Studies On The Mechanism Of Apoptosis In A549 Cells Induced By Phellamurin And Chlorogenic Acid

Cancer has been a human health killer,in recent years,lung cancer,whose incidence is after breast cancer and prostate cancer,has become the leading cause of cancer deaths.Phellamurin and chlorogenic acid are active ingredients of leaves of Phellodendron amurense.As natural products,they have low toxicities and little side effects.The aim of this study is to investigate the antitumor activity of phellamurin and chlorogenic acid and its molecular mechanism,and to find a new approach for cancer treatment.A549 cells were treated with different concentrations of phellamurin and chlorogenic acid respectively.The inhibition ratio of cell growth was detected by MTT assay.Cell cycle distribution and cell apoptosis were examined through flow cytometry.SOD activity,GSH content,MDA level and caspase 3 activity in A549 cells were determined via colorimetry.In addition,mitochondrial membrane potential(MMP)was detected by way of JC-1 staining and the expressions of apoptosis related proteins were measured through western blotting.The result of MTT assay showed that when the drug concentration was 2000 ?M,the inhibition ratio caused by phellamurin was 70.87%,while it was 29.01% caused by chlorogenic acid.The results of flow cytometry indicated that phellamurin induced G2/M phase arrest which was accompanied by a cell apoptosis ratio of 84.32%,while chlorogenic acid caused an S phase arrest with an apoptosis ratio of 30.10%.JC-1staining showed that both phellamurin and chlorogenic acid could decrease the MMP level of A549 cells within 24 hours.In addition,after treated with phellamurin,the activity of SOD in A549 cells decreased by 50%,the content of GSH was decreased by 30%,and the level of MDA increased by 60%,but chlorogenic acid had no significant effect on them.Furthermore,the activity of caspase 3 in A549 cells was increased by 31% and 25%,respectively.The results of western blotting demonstrated that phellamurin made the level of Bax / Bcl-2 up to 33 times of that in control group,while chlorogenic acid made it up to 1.8 times of that in control groupIn this study,we found that both phellodendron and chlorogenic acid could blockthe cell cycle,damage the cytogenetic material,up-regulate the expression of Bax,down-regulate the expression of Bcl-2 and activate caspase 3 to induce apoptosis of A549 cells.