Zandong Yang

Activation of 12-lipoxygenase in proinflammatory cytokine-mediated beta cell toxicity

Aims/hypothesisBeta cell inflammation and cytokine-induced toxicity are central to autoimmune diabetes development. Lipid mediators generated upon lipoxygenase (LO) activation can participate in inflammatory pathways. 12LO-deficient mice are resistant to streptozotocin-induced diabetes. This study sought to characterise the cellular processes involving 12LO-activation lipid inflammatory mediator production in cytokine-treated pancreatic beta cells.MethodsIslets and beta cell lines were treated with a combination of IL-1β, IFN-γ and TNF-α, or the 12LO product 12(S)-hydroxyeicosatetraenoic acid (HETE). Insulin secretion was measured using an enzyme immunoassay, and cell viability was evaluated using an in situ terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling assay. 12LO activity was evaluated and 12LO protein levels were determined using immunoblotting with a selective leucocyte type 12LO antibody. Cellular localisation of 12LO was evaluated using immunocytochemistry.ResultsBasal expression of leucocyte type 12LO protein was found in human and mouse islets and in several rodent beta cell lines. In mouse β-TC3 cells, and in human islets, cytokines induced release of 12-HETE within 30xa0min. Cytokine addition also induced a rapid translocation of 12LO protein from the cytosol to the nucleus of β-TC3 cells as shown by subcellular fractionation and immunostaining. Cytokine-induced cell death and inhibition of insulin secretion were partially reversed by baicalein, a 12LO inhibitor. 12(S)-HETE inhibited β-TC3 cell insulin release in a time- and concentration-dependent manner. Incubating β-TC3 cells with 100xa0nmol/l of 12(S)-HETE resulted in a 57% reduction in basal insulin release (6xa0h), and a 17% increase in cell death (18xa0h) as compared with untreated cells. 12(S)-HETE activated the stress-activated protein kinase c-Jun N-terminal kinase and p38 within 15xa0min, as judged by increased kinase protein phosphorylation.Conclusions/interpretationThe data suggest that inflammatory cytokines rapidly activate 12LO and show for the first time that cytokines induce 12LO translocation. The effects of 12-HETE on insulin secretion, cytotoxicity and kinase activation were similar to the effects seen with cytokines. The results provide mechanistic information of cytokine-induced toxic effects on pancreatic beta cells and support the hypothesis that blocking 12LO activation could provide a new therapeutic way to protect pancreatic beta cells from autoimmune injury.

The immune modulator FYT720 prevents autoimmune diabetes in nonobese diabetic mice.

FTY720 is a novel immune regulatory drug derived from the fungal sphingosine analog ISP-1 (myriocin). FTY720 causes a redistribution of lymphocytes from circulation to secondary lymphoid tissues. Type 1 diabetes is an autoimmune disorder caused by cellular-mediated destruction of insulin-producing pancreatic beta cells in the islets of Langerhans. Indeed, local infiltration of islets by mononuclear cells is the hallmark of Type 1 diabetes. Based on both FTY720s action and the involvement of cellular infiltration in the disease progression, we tested FTY720 for its ability to prevent autoimmune diabetes in diabetes-prone, nonobese diabetic (NOD) mice. We found that treatment with FTY720 completely prevented NOD mice from developing autoimmune diabetes. The FTY720-treated animals showed both reduced numbers of circulating lymphocytes and sharply diminished cellular infiltration of pancreatic islets. These results suggest that FTY720 may be effective in prevention of autoimmune diabetes or in slowing its progression.

The anti-inflammatory compound lisofylline prevents Type I diabetes in non-obese diabetic mice.

AbstractAims/hypothesis. Pro-inflammatory cytokines are increased during the active stages of Type I (insulin-dependent) diabetes mellitus. The aim of this study was to investigate the applicability of using a new anti-inflammatory compound, Lisofylline, to prevent diabetes in non-obese diabetic (NOD) mice. Lisofylline has previously been shown to block Th1 cell differentiation and to reduce IL-1β-induced dysfunction in rat islets.n Methods. Lisofylline was added to isolated NOD islets in vitro, with or without IL-1β. Insulin secretion and DNA damage of the islets was assessed. Lisofylline was administered to female non-obese diabetic mice starting at 4, 7 and 17 weeks of age for 3 weeks. Cytokines and blood glucose concentrations were monitored. Histology and immunohistochemistry were carried out in pancreatic sections. Splenocytes isolated from donor mice were intravenously injected into immunodeficient NOD (NOD.scid) mice.n Results. In vitro, Lisofylline preserved beta-cell insulin secretion and inhibited DNA damage of islets in the presence of IL-1β. In vivo, Lisofylline suppressed IFN-γ production, reduced the onset of insulitis and diabetes, and inhibited diabetes after transfer of splenocytes from Lisofylline-treated donors to NOD.scid recipients. However, cotransfer of splenocytes from both Lisofylline-treated and diabetic NOD donors did not suppress diabetes in recipient mice.n Conclusion/interpretation. Lisofylline prevents the onset of autoimmune diabetes in NOD mice by a mechanism that does not seem to enhance the function of regulatory T cells, but could be associated with suppression of proinflammatory cytokines and reduction of cellular infiltration in islets. This study suggests that Lisofylline could have therapeutic benefits in preventing the onset of Type I diabetes.