Effect of Dexamethasone and Diclofenac on the Proliferation of Human Corneal Keratocytes

Lee, Ji Eun; Lee, Seung Uk; Kim, Jong Hwan; Lee, Jong Soo

Abstract

PURPOSE
To evaluate the inhibitory effect of dexamethasone and diclofenac on the proliferation of cultured human keratocytes, and to investigate the apoptotic response and the cellular morphologic changes associated with dexamethasone and diclofenac in vitro. METHODS: Human corneal keratocytes were exposed to 0.05, 0.1, 0.2, 0.4, 0.8, 1.6, and 3.2 mM concentration of dexamethasone and diclofenac for 4, 24, and 48 hours. MTT based calorimetric assay, flow cytometric analysis, fluorescent micrograph, inverted phase-contrast micrograph, and electron microscopy were used to evaluate the results. RESULTS: The inhibitory effect of human keratocyte proliferation increased at higher concentrations and longer exposure times of dexamethasone and diclofenac (p<0.05). In flow cytometry, the maximal apoptotic response developed at 0.4 mM concentration of dexamethasone and 1.6 mM concentration of diclofenac after 4 hours. Apoptotic cells were demonstrated in fluorescent micrograph. Dexamethasone-treated cases showed a more damaged appearance, more swollen rather than spindle shaped, with greater detachment from the bottom of the dish and the chromatin of the nuclear remnant condensed along the nuclear periphery with cytoplasmic vesication and cytoplasmic blebs formation, and partial disruption of the nuclear membrane compared with diclofenac. CONCLUSIONS: The apoptotic response of dexamethasone and diclofenac is associated with the inhibitory effect of human corneal keratocyte proliferation. For inhibition of cellular proliferation of human corneal keratocytes, dexamethasone may be more effective at lower concentration and shorter exposure time than diclofenac.