Methionine sulfoxide reductase A: Structure, function and role in ocular pathology

Figure 1 Scheme depicting the role of methionine sulfoxide reductase A and α-crystallin in methionine metabolism. Note the reversible nature of oxidation-reduction of methionine and its influence on the chaperone properties of α-crystallin. The inhibition of the apoptotic events from oxidative stress by αB crystallin is also presented in the figure. ROS: Reactive oxygen species.

Figure 2 Immunohistochemical localization of αA-crystallin (red), αB-crystallin (red) and methionine sulfoxide reductase A (green) in the retina of patients with AMD. Retinal cryosections were air-dried, fixed, and processed as described[18] using αA-crystallin and αB-crystallin rabbit polyclonal antibodies (Stressgen, Ann Arbor, MI, USA), and mouse monoclonal methionine sulfoxide reductase (Msr) A antibody (Novus Biologicals, Littleton, CO, USA). Sections were viewed under a confocal microscope (Carl Zeiss, Thornwood, NY, USA). Arrows indicate co-localization (yellow) of αA-crystallin or αB-crystallin and MsrA. AMD:Age-related macular degeneration; *: Drusen; Scale bar = 50 μm.

Figure 3 Regulation of methionine sulfoxide reductase A and B2 in retinal pigment epithelial subjected to chemically induced hypoxia. Hypoxia-inducible factor (HIF)-1α (A) and vascular endothelial growth factor (VEGF) (B and C) were used to validate hypoxia from CoCl₂. Confluent human fetal retinal pigment epithelial (RPE) cells were serum starved overnight and treated with 100 μmol/L CoCl₂ for 4 h. Western blot analysis shows HIF-1α (mouse monoclonal, Novus Biologicals) upregulation in nuclear extracts of CoCl₂-treated RPE cells (A). VEGF expression was significantly upregulated both at the mRNA (B) and protein levels (C). Real-time polymerase chain reaction was performed as described[19] in a light cycler (Roche, IN, USA) using β-actin as normalizing gene. A polyclonal antibody (Santa Cruz Biotechnology, Santa Cruz, CA, USA) was used for the detection of VEGF. Methionine sulfoxide reductase (Msr) A and MsrB showed initial upregulation but, at 4 h, showed significant downregulation (D and E). Data presented are mean ± SE. ^aP < 0.05 vs control.