Dominance of cytokine- over FasL-induced impairment of the mitochondrial transmembrane potential (Deltapsim) in the pancreatic beta-cell line NIT-1.
Mitochondria of pancreatic beta-cells are potential targets of intrinsic and extrinsic apoptotic pathways in the autoimmune pathogenesis of type 1 diabetes. We aimed to investigate whether cytokine- and FasLigand (FasL)-induced apoptosis is associated with impaired mitochondrial transmembrane potential (Deltapsim) in the pancreatic beta-cell line NIT-1. NIT-1 cells were exposed to the interleukin-1beta/interferon-gamma (IL-1beta/IFN-gamma) cytokine combination to induce apoptosis in vitro. Low concentrations of cytokines resulted in Deltapsim impairment, and increasing concentrations had only a minor additional effect. Treatment with the inducible nitric oxide synthase (iNOS) inhibitor Nw-nitro-L-arginine methyl ester hydrochloride (L-NAME) prevented cytokine-mediated Deltapsim impairment, implying that cytokines affect Deltapsim via nitric oxide. The broad-spectrum caspase inhibitor Z-VAD(Ome)-FMK (ZVAD) revealed dichotomic actions. In the presence of ZVAD, cytokine-induced nitrite generation was increased but cell death and Deltapsim impairment were reduced. Deltapsim impairment was also reduced by inhibitors of caspases 1, 6 and 8. Induction of Fas by IL-1beta/IFN-gamma coupled with activation by Super-FasL augmented cytokine-induced cell death. We observed a clear dominance of cytokine- over FasL-induced effects on Deltapsim. Our findings show that IL-1beta/IFN-gamma cytokines have a strong effect to impair Deltaym and prime beta-cells for apoptosis via the intrinsic pathway mediated by iNOS and caspases. Furthermore, at least in NIT-1 cells, the extrinsic FasL/Fas pathway has only a minor additive effect on cytokine-induced Deltapsim impairment.
Augstein
Petra
P
Institute of Diabetes "Gerhardt Katsch" Karlsburg e.V., Karlsburg, Germany. augstein@diabetes-karlsburg.de
Heinke
Peter
P
Salzsieder
Eckhard
E
Grimm
Rita
R
Giebel
Jürgen
J
Salzsieder
Christel
C
Harrison
Leonard C
LC
D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D000818:Animals; D017209:Apoptosis; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism; D002460:Cell Line; D004791:Enzyme Inhibitors / Q000494:pharmacology; D053222:Fas Ligand Protein / Q000378:metabolism*; D050417:Insulin-Secreting Cells / Q000187:drug effects / Q000201:enzymology / Q000276:immunology* / Q000473:pathology; D007371:Interferon-gamma / Q000378:metabolism*; D053583:Interleukin-1beta / Q000378:metabolism*; D053078:Membrane Potential, Mitochondrial; D051379:Mice; D008928:Mitochondria / Q000187:drug effects / Q000201:enzymology / Q000276:immunology* / Q000473:pathology; D019331:NG-Nitroarginine Methyl Ester / Q000494:pharmacology; D009569:Nitric Oxide / Q000378:metabolism; D052247:Nitric Oxide Synthase Type II / Q000037:antagonists & inhibitors / Q000378:metabolism; D009573:Nitrites / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D013997:Time Factors
D000590:Amino Acid Chloromethyl Ketones; D061945:Caspase Inhibitors; D004791:Enzyme Inhibitors; D053222:Fas Ligand Protein; C505658:Fasl protein, mouse; D053583:Interleukin-1beta; D009573:Nitrites; D011994:Recombinant Proteins; C476093:z-Val-Ala-Asp(Ome)-fluoromethylketone; D009569:Nitric Oxide; D007371:Interferon-gamma; D052247:Nitric Oxide Synthase Type II; C496317:Nos2 protein, mouse; D020169:Caspases; D019331:NG-Nitroarginine Methyl Ester