title: Evidence for the involvement of cGMP and protein kinase G in nitric oxide-induced apoptosis in the pancreatic B-cell line, HIT-T15.
abstract: Intracellular production of nitric oxide (NO) is thought to mediate the pancreatic B-cell-directed cytotoxicity of cytokines in insulin-dependent diabetes mellitus, and recent evidence has indicated that this may involve induction of apoptosis. A primary effect of NO is to activate soluble guanylyl cyclase leading to increased cGMP levels and this effect has been demonstrated in pancreatic B-cells, although no intracellular function has been defined for islet cGMP. Here we demonstrate that the NO donor, GSNO, induces apoptosis in the pancreatic B-cell line HIT-T15 in a dose- and time-dependent manner. This response was significantly attenuated by micromolar concentrations of a specific inhibitor of soluble guanylyl cyclase, ODQ, and both 8-bromo cGMP (100 microM) and dibutyryl cGMP (300 microM) were able to fully relieve this inhibition. In addition, incubation of HIT-T15 cells with each cGMP analogue directly promoted cell death in the absence of ODQ. KT5823, a potent and highly selective inhibitor of cGMP-dependent protein kinase (PKG), abolished the induction of cell death in HIT cells in response to either GSNO or cGMP analogues. This effect was dose-dependent over the concentration range of 10-250 nM. Overall, these data provide evidence that the activation of apoptosis in HIT-T15 cells by NO donors is secondary to a rise in cGMP and suggest that the pathway controlling cell death involves activation of PKG.
authors.affiliation: Cellular Pharmacology Group, Department of Biological Sciences, Keele University, Staffordshire, UK.
authors.forename: A C
authors.initials: AC
authors.lastname: Loweth
authors.forename: G T
authors.initials: GT
authors.lastname: Williams
authors.forename: J H
authors.initials: JH
authors.lastname: Scarpello
authors.forename: N G
authors.initials: NG
authors.lastname: Morgan
mesh_terms: D000470:Alkaloids / Q000494:pharmacology; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D002227:Carbazoles; D002460:Cell Line; D006152:Cyclic GMP / Q000031:analogs & derivatives / Q000378:metabolism* / Q000494:pharmacology; D017869:Cyclic GMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D003995:Dibutyryl Cyclic GMP / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005978:Glutathione / Q000031:analogs & derivatives / Q000494:pharmacology; D007211:Indoles; D007515:Islets of Langerhans / Q000166:cytology*; D009603:Nitroso Compounds / Q000494:pharmacology; D010069:Oxadiazoles / Q000494:pharmacology; D011810:Quinoxalines / Q000494:pharmacology; D026422:S-Nitrosoglutathione
chemical_list: C095284:1H-(1,2,4)oxadiazolo(4,3-a)quinoxalin-1-one; D000470:Alkaloids; D002227:Carbazoles; D004791:Enzyme Inhibitors; D007211:Indoles; D009603:Nitroso Compounds; D010069:Oxadiazoles; D011810:Quinoxalines; C073601:KT 5823; C016276:8-bromocyclic GMP; D003995:Dibutyryl Cyclic GMP; D026422:S-Nitrosoglutathione; D017869:Cyclic GMP-Dependent Protein Kinases; D005978:Glutathione; D006152:Cyclic GMP