Fig. 4: The synergism of the HPA-12 and Creno combination induces pro-cell death Cer generation.

a, b Stacked bar chart of the relative abundance statistics of ceramide determined by LC‒MS. n = 3 independent experiments. a MV4-11 cells were treated with HPA-12 (80 µM) plus Creno (6 µM) (red bars) or vehicle control (0.1% DMSO; blue bars) for 48 h. b MV4-11/CERT-KD cells were treated with Creno (6 µM) (red bars) for 48 h. Transmission electron microscopy images of the endoplasmic reticulum (c) and lipid accumulation (d) in treated MV4-11 cells. The cells were treated as described in (a, b) and harvested for TEM sample preparation following 48 h of treatment. Blue arrows indicate flat vesicular structures representing the endoplasmic reticulum. The purple arrows indicate vacuole-like transparent lipid droplets. Scale bar, 2 μm. e Representative immunofluorescence images of ceramide distribution in the ER (PDI, green) and Golgi. Scale bar, 50 μm. f Statistical analysis of the green MFI representing the ceramide in (e). MFI, mean fluorescence intensity. g Statistical analysis of the colocalization of ceramide (green) with the Golgi (red). h Statistical analysis of the colocalization of ceramide (green) with the ER (yellow). One field per slide and three slides per group were chosen for analysis with ImageJ and subsequent statistical tests. n = 3 independent experiments. The data are presented as the means ± SDs, and differences were compared by 2-tailed Student’s t tests. Multiple groups were analyzed with one-way ANOVA. *P < 0.05; **P < 0.01; ***P < 0.001. Source data are provided as a source data file.