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Vol. 293, Issue 3, 973-981, June 2000
Department of Dermatology, University of California, Davis,
California (S.Z., A.N., S.A.G.); Department of Dermatology, University
of Minnesota, Minneapolis, Minnesota (T.X.L.); and Research & Diagnostic Antibodies, Richmond, California (R.J.W.)
Early stages of wound healing rely on the ability of keratinocytes
(KCs) to move over the denuded dermis to re-epithelialize the defect.
The agarose gel keratinocyte outgrowth system (AGKOS) is an in vitro
model of skin re-epithelialization designed to study the migratory
function of KCs. Endogenously secreted acetylcholine controls crawling
locomotion of KCs in AGKOS by binding to the cholinergic receptors of
both the nicotinic and the muscarinic classes that are expressed by
KCs. In this study, we used AGKOS to elucidate the nicotinic pathway of
cholinergic control of keratinocyte migration. Activation of the
nicotinic acetylcholine receptors decreased the migration distance of
KC in a dose-dependent fashion without altering cell viability.
Nicotine also increased in a dose-dependent manner transmembrane influx
of 45Ca2+, and caused a transient rise in the
concentration of [Ca2+]i. Perfect correlation
between concentration responses found in the migration and
45Ca2+ influx assays suggested that
nicotine-induced inhibition of crawling locomotion relies on modulation
of Ca2+ metabolism in KCs. The effects of nicotine could be
mediated by the 
3- and the 7-containing nicotinic receptors
visualized on KCs by immunostaining. Long-term incubation with nicotine
up-regulated 7 and down-regulated 3 expression. Thus, nicotine
exerts inhibitory effects on keratinocyte migration, and
Ca2+ serves as a second messenger in the signaling pathway.
These results help explain deleterious effects of nicotine on wound re-epithelialization, and suggest that smoking may delay wound healing
via nicotinic receptor-mediated pathway.
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