Substance P-induced cadherin expression and its signal transduction in a cloned human corneal epithelial cell line.
Although the absence of Substance P (SP), a neurotransmitter in the trigeminal nerve, has been speculated as a cause for developing neurotrophic keratitis, its exact pathogenesis is still not clarified. In a previous report, we showed with electron microscopic examination that epithelial cell attachment was weakened in denervated corneas. In this study, SV40-transformed human corneal epithelial cells (HCE-Ts) were used to explore the molecular mechanisms responsible for mediating regulation of E-cadherin expression in response to Substance P receptor stimulation. Expression of the mRNAs for specific SP receptors, neurokinin (NK)-1R, NK-2R, and NK-3R, was demonstrated with RT-PCR. The cells were treated with various concentrations of SP in vitro, and the expression of an adhesion molecule E-cadherin was analyzed by immunofluorescence, immunoblotting, and enzyme-linked immunosorbent assay (ELISA) using an anti-E-cadherin antibody. E-cadherin expression was increased by SP in a dose-dependent manner both in the cytosolic fraction and in the cell membrane fraction. This increase in E-cadherin expression was completely inhibited by Calphostin C (PKC inhibitor) and KN-62 (CaMK inhibitor), but not by H-89 (PKA inhibitor), indicating that SP-induced E-cadherin expression involves the activation of protein kinase C (PKC) and calmodulin kinase (CaMK). SP did not affect cell proliferation at all. All these findings indicate that SP induced E-cadherin expression through PKC and CaMK activation and suggest that a lack of SP may account in part for the pathogenesis of neurotrophic keratitis.
Araki-Sasaki
K
K
Department of Ophthalmology, Osaka University School of Medicine, Osaka, Japan. sasakis@sa2.so-net.ne.jp
Aizawa
S
S
Hiramoto
M
M
Nakamura
M
M
Iwase
O
O
Nakata
K
K
Sasaki
Y
Y
Mano
T
T
Handa
H
H
Tano
Y
Y
D015820:Cadherins / Q000378:metabolism*; D002455:Cell Division / Q000187:drug effects; D002460:Cell Line; D019573:Epithelium, Corneal / Q000166:cytology / Q000378:metabolism*; D006801:Humans; D012333:RNA, Messenger / Q000378:metabolism; D018039:Receptors, Tachykinin / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects*; D013373:Substance P / Q000494:pharmacology*
D015820:Cadherins; D012333:RNA, Messenger; D018039:Receptors, Tachykinin; D013373:Substance P