Apoptosis Of Lung Cancer Cells Induced By Curcumol Through ROS/JNK Pathway

Lung cancer is the most common malignant tumor all over the world,with high metastasis and recurrence rate of advanced lung cancer.Despite there are some new therapies and novel drugs for lung cancer in recent years,however,the 5-year-survival rate is low.The targeted drugs are only for certain patients,which made the lung cancer is still a major problem for all the drug developers.Traditional Chinese medicine,as a traditional treasure of the China,has a long history and a wide range of applications.With the development of modern medical technology,the research of traditional Chinese medicine is more and more in-depth,and there are a great number of reports about the anti-tumor effect of traditional Chinese medicine monomers both here and abroad.Considering the advantages of traditional Chinese medicine as long-effect,multi-target,low toxicity and side effects,it can effectively improve the survival rate and the quality of life of patients.Curcumol is a sesquiterpene compound extracted and isolated from the volatile oil of Rhizoma Curcumae,which is a traditional anti-tumor medicine,with the pharmacological function of antitumor.Although so many different theories about the antitumor mechanism of curcumol including IGF-1R,MAPK,AKT,NF-kappa B or some another apoptosis proteins or cyclins had been reported,there is no final conclusion has yet been reached on this matter.Therefore,the research discussed the effect and mechanism of curcumol against lung cancer.There are 4 experimental section involved in this paper.The first part: select the suitable organic solvent and non-toxic dose range to cellsThis part is mainly purpose to selecting a suitable organic solvent to ensure theefficacy of curcumol.The MTT assay was used to test the cytotoxicity of the organic solvent(DMSO and ethanol)on 6 different kinds of lung cancer cells,and ethanol with a concentration of 1.6%(V/V)was selected as the highest concentration for the following experiments for its adequate solubility for curcumol and its low cytotoxicity for lung cancer cells.The second part: the antitumor effect of curcumol in vitroThe proliferation inhibition effect of curcumol with a time-and dose-dependent manner was confirmed by MTT assay.Compared with the result of MRC-5 MTT assay,we believed the inhibition of curcumol on lung cancer cells was selective.The third part: the anti-tumor effect of curcumol in vivoNude mice with subcutaneous tumor was treated with curcumol and cisplatin,respectively.The body weight and the tumor volume were measured every other day,and all the data showed that curcumol was effective in vivo,and more safe than cisplatin.The fourth part: the anti-tumor mechanism of curcumolIn this part,cell apoptosis was observed by Hoechst 33258 staining.The mitochondrial membrane potential change was detected by JC-1 probe.By FITC-AnnexinV/PI double staining,we found the cell apoptosis induced by curcumol was in a dose-dependent manner.Western blotting results showed that curcumol could activate JNK protein.DCFH-DA probe was used to detect the ROS,and the results showed that curcumol could induce intracellular ROS increasing in a dose-dependent manner.The increase of ROS induced by curcumol could be reduced when ROS inhibitor was existed.The apoptosis rate caused by curcumol could be reduced by the addition ROS inhibitor and JNK inhibitors.The results of western blotting showed that both ROS inhibitors and JNK inhibitors could inhibit the activation of JNK caused by curcumol.The results of cell cycle detected by PI staining showed that curcumol could arrestcell cycle in G2/M phase.In this study we found that curcumol could induce the cell apoptosis,result in the increase of ROS in cells,and activate JNK protein.The phenomenon can be greatly reduced by adding ROS inhibitor and JNK inhibitor.It is confirmed that curcumol induced lung cancer cell apoptosis through the ROS/JNK pathway.Meanwhile,the cell cycle arrest in G2/M phase was also found in cells treated by curcumol.In nude mice experimental,the tumor volume growth of curcumol group was obviously slower compared with control group,and the tumor mass inhibition rate was similar to the cisplatin group,which means curcumol has favorable anti-tumor effect in vivo as well as in vitro.