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 trigemi nal nerve, has been speculated as a cause for developing neurotrophic kerat itis, its exact pathogenesis is still not clarified. In a previous report, we showed with electron microscopic examination that epithelial cell attach ment was weakened in denervated corneas. In this study, SV40-transformed hu man corneal epithelial cells (HCE-Ts) were used to explore the molecular me chanisms responsible for mediating regulation of E-cadherin expression in r esponse to Substance P receptor stimulation. Expression of the mRNAs for sp ecific 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 v itro, 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 increa sed 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 comp letely inhibited by Calphostin C (PKC inhibitor) and KN-62 (CaMK inhibitor) , but not by H-89 (PKA inhibitor), indicating that SP-induced E-cadherin ex pression involves the activation of protein kinase C (PKC) and calmodulin k inase (CaMK). SP did not affect cell proliferation at all. All these findin gs indicate that SP induced E-cadherin expression through PKC and CaMK acti vation and suggest that a lack of SP may account in part for the pathogenes is of neurotrophic keratitis. (C) 2000 Wiley-Liss, Inc.