The role of NF-kappaB, IRF-1, and STAT-1alpha transcription factors in the iNOS gene induction by gliadin and IFN-gamma in RAW 264.7 macrophages.
Nitric oxide (NO) plays an important role in the pathogenesis of celiac disease. We have examined the involvement of nuclear factor-kappaB (NF-kappaB), interferon regulatory factor-1 (IRF-1), and signal transducer and activator of transcription-1alpha (STAT-1alpha) on the synergistic induction of inducible nitric oxide synthase (iNOS) gene expression by gliadin (G) in association with interferon-gamma (IFN-gamma) in RAW 264.7 macrophages. We found that IFN-gamma was efficient in enhancing the basal transcription of the iNOS promoter at 1, 6, and 24 h, whereas G had no effect. The G plus IFN-gamma association caused an increase in iNOS promoter activity which was inhibited by pyrrolidine dithiocarbammate (PDTC) at 6 and 24 h as well as by genistein (Gen) and tyrphostine B42 (TB42) at 1 h, inhibitors of NF-kappaB, IRF-1, and STAT-1alpha activation, respectively. Similarly, the IFN-gamma and G combination treatment led to a higher increase in iNOS mRNA levels at 1, 6, and 24 h compared with IFN-gamma alone. Gen and TB42 inhibited iNOS mRNA levels at 1 h, whereas PDTC inhibited iNOS mRNA levels at 6 and 24 h. In addition, the synergistic induction of iNOS gene expression by G plus IFN-gamma correlated with the induction of NF-kappaB, IRF-1, and STAT-1alpha/DNA binding activity and mRNA expression. In conclusion, our study, which provides evidence that the effect of G on iNOS gene transcription in IFN-gamma-stimulated RAW 264.7 cells can be ascribed to all three transcription factors, may contribute to lead to new insights into the molecular mechanisms governing the inflammatory process in celiac disease.
De Stefano
Daniela
D
Dipartimento di Farmacologia Sperimentale, Via D. Montesano 49, University of Naples Federico II, 80131 Naples, Italy.
Maiuri
Maria Chiara
MC
Iovine
Barbara
B
Ialenti
Armando
A
Bevilacqua
Maria Assunta
MA
Carnuccio
Rosa
R
D000818:Animals; D000975:Antioxidants / Q000494:pharmacology; D002446:Celiac Disease / Q000378:metabolism; D002460:Cell Line; D004791:Enzyme Inhibitors / Q000494:pharmacology; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D019833:Genistein / Q000494:pharmacology; D005903:Gliadin / Q000378:metabolism*; D006801:Humans; D050856:Interferon Regulatory Factor-1 / Q000378:metabolism*; D007371:Interferon-gamma / Q000378:metabolism*; D008264:Macrophages / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D051379:Mice; D016328:NF-kappa B / Q000037:antagonists & inhibitors / Q000378:metabolism*; D052247:Nitric Oxide Synthase Type II / Q000235:genetics / Q000378:metabolism*; D009573:Nitrites / Q000378:metabolism; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D011759:Pyrrolidines / Q000494:pharmacology; D050794:STAT1 Transcription Factor / Q000235:genetics / Q000378:metabolism*; D013859:Thiocarbamates / Q000494:pharmacology; D015533:Transcriptional Activation; D020032:Tyrphostins / Q000494:pharmacology
D000975:Antioxidants; D004791:Enzyme Inhibitors; D050856:Interferon Regulatory Factor-1; D016328:NF-kappa B; D009573:Nitrites; D020033:Protein Isoforms; D011759:Pyrrolidines; D050794:STAT1 Transcription Factor; C494080:Stat1 protein, mouse; D013859:Thiocarbamates; D020032:Tyrphostins; C095512:alpha-cyano-(3,4-dihydroxy)-N-benzylcinnamide; C020972:pyrrolidine dithiocarbamic acid; D007371:Interferon-gamma; D005903:Gliadin; D019833:Genistein; D052247:Nitric Oxide Synthase Type II; C496317:Nos2 protein, mouse