Figure 10.

Hyd, when instilled topically in mouse eyes in vivo, enhanced Nrf2/ARE-mediated antioxidant genes. (A) Three groups of 16 M old C57BL/6 mice (each group; n = 4, 8 lenses) were used for the experiment. Hyd or buffered saline was instilled topically once daily for 7 days and 15 days as follows: Group 1, 2, and 3, vehicle control (left eye, buffered saline, pH 7.2) and Group 1, 2, and 3, Hyd treated (right eye, 25 µM/5 µL or 50 µM/5 µL or 100 µM/5 µL each eye, respectively). On day 8 or 16, mice were sacrificed, and total RNA (7 or 15-days treatment) and protein (15-days treatment) were isolated from the lenses. RT-qPCR (B–G) and immunoblotting (H) analyses were performed, and data were presented. (B–G) Hyd treatment significantly increased the expression of Nrf2 (B), and Nrf2 target antioxidant genes (C–G), as shown. The data presented as mean ± S.D. values obtained from three independent experiments. Control vs. Hyd-treated, * p < 0.05; ** p < 0.001. ns, not significant. (H) An equal amount of protein was loaded onto SDS-PAGE gel and immunoblotted with Nrf2, Prdx6, and β-actin antibodies, as shown. Nrf2 and Prdx6 protein bands were scanned and quantified, and the obtained values were normalized with corresponding β-actin band values. Relative densities were presented as histograms below the protein bands. * p < 0.05; ** p < 0.001.