FGL1 Regulates The Sensitivity Of Lung Adenocarcinoma Cells To Docetaxel Through The ERK1/2 Signaling Pathway

Objective: To investigate the effect of fibrinogen protein 1(FGL1)on docetaxel sensitivity of lung adenocarcinoma cells and malignant biological behavior of tumor cells,and to explore the specific mechanism of FGL1 regulating docetaxel sensitivity of lung adenocarcinoma cells.Methods: 1.Download relevant clinical data from the TCGA data portal and analyze the expression level of FGL1 in a variety of tumors,and the online Kaplan Meier Plotter was used to assess the survival and prognosis of the sample information of lung adenocarcinoma patients in the GEO data portal.2.Western Blot was used to detect the expression of FGL1 protein in human lung epithelial cells BEAS-2B and human lung adenocarcinoma cell lines H1975,PC9 and A549,and cell lines with high level expression of FGL1 were selected for subsequent studies.3.The human lung adenocarcinoma cells PC9 and A549 were selected as the research objects,si RNA was used to knock down the FGL1 expression in PC9 and A549 cells.the transfection efficiency were measured by q RT-PCR and Western Blot.4.The CCK-8 assay was use to detect inhibition of cell p-roliferation and the corresponding half-maximal inhibitory concentration treated with different concentrations docetaxel(0,5,10,20,40,80 μmol/L)on PC9 and A549 cells,We selected the appropriate concentration according to the results of the CCK8 assay for subsequent experiments.5.To examine the effect of FGL1 knockdown on the proliferation,apoptosis and malignant biological behavior of docetaxel-induced lung adenocarcinoma cells.There were five blocks in the experiment.: Control group,NC group,Docetaxel group,NC + Docetaxel group,si FGL1 group and Docetaxel + si FGL1 group,cell proliferation,cell cycle,apoptosis,migration and invasion were then evaluated using an CCK-8 assay,flow cytometry,scratch test and Transwell assay,respectively.6.The expression levels of ERK1/2 signaling pathway key proteins(ERK1/2,pERK1/2)and apoptosis-related proteins(Bax,Bcl-2,Cleaved-caspase3)were detected by Western Blot.A specific inhibitor of ERK1/2 pathway(U0126)and the specific agonist(EGF)were used to further determine the specific mechanism of FGL1-ERK1/2 pathway in docetaxel-induced apoptosis of lung cancer cells.Results: 1.The data from the the Gene Expression Omnibus(GEO)database and The Cancer Genome Atlas(TCGA)database demonstrated that FGL1 expression is significantly up-regulated in a variety of tumor tissues,and its expression is significantly increased in lung adenocarcinoma and prostate cancer(P<0.01).Survival prognosis displayed that the prognosis of FGL1 high expression group was worse(P<0.05).2.The Western Blot experiment confirmed that,the level of FGL1 was higher in lung adenocarcinoma cells PC-9 and A549 as compared with normal lung epithelial cells.(P<0.01).3.Compared with the Control group and the NC group,the proliferation,invasion and migration abilities of the Docetaxel group,the si FGL1 group and the combined group all decreased to varying degrees(P<0.05),and the apoptosis rate was increased(P<0.05),and the changes are the most obvious in the combined group.Compared with the Control group,the Docetaxel group and the NC+Docetaxel group,the NC group showed no significant difference in proliferation,invasion,migration and cell apoptosis rate(P>0.05).4.Western Blot analysis showed that knockdown of FGL1 combined with docetat not only significantly reduced ERK and p-ERK levels,but also further up-regulated phosphorylated ERK(p-ERK)and apoptotic protein Bax and cleaved caspase-3 levels and decreased apoptotic inhibitory protein Bcl-2 levels,promoting apoptosis.The use of ERK1/2 pathway inhibitor U0126 and agonist EGF partially enhanced and reversed the effect of FGL1 knockdown on the expression of docetaxel-induced apoptotic proteins and apoptotic suppressor proteins,respectively(P<0.05).Conclusion: 1、FGL1 was upregulated in lung adenocarcinoma cells,and the overall survival of patients with high FGL1 expression was significantly lower than that of patients with low FGL1 expression.2、Knockdown of FGL1 expression significantly inhibited the proliferation,invasion and migration activities of lung adenocarcinoma cells,promoted the apoptosis of tumor cells,and increased the sensitivity of lung adenocarcinoma cells to docetaxel.3、FGL1 may affect the sensitivity of lung adenocarcinoma cells to docetaxel by regulating the phosphorylation level of ERK1/2,and further regulating the levels of apoptotic proteins Bax and cleaved caspase-3 and apoptotic inhibitor Bcl-2.