The Role Of FOXC1 In The Migration And Invasion Of Lung Adenocarcinoma A549 Cells

BackgroundCurrently,lung cancer has the highest mortality rate of malignant tumor in the world,and the 5-year survival rate is only 18%.According to different pathological types,lung cancer is mainly divided into four categories:small cell lung cancer,lung squamous cell carcinoma,lung adenocarcinoma and other rare types.Since 1970s,the incidence of lung adenocarcinoma has increased rapidly,and according to reports in the literature that lung adenocarcinoma accounts for about 40%of the total number of lung cancers,which has replaced the lung squamous cell carcinoma becoming the most common pathological type.Clinical treatments for lung adenocarcinoma includes surgical treatment,systemic chemotherapy,epidermal growth factor receptor tyrosine kinase inhibitors and so on.Although the standard treatment for advanced lung adenocarcinoma is combination chemotherapy with platinum,the efficacy of chemotherapy has entered the plateau stage,and patients with EGFR mutations who receive EGFR-TKIs for about 6-12 months may also develop drug resistance.The poor prognosis of lung cancer is closely related to the biological characteristics of invasion and metastasis,and they are not only a sign of deterioration of the disease,but also an important reason for the failure of treatment.It is a problem to be solved urgently to seek for a new way to treat lung adenocarcinoma.The occurrence and development of lung cancer is a multi-step,multi-gene involved process.Epithelial-mesenchymal transition is one of the key steps in the invasion and metastasis of epithelial malignant tumors.EMT is a phenomenon in which epithelial cells are transformed into mesenchymal cells under certain physiological and pathological conditions.During this process,epithelial cells gradually lose their polarity,adhesion and tight connections,eventually become cells with the shape and characteristics of the interstitial cells,thus gaining the ability of infiltration and migration,which is characterized by the loss of epithelial index protein and the acquisition of vimentin and fibronectin.Transcription factor is one of the motility factors of EMT,and it is also affected by the activation of various signaling pathways including TGF-?,Notch,Wnt,STAT3 and so on,which promote tumor cell metastasis.With regard to transcription factors,Snail,Twist and ZEB family members are the most studied.In recent years,the Forkhead Box?FOX?series has drawn increasing attention.FOX is a kind of ubiquitous transcription factor.In recent years,FOX protein has been involved in many biological processes such as embryonic development,cell cycle regulation,tissue specific gene expression,immune regulation,cell proliferation,differentiation,metastasis and apoptosis.The gene mutation or abnormal expression have something to do with developmental deformity and tumorigenesis.More and more studies have revealed that FOXCl also plays an important role in the carcinogenesis and progression of tumors.The study confirmed that in malignant melanoma,gastric cancer,liver cancer,esophageal cancer,pancreatic cancer and other cancerous tissues FOXC1 expression is significantly higher than the paracancerous tissue,and the higher the level of expression the prognosis worse.However,the expression of FOXC1 in lung adenocarcinoma was rarely reported.High expression of FOXC1 can induce EMT and promote the metastasis and invasion of endometrial cancer cells,breast cancer cells and esophageal cancer cells.It is remains unknown that whether FOXC1 can regulate the occurrence of EMT to increase the ability of invasion and metastasis of lung adenocarcinomar cells and affect the prognosis of patients.Therefore,study the role and mechanism of FOXCl in lung adenocarcinoma invasion and invasion,which may provide new ideas and theoretical basis for clinical treatment of lung adenocarcinoma.ObjectiveTo investigate the role of FOXCl in the migration and invasion of lung adenocarcinoma by detecting the expression level of E-cadherin and Vimentin and the migration and invasion ability of A549 cells after FOXCl silencing,and to provide theoretical basis for clinical treatment for lung adenocarcinoma.Materials and methods1.materialsHuman lung adenocarcinoma A549 cells were purchased from Shanghai Institutes for Biological Sciences,Chinese Academy of Sciences.FOXC1 small interfering RNA?si RNA-FOXC1?and negative control?Negative control,NC.siRNA?were synthesized by Shanghai Jima Pharmaceutical Technology Co.,Ltd.2.Test methods2.1 A549 cells were cultured in RPMI1640 culture medium containing 10%fetal bovine serum,placed in 37?,containing 5%CO₂ incubator,routine liquid change,digestion and passage.Si RNA-FOXC1,NC.siRNA-FOXC1 were transfected into A549 cells,respectively,and were recorded as experimental group and negative control group.The blank control group was treated without any treatment.2.2 The relative mRNA expression of FOXC1,E-Cadhenrin and Vimentin in three groups were detected by RT-qPCR.Correspondingly,the expression of protein of FOXC1,E-Cadhenrin and Vimentin were detected by Western Blot.Gapdh was the internal reference gene2.3 Transwell invasion assay was used to determine the cell invasion ability in three groups.Scratch test was used to detect the migration ability of thethree groups of cells.Statistical analysisSPSS 21.0 was used for statistical analysis of the data,measurement data described by x±s.The comparison of multiple sets of quantitative data was performed using a one-way ANOVA with?=0.05 as the test level.The LSD-t test was used for the comparison between groups with normal distribution and variance.Otherwise,Kruskal-Wallis test was used.?=0.05 was taken as the test level.Results1.The expression level of FOXC1 m RNA and protein was significantly lower compairing experimental group A549 cells to blank control group and negative control group??<0.001?.There was no statistically difference in FOXC1 mRNA expression and protein expression level between negative control group and blank control group,?>0.05.2.The expression level of E-cadhenrin m RNA and protein was significantly higher compairing experimental group A549 cells to blank control group and negative control group??<0.001?.And there was no statistically significant difference between the negative control group and the blank control group about it.The expression of Vimentin mRNA and protein was significantly lower compairing experimental group A549 cells to blank control group and negative control group??<0.001?.Also,there was no statistically significant difference between the negative control group and the blank control group??>0.05?too.3.The scratch test showed that the migration rate of A549 cells in the experimental group was significantly lower than that of the negative control group and the blank control group??<0.001?.And there was no statistically significant difference between the negative control group and the blank control group??>0.05?.4.Transwell invasion test results showed that the number of invasive A549 cells in the experimental group was significantly lower than that in the negative control group and the blank control group??<0.001?.And there was no statistically significant difference between the negative control group and the blank control group??>0.05?.ConclusionSilencing FOXC1 inhibited the occurrence of epithelial-mesenchymal transition and attenuated the migration and invasion of lung adenocarcinoma A549 cells,providing a theoretical basis for clinical treatment of lung adenocarcinoma.