ALDH2 knockdown blocks isoflurane-induced protection against hypoxia/reoxygenation.

<p>A. Schematic representation of adenoviruses encoding RFP (Ad-RFP) and ALDH2shRNA (Ad-Si-ALDH2-RFP, left panel); right panel, immunoblotting analysis of ALDH2 protein level. β-actin was used as a control. B. ALDH2 down-regulation inhibited attenuation of TUNEL positive staining level by isoflurane. (a) Apoptotic cells were examined using TUNEL assay for DNA fragmentation. Cardiomyoctes were photographed by fluorescence microscopy after 24 hours of hypoxia followed by 12 hours of reoxygenation. TUNEL-positive nuclei are shown in green and transfection efficiency of adenoviruses in red. (b) Quantitative analysis (percentage of apoptotic cells versus total) is shown in histogram. C. Representative Western blots of cleaved caspase-3 and full length caspase-3 (FL caspase-3) in (a), and 3 independent experiments were quantitated in (b). β-actin was used as a loading control. D. Cell lysates under each condition were quantitatively assayed for caspase-3 activity. E. LDH concentrations in cell culture media were analyzed. Values are means ± S.E.M., n = 5 in each group. *P<0.05, **P<0.01 vs. Control group, and ^#P<0.05, vs. the corresponding Ad-RFP group.