Ursolic Acid Inhibits Proliferation And Induces Apoptosis In Human Gastric Carcinoma Cell Line BGC-823

Objective: To investigate whether ursolic acid can inhibit proliferation of human gastric carcinoma cell line BGC-823 and induce apoptosis, and its primary mechanisms.Methods: The concentrations of UA used in cell culture system were 0, 20, 30, 40, 50, 60 μmol/L and 12, 24, 36, 48 hours were selected as the termination point of cell culture. Inhibition of cell proliferation was determined by MTT assay. Nuclear morphological changes were observed by Hoechst33258 fluorescent staining. Cell cycle and apoptosis rate were analysed by flow cytometry (FCM). Activation of caspase-3 and expression of cyt-c and survivin were detected by Western blotting.Results: MTT assay showed UA had a moderate anti-proliferation effect on BGC-823 cells at the concentration above 20 μmol/L after 24 hours in time-and-dose-dependant manner. The highest dose (60 μmol/L) of UA resulted in almost all inhibition in tumor cell growth after 48 hours and the IR was 85.2%. Low values of IC50 were measured: 58.83 μmol/L for 12 hours, 43.10 μmol/L for 24 hours, 37.67 μmol/L for 36 hours, 31.27 μmol/L for 48 hours. After treatment with UA, nuclei shrunk and the chromosomes were condensed and decomposed into fragments, which showed by intensive fluorescence. After exposure to different concentrations (20, 30, 40, 50, 60 uM) of UA for 24 hours, BGC-823 cells showed G1 phase declined and S phase arrested: cells at S phase were 32.2%, 25.0%, 40.1%, 45.4%, 47.9%, 77.9% in control group and 20, 30, 40, 50, 60 uM-treated groups respectively. What's more we observed the obvious "subG1" peak after treatmemt in DNA frequency histogram. the percentage of apoptotic cells were differently 6.1%, 13.9%, 33.6%, 46.5%, 68.6% in UA-treated groups, while 0.9% in control group (P<0.05). 20~60 μmol/L of UA for 24 hours caused up-regulation of cyt-c protein (15 kD) and down-regulation of survivin protein (16.5 kD) in BGC-823 cells. Compared with the control group, inactive form of caspase-3, which was a 32 kD a protein decreased, while 17 kD and 19 kD active cleaved fragments increased in UA-treated groups, detected by western blotting.Conclusion: Overall, our findings provided powerful evidence that ursolic acid exhibited both anti-proliferation and apoptosis-induced effects on human gastric adenocarcinoma BGC-823 cells. The increased accumulation of cyt-c and the depression of survivin both activated caspase-3 protease, which may mainly contribute to ursolic acid-induced apoptosis in BGC-823 cells.