Nitric oxide production in Caco-2 cells exposed to different inducers, inhibitors and natural toxins.
The involvement of the NO pathway in several intestinal inflammatory diseases is under investigation. In vitro models may provide a useful approach to better characterise this pathway at the cellular level. For this purpose, we have used Caco-2 cells, which are able to spontaneously differentiate in long-term culture to small intestine enterocytes. The effect of different NO pathway inducers [gamma-interferon (IFN-gamma) and phorbol myristate acetate (PMA)] has been studied. Our results demonstrate that Caco-2 cells constitutively express NOS at very low levels, while the induction with PMA+IFN-gamma triggers the expression of the inducible isoform with a stronger effect starting from day 14 of differentiation. The use of specific inhibitors of gene expression, at transcriptional and translational level, suggests that new synthesis of iNOS mRNA is required, through direct activation of the gene or new synthesis of transcription-required factors, as indicated by CHX inhibition. The morphological alteration induced by PMA+IFN-gamma is reversed by iNOS inhibitor, suggesting that the NO pathway may be involved in the cytoskeletal alterations. The DSP toxins, OA and DTX-1, induce NO production at levels corresponding to their different toxicity, previously detected in Caco-2 cells.
Vignoli
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AL
Istituto Superiore di Sanità, Lab. T.C.E., Viale Regina Elena 299, 00161 Rome, Italy.
Srivastava
R C
RC
Stammati
A
A
Turco
L
L
Tanori
M
M
Zucco
F
F
D015153:Blotting, Western; D018938:Caco-2 Cells / Q000187:drug effects* / Q000378:metabolism* / Q000473:pathology; D002470:Cell Survival / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D004338:Drug Combinations; D004790:Enzyme Induction / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D006801:Humans; D007371:Interferon-gamma / Q000494:pharmacology; D008387:Marine Toxins / Q000494:pharmacology*; D019331:NG-Nitroarginine Methyl Ester / Q000494:pharmacology; D009569:Nitric Oxide / Q000096:biosynthesis*; D019001:Nitric Oxide Synthase / Q000096:biosynthesis / Q000235:genetics; D052247:Nitric Oxide Synthase Type II; D052250:Nitric Oxide Synthase Type III; D019319:Okadaic Acid / Q000494:pharmacology / Q000031:analogs & derivatives; D011714:Pyrans / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D019323:omega-N-Methylarginine / Q000494:pharmacology
D004338:Drug Combinations; D004791:Enzyme Inhibitors; D008387:Marine Toxins; D011714:Pyrans; D012333:RNA, Messenger; D019319:Okadaic Acid; D019323:omega-N-Methylarginine; D009569:Nitric Oxide; C051904:dinophysistoxin 1; D007371:Interferon-gamma; C496319:NOS2 protein, human; C496314:NOS3 protein, human; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; D052250:Nitric Oxide Synthase Type III; D013755:Tetradecanoylphorbol Acetate; D019331:NG-Nitroarginine Methyl Ester