Cytokine-induced human islet cell death in vitro correlates with a persistently high phosphorylation of STAT-1, but not with NF-κB activation

Abstract

"Cytokine-induced human islet cell death in vitro correlateswith a persistently high phosphorylation of STAT-1, but not with Access to the published version may require subscription. ABSTRACTStudies of insulin producing -cells have reported conflicting responses to NF-κB activation, encompassing both pro-and anti-apoptotic effects, possibly reflecting the use of -cells from different species. Therefore, the aim of this study was to compare the temporal activation of NF-κB in rat and human insulin producing cells and relate this to the dynamics of cell death, STAT-1 activation and the production of nitric oxide (NO). Rat RIN5AH and human islet cells were exposed to the cytokines IL-1 and IFN- and the NOS inhibitor aminoguanidine. Cell death, NO production,  phosphorylation, p65 methylation, STAT-1 phosphorylation and cIAP-2 levels were analyzed at different time-points. Cytokine-induced RIN5AH cell death occurred on day 1, and this was paralleled by NF-B activation, STAT-1 phosphorylation and production of NO. On the other hand, the human islet cells instead died by an NO-independent mechanism on day 3and 5. This later occurring cell death was associated with a gradual decrease in  phosphorylation and p65 methylation, and a lowered expression of the NF-B target genes  and cIAP-2. STAT-1 phosphorylation was persistently high during the entire cytokine exposure period in human islet cells. The results favor a pro-survival role of NF-B and a pro-apoptotic role of STAT-1 in human islet cells. Thus, rodent insulin producing cells may not be suitable as models for human -cells in the context of cytokine-induced damage.

Sensitivity profile of the human EndoC-βH1 beta cell line to proinflammatory cytokines

“…The vulnerability of EndoC-βH1 beta cells towards proinflammatory cytokines could be significantly enhanced by a three-dimensional configuration of EndoC-βH1 beta cells as pseudoislets, which can be attributed to improved cell-cell contact. Our results in EndoC-βH1 beta cells are in line with previous observations in human islets [20,[22][23][24].…”

“…This NFκB activation was primarily dependent on IL-1β, which is in line with a corresponding observation in human islets [24]. The weak cytokine-mediated activation of NFκB in EndoC-βH1 beta cells, as in human islets, failed to induce the iNOS pathway [22,24,33], which is at variance to rodent beta cells [7,13,19,21,[34][35][36][37][38][39]. Our results confirm recent studies in EndoC-βH1 beta cells that reported virtually no expression of iNOS [4,40].…”

“…In addition, we documented a lack of expression of other NOS isoforms: endothelial NOS and neuronal NOS. Thus, similar to human islets [24,41], EndoC-βH1 beta cells are apparently not capable of producing NO.…”

“…This difference between the in vitro and in vivo situation might explain the weaker in vitro toxicity of the cytokines towards the EndoC-βH1 beta cells, as compared with rodent beta cells. The lack of NO production in human beta cells [22,24] as well as in EndoC-βH1 beta cells thus emphasises the role of the beta cell as an innocent bystander, rather than an active participant, in the process of autoimmune-mediated beta cell destruction in patients with type 1 diabetes mellitus.…”

Evidence of Contribution of iPLA2β-Mediated Events During Islet β-Cell Apoptosis Due to Proinflammatory Cytokines Suggests a Role for iPLA2β in T1D Development

“…Islets (500) were treated with either vehicle (DMSO, 1 L/mL) alone or with IL-1␤ (100 U/mL) ϩ IFN␥ (300 U/mL) and cultured for up to 48 hours. Because of limited availability of human islets, our studies were confined to testing a combination of proinflammatory cytokines (IL-1␤ϩIFN␥, designated CTK) that has previously been shown to be sufficient in impacting human islet ␤-cell function/survival and promoting insulitis in T1D (25)(26)(27)(28)(29)(30). In some experiments, the islets were pretreated with inhibitors of iPLA 2 ␤ (S-BEL, 10 mol/L) for 30 minutes or NSMase2 (GW4869, 20 mol/L) for 1 hour, prior to treatment with DMSO or CTK.…”