Mechanism Of Apoptosis Induced By Ursolic Acid In Human Gastric Carcinoma BGC-803 Cells

Object The present study was designed to investigate the anti-tumor effect and apoptosis- inducing mechanism of UA on human gastric carcinoma BGC- 803 cells.Methods Based on the culture of human gastric carcinoma BGC-803 cells in vitro,the effect of ursolic acid on growth of human gastric carcinoma BGC- 803 cells was evaluated by MTT. To observe occurrence of apoptosis in the early, intermediate and advanceed stage , the morphology of apoptosis , DNA fragmentation and phos- phatidylserine (PS) residues were detected by HE staining,agarose gel electropho- resis and ann exin V/PI staining in flow cytometry , respectively. Procaspase- 3, 8, 9, bax and bcl-2 proteins expression obtained by Western blot analysis. Results MTT showed UA had a significant antiproliferative effect on human gastric carcin- oma BGC-803 cells in a dose- and time-dependent manner. The IC50 value of UA at 12 h, 24 h, 36 h and 48 h were 61.29μM, 43.78μM, 35.94μM, and 24.95μM, respec- tively. Oligonucleosomal-sized DNA fragmentation resulted from the continuous exposure of BGC-803 cells to ursolic acid for 24h and 36h. Result of flow cyto- metry showed apoptosis rate was increased after treatment with UA for 36h(F= 72.579 ;P < 0.05). HE staining showed Morpholgical changes characteristic of apoptosis appeared after treatment with UA for 24h and 36h , such as apoptotic bodies and nuclei condensation. Western blot analysis indicated that expression level of procaspase-3, procaspase-8, procaspase-9 and Bcl-2 proteins was decreaced , while expression level of bax didn't change , after treatment with UA for 36h. Conclusion UA had a significant antiproliferative effect on human gastric carcinoma BGC-803 cells in a dose- and time-dependent manner. UA-induced apoptosis was proved by HE staining,agarose gel electrophoresis and annexin V/PI staining in flow cytometry , respectively, Expression change of procaspase-3, procaspase-8, procaspase-9, bax and Bcl-2 proteins after treatment with UA for 36h suggested UA could induce apoptosis in BGC-803 cells via intrinsic and extrinsic pathways. Treatment with UA could pro- duce the membrane death signal and mitochondrion death signal , which triggered the extrinsic and intrinsic pathways, procaspase -8 and -9 were cleaved and activated , then procaspase-3 was activated and directly induced apoptosis in BGC -803 cells. Meanwhile , the cleavage of bcl-2 resulted in the ration of bax to bcl-2 increased, the integrity of Mitochondrion membrane was damaged and proapoptosis proteins were released which intensified the intrinsic pathway of apoptosis.