Screening Of Differential Expression Profiles And Related Gene Function In Thyroid Carcinoma

Objective: To explore the differential expression profiles of papillary thyroid carcinoma by second-generation sequencing technology,and to screen and explore the related roles of differentially expressed genes in papillary thyroid carcinoma.Methods:(1)To obtain differentially expressed transcriptome map using second-generation sequencing technology by the culture of papillary thyroid carcinoma cell lines BCPAP,TPC-1,K1 and human normal thyroid cell line Nthy-1,and to screen differences expressed genes(C0L1A2,PDGFRA,THBS-1,TNC,ITGB4,PDGFB,FGFR4)for further study based on sequencing results and literature reference;(2)To culture thyroid papillary carcinoma cell line BCPAP,TPC-1,K1 and human thyroid normal cell line Nthy-1 by collection of 54 cases clinical tissue samples from patients with papillary thyroid carcinoma(cancer tissue and adjacent to 3-5 cm adjacent to cancer),and to verify the level of the screened gene on the tissue level and cell level by real-time fluorescent quantitative PCR technology,and to screen differentially expressed genes(ITGB4)combined with the results of the verification and sequencing.(3)To extract the total protein of thyroid papillary carcinoma cell line(TPC-1,K1,BCPAP)and normal thyroid cell line(Nthy-1),and to identify the target gene ITGB4 according to the test results by Western blot analysis of protein expression of ITGB4 at the cellular level.(4)The ITGB4-RNAi vector was used to knock down the expression of ITGB4 in BCPAP,TPC-1 and K1 cells and to detecte the knockdown effect by fluorescence microscopy,real-time PCR and Western blot.(5)To investigate the effects of ITGB4 knockdown on the proliferation,migration and invasion of thyroid papillary carcinoma cells by CCK-8 assay,cell scratch assay and Trans Well cell invasion assay.Results:(1)There were 1961 known genes differentially expressed in the thyroid papillary carcinoma cell lines BCPAP,TPC-1,and K1 compared with the human thyroid normal cell line Nthy-1 according to the results of transcriptome sequencing,in which 1179 genes were significantly up-regulated and 782 genes were significantly down-regulated.(2)Comprehensive sequencing data and literature reference,preliminary screening of 7 differentially expressed genes,in which COL1A2,PDGFRA,THBS-1,TNC were significantly down-regulated,ITGB4,PDGFB,FGFR4 were significantly up-regulated,and further validation on 7genes in papillary thyroid carcinoma was performed in cell lines and patient tissue samples,and ITGB4 was for the target gene.(3)The ITGB4-RNAi vector successfully transfected K1 cells and BCPAP cells,and the expression level of ITGB4 in K1 cells and BCPAP was significantly knocked down.(4)After knockdown,the proliferation,migration and invasion ability of BCPAP were significantly inhibited,and there was no significant change in proliferation,migration and invasion ability of K1 cells after knockdown.Conclusion:(1)COL1A2,PDGFRA,THBS-1,and TNC were significantly down-regulated in the differentially expressed genes of thyroid papillary carcinoma cell lines and normal cell lines,and ITGB4,PDGFB,and FGFR4 were significantly up-regulated.(2)ITGB4-sh RNA knocked down the expression of ITGB4 in BCPAP,TPC-1 and K1,and BCPAP and K1 cell lines were effectively knocked down.After knockdown,BCPAP proliferation,migration and invasion ability were significantly inhibited.It indicates that the abnormal expression of ITGB4 has an effect on the proliferation,migration and invasion of papillary thyroid carcinoma.