Figure 3. MiR-145 inhibited H₂O₂-induced apoptosis in cultured NRVMs.

For both DNA ladder ELISA and TUNEL assay, cultured NRVMs were transduced with Ad-LacZ or Ad-miR-145 (MOI = 100) and transfected with either 0.5 µg Bnip3 CDS or Bnip3 CDS/3′UTR or rAd-psilence empty vector per 10⁶ cells; and then transfected with 10 nM miR-145 inhibitor or control inhibitor per 10⁶ cells. Cells were treated with vehicle or 50 µM H₂O₂ for 2 hours. In DNA ladder ELISA (A), apoptosis of cardiomyocytes in different group was evaluated by calculating A405 nm-A490 nm values. The result shows that miR-145 protected against H₂O₂-induced cardiomyocytes apoptosis, which was significantly enhanced by Bnip3. In TUNEL staining assay (B), the percentage of cell apoptosis was calculated by the following formula: number of TUNEL (red)-positive cells/number of DAPI (blue)-positive cells. The rates were calculated in 10 randomly chosen fields from each group. Scale bar indicates 50 µm. C, the quantitative analysis of cardiomyocyte apoptosis was determined by TUNEL staining assay. Data are presented as means ± SEM. Significance is indicated as *P<0.05 and **P<0.01, as determined by student’s t test.