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Effect of ERK inhibition on H 2O 2induced cytotoxicity in ARPE-19
H 2 O 2 induced cytotoxicity in RPE cells appears to be affected by the density of cells in cultures [ 29 ] . To study this phenomenon, ARPE-19 cells plated in 96-well plates with densities of 10,000, 20,000 or 40,000 cells/well were subjected to H 2 O 2 treatment.
Since ERK activation appeared to be an early event associated with H 2 O 2 induced cytotoxicity, experiments were performed to determine if inhibitors of ERK activation could affect the cytotoxicity caused by H 2 O 2 . ARPE-19 cells were pretreated with 10 μM PD98059 (a MEK inhibitor which functions upstream of ERK [ 33 ] ) followed by H 2 O 2 challenge.
To analyze morphological changes of RPE cells treated with a cytotoxic concentration of H 2 O 2 , ARPE-19 cells were treated with 1.5 mM H 2 O 2 for various periods of time and then processed for actin staining.
Though the H 2 O 2 titration curve in ARPE-19 was not as steep as observed in primary RPE, one-day treatment of these cells with 1.8 mM H 2 O 2 could decrease the cellular viability to ~10% of control while 1 mM H 2 O 2 did not affect cellular viability (Fig.