Abstract
Purpose: :
To evaluate the expression and protein levels of antioxidant enzymes in the rat retina exposed to oxidative stress induced by ischemia–reperfusion injury.
Methods: :
Retinal ischemia was induced in female Wistar rats by ligation of the optic nerve and vessels behind the left eye bulb, and was followed by reperfusion for 0, 3, 6, or 24 hours. The right eye served as control. RNA and protein were extracted simultaneously from each retina. Expressions of the endogenous antioxidant enzymes GPx1, CAT, CuZnSOD, MnSOD, and GCLc were analyzed with Real Time RT–PCR and related to the endogenous control Cyp B. Protein levels were measured with Western blot.
Results: :
During the early phase (0 or 3 hours) of reperfusion no changes were seen in enzyme expression. After 6 hours, GCLc expression increased by a factor of 1.14 (p=0.034), followed by a decline of 0.80 after 24 hours (p=0.00004), according to the comparative Ct–method. After 24 hours of reperfusion, GPx1 expression increased by a factor of 1.14 (p=0.028), and CAT had decreased by 0.82 (p=0.022). Expressions of CuZnSOD and MnSOD showed a tendency towards a decrease by factors of 0.86 (p=0.055) and 0.88 (p=0.053), respectively after 24 hours. Protein levels did not differ for any of the antioxidants, regardless of reperfusion time.
Conclusions: :
The slightly increased mRNA expression of GPx1 after 24 hours of reperfusion with a concomitant very modest decrease in CAT and GCLc expression and no change in protein levels indicate a very modest, if any, response to oxidative stress generated by ischemia followed by reperfusion in rat retina.
Keywords: ischemia • oxidation/oxidative or free radical damage • retina