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Science 327 (5962): 217-220

Copyright © 2010 by the American Association for the Advancement of Science

Overexpression of Alpha2A-Adrenergic Receptors Contributes to Type 2 Diabetes

Anders H. Rosengren1, Ramunas Jokubka1,*, Damon Tojjar1,*, Charlotte Granhall1, Ola Hansson1, Dai-Qing Li2, Vini Nagaraj1, Thomas M. Reinbothe1, Jonatan Tuncel3, Lena Eliasson1, Leif Groop1, Patrik Rorsman4, Albert Salehi1, Valeriya Lyssenko1, Holger Luthman1, and Erik Renström1,{dagger}

1 Lund University Diabetes Centre, Malmö, SE-20502 Malmö, Sweden.
2 Key Laboratory of Hormones and Development, Ministry of Health, China, Tianjin Metabolic Diseases Hospital, Tianjin Medical University, China.
3 Section for Medical Inflammation Research, Department of Medical Biochemistry and Biophysics, Karolinska Institutet, SE-17177 Stockholm, Sweden.
4 Oxford Centre for Diabetes Endocrinology and Metabolism, University of Oxford, Churchill Hospital, Oxford, UK.


Figure 1
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Fig. 1. Characterization of congenic rat strains. (A) Solid bars show known GK-derived genetic segments; open ends designate intervals containing the recombinant end points. (B) Insulin secretion from control, N1I11, and N1I5 islets (n = 5 to 9 batches per strain). (C) Increases in cell capacitance ({Delta}C) in N1I11 and N1I5 β cells after 10 depolarizations. The histogram shows total exocytosis evoked by the train stimulus ({Delta}CTOT) (n = 12, 8, 11, and 21 cells from control, N1I11, N1I5, and N1I12, respectively. (D) {Delta}C evoked by intracellular infusion of a Ca2+-containing solution in N1I11 (blue) and N1I5 (red) β cells). The arrow indicates the establishment of the standard whole-cell configuration. (E) Maximal change in fluorescence (F) ratio ({Delta}F ratio) relative to baseline after the elevation of glucose to 20 mM or KCl to 60 mM (n = 9 to 11 islets for glucose, and n = 5 islets for K+). (F) The percentage of rat β cell granules docked at the plasma membrane (n = 25 cells per strain). (G) Islet mRNA expression of indicated genes normalized to Hprt (n = 8 to 9 rats per group). (H) Immunoblots of total protein from rat islets using polyclonal alpha(2A)AR antisera. (I) Glucose and insulin levels during IPGTT with or without yohimbine or clonidine as indicated (n = 7 to 20 rats per group). Time scale expanded at 0 to 30 min. *P < 0.05; **P < 0.01; ***P < 0.001.

 

Figure 2
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Fig. 2. Analysis of alpha(2A)AR signaling in rat pancreatic islets. (A) Insulin secretion from N1I11 and N1I5 islets at different glucose concentrations with or without clonidine or yohimbine as indicated (n = 3 to 8 batches per group). (B) Immunoblots of total protein from N1I5 islets transfected with small interfering RNA (siRNA) active against Adra2a or inactive siRNA. The histogram shows average alpha(2A)AR signal normalized for β-actin. Data from four blots. (C) Insulin secretion at 20 mM glucose from islets that are nontransfected (N.T.) or transfected with inactive or active siRNA [n = 6 to 13 batches per transfection condition for N1I11, N1I5, or N1I5 with 1 µM clonidine (C)]. (D) Depolarization-evoked capacitance increase in N1I5 β cells transfected with inactive or active siRNA. Average total exocytosis ({Delta}CTOT) from 7 to 9 cells per group. (E) Insulin release from N1I5 islets at 20 mM glucose with active siRNA, deltamethrin, or clonidine as indicated (n = 4 to 6 batches per group). *P < 0.05; **P < 0.01; ***P < 0.001.

 

Figure 3
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Fig. 3. Association of ADRA2A rs553668 with insulin secretion in humans. (A) Effects of rs553668 genotype on insulin secretion during IVGTT in 799 individuals. Data are means ± SEM. P values were obtained using an additive model. (B) Immunoblots of total protein from human islets from eight individuals using polyclonal alpha(2A)AR antisera. The histogram shows average alpha(2A)AR signal normalized for β-actin from four blots from a total of 11 GG, 7 GA, and 1 AA carriers. (C) Islet ADRA2A mRNA expression in 24 GG, 7 GA, and 1 AA carriers. P < 0.05 for GG versus GA/AA, or P < 0.05 for linear regression of expression versus number of risk alleles. (D) Islet insulin secretion at 2.8 or 20 mM glucose with or without alpha(2A)AR antagonist. (E) Electron micrographs of human islet sections from GG and GA carriers at 20 mM glucose without alpha(2A)AR antagonist showing β cells with insulin granules, recognized by the central dense core and surrounding halo. The border to adjacent cells is indicated. The histogram shows the distribution of insulin granules located in 0.2-µm concentric shells in the first 2 µm below the plasma membrane after incubation at 20 mM glucose with or without alpha(2A)AR antagonist. Total granule numbers per section did not differ between GG (104 ± 9) and GA (93 ± 15) [n for (D) and (E) is specified in (10)]. *P < 0.05; **P < 0.01; ***P < 0.001.

 


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