : American Association for the Advancement of Science (AAAS). - 1095-9203 .- 0036-8075. ; 327:5962, s. 217-20
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Tidskriftsartikel (refereegranskat)abstract
- Several common genetic variations have been associated with type 2 diabetes, but the exact disease mechanisms are still poorly elucidated. Using congenic strains from the diabetic Goto-Kakizaki rat, we identified a 1.4-megabase genomic locus that was linked to impaired insulin granule docking at the plasma membrane and reduced beta cell exocytosis. In this locus, Adra2a, encoding the alpha2A-adrenergic receptor [alpha(2A)AR], was significantly overexpressed. Alpha(2A)AR mediates adrenergic suppression of insulin secretion. Pharmacological receptor antagonism, silencing of receptor expression, or blockade of downstream effectors rescued insulin secretion in congenic islets. Furthermore, we identified a single-nucleotide polymorphism in the human ADRA2A gene for which risk allele carriers exhibited overexpression of alpha(2A)AR, reduced insulin secretion, and increased type 2 diabetes risk. Human pancreatic islets from risk allele carriers exhibited reduced granule docking and secreted less insulin in response to glucose; both effects were counteracted by pharmacological alpha(2A)AR antagonists.
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