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Erschienen in:
01.03.2010 | Original Research Paper
Nitric oxide synthase 1 and cyclooxygenase-2 enzymes are targets of muscarinic activation in normal and inflamed NIH3T3 cells
Erschienen in: Inflammation Research | Ausgabe 3/2010
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Objective
Fibroblasts are sentinel cells that could serve as intermediaries in the immune reaction in the inflammatory process. In this work, we investigate the action of the muscarinic agonist carbachol (CARB) on the expression and function of nitric oxide synthase (NOS) and cyclooxygenase (COX) in fibroblasts under normal or inflammatory conditions.
Methods
The normal fibroblast cell line, 3T3, from NIH swiss mouse embryo, was used. The inflammatory milieu was mimicked with lipopolysaccharide (LPS) (10 ng/ml) plus interferon gamma (IFNγ) (0.5 ng/ml). Nitric oxide (NO) and prostaglandin E2 (PGE2) production were measured by Griess reagent and radioimmunoassay, respectively. NOS, COX, and nuclear transcription factor kappa B (NF-κB) were studied by Western blot.
Results
CARB increased NO synthesis by 57 ± 5%, while a 150 ± 10% increase in NO liberation was triggered by LPS plus IFNγ treatment. CARB added to LPS plus IFNγ potentiated NO synthesis by 227 ± 19%. CARB also upregulated NOS1 protein expression via NF-κB activation. In addition CARB and LPS plus IFNγ stimulated PGE2 synthesis by 72 ± 9 and 42 ± 4%, respectively, while CARB added to LPS plus IFNγ treated cells produced a synergism in PGE2 liberation (130 ± 12%) via COX-2.
Conclusion
Activation of muscarinic acetylcholine receptors can mimic mild inflammatory conditions or can deepen pre-existing inflammation, establishing a fine-tuned set-up on fibroblasts that in turn could be alerting the immune system.