A2E Selectively Induces COX-2 in ARPE-19 and Human Neural Cells

Abstract

Purpose: To investigate the expression of cyclooxygenase (COX)-1, -2, and -3 RNA and protein in retinal pigment epithelial (ARPE-19) cells and in human neural (HN) cells exposed to the stress-inducing cytokines IL-1β and TNF-α, the oxidizing peroxide H2O2, the combination of TNF-α + H2O2, and the lipofuscin fluorophore A2E. Methods: Three-week-old ARPE-19 and HN cells were incubated with IL-1β (10 ng/ml), TNF-α (10 ng/ml), H2O2 (0.6 μM), TNF-α + H2O2 (10 ng/ml and 0.6 μM), or A2E (10 μM) for 8 hr, after which total RNA and whole cellular proteins were isolated. Cyclooxygenase-1, -2, and -3 RNA and protein levels were quantified using Northern and Western immunoassay. Results: IL-1β-, H2O2-, TNF-α-, TNF-α + H2O2-, or A2E-stressed ARPE-19 or HN cells displayed no significant upregulation in COX-1 or COX-3 RNA message abundance; however, significant upregulation was observed in COX-2 RNA message and protein abundance. A2E treatment of HN cells resulted in modest increases in COX-3 protein, an effect that was not observed in ARPE-19 cells. Conclusions: COX-2 RNA levels were induced in cytokine-, peroxide-, and A2E-stressed ARPE-19 and HN cells. Lack of induction of COX-3 RNA message by A2E, coupled with increases in COX-3 protein under identical treatment conditions, suggest that significant post-transcriptional or post-translational controls may regulate COX-3 gene expression in HN cells. Stress-induced upregulation of COX-2 gene expression in ARPE-19 and HN cells may play a mechanistic role in promoting proinflammatory and/or pro-oxidative pathology in these tissues.