Fig. 2

miR-941 overexpression protects human endometrial cells from OGDR. Stable T-HESC cells with lentiviral pre-microRNA-941 (“lv-pre-miR-941”, two lines) or the lentiviral non-sense microRNA (“lv-miRC”) were subjected to OGD exposure for 4 h, followed by re-oxygenation (“OGDR”) for applied time periods, ROS production (superoxide and lipid peroxidation contents, a), mitochondrial CypD-ANT1-p53 association (tested by mito-IP assay, b) as well as mitochondrial depolarization (JC-1 green fluorescence accumulation, c) and cytochrome C release (d, testing cytosol proteins) were tested; Cell survival and necrosis were tested by CCK-8 assay (e) and LDH release assay (f), respectively. T-HESC cells were transfected with 500 nM of non-sense microRNA control (“miRC”), the wild-type (“WT”) or the mutant miR-941 mimics for 48 h, followed by the same OGDR stimulation for another 24 h, cell survival (g) and necrosis (h) were tested. The primary human endometrial cells, with lv-pre-miR-941 or lv-miRC, were subjected to the same OGDR procedure, ROS production (superoxide contents, i), mitochondrial depolarization (j) and cytosol cytochrome C release (k) were tested similarly, with cell necrosis tested by LDH release assay (l). For mito-IP assay, CypD-bound ANT1 and p53 were quantified (b), with total levels of CypD, ANT1 and p53 tested as the “Input” control (b). For the cytochrome C release assay, relative cytosol cytochrome C level was quantified (d and k). Data were presented as mean ± SD (n = 5), and results were normalized. “Mock” stands for non-OGDR treatment (same for all Figures). * P < 0.05 vs. “Mock” treatment in “lv-miRC”/“miRC” cells. ^#P < 0.05 vs. OGDR treatment in “lv-miRC”/“miRC” cells. Experiments in this figure were repeated five times with similar results obtained. Bar = 50 μm (c and j)