Combination Of Celastrol With Wogonin Exhibits Anti-tumor Efficacy And Affects Energy Metabolism In Tumor Cells

With the progress of science and technology and the growingenvironment pollution,cancer has become a threat to human existence. The principal means of cancer treatment include radiotherapy,chemotherapy and surgery, all these treatments will influence normal tissue in a certain degree.In recent years, traditional Chinese medicine for the prevention and treatment of tumor prevention has get the wide attention of researchers.Celastrol and wogonin are the active ingredient of traditional Chinese medicine (TCM),both have the effect of anti-cancer,anti-inflammatory,antiviral and antibacterial, etc.But the cytotoxic effect of celastrol is stronger,the dose of wogonin is higher,which affect their in clinical application.In this topic we joint the two medicine to lower the cytotoxic effect and reduce the drug dose, which can increase the activity of anti-cancer.Objective:To study activity and the influence of energy metabolism of joint application or individual of Celastrol and Wogonin to three kinds of cells,epithelial cells of thehuman umbilical vein (ECV304),human gastric cancer cells(SGC-7901)and human lung cancer cells(A549).Risult display compare with the individual,the combination is better to lower the cytotoxic effect and reduce the drug dose whiche can also increase the drugs'activity of anti-cancer.Methods:1.MTT method is used to determine the inhibition rate of tumor cells and human umbilical vein cells;2.The cell growth curve is used to determine tumor cells and human umbilical vein cells growth trend;3.HE staining method is used to observe the morphological changes of tumor cells and human umbilical vein cells;4.Useing the spectrophotometric method for determination of the enzyme include hexokinase,pyruvate kinase and lactate dehydrogenase in glycolytic pathway within the tumor cells and endothelial cells,the marker enzyme in the Krebs cycle succinate dehydrogenase,and the end-products in energy metabolism-ATP levels;5.The Weste-rn blot method is used to determine expression level of hif-1?(HIF-1?)and single carboxyl transporter(MCF-4).Result:1.The best concentration is 15?g mL-1 of Wogonin and 3?g·mL-1 of Celastrol analyzed by MTT;2.HE staining and cell growth curve results showed that proliferation of SGC-7901?ECV304 cells was markedly inhibited in combined drug group,especially for SGC-7901 but low profile for A549. Appeared to volume reduction.roundout.cytoplasmic condensation and color burning and cell concentration decreased significantly.3.The growth and proliferation of tumor cells due to hypoxia microenvironment,glycolytic reaction which provides energy for life is very important to tumor cells.Experiments to detect content variation of three enzyme in the glycolytic pathway called hexokinaseand?pyruvate kinase and lactate dehydrogenase.Results showed different inhibition strength to different cancer cells for conbined administration.The inhibition ratio to hexokinase is that ECV304 and SGC-7901 more upper,but A549 cells obscure.It is non-influence for pyruvate kinase.The content of lactic acid influence the migration and invasion of cancer, the results of lactate dehydrogenase activity assay confirmed combination significantly reduced the acid dehydrogenase vitality,The results demonstrated that the change of acidic microenvironment in tumor cells would suppress their formation and development.Succinic dehydrogenase is the symbol of the mitochondria.Experiments result display that combination is difference impact in three cells oxidative phosphorylation energy metabolism. The inhibition ratio to succinic is that SGC-7901 higher than ECV304 and A549 cells obscure.ATP is primary of enery supply.The result of ATP content illustrated that combination can lower the energy supply of ECV304 and SGC-7901 but is invalid to A549 cells.4.HIF-la is the important regulate protein in cell proliferation and glycolysis.MCT-4 is the leading protein to transport lactate out of cell and keep the acidness environment. Combined drug can significantly reduce the expression level of HIF-la and MCT-4 in SGC-7901 and ECV304 cell,but noneffective to A549 cells.Conclusion:The drug combination of celastrol and wogonin compare individual more stronger in inhibition ECV304?SGC-7901 cells expression level of HIF-la and MCT-4,disturb energy metabolise.Then the drug combination is more effective to inhibition enzyme in energy metabolism,reduce ATP content. The result show cell multiplication of two cells are inhibition but noneffective to A549 cells.The drug combination effect strong to cancer cells human umbilical and also has result to vein endothelial cells,can inhibit human umbilical grow and vascellum generate.