To evaluate the in vitro effects of 1-mM rebamipide ophthalmic solution on the expression of inflammatory cytokines and MUC5AC in Cu, Zn-superoxide dismutase-1 (SOD-1) knock-down conjunctival epithelium.
Conjunctival epithelium from C57BL/6 wild-type mice was cultured and treated with rebamipide ophthalmic solution. Using cytometric bead array, we examined the levels of interleukin-(IL)-6, IL-10, IL-17, monocyte chemoattractant protein-1, interferon-γ (INF-γ), tumor necrosis factor, and IL-12p70 in the culture supernatants. The culture supernatants were obtained from the culture medium of nontreated or SOD-1 knock-down conjunctival epithelium using small interfering RNA (siRNA). In addition, ELISA was performed to ascertain the MUC5AC concentration in the culture medium.
After rebamipide ophthalmic solution was applied, IL-6 concentration in the supernatants of conjunctival epithelial cells treated with and without siRNA showed a significant timewise decrease from 0 to 24 hr (963±42 to 0.07±0.05 pg/mL and 932±168 to 2.2±0.05 pg/mL, respectively) (P<0.001). Compared with baseline values, MUC5AC concentrations significantly increased 24 hr after rebamipide application to the conjunctival cultures—both with and without SOD-1 siRNA treatment (P<0.05 in both cases).
Rebamipide seems to increase MUC5AC levels and suppress inflammation by decreasing IL-6 levels in mouse conjunctival epithelial cell cultures. SOD-1 siRNA-treated mouse conjunctival epithelial cell culture is a practical method for investigating changes in mucosa-associated mucins and proinflammatory cytokines in response to therapeutic interventions.
Department of Ophthalmology (M.O., A.I., M.D., J.S.), Tokyo Dental College, Ichikawa General Hospital, Chiba, Japan; and Department of Ophthalmology (C.S., T.K., T.N., T.K., M.D., K.T.), Keio University School of Medicine, Tokyo, Japan.
Address correspondence to Murat Dogru, M.D., Ph.D., Department of Ophthalmology, Keio University School of Medicine, Shinanomachi 35, Shinjuku-ku, Tokyo 160-8582, Japan; e-mail: firstname.lastname@example.org
None of the authors have any conflicts of interest to declare.
Supported by a grant from Otsuka Pharmaceuticals and presented at the Japan Cornea Conference, which took place on January 30, 2014–February 1, 2014, in Ginowan city, Okinawa, Japan.
Accepted September 24, 2018