Emerging functional crosstalk between the Grx system and Nrf2 pathway: evidence from UV radiation-induced cataract formation

Glaucoma, cataracts, age-related macular degeneration (AMD), are linked to oxidative stress by the external and internal environment. Ocular tissues are more susceptible to oxidative stress due to daily exposure of UV light and high oxygen consumption. The human body has several antioxidant enzymes such as catalase, superoxide dismutase (SOD), and thioredoxin. Nuclear factor erythroid 2-related factor 2 (Nrf2) is an antioxidant enzyme transcription factor that regulates the downstream antioxidant genes. It also has glutathione (GSH) that searches for the free radicals in our body and oxidizes to form glutathione mixed disulfide (GSSG). As the GSSG levels increase, it naturally adds to other proteins causing protein glutathionylation (PSSG). PSSG is an important post-translational modification linked to oxidative stress. Research has shown that the glutaredoxin (Grx) system is capable of reversing PSSG formation which can be assumed to cause less oxidative stress. To take a closer look at the function of the Grx system in protecting the lens against ultraviolet (UV)- induced cataract formation, glutaredoxin (Grx1) and glutaredoxin 2 (Grx2) is studied in a Grx1/Grx2 double knockout (DKO) mice model. By intercrossing Grx1 knockout (KO) and Grx2 KO mice, Grx1/Grx2 DKO mice resulted. The study population was half male and half female, one month old Grx1/Grx2 DKO and age-matched wild type (WT) mice. They were exposed to 20.6 kJ/m2 UV radiation for 15 mins to induce cataracts. Mice were euthanized at 4 days post-exposure. The degree of the cataract and lens morphology were evaluated under a dissecting microscope. Glutathione (GSH), free protein thiol (PSH), and protein glutathionylation (PSSG) levels were measured as general markers of oxidative damage. To further define the crosstalk between the Grx system and nuclear factor erythroid 2-related factor 2 (Nrf2) antioxidant pathway, Nrf2 and its downstream target proteins were examined by using Western blot analysis. The results showed that UV radiation caused more severe anterior subcapsular cataract in Grx1/Grx2 DKO than that of WT mice. The opacity of the lenses in DKO mice, appeared to extend deeper into the cortical and even nuclear regions. Lenses of Grx1/Grx2 DKO mice contained significant lower levels of GSH and PSH. On the other hand, the accumulation of PSSG, a marker for protein thiol oxidation, was much higher in Grx1/Grx2 DKO group. Deletion of Grx1 and Grx2 also decreased the expression of antioxidant enzyme transcription factor regulator, Nrf2, and its downstream antioxidant genes, including catalase, superoxide dismutase (SOD), and another redox regulator of thioredoxin (Trx). The Nrf2 dependent antioxidant response can no longer function with combined Grx1 and Grx2 deletion. This will cause more oxidative stress and increase the lens susceptibility to UV-induced damage.