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Yoshiaki Tagawa, Kousuke Noda, Ken-ichi Otsuguro, Erdal Tan Ishizuka, Atsuhiro Kanda, Susumu Ishida; Mechanism in suppression of CCK pathway by rebamipide in trigeminal neurons.. Invest. Ophthalmol. Vis. Sci. 2017;58(8):467.
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© ARVO (1962-2015); The Authors (2016-present)
Recently, corneal hypersensitivity has been reported to participate in the pathogenesis of dry eye disease. Rebamipide ophthalmic suspension, which is widely used in the treatment of dry eye disease, is known to repair the milieu of keratoconjunctival epithelia and increase mucin production in conjunctival goblet cells. In addition to the restoration of ocular surface, we also demonstrated that rebamipide ameliorated the pain sensation of dry eye patients and that rebamipide reduced intracellular calcium concentration caused by activation of transient receptor potential channel subfamily member V1 (TRPV1), a pain receptor, through inhibition of neuropeptide cholecystokinin (CCK) pathway. In this study, we further explored the detailed pharmacological mechanism by which rebamipide inhibits CCK pathway.
After euthanasia, trigeminal ganglion (TG) cells were harvested from Wistar rats (1- to 3-week-old). We examined the expression of CCK and CCK receptors (CCK-1R and -2R) by reverse transcriptional PCR and in situ hybridization. The effect of CCK-1R and -2R antagonists in the cultured TG cells treated with CCK (10-7M) was evaluated by intracellular calcium concentration measurement. Additionally, we quantified the binding affinity between rebamipide and CCK receptors using radioisotope-labelled ligand binding assay.
We confirmed the expression of CCK, CCK-1R and -2R in rat TG cells. The intracellular calcium concentration of rat TG cells increased by CCK stimulation (41.2±10.7nM, n=32). CCK-1R antagonist did not suppress the response (38.3±12.5nM, n=32, P=0.47). By contrast, CCK-2R antagonist suppressed the increase of intracellular calcium concentration caused by CCK stimulation (1.1±0.14nM, n=32, P<0.01). However, the binding affinity of rebamipide to CCK-1R (IC50=4.70x10-5M) was higher than to CCK-2R (IC50>1x10-3M).
The current data demonstrate that rebamipide preferably binds to CCK-1R; however, CCK induces the increase of intracellular calcium concentration through CCK-2R signaling in rat TG cells.
This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.
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