Figure 5 | Cell Death & Disease

Figure 5

From: Essential role of PH domain and leucine-rich repeat protein phosphatase 2 in Nrf2 suppression via modulation of Akt/GSK3β/Fyn kinase axis during oxidative hepatocellular toxicity

Figure 5

PHLPP2-silencing restores Nrf2 signaling by promoting Akt-induced Fyn kinase deactivation during tBHP exposure. Normal and PHLPP2-silenced hepatocytes were challenged with 250 μM tBHP for 90 min. (a) Shows immunoblot detection of key proteins involved in Nrf2 and Akt pathway. (b) Graph representing change in ratio of phosphorylated/total Akt and Fyn kinase in normal and PHLPP2-silenced hepatocytes treated with tBHP. (c) Western blotting images of PHLPP2, pAkt(Ser473), Nrf2 and Fyn kinase in nuclear and cytosolic extracts. β-Actin was used as an endogenous control to normalize protein expression in cytosolic extracts while lamin b was used as a reference protein for nuclear extract. (d) Immunofluorescence staining of hepatocytes for Nrf2 (green) and Hoechst (blue) illustrating nuclear translocation of Nrf2 (magnification × 40). (e) Levels of ubiquitinated Nrf2 were analyzed by immunoprecipitating Nrf2 protein followed by western blotting detection with anti-ubiquitin antibody. (f) Estimation of enzyme-linked immunosorbent assay-based Nrf2-ARE binding affinity in nuclear lysates obtained from normal and PHLPP2-silenced hepatocytes treated with tBHP. The data are presented as mean±S.E. of at least three independent experiments. *P<0.05 compared with control; #P<0.05 compared with tBHP

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