To evaluate the expression levels of miR-23a in RPE cells from the macula of AMD and normal age-matched donor eyes, qRT-PCR was used. miR-23a was significantly downregulated in AMD compared with normal (
Fig. 1A). The results were confirmed by Northern blot analysis (
Fig. 1B). Collectively, aberrant expression of miR-23a was a remarkable characteristic in the RPE cells of human AMD, suggesting the possibility that it could function as a mediator in the process of AMD development.
N-(4-hydoxyphenyl)-retinamide (4HPR), a retinoic acid, has been reported to induce ROS generation and increase the expression level of miR-23a in ARPE-19 cells.
27 Our results showed that the expression level of miR-23a in primary cultured RPE cells increased when exposed to H
2O
2 at 100 to 200 μM, but decreased at 300 to 500 μM (
Fig. 1C). The same pattern was followed in ARPE-19 cells (
Fig. 1D). Previous studies suggest that 200 μM H
2O
2 does not affect ARPE-19 cell viability.
28,29 Persistent elevation of phosphorylated-Akt induced by 200 μM H
2O
2 may enhance the ARPE cell's ability to resist the damaging effects of low level oxidative stress.
28 Preconditioning the cells by low dose of H
2O
2 induces resistance to killing,
30 while high doses of H
2O
2 produced toxicity through oxidative stress.
31 Different enzyme activities or signaling pathways are triggered in cells treated with different doses of H
2O
2.
30,31 Our results suggest that H
2O
2 causes differential expression of miR-23a based on the dose. The increased level of miR-23a induced by 200 μM in our study indicated that miR-23a may also play a role in cell survival. However, higher concentrations of H
2O
2 may induce destruction of RPE cells. The combined effect of apoptosis and necrosis may decrease the level of miR-23a.