Abstract
The aim of the present study was to evaluate the effect of fluoxetine on activation of the mitogen-activated protein kinase (MAPK) signaling pathway and the expression of apoptosis-associated factors in human conjunctival epithelial cells (HConEpiCs) in culture. HConEpiCs were isolated, cultured and characterized by immunostaining. HConEpiC cells at passage 3-4 were cultured with fluoxetine at different dosages (0, 1, 2.5, 5, 10, 20 and 40 µM) and proliferation rates were determined using a Cell Counting Kit-8 assay. Subsequently, Transwell assays were performed to evaluate the effect of fluoxetine (5 µM) on the invasion and migration capacities of HConEpiCs. ERK1/2 and phosphorylated (p-)ERK1/2 levels were also evaluated in control and fluoxetine-treated groups of HConEpiCs via immunostaining. Finally, western blot assays were performed to evaluate the intracellular protein levels of ERK, p-ERK, Bcl-2, Bax and matrix metalloproteinases (MMPs) in HConEpiCs. It was identified that, as the fluoxetine concentration increased, proliferation rates of HConEpiCs gradually decreased and 5 µΜ fluoxetine was selected for further evaluation. The results of Transwell assays indicated that fluoxetine treatment significantly repressed cell migration and invasion. Immunostaining suggested that there was no significant difference in fluorescence intensity of ERK1/2 between the control and fluoxetine-treated groups, while p-ERK1/2 was significantly enhanced in the fluoxetine-treated group. This result indicated that fluoxetine promoted ERK1/2 activation without affecting its expression. Similarly, western blot analysis revealed no significant difference in ERK1/2 and MMP levels between fluoxetine-treated and control groups, but p-ERK1/2 and Bax were upregulated and Bcl-2 was decreased in the fluoxetine-treated group. In conclusion, fluoxetine induces apoptosis of HConEpiCs in culture via activating the MAPK-ERK1/2 signaling pathway.