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Sebastian Di Cesare, Tamara J. Granner, Matthew Balazsi, Luca A. Petruccelli, Bruno F. Fernandes, Miguel N. Burnier, Jr.; The Characterization of General Cell Stress Response Signals in the ARPE-19 Cell Line when Treated with Various Concentrations of Blue Light. Invest. Ophthalmol. Vis. Sci. 2011;52(14):2321. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
Age Related Macular Degeneration (AMD) is a leading cause of central visual loss in patients >50 years of age. The primary cause of AMD is known to be attributed to the atrophying and environmental damaging effects of retinal pigment epithelial cells. We sought to quantify and characterize specific cell stress responses that may occur in a retinal pigment epithelial cell line (ARPE-19) when treated with various concentrations of blue light.
The APRE-19 cell line was exposed to blue light at varying concentrations (5000 lux, 1000 lux, 500 lux, 100 lux) for 2 hours in order to determine a sub-lethal concentration of blue light. Cells covered by aluminum foil were used as a control. A set of cells (3.0x106) was immediately harvested post-exposure while the second set was allowed to recover overnight (12h) in an incubator (37oC, 5% CO2). The standard method protocol for propidium iodide staining was used in order to process the cells and the percentage of apoptotic cell bodies within each sample population was determined via computer analysis software (FlowJo). This experiment was performed in triplicate. Once the sub-lethal dose of blue light was determined, we proceeded to treat with the optimal concentration of blue light and extract RNA. The RNA was later processed for an RT2 ProfilerTM PCR Array (SA Biosciences) that determines pathway-focused gene expression profiling for specific cellular stress and toxicity genes (84 genes). This experiment was performed in duplicate.
There was no significant apoptosis detected in any of the blue light doses after 2h post-treatment or the 12h recovery phase(<2.5% of cells, Sub-Go). Although, there was an average significant increase in cell cycle arrest in cells treated with blue light (52.40% of cells, G2-M) when compared to the control (35.84% of cells, G2-M) in both treatment groups. Due to these results we chose the sub-lethal dose of blue light to be 5000 lux. The PCR array revealed a general up-regulation (>2-fold) of most cellular stress and toxicity genes (24 genes). Interestingly, Cytochrome P450 Reductase (POR) mRNA had the greatest fold change (>3-fold) for all genes tested.
To the best of our knowledge this is the first study to profile ARPE-19 cells for cellular stress response genes when treated with a sub-lethal concentration of blue light. This study clearly demonstrates the short-term cellular stress and toxicity effects blue light has on the ARPE-19 cell line.
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