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Vol. 295, Issue 2, 601-606, November 2000
Departments of Surgery (B.H.T., S.R.S., V.M.M.), Internal Medicine
(R.D.H.), Nicotine Research Center (R.D.H.), and Physiology and
Biophysics (V.M.M.), Mayo Clinic and Foundation, Rochester,
Minnesota
Effects of nicotine on arterial endothelium-dependent relaxations
mediated by nitric oxide are controversial. Experiments were designed
to test the hypothesis that nicotine can directly alter activity of
endothelial nitric-oxide synthase (eNOS). NOS from aortic endothelial
cells of untreated dogs and recombinant eNOS, neuronal NOS, and
inducible NOS were used for these experiments. NOS activity was
determined as conversion of L-[3H]arginine to
L-[3H]citrulline in the absence or presence
of nicotine (10
7-103 M) in vitro. In
separate assays, concentrations of cofactors NADPH, FAD, and
tetrahydrobioprotein were reduced by half to assess for possible
interaction with nicotine. With enzyme from aortic endothelial cells,
total and calcium-dependent accumulation of citrulline increased by
30% in the presence of 105 M nicotine. Nicotine dose
dependently also increased citrulline accumulation by recombinant eNOS
and neuronal NOS but not inducible NOS. Effects of nicotine on
accumulation of citrulline by isolated eNOS and recombinant eNOS were
further modulated by changes in the concentration of NADPH in the
incubation solution. Our data demonstrate a significant effect of
nicotine on eNOS-mediated citrulline accumulation. These results
suggest that effects of nicotine on production of nitric oxide may
depend on NADPH or oxygen radical interactions with NOS and thus may
explain, in part, inconsistent findings of changes in production of
endothelium-derived nitric oxide with nicotine administration.
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