Double-stranded ribonucleic acid (RNA) induces beta-cell Fas messenger RNA expression and increases cytokine-induced beta-cell apoptosis.
Type 1 diabetes mellitus (T1DM) is an autoimmune disease caused by progressive destruction of insulin-producing pancreatic beta-cells. Both viral infections and the cytokines interleukin-1beta (IL-1beta) and interferon-gamma (IFN-gamma) have been suggested as potential mediators of beta-cell death in early T1DM. We presently investigated whether the viral replicative intermediate double stranded RNA [here used as synthetic polyinosinic-polycytidylic acid (PIC)] modifies the effects of IL-1beta and IFN-gamma on gene expression and viability of rat pancreatic beta-cells. For this purpose, fluorescence-activated cell sorting-purified rat beta-cells were exposed for 6-16 h (study of gene expression by RT-PCR) or 6-9 days (study of viability by nuclear dyes) to PIC and/or IL-1beta and IFN-gamma. PIC increased the expression of Fas and Mn superoxide dismutase messenger RNAs by 5- to 10-fold. IL-1beta and a combination of PIC and IFN-gamma (but not PIC or IFN-gamma alone) induced expression of inducible nitric oxide (NO) synthase (iNOS) and consequent NO production. Induction of iNOS expression by PIC and IFN-gamma requires nuclear factor-kappaB activation, as suggested by transfection experiments with iNOS promoter-luciferase reporter constructs into primary beta-cells. Combinations of IL-1beta plus IFN-gamma, PIC plus IFN-gamma, or PIC plus IL-1beta induced a 2- to 3-fold increase in the number of apoptotic beta-cells. Blocking of iNOS activity significantly decreased PIC- plus IL-1beta-induced, but not PIC- plus IFN-gamma-induced, apoptosis. In conclusion, PIC alone or in combination with cytokines modifies the expression of several genes in pancreatic beta-cells. Two of these genes, Fas and iNOS, may contribute to beta-cell death. The transcription factor nuclear factor-kappaB is required for PIC-induced iNOS expression. PIC has an additive effect on cytokine-induced beta-cell death by both NO-dependent (in the case of IL-1beta) and NO-independent (in the case of IFN-gamma) mechanisms. These findings suggest that viral intermediates in synergism with local cytokine production may play an important role in beta-cell apoptosis in early T1DM.
Liu
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Gene Expression Unit, Diabetes Research Center, Vrije Universiteit Brussel, B-1090 Brussels, Belgium.
Darville
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Eizirik
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D000818:Animals; D017209:Apoptosis; D002478:Cells, Cultured; D016207:Cytokines / Q000494:pharmacology*; D005434:Flow Cytometry; D015870:Gene Expression / Q000187:drug effects*; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000235:genetics; D007371:Interferon-gamma / Q000494:pharmacology; D007375:Interleukin-1 / Q000494:pharmacology; D007515:Islets of Langerhans / Q000166:cytology / Q000378:metabolism*; D008156:Luciferases / Q000235:genetics; D008297:Male; D016328:NF-kappa B / Q000502:physiology; D009569:Nitric Oxide / Q000096:biosynthesis; D019001:Nitric Oxide Synthase / Q000235:genetics; D052247:Nitric Oxide Synthase Type II; D011070:Poly I-C / Q000494:pharmacology; D011401:Promoter Regions, Genetic; D012330:RNA, Double-Stranded / Q000494:pharmacology*; D012333:RNA, Messenger / Q000032:analysis*; D051381:Rats; D017208:Rats, Wistar; D011993:Recombinant Fusion Proteins; D013482:Superoxide Dismutase / Q000235:genetics; D014162:Transfection; D019014:fas Receptor / Q000235:genetics*
D016207:Cytokines; D007375:Interleukin-1; D016328:NF-kappa B; D012330:RNA, Double-Stranded; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D019014:fas Receptor; D009569:Nitric Oxide; D007371:Interferon-gamma; D008156:Luciferases; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; C496318:Nos2 protein, rat; D013482:Superoxide Dismutase; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases; D011070:Poly I-C