Cigarette smoke oxidant, hydroquinone, downregulates p62 and Nrf2 protein expressions inhuman RPE and induces cell death

Cigarette smoking is a major risk factor for age-related macular degeneration. The cigarette smoke oxidant, hydroquinone (HQ), has been implicated in the death of RPE, an important process that could lead to retinal degeneration. However, the molecular mechanism associated with HQ-induced retinal degeneration is not well understood. Normally under conditions of oxidative stress, there is an activation of the p62-Nrf2 protein expressions for protection and cell survival. It was, therefore, hypothesized that HQ-induced RPE damage was related to an inhibition of the p62-Nrf2 pathway. To evaluate mechanisms related to HQ-induced injury in the retina, human ARPE-19 cells were cultured and exposed to varying concentrations of HQ (from 0 - 90 µM). Trypan blue assay was performed to assess cell viability in the ARPE-19 cells following treatment with HQ for 4 hours. Cell lysates from the ARPE-19 cells treated with sub-lethal doses of HQ (0-75µM) for 2 hrs were prepared and p62 and Nrf2 protein expressions determined by Western blotting. Also, the level of oxidative damage in cell lysates was quantified by ELISA following. One-way ANOVA was used to determine differences in effect among the varying doses of treatments on ARPE-19 cells. Viability of the human ARPE-19 cells was significantly reduced following treatment with increasing concentrations of HQ. ARPE-19 cells treated with the highest concentration of 90 µMHQ had the least cell viability percentage (15.0 ± 2.1; P <0.001), showing that cell death by HQ treatment was dose-dependent. Densitometric analyses of Western blot results demonstrated that HQ treatment resulted in significant downregulation of p62 and Nrf2 protein expressions dose-dependently (P < 0.5 - 0.001). Also, HQ treatment resulted in a dose-dependent increase in the protein carbonyl level, an indication of oxidative stress. Human retinal pigment epithelium death due to hydroquinone is associated with oxidative damage and downregulation of the p62-Nrf2 signaling pathway. Treatments targeting the activation of p62-Nrf2 pathway in the RPE might prove effective in the management of age-related macular degeneration. This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.