Figure 4. Forced activation of Nrf2 by Keap1 KO mimics and nullifies GRh3-induced endometrial cell protection against OGDR. Expression of listed proteins in total cell lysates of stable T-HESC cells with the CRISPR-Cas9-Keap1-KO construct (“ko-Keap1”) or the empty vector (“Vector”) was shown (A). Cells were pretreated with GRh3 (10 μM) for 2h, followed by OGD (4h)-reoxygenation (“OGDR”) for applied time periods, then mitochondrial depolarization (JC-1 green intensity, B) and ROS production (superoxide contents, (C) were tested, with cell viability and necrosis examined by CCK-8 (D) and LDH release (E) assays respectively. Expression of the listed proteins was quantified, after normalizing to the loading control protein (A). Error bars stand for mean ± standard deviation (SD, n=5). #p<0.05. Each experiment was repeated three times with similar results obtained. Bar=100 μm (B).