FIG 4.

The E2 protein is involved in cell viability. (A) OL and HeLa cells were transfected with antisense oligonucleotides (ASOs) against hsEBLN-2 RNA (ASO 1 and 2) and control ASO (Ctrl), and relative levels of hsEBLN-2 RNA were quantified by real-time reverse transcription-PCR (RT-PCR). Values are presented as the mean ± standard error (SE) of three independent experiments. (B and C) OL and HeLa cells were transfected with each ASO, and PARP and caspase-3 cleavages were detected with an anti-PARP antibody and an anti-caspase-3 antibody. Relative expression levels of cleaved proteins were calculated by the band intensity based on Western blotting. Values are presented as the mean ± SE of three independent experiments. Statistical significance was analyzed by two-tailed t test. *, P < 0.05’ **, P < 0.01. (D) Knockdown of hsEBLN-2 RNA reduces cell viability. OL and HeLa cells were transfected with ASOs, and cell viability was measured by the WST-1 assay. Values are presented as the mean ± SE of three independent experiments. Statistical significance was analyzed by two-tailed t test. *, P < 0.05; **, P < 0.01. (E) E2 overexpression increases stress resistance. OL and HeLa cells were transfected with E2 expression (E2) and control empty (Ctrl) plasmids, and 24 h after transfection, the cells were treated with 400 μM H₂O₂ for 24 h. Cell viability was measured by the WST-1 assay. Values are presented as the mean ± SE of six or three independent experiments. Statistical significance was analyzed by two-tailed t test. *, P < 0.05. (F) OL cells were transfected with E2 or control empty plasmid (Ctrl) and treated with 100 μg/ml cycloheximide (CHX) for 2, 3, and 4 h. Expression levels of HAX-1 and tubulin were detected by Western blotting, and band intensities were measured by ImageJ. Values are presented as the mean ± SE of three independent experiments.