Screening And Validation Of Differential Expression Of CircRNA In Lung Adenocarcinoma Tissues

Objectives:Lung cancer has gradually become one of the malignant tumors with the highest morbidity and mortality in the world.The 5-year survival rate of lung cancer is relatively low due to insufficient research on the pathogenesis of lung cancer and lack of biomarkers for early diagnosis.In recent years,studies have confirmed that circ RNA is widely expressed in organisms as non-coding RNA and is closely related to the occurrence and development of various tumors.In this paper,circ RNA differentially expressed in lung adenocarcinoma and adjacent tissues was screened by gene chip technology.According to the ce RNA regulatory network and the screening principle of circ RNA for subsequent research purposes,the target circ RNA was screened and verified.Objective To investigate the role of circ RNA in the occurrence and development of lung adenocarcinoma,to provide new biomarkers for the diagnosis and prognosis of lung adenocarcinoma,and to provide new ideas for targeted therapy research.Methods:1.Lung adenocarcinoma sample collection: 4 pairs of surgically resected lung adenocarcinoma and adjacent tissue samples were collected.2.Detection of differential gene expression in lung adenocarcinoma and adjacent tissues by gene chip technology.3.Processing and analysis of original data: normalize the original data of the chip,and then compare and analyze the data of lung adenocarcinoma and adjacent tissues.(1)Sample cluster analysis,sample correlation analysis and principal component analysis are used for comparative analysis among samples.(2)Screen differentially expressed RNA.The screening criteria : FC(fold change)≥2 or FC ≤ 0.5,P < 0.05.Scatter plot,volcano plot,and heatmap can be used to visualize the distribution and expression of differentially expressed RNA.(3)Predict the target mi RNA and target gene of circ RNA and establish ce RNA regulatory network.4.According to the prediction results of ce RNA regulatory network and the screening principle of differentially expressed circ RNA,the circ RNA for subsequent research was screened and further verified in lung adenocarcinoma cell lines and 40 pairs of lung adenocarcinoma tissues.5.According to the function of the target gene predicted by the target circ RNA in the occurrence and development of tumor,the function of the target circ RNA in the occurrence and development of lung adenocarcinoma is speculated.Results:1.After normalization of the original data,comparative analysis between samples shows that the two groups of samples are grouped reliably.Compared with adjacent tissues,there were 2768 up-regulated circ RNA and 2935 down-regulated circ RNA in lung adenocarcinoma tissues(FC≥2 or FC≤0.5,P<0.05)2.Predicted the target mi RNA and target gene that differentially express circ RNA,and established ce RNA regulatory network.3.According to the prediction results of ce RNA regulatory network and the screening principle of differentially expressed circ RNA,the down-regulated hsa＿circ＿0024888,hsa＿circ＿0026554,hsa＿circ＿0060184,hsa＿circ＿0056510 were selected from differently expressed circ RNA as the following research objects.4.RT-q PCR results of lung adenocarcinoma cell lines and lung adenocarcinoma tissue samples show that the relative expression of target circ RNA: hsa＿circ＿0024888,hsa＿circ＿0026554,hsa＿circ＿0060184,hsa＿circ＿0056510 in lung adenocarcinoma cell lines(A549,H1299,H1975,H1793,H1993)and normal lung epithelial cells(BEAS-2B)is consistent with the results of lung adenocarcinoma tissue gene chip.More than half of the relative expressions of the above four circ RNA in 40 pairs of lung adenocarcinoma and adjacent tissues are consistent with the results of gene chip in lung adenocarcinoma tissues.5.According to the down-regulated target gene of circ RNA in ce RNA prediction results,the target gene of circ RNA is down-regulated in lung cancer or other malignant tumors and has cancer inhibiting effect,and it is presumed that the four down-regulated target circ RNA have cancer inhibiting effect.Conclusion:1.Through gene chip and bioinformatics technology,abnormal expression of circ RNA was found in lung adenocarcinoma.The target mi RNA and target genes differentially express circ RNA are predicted,and the ce RNA regulatory network is established.2.For the first time,hsa＿circ＿0024888,hsa＿circ＿0026554,hsa＿circ＿0060184,hsa＿circ＿0056510 were found to have low specific expression in lung adenocarcinoma tissues.3.Cell experiments verified that the expression of circ RNA in lung adenocarcinoma cell lines was consistent with the results of gene chip in lung adenocarcinoma tissues.Verification test results of 40 lung adenocarcinoma tissue samples show that more than half of the relative expressions of the above 4 circ RNA in lung adenocarcinoma tissues are consistent with the results of tissue gene chip.4.According to the anti-cancer effect of the target gene of circ RNA predicted by ce RNA regulatory network,it is presumed that the down-regulated circ RNA has cancer inhibiting effect.