The Mechanism Of Kaempferol In Promoting Autophagy And Apoptosis Of Non-small Cell Lung Cancer

ObjectiveTo explore the effect and molecular mechanism of kaempferol in promoting cell autophagy and apoptosis of non-small cell lung cancer cell(NSCLC).Methods1.In vitro,NSCLC cell line A549,H1299 and A549 were used as the research subjects.MTT assay and EdU assay were used to observe the effect of kaempferol on the NSCLC cell proliferation.Flow cytometry of cell apoptosis was used to detect the apoptosis of A549 and H1299 when treated with different dosages of kaempferol.Western blot was used to detect the effect of kaempferol on protein expression,including Caspase 3(Cleaved-Caspase 3),Caspase 7(Cleaved-caspase 7),and Bcl-2 family(Bcl-2,Bax,Bad).2.mCherry-EGFR-LC3 assay,electron microscopy assay,and western blot assay were used to explore the autophagy of NSCLC when treated with Kaempferol.Meanwhile,Autophagy inhibitors were also used to explore the effect of kaempferol-induce-autophagy in NSCLC cell proliferation.And western blot assay is the main way to explore the relative protein express in non-small cell lung cancer cell.In addition,the overexpress Met gene plasmid was transiently transfected in A549 cells to explore the difference when kaempferol treated.3.In vivo,a subcutaneously implanted tumor model in BALB/C nude mice was found by use human NSCLC cells A549-Luc.High and low dose of kaempferol was given to nude model to explore the kaempferol effect in inhabiting NSCLC growth by measure the tumor bioluminescence intensity in the first and the last administration.At the end of experiment,mise was euthanized and subcutaneous tumor was remove and weighted.In addition,western blot was used to detect the effect of kaempferol on relative protein expression in mice subcutaneously tumor tissues.Results1.Kaempferol could inhibit the growth of NSCLC cells in vivo and vitro.2.Kaempferol could promote apoptosis in NSCLC cells.Kaempferol increased the protein expression of Cleaved-Caspase 3,Cleaved-Caspase 7,Bax,Bad,while decreased the protein expression of Bcl-2.3.Kaempferol could induce autophagy in NSCLC cells.Kaempferol increased the protein expression of LC3B,Beclinl,while decreased the protein expression of P62.And kaempferol combined with autophagy inhibitors increased the viability of A549 cells.Kaempferol decreased the expression of Met in protein and mRNA level,and decreased the phosphorylation of PI3K、Akt and m-TOR protein.The mechanism may though Met inducing autophagy,and PI3K/AKT/m-TOR signal pathway inducing autophagy.Transient overexpression of Met reduced the anti-tumor activity of kaempferol,and relieving kaempferol-induced-autophagy in A549 cells.ConclusionKaempferol may regulate Bcl-2/Caspase 3 pathway to promote apoptosis in NSCLC proliferation.And kaempferol may through Met and PI3K/AKT/m-TOR signal pathway to promote death-relative-autophagy in NSCLC growth.