Inhibition of NOS-2 expression in macrophages through the inactivation of NF-kappaB by andalusol.
1. Andalusol, ent-6alpha,8alpha,18-trihydroxy-13(16),14-labdadiene, is a naturally occurring diterpene, isolated from Sideritis foetens (Lamiaceae). This compound exhibited therapeutic activity when evaluated in in vivo models of paw and ear inflammation (Navarro et al., 1997: Z. Naturforsch., 52, 844-849). The pharmacological effects of this diterpene have been analysed on the activation of the macrophage cell line J774 with lipopolysaccharide (LPS) and interferon-gamma (IFN-gamma). 2. Incubation of J774 macrophages with andalusol (0.1 - 100 microM) inhibited the synthesis of nitrite caused by LPS (1 microg ml-1) in concentration and time-dependent manners. The maximal inhibition was observed when andalusol was added 30 min before LPS stimulation and decreased progressively as the interval between andalusol and LPS challenge increased up to 14 h. 3. Incubation of J774 cells with LPS resulted in the expression of NOS-2 protein (130 kDa) as identified by Western blot analysis. The levels of this enzyme decreased significantly in the presence of andalusol (IC50=10.5 microM), suggesting that this diterpene inhibited NOS-2 expression. 4. Andalusol inhibited nuclear factor kappaB activation, a transcription factor necessary for NOS-2 expression in response to LPS and IFN-gamma. This compound also inhibited the degradation of IkappaBalpha favouring the retention of the inactive NF-kappaB complexes in the cytosol. 5. Related compounds to andalusol but lacking the polyol groups were less effective inhibiting NOS-2 expression in LPS-activated macrophages. The present findings provide a mechanism by which the anti-inflammatory properties of this diterpene could be mediated.
de las Heras
B
B
Departamento de Farmacología. Facultad de Farmacia, Universidad Complutense, 28040 Madrid, Spain.
Navarro
A
A
Díaz-Guerra
M J
MJ
Bermejo
P
P
Castrillo
A
A
Boscá
L
L
Villar
A
A
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D004247:DNA; D004224:Diterpenes / Q000494:pharmacology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D008264:Macrophages / Q000187:drug effects* / Q000201:enzymology; D051379:Mice; D016328:NF-kappa B / Q000378:metabolism*; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors*; D009573:Nitrites / Q000378:metabolism
D004224:Diterpenes; D004791:Enzyme Inhibitors; D016328:NF-kappa B; D009573:Nitrites; C110596:ent-13(16),14-labdadiene-6,8,18-triol; D004247:DNA; D019001:Nitric Oxide Synthase