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-celliine HIT-TI5 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 ~) and dibutyryl cGMP (300 ~) 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 fiT 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-TI5 cells by NO donors is secondary to a rise in cGMP and suggest that the pathway controlling cell death involves activation of PKG.

Imidazoline compounds protect against interleukin 1beta-induced beta-cell apoptosis.

“…These results are in line with the suggestion that PKG may be involved in NO-induced apoptosis in insulin-secreting HIT T15 cells based on the data showing that the NO donor S-nitrosoglutathione and the permeable analog of cGMP stimulated apoptosis, an effect that could be inhibited by KT5823 (34). Mechanisms involving PKG in ␤-cells are poorly understood; our data show that they do not involve activation of Ca 2+ channels.…”

Factors mediating the transdifferentiation of islets of Langerhans to duct epithelial-like structures

“…An increase in intracellular cAMP is of interest too, because a rise in cAMP may form part of the effector system controlling apoptosis in pancreatic -cells (Loweth et al 1997). It is therefore noteworthy that cell loss due to apoptosis is the first step we observed in the process of islet-to-cyst transformation.…”

“…cAMP-mediated information flow, however, appears to play an important role (Osaka & Sabban 1997, Ghee et al 1998, Yarwood et al 1998. This is of particular interest, because a rise in cAMP has recently been reported to form part of the effector system controlling apoptosis in pancreatic -cells (Loweth et al 1997), and cell loss is the first stage observed in the transformation of islets to duct epithelial structures (Yuan et al 1996). Moreover, during this transition, islets are maintained in medium supplemented with cholera toxin (CT), which is known to elevate intracellular cAMP.…”

Cyclic nucleotide phosphodiesterases in pancreatic islets

“…8-bromoguanosine-cyclic GMP was found to augment glucose-induced Ca 2+ oscillations in mouse islets and to inhibit K ATP channel activity in mouse-intact beta cells [51]. Cyclic GMP was reported to mediate nitric oxide-induced apoptosis in the insulin-secreting cell line HIT-T15 [52].…”