thapsigargin and Diabetes-Mellitus--Type-1

Pancreatic islets are exposed to strong pro-apoptotic stimuli: inflammation and hyperglycemia, during the progression of the autoimmune diabetes (T1D). We found that the. We studied T1D pathogenesis in NOD mice hemideficient for CDK11 (N-HTZ), and, in N-HTZ deficient for Cyclin D3 (K11HTZ-D3KO), in comparison to their respective controls (N-WT and K11WT-D3KO). Moreover, we exposed pancreatic islets to either pro-inflammatory cytokines in the presence of increasing glucose concentrations, or Thapsigargin, an Endoplasmic Reticulum (ER)-stress inducing agent, and assessed apoptotic events. The expression of key ER-stress markers (. N-HTZ mice were significantly protected against T1D, and NS-HTZ pancreatic islets exhibited an impaired sensitivity to cytokine-induced apoptosis, regardless of glucose concentration. However, thapsigargin-induced apoptosis was not altered. Furthermore, CDK11 hemideficiency did not attenuate the exacerbation of T1D caused by Cyclin D3 deficiency.. This study is the first to report that CDK11 is repressed in T1D as a protection mechanism against inflammation-induced apoptosis and suggests that CDK11 lies upstream Cyclin D3 signaling. We unveil the CDK11/Cyclin D3 tandem as a new potential intervention target in T1D.

Topics: Activating Transcription Factor 4; Animals; Apoptosis; Autoimmunity; Blood Glucose; Cyclin D3; Cyclin-Dependent Kinases; Cytokines; Diabetes Mellitus, Type 1; Disease Models, Animal; Endoplasmic Reticulum Chaperone BiP; Endoplasmic Reticulum Stress; Heat-Shock Proteins; Inflammation; Insulin-Secreting Cells; Mice, 129 Strain; Mice, Inbred C57BL; Mice, Inbred NOD; Mice, Knockout; Protein Serine-Threonine Kinases; Thapsigargin; Tissue Culture Techniques; Transcription Factor CHOP

Topics: Animals; Cell Line; Cell Survival; Diabetes Mellitus, Type 1; Endoplasmic Reticulum Stress; Insulin-Secreting Cells; Mice; Mice, Knockout; S100 Calcium Binding Protein G; Taurochenodeoxycholic Acid; Thapsigargin

The effects of cytokine and fatty acid treatment on signal transduction in dermal fibroblasts from type 1 diabetics and matched controls were compared. Chronic exposure to TNF, accentuated Ca(2+) mobilization in response to bradykinin (BK) in cells from both controls and diabetics; responses were three-fold greater in cells from diabetics than in controls. Similarly, with chronic exposure to IL-1β, BK-induced Ca(2+) mobilization was accentuated in cells from type 1 diabetics compared to the controls. Pretreatment with the protein synthesis inhibitor cycloheximide or the protein kinase C inhibitor calphostin C prior to the addition of TNF completely abrogated the TNF-induced increment in peak bradykinin response. Ca(2+) transients induced by depleting endoplasmic reticulum (ER) Ca(2+) with thapsigargin were also greater in TNF treated fibroblasts than in untreated cells, with greater increases in cells from diabetics. Exposing fibroblasts for 48 hours to 2 mM oleate also increased both the peak bradykinin response and the TNF-induced increment in peak response, which were significantly greater in diabetics than controls. These data indicate that cells from diabetic patients acquire elevated ER Ca(2+) stores in response to both cytokines and free fatty acids,and thus exhibit greater sensitivity to environmental inflammatory stimuli and elevated lipids.

Topics: Blotting, Western; Bradykinin; Calcium; Case-Control Studies; Cells, Cultured; Cytokines; Dermis; Diabetes Mellitus, Type 1; Endoplasmic Reticulum; Enzyme Inhibitors; Fatty Acids, Nonesterified; Fibroblasts; Humans; Interleukin-1; Receptors, Bradykinin; Siblings; Signal Transduction; Thapsigargin; Tumor Necrosis Factor-alpha

Pancreatic beta cell destruction in type 1 diabetes may be mediated by cytokines such as IL-1β, IFN-γ and TNF-α. Endoplasmic reticulum (ER) stress and nuclear factor-κB (NFκB) signalling are activated by cytokines, but their significance in beta cells remains unclear. Here, we investigated the role of cytokine-induced ER stress and NFκB signalling in beta cell destruction.. Isolated mouse islets and MIN6 beta cells were incubated with IL-1β, IFN-γ and TNF-α. The chemical chaperone 4-phenylbutyric acid (PBA) was used to inhibit ER stress. Protein production and gene expression were assessed by western blot and real-time RT-PCR.. We found in beta cells that inhibition of cytokine-induced ER stress with PBA unexpectedly potentiated cell death and NFκB-regulated gene expression. These responses were dependent on NFκB activation and were associated with a prolonged decrease in the inhibitor of κB-α (IκBα) protein, resulting from increased IκBα protein degradation. Cytokine-mediated NFκB-regulated gene expression was also potentiated after pre-induction of ER stress with thapsigargin, but not tunicamycin. Both PBA and thapsigargin treatments led to preferential upregulation of ER degradation genes over ER-resident chaperones as part of the adaptive unfolded protein response (UPR). In contrast, tunicamycin activated a balanced adaptive UPR in association with the maintenance of Xbp1 splicing.. These data suggest a novel mechanism by which cytokine-mediated ER stress interacts with NFκB signalling in beta cells, by regulating IκBα degradation. The cross-talk between the UPR and NFκB signalling pathways may be important in the regulation of cytokine-mediated beta cell death.

Topics: Animals; Antineoplastic Agents; Cell Death; Cell Line, Tumor; Cell Survival; Cytokines; Diabetes Mellitus, Type 1; Endoplasmic Reticulum Stress; Enzyme Inhibitors; Insulin-Secreting Cells; Interferon-gamma; Interleukin-1beta; Mice; Mice, Inbred C57BL; NF-kappa B; Pancreatic Neoplasms; Phenylbutyrates; RNA, Small Interfering; Signal Transduction; Thapsigargin; Transcription Factor CHOP; Tumor Necrosis Factor-alpha; Tunicamycin; Unfolded Protein Response