To verify whether ESW-elicited enhancement of nNOS
activity resulted in an increase of the NO synthesis in
ESW-treated C6 cells, the intracellular NO production was
measured using the DAF-2DA detection system. When the
cells were treated with ESW, DAF-2T fluorescence was sig-
nificantly enhanced above the background level in C6 cells
(Fig. 2). As expected, this fluorescence response was pre-
vented in cells treated with 1 mM N-nitro-L L-arginine methyl
ester (L L-NAME) for 30 min before ESW treatment (Fig. 2),
thus, indicating that the increase of DAF-2T fluorescence
was consequent to the activation of the L L-arginine-NO path-
way.
The ESW-induced enhancement of constitutive NOS-depen-
dent NO levels is worth of interest when we take into account
the recent notion that physiological, low levels of NO, similar
to those produced by the basal activity of nNOS and/or eNOS,
prevent induction of iNOS mRNA expression (but also of
other NF-jB-dependent genes, including TNF-a and cycloox-
ygenase-2) through the suppression of NF-jB activation
[17,19–23]. As a consequence, iNOS gene expression takes
place after LPS/cytokine stimulation, provided that the
cNOS-generated NO is reduced below a threshold value in a