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et al., 1998; Herring and Kaminski, 1999). The possibility
that experimental and methodological factors prevent the
observation of the effect of cannabimimetic lipophilic sub-
stances on NF-B at submicromolar concentrations should be
taken into account. Furthermore, the effect of AEA on VR1
was also shown to occur at Ͼ1 M concentrations, but further
experiments have shown that the threshold for VR1 activa-
tion by AEA can be sensibly lowered by several regulatory
factors (Di Marzo et al., 2001). For example, the action of
AEA at both VR1 and T-type channels is exerted at a site on
the cytosolic side of these membrane proteins and is con-
trolled by AEA-facilitated transport into the cell (Chemin et
al., 2001; De Petrocellis et al., 2001). It is therefore possible
that the AEA membrane transporter is lacking, or at least
not very active, in the cells used in our assays, and that
higher extracellular concentrations of AEA are required for
the observation of its intracellular effects. Furthermore, the
relatively low potency of AEA against NF-B activity should
not diminish the potential physiological and pathological im-
portance of our observations. In fact, although nanomolar
extracellular concentrations of AEA are expected to occur
under physiological conditions, the intracellular concentra-
tions might be much higher, particularly under pathological
conditions leading to inflammation and apoptosis, such as
cell injury and tissue damage (Berdyshev et al., 2000, 2001a).
Taken together, our observations show that AEA exhibits
NF-B inhibitory activity and that this effect is not mediated
by the interaction of AEA with cannabinoid or vanilloid re-
ceptors. This activity might underlie some of the known
anti-inflammatory and proapoptotic effects of AEA and pro-
vides a rationale for the synthesis of AEA analogs endowed
with selective anti-inflammatory and anticancer properties.
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Acknowledgments
We thank Dr. Nicole Israe¨l (Institute Pasteur, Paris, France) for
the 5.1 cells; Dr. Alain Israe¨l (Institute Pasteur) for the anti-p50 and
anti-p65 antisera; Dr. R. T. Hay (CBMS, University of St. Andrews,
Fife, Scotland) for the mAb 10B; and colleagues Drs. D. Goeddel (San
Francisco, CA), D. Wallach (Rehovot, Israel), and L. Schmitz (Bern,
Switzerland) for providing plasmids. We are grateful to Dr. Filomena
Fezza (Instituto di Chimica Biomolecolare) and to Dr. Nives Dadd-
ario (Dipartimento di Scienze Chimiche Alimentari Farmaceutiche e
Farmacologiche) for the synthesis of N-acylethanolamines and N-
acylvanillamines, respectively. Finally, we thank Carmen Cabrero-
Doncel for assistance with the manuscript.
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