Fig. 5

NOX4 activation is required for ASMase-mediated cardiac hypertrophy and apoptosis. a–b Representative Western blot images and quantitative analysis of protein expression of ASMase, NOX2, and NOX4 in six groups of H9c2 cells treated with vehicle, glucose + PA, ceramide, apocynin, apocynin + glucose + PA, apocynin + ceramide, respectively. β-actin was used as a loading control. Three independent experiments were performed to calculate the means. c Quantitative analysis of NOX1, NOX2 and NOX4 mRNA expression by qPCR in H9c2 cardiomyocytes. Five independent experiments were performed to calculate the means. d Representative fluorescence images of rhodamine-phalloidin staining to visualize cell size, scale bar:100 μm. e Quantitative analysis of pro-hypertrophic markers, Nppb and Myh7 mRNA expression by qPCR in H9c2 cardiomyocytes. Five independent experiments were performed to calculate the means. f–g Representative fluorescence images and quantitative analysis of DCFH-DA staining for ROS measurement in H9c2 cells, scale bar:100 μm. Three independent experiments were performed to calculate the means. h Representative Western blot images of protein expression of cleaved caspase-3, β-actin was used as a loading control. i–j Representative fluorescence images and quantitative analysis of TUNEL staining, scale bar:100 μm. Five independent experiments were performed to calculate the means. Data were presented as mean ± SEM, n = 3. ^*P < 0.05, ^**P < 0.01, ^***P < 0.001, ^****P < 0.0001