The Mechanisms Of Gallic Acid-induced Lung Cancer Cell Apoptosis And The Study Of Its Application

Gallic acid （GA） as a polyhydroxylphenolic compound is widely distributed invarious plants, fruits and foods. Tea is rich in GA which is very well absorbed inhumans. It has been reported that GA has various biological activities. The majorinterest in GA is related to its antitumoral activity. In fact, the antitumoral activity ofGA has been reported in various cancer cells, such as liver, colon and breast cancer.Lung cancer is a major cause of cancer death in developed countries. Studies ofthe molecular mechanisms of cytotoxic drug action have shed light on the treatment oflung cancer, and novel agents that target specific intracellular pathways related to thedistinctive properties of cancer cells continue to be developed. There are severalreports have proved that GA can induce cancer cell death. Hwever, the mechanismswere not very clear. In the present study, we selected three kinds of lung cancer cellsto to analyze the effect of GA on cell growth and investigated the mechanisms of GAwhich act as pro-oxidant to kill cells.The MTT results showed that GA can inhibit cell viability of lung cancer cells,including95-D cells,549cells and NCI-H460cells. The percent viability of cellsdecreases with GA treatment in a concentration-dependent manner. The results ofenvironmental scanning electron microscope and confocal microscope showed thatGA can induce cell death via apoptosis.The mechanisms of GA-induced cell apoptosis were not clear. The pro-oxidativeeffects can make cells to produce reactive oxygen species （ROS） which is related toanti-tumor effect of GA. ROS is a multifunctional molecule which includes hydrogenperoxide （H2O2）, peroxynitrite （ONOO·）, superoxide anion （）, and hydroxyl radical（·OH）. It participates in a variety of biological processes in cell apoptosis.In order to analyze the mechanisms of GA-induced cell apoptosis, we tested theproduction of ROS-triggered by GA. The results of flow cytometry showed that GAcan induce ROS production, and promote the generation of H2O2and. GA acts asa pro-oxidant to induce cell apoptosis.The production of ROS can activate endocytoplasmic reticulum, extrinsic and intrinsic signaling pathways. Thus, we tested the signal molecules which are proved toinvolve in these pathways. The results showed that GA decreased mitochondrialmembrane potential （MMP; ΔΨm） and induced cytochrome c release to cytosol. ThenGA caused mitochondrial translocation of Bax and up-regulated the expression of Bax.The expression of Bcl-2and Bcl-XLwere drastically changed following GA treatment.The caspase-9was activated and intrinsic signaling pathway activated. GA inducedup-regulation of Fas, FasL and activated caspase-8. The extrinsic signaling pathwaywas activation. GA inhibited the expression of TGF-β1which up-regulated theexpression of Fas and FasL. The extrinsic signaling pathway was activation. Theactivations of caspase-9and caspase-8led to activate caspase-3, resulting to apoptosis.The intracellular Ca²⁺elevation led to endocytoplasmic reticulum signaling pathwayactivated. The mitochondria were destroyed when Ca²⁺elevation in mitochondria.The intrinsic signaling pathway activated.In addition, Akt signal pathway was also involved in the process of cell apoptosis.We also analyzed the signal factors which involved in Akt signal pathway. The resultsof wesrern blot and confocal microsope showed that the down-regulation of Akt led toinactive of NF-κBp65and COX2. The down-regulation of Akt also induced theexpression of p53. Cell apoptosis induced by Akt signaling pathway activation.Furthermore, GA can also induce caspase-6activation and cytoskeleton damage.Cell apoptosis were induced.In order to better understand the cytotoxicity of GA, we also observed thecytotoxicity of GA combined with cisplatin （DDP） and the effects of GA on cellmigration.Breast cancer is an epithelialtumor. The high morbidity and mortality are themajor cause of women death. In order to reduce the tolerance of tumors, several drugsare advised to apply together. DDP is a kind of antitumoral drugs which we use toapply with GA to observe the effect on MCF-7cells. The results showed that GAcombined with DDP can induce more MCF-7cell death than GA group and DDPgroup. The mechanisms of GA-co-DDP-induced cell death are similar to GA. Theextrinsic and intrinsic apoptotic pathways were activated by GA combined with DDP.GA induced cell apoptosis by down-regulation of NF-κBp65.The cells near the wound edges play an important role in the process of cellmigration. The microfluidic device can avoid the disadvantages which the cells nearthe edges were damaged by scrapping method. We analyzed the effection of GA on cell migration by microfluidic method. The results showed that GA can inhibitfibroblasts migration. The average migration distance was lower that control.