Study On The Regulation Of Proliferation And Migration Of Hepatocellular Carcinoma Cells By LncRNA CYTOR And EGFR

Background and objectiveLong non-coding RNA(Long non-coding RNA,lncRNA)is a non-coding RNA whose transcript length is greater than 200 nucleotides.It does not code for protein.It can competitively bind to miRNA and participate in cell cycle regulation and chromatin in the form of RNA.Remodeling,protein modification and other biological functions.Existing research has overturned the argument that LncRNA is the "noise" of the gene transcriptome,and found that LncRNA plays an important regulatory role in tumor triggering and progression.At present,no immune-related IncRNAs prognostic risk model has been established in patients with hepatocellular carcinoma.Therefore,this study intends to use potential molecular biomarkers to construct models to predict the prognosis of patients with hepatocellular carcinoma,so as to facilitate the implementation of personalized treatment plans.At the same time,One of the lncRNAs was selected for molecular biology to verify its biological role in the proliferation and apoptosis of hepatocellular carcinoma cells.In addition,a star molecule EGFR was also selected for molecular biology experiments,and tried to explore the relationship between the two.Materials and methodsIn this study,we downloaded the gene transcript data of hepatocellular carcinoma and the patient’s clinical data from the TCGA(The Cancer Genome Atlas)database,obtained the gene expression profile matrix and extracted the patient’s clinicopathological feature information;download and download from the GSEA database All related immune genes IMMUNE_RESPONSE and IMMUNE_SYSTEM_PROCESS,get the expression of all immune genes;use the"limma" package in the R language to convert the obtained data to screen out the differentially expressed immune-related LncRNA,and then extract the survival time greater than 90 Days of clinical patient information,single-factor COX regression is used to screen immune IncRNAs related to hepatocellular carcinoma OS,these candidate IncRNAs are included in the multi-factor COX regression model,and further screening is analyzed and finally immune LncRNAs related to patient survival prognosis are obtained.Based on these genes,a hepatocellular carcinoma prognostic risk scoring model related to the prognosis of immune LncRNA was constructed,and the area under the ROC curve of the model was calculated to verify the validity and accuracy of the model.Perform immune signal,KEGG pathway and enrichment analysis on the screened out IncRNAs significantly related genes with GSEA software.Finally,select a key LncRNA CYTOR and EGFR gene for molecular biology verification.1.Use qRT-PCR to detect the expression of key molecules CYTOR and EGFR in hepatocellular carcinoma cell lines and normal cell lines to verify whether the IncRNA is related to the occurrence and development of hepatocellular carcinoma;2.Transfect SMMC-7721 hepatocellular carcinoma cell line;3.Perform functional experiments after silencing IncRNA and EGFR in 7721 cells:CCK8 experiment to detect cell proliferation;cell scratch experiment to detect cell migration ability;Results1.A total of 424 expression profiles were obtained based on the TCGA database,including 50 normal genes,374 tumor genes,and 377 clinical sample information.2.Single factor and multivariate regression analysis obtained 12 immune LncRNA closely related to survival(PRRT3-AS1,AC099850.3,AL031985.3,AL117336.3,SREBF2-AS1,SNHG3,THUMPD3-AS1,CYTOR,MSC-AS1,AC124798.1,AC009005.1,AC026401.3)(P<0.001),used to construct a risk scoring model.3.Based on the median risk score,patients are divided into high and low risk groups;Kaplan-Meier method is used to analyze the results,and survival curves are drawn based on this,and the difference analysis uses Log-Rank test,according to the survival curve,high and low risk groups Survival difference is significant(P<0.05)The five-year survival rate of low-risk patients is 66%,and the five-year survival rate of high-risk patients is 36%;in addition,according to the ROC curve,it can be seen that the area under the risk score curve of the model is higher than that of patients Other clinical traits(riskscore AUC=0.808;stage AUC=0.750;gender AUC=0.544;age AUC=0.447),suggesting that this prognostic model has high predictive value and stability.4.The clinical correlation analysis shows that the patient’s risk score is closely related to the patient’s prognosis(P<0.001),and the patient’s age,gender,stage,and grade are not related to the patient’s prognosis(P>0.05).5.Principal component analysis(PCA)confirmed that the risk scoring model can distinguish high-risk and low-risk immune LncRNAs well.The gene enrichment results showed that when the core LncRNA molecules were up-regulated,the cell cycle,proteolysis caused by ubiquitin,purine metabolism,meiosis and other pathways were activated(P<0.05,FDR p<0.25,|NES|≥1).6.Select CYTOR and EGFR genes for molecular biology verification,and found that the content of CYTOR and EGFR in normal liver cell lines is significantly lower than that in hepatocellular carcinoma cell lines;after transfecting hepatocellular carcinoma cell lines with silenced CYTOR and EGFR,CCK8 showed a significant decrease in cell proliferation ability,cell scratch test shows that the migration ability is significantly reduced;Conclusions1.This study uses bioinformatics methods to screen out 12 key LncRNAs,and construct a prognostic model of immune-related LncRNAs for hepatocellular carcinoma.This prognostic model is capable of predicting OS in patients with hepatocellular carcinoma;2.These new lncRNAs may serve as potential therapeutic targets.3.CYTOR and EGFR can promote the proliferation and migration of hepatocellular carcinoma cells.