Research Paper Volume 12, Issue 3 pp 3010—3024

microRNA-1203 targets and silences cyclophilin D to protect human endometrial cells from oxygen and glucose deprivation-re-oxygenation

Figure 4. miR-1203 inhibition can exacerbate OGDR-induced cytotoxicity in human endometrial cells. Stable T-HESC cells with the pre-miR-1203 anti-sense lentivirus (“lv-antagomiR-1203-L1/L2”, two lines) or the microRNA anti-sense control lentivirus (“anta-C”) were subjected to OGDR for applied time periods, ROS production (DCF-DA intensity, (A) mitochondrial depolarization (JC-1 green fluorescence accumulation, (B) cytochrome C release (C) testing cytosol proteins) were tested by the assays mentioned in the text; Cell survival and necrosis were tested by CCK-8 and medium LDH release assays (D). Stable T-HESC cells with the pre-miR-1203 anti-sense lentivirus (“lv-antagomiR-1203-L1”) were pretreated with cyclosporin A (CsA, 10 μM) for 1h, followed by the OGDR stimulation for 24h, cell viability and necrosis were tested similarly (E). The primary human endometrial cells were infected with lv-antagomiR-1203 or anta-C lentivirus for 48h, followed by OGDR procedure for the applied time periods, ROS production (F), mitochondrial depolarization (G), cytochrome C release (H, testing cytosol proteins) and cell necrosis (I) were tested. For the cytochrome C release assay, relative cytosol cytochrome C level (vs. Tubulin) was quantified (C and H). Data were presented as mean ± SD (n=5). * P <0.05 vs. “Mock” treatment in “anta-C” cells. #P <0.05 vs. OGDR treatment in “anta-C” cells. $P <0.05 (E). Experiments in this figure were repeated five times with similar results obtained. Bar= 50 μm (B and G).