Molecular Mechanisms Raddeanin A Inducing Apotosis And In Human Colorectal Cancer Cells

Objective:To study Raddeanin A on human colon cancer cell proliferation in vitro, and to explore its molecular mechanism of induceding apoptosis.Methods:Evaluation of cell viability induced by Raddeanin A of the different drug concentration treatment (48 h) to human colon cancer cell SW480,HCT116 and LOVO by MTT and 0.1% DMSO as a negative control calculation raddeanin A on cell growth inhibition rate Half inhibitory concentration IC50; Then to draw LOVO cell growth curve after human colon cancer cells LOVO were treated with a different concentration gradient raddeanin A inl d,3 d,5 d; The cell cycle phase distribution by flow cytometry and PI staining after Raddeanin A of the different drug concentration (three concentration gradient) treatment(24 h) to human colon cancer cell LOVO; The morphological changes of apoptosis induced by Raddeanin A treatment to human colon cancer cell LOVO and apoptosis observed changes in nuclear chromatin by DAPI staining; Caspase-3 activity changed by Raddeanin A treatment (24 h) to human colon cancer cell LOVO were measured by spectrophotometry;The total protein of Raddeanin A treatment to human colon cancer cell LOVO were extraction by RIPA lysis method and the activity of PARP,Bim,P-44/42,P-JNK,XIAP,Survivin,Bcl-2,Bcl-x1,P-STAT3 and the expression of cytochrome C in mitochondria and in cytosolic were measured by Western blot; The plasmid containing the Survivin gene were transiently transfected into LOVO cells,and to detected cell viability by MTT; The certain concentration of P-JNK inhibitor SP10065 and raddeanin A were joined in the medium and to detected human colon cancer cell LOVO viability by MTT;.Results:Raddeanin A significant inhibition of human colon cancer cells SW480, HCT116 and LOVO proliferation,48h half inhibitory concentration IC50 was 1.783μM (SW480), 1.338μM (HCT116),0.156μM (LOVO) (P<0.05), and In 14.00μM concentration has reached about 90% inhibition (P<0.05) and with the increase of drug concentration and time of its inhibitory effect is more significant;The morphological changes of apoptosis induced by Raddeanin A of the different drug concentration treatment (24 h) treatment to human colon cancer cell LOVO is clearly, the chromatin significant shrinkage and apoptotic debris were visible by DAPI staining; Raddeanin A induce the G0/G1 phase proportion increased significantly in human colon cancer cell LOVO; Compared with the control, the role of raddeanin A LOVO in human colon cancer cells significantly activated PARP, Bim, caspase-3, P-44/42, P-JNK, and reduce the activity of XIAP,survivin,Bcl-2,Bcl-x1, P-STAT3 and the activity of cytochrome C in cytoplasm was higher than in mitochondria, and the difference is significant; Detected by MTT, the activity of LOVO cells transfected with survivin gene was significantly higher than the control; and the apoptosis of human colon cancer cells LOVO is inhibited after joining the P-JNK inhibitor SP10065.Conclussion:Raddeanin A could inhibit the growth of human colon cancer cells and induced apoptosis, which may be by making the tumor cells were arrested in G0/G1 phase and induced by activating the mitochondrial pathway to apoptosis of colon cancer cells...