The Study On Inhibition Of Non-small Cell Lung Cancer Growth By Interfering With Aerobic Glycolysis Of α-Hederin

Object: To study the intervention effect of the active ingredient α-hederin of Pulsatilla saponin on the glycolysis of Non-small cell lung cancer and its mechanism,and elucidate the anti-tumor mechanism of α-hederin,and lay a foundation for further research.Part Ⅰ Effect of α-hederin on Proliferation of Human Lung Cancer CellsMethod: Human lung cancer A549,NCI-H292,NCI-H460 cells were cultured,and α-hederin was administered to intervene.CCK8 cell proliferation-toxicity assay was used to study α-hederin on non-small cell lung cancer A549,NCI-H292,NCI-H460 cells.By using plate clone formation experiment to study the effect of the α-hederin on the proliferation of non-small cell lung cancer cell A549.Result: The results of CCK8 showed that α-hederin has an inhibitory effect on non-small cell lung cancer A549,NCI-H292 and NCI-H460 cells.The IC50 of α-hederin on Non-small cell lung cancer A549,NCI-H292 and NCI-H460 cells were 9.53 μg/ml,12.52 μg/ml,and 12.86 μg/ml.The results of plate clone formation showed that α-hederin inhibited non-small cell lung cancer A549 cells in a dose-dependent manner.Part Ⅱ Intervention of α-hederin on Glycolysis of Non-small Cell Lung Cancer A549 Cells and its MechanismMethod: The contents of GLU,LD and ATP in the cell culture medium were detected by kit,and the intracellular protein levels were detected by Western Blot.Conclude the key enzymes of glycolysis GLUT1,HK2,PKM2,LDHA,MCT4 and glycolytic control factors c-Myc,HIF-1α,p53,Akt.Result: The results showed that α-hederin significantly decreased glucose uptake in A549 cells compared with the control(P<0.05),increased lactic acid production(P<0.05),and decreased intracellular ATP level(P<0.05);Western Blot results showed that α-hederin significantly down-regulated the expression of key enzymes GLUT1,HK2,PKM2,LDHA,MCT4,c-Myc,HIF-1α.Part Ⅲ The inhibitory effect of α-hederin on transplanted tumor of non-small cell lung cancer cell A549 in nude mice and its mechanismMethod: Through the establishment of human non-small cell lung cancer cell A549 in nude mice xenograft model,the tumor weight and tumor volume were measured to observe the inhibition of α-hederin on the tumor;using the kit to detect tumor tissue GLU,LD and intracellular ATP level;immunohistochemical(IHC)was used to determine the expression of key enzymes and regulatory factors GLUT1,HK2,PKM2,LDHA and MCT4 in transplanted tumors.Result: The results of xenograft tumors in nude mice showed that α-hederin group has a significant inhibitory effect on Non-small cell lung cancer A549 cells transplanted in nude mice(P<0.05),and the safety was good;GLU,LDand ATP kit showed that the levels of LD and ATP in the α-hederin 5 mg/kg group and the 10 mg/kg group were significantly lower(P<0.05),and the GLU level was significantly increased(P<0.05);IHC results showed that by administration of α-hederin,the expression of GLUT1,HK2,PKM2,LDHA and MCT4 was significantly down-regulated in the tumor.Conclusion: α-hederin inhibits the growth and proliferation of Non-small cell lung cancer A549 by inhibiting aerobic glycolysis in vivo and in vitro.Its mechanism is α-hederin inhibit the expression of GLUT1,HK2,PKM2,LDHA and MCT4 by inhibiting c-Myc and HIF-1α.It has been proved that α-hederin inhibit tumor glycolysis by acting on c-Myc,and its effect on tumor glycolysis by affecting HIF-1α needs to be verified...