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Apoptosis induced by ceramide in hepatocellular carcinoma Bel7402 cells
1Department of Therapeutic Basis, Cancer Center, Sun Yat-sen University of Medical Sciences, Guangzhou 510060, China. xfzhu@gzsums.edu.cn
Aim:
To study the biological function of ceramide signaling in Bel7402 cells.
Methods:
Inhibition of cell growth was assayed using MTT method. Morphologic assessment of apoptosis was performed with fluorescence microscope. DNA fragmentation was detected by electrophoresis and flow cytometry. The levels of protein p53, Bcl-2, and Bax were measured with Western blot.
Results:
Bel7402 cells treated with C2-ceramide underwent cell proliferation inhibition. IC50 value was 14.28 mumol.L-1. After treatment of Bel7402 with ceramide, the morphologic changes including reduction in volume, nuclear chromatin condensation, fluorescence strength were observed. SubG1 peaks were detected on flow cytometry (FCM). Agarose gel electrophoresis of DNA from cells treated with ceramide revealed "ladder" pattern. The Western blot assay from cell extracts showed that the levels of protein p53 were decreased after ceramide treatment. The levels of protein Bcl-2 were decreased also. But the levels of Bax protein showed no difference between untreated cells and treated cells.
Conclusion:
Ceramide induces apoptosis in Bel7402 cells, related to Bcl-2 down-regulation.
Insights
Ceramide triggers apoptosis in Bel7402 cells by inhibiting proliferation and altering cell morphology. This process is linked to decreased Bcl-2 protein levels, indicating its role in programmed cell death.
Area of Science:
- Cell biology
- Molecular signaling
Background:
- Ceramide signaling plays a crucial role in cellular processes.
- Understanding ceramide's function in cancer cells is vital for therapeutic development.
Purpose of the Study:
- To investigate the biological role of ceramide signaling in Bel7402 cells.
- To elucidate the mechanisms by which ceramide affects Bel7402 cell viability and apoptosis.
Main Methods:
- Cell proliferation was assessed using the MTT assay.
- Apoptosis was evaluated through morphological analysis, DNA fragmentation detection (electrophoresis, flow cytometry), and Western blot analysis of key proteins (p53, Bcl-2, Bax).
Main Results:
- Ceramide treatment inhibited Bel7402 cell proliferation with an IC50 of 14.28 μmol/L.
- Morphological changes indicative of apoptosis, such as cell shrinkage and nuclear condensation, were observed.
- DNA fragmentation and SubG1 peaks in flow cytometry confirmed apoptosis induction.
- Western blot analysis revealed decreased levels of p53 and Bcl-2, while Bax levels remained unchanged.
Conclusions:
- Ceramide induces apoptosis in Bel7402 cells.
- The observed apoptosis is associated with the down-regulation of Bcl-2 protein.