ABSTRACT
OBJECTIVE: To investigate the apoptosis of the auction and mechanism of the hepatoma BEL-7402 cells induced by the ginseng polysaccharides (GPS). METHODS: The hematoma cells BEL-7402 were incubated with GPS, cell viability was measured by CCK8, cell cycle distribution was assessed by fluorescence-activated cell sorting (FACS), the cell morphological changes were traced with TUNEL and scanning electron microscope, TNFRl, BCL-2 and Bax protein expression and ERK phosphorylation are tested by Western blot. RESULTS: CCK8 results showed that GPS inhibited cells growth of BEL-7402 in dose-dependent and time-dependent. Flow cytometry found that S phase arrest was increased upon GPS concentrations. Obviously apoptotic sub-g peak was also found. And the peak was gradually enhanced with the concentration increased. TUNEL staining and SEM results showed that GPS led to significant cell morphology changes in hepatoma cells BEL-7402. And the apoptosis effect was increased upon the GPS concentrations. Western blot showed that level of apoptosis related protein Bax was increased, the expression of apoptosis-antagonizing protein Bcl-2 was decreased and the expression of death receptor TNFRl appeared with GPS concentrations increased gradually, raise ERK phosphorylation. CONCLUSION: GPS can induce apoptosis of hepatoma cells BEL-7402 by the mitochondrial pathway and the death receptor dependent pathway and ERK pathway.
ABSTRACT
It has been well known that apoptosis induction and cell cycle arrest are typical biological effects observed in cancer cells after proteasome inhibition. TH2 is a new natural xanthone analogue isolated from the resin of Garcinia hurburyi tree. Here, the cell growth inhibition of TH2 on human hepatocellular carcinoma cell line (Bel-7402) was evaluated in vitro using SRB assay. The treatment of 10 ?mol/L TH2 reduced the surviving fraction from 86% (12 h) to 17.2% (48 h). To assess whether TH2 induce apoptosis, the appearance of sub-G1 peak, a specific fraction for apoptosis was detected by flow cytometry analysis. Progressive increase in the percentage of apoptotic population was observed in a dose-and time-dependent manner. Furthermore, a cleavage of poly (ADP-ribose) polymerase (PARP), a marker of early apoptosis, was observed clearly when the cells exposed to 10 ?mol/L of TH2 for 24 h by immunoblotting analysis. In vitro activities of 20 S proteasome purified from human erythrocytes on fluorogenic peptide substrates revealed that TH2 inhibited the trypsin-like, chymotrypsin-like and peptidylglutamyl peptide hydrolyzing activities in dose-dependent manner. Moreover, the turnover of tumor suppressor p53, a sign of deregulation of cell cycle progression and apoptosis induction by classical proteasome inhibitors, was disrupted in Bel-7402 cells. All these data indicate that TH2 had inhibitory effect on the proliferation of Bel-7402 cells and induction of apoptosis, which might be related to its inhibition of proteasome.
ABSTRACT
Aim To investigate the effects of curcumin on expression of HIF-1? in human hepatocellular carcinoma BEL-7402 cells.Methods Different concentrations of 0,2.5,5,10,15 and 20 ?mol?L~(-1) curcumin-treated human hepatocellular carcinoma BEL-7402 cells were cultured under hypoxia for 6 hours.The cell proliferation and cell viability were assayed by WST-8 and the trypan blue dye exclusion method.The expression level of HIF-1? in human hepatocellular carcinoma BEL-7402 cells was checked by RT-PCR and Western Blot;0,10 ?mol?L~(-1) curcumin,10 ?mol?L~(-1) MG-132,and 10 ?mol?L~(-1) curcumin +10 ?mol? L~(-1) MG-132-treated human hepatocellular carcinoma BEL-7402 cells were cultured under hypoxia for 6 hours.The protein expression level of HIF-1? protein in human hepatocellular carcinoma BEL-7402 cells was checked by Western Blot.Results ① No significant difference was found in the cell proliferation rate and cell viability between control and different concentrations of curcumin.② Curcumin can down-regulate the expression of HIF-1? protein with the increasing concentrations.③ the effect of curcumin on expression of HIF-1?mRNA had no significant difference between control and different concentrations.④ MG-132 can reverse the curcumin-mediated decrease of HIF-1? protein.Conclusion Inhibitory effect of curcumin on expression of HIF-1? in human hepatocellular carcinoma BEL-7402 cells was acted through posttranscriptional mechanisms and the proteasome pathway.