Skip to main content
Figure 6 | Molecular Cancer

Figure 6

From: Withanolide D induces apoptosis in leukemia by targeting the activation of neutral sphingomyelinase-ceramide cascade mediated by synergistic activation of c-Jun N-terminal kinase and p38 mitogen-activated protein kinase

Figure 6

Effect of Fumonisin B1, (GW4869) and N-SMase silencing on ceramide production, activation of MKK group of kinases and regulation of WithaD-mediated apoptosis. (A) Effect of Fumonisin B1 (10 μM) and GW4869 (10 μM) on ceramide level induced by WithaD. Cells were pretreated with inhibitors for 1 hr; incubated further 1 hr with WithaD and ceramide levels were measured by FACS. Each column represented the mean ± S.D. in duplicate in three independent experiments. *indicates statistically significant difference (P < 0.005). (B) Effect of SMPD3 silencing in WithaD mediated ceramide accumulation. Cells were transfected with SMPD3 specific siRNAs as described in materials and methods. Ceramide level were measured by DAG kinase assay as described in Materials and methods. Results are the mean ± S.D. of two independent experiments. * indicates statistical significant difference P < 0.05. (C) and (D) Consequence of N-SMase inhibition by GW4869 and N-SMase silencing by siRNA1 on phosphorylation of MKK4, MKK3/6 and MKK7. K562 and MOLT-4 cells were pretreated with GW4869 (10 μM) for 1 hr and further incubated for 2 hr with 3 μM and 1 μM WithaD respectively. (E) Effect of Fumonisin B1 (10 μM) and GW4869 (10 μM) on cell death. Cells were pretreated separately with the inhibitors for 1 hr and after 48 hr apoptosis were measured. Each column represented the mean ± S.D. in duplicate in three independent experiments. * indicates statistically significant difference (P < 0.005). ** indicates difference (p = 0.1250). Etoposide was used as positive control. (F) Effect of SMPD3 silencing in WithaD treated cells. After 48 hr of transfection, apoptosis was measured. Results are representative of three independent experiments. * indicates statistical significant difference, where P < 0.05.

Back to article page