A. ALDH2 O-GlcNAcylation in Con, Con+PUGNAc and Con+DON, and HG, HG+PUGNAc and HG+DON groups in vitro. Cell lysates were immunoprecipitated with specific anti-ALDH2 antibody and then subjected to western blot using anti-O-GlcNAc (CDT110.6) antibody. B. ALDH2 dehydrogenase activity in Con, Con+PUGNAc and Con+DON, and HG, HG+PUGNAc and HG+DON groups in vitro. ALDH2 activity was determined by measuring the conversion of NAD+ to NADH. C. ALDH2 O-GlcNAcylation in Con+H/R, Con+PUGNAc+H/R and Con+DON+H/R, and HG+H/R, HG+PUGNAc+H/R and HG+DON+H/R groups in vitro. D. ALDH2 dehydrogenase activity in Con+H/R, Con+PUGNAc+H/R and Con+DON+H/R, and HG+H/R, HG+PUGNAc+H/R and HG+DON+H/R groups in vitro. Mannitol was used to keep the osmolarity consistent among different glucose concentrations in vitro. In each graph, the mean value of Con or Con+H/R was expressed as 1 or 100%. IP, immunoprecipitation; IB, immunoblotting; Con: normal glucose; HG: high glucose; PUGNAc: a specific O-GlcNAc modification enhancer; DON: a specific O-GlcNAc modification inhibitor; H/R: hypoxia/reoxygenation. Data are means ± SEM of 4 to 5 samples in each group. *P<0.05 vs Con group, †P<0.05 vs HG group in normal oxygen conditions or H/R conditions.