Substance P-induced cadherin expression and its signal transduction in a cloned human corneal epithelial cell line

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Abstract
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. J. Cell. Physiol. 182:189–195, 2000.

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