Study On The Effect And Mechanism Of Biejiajianwan On Hepatocellular Carcinoma Cells

Objective:To prove the inhibitory effect of Biejiajianwan on hepatocellular carcinoma cells,and discuss the molecular mechanism of Biejiajianwan on hepatocellular carcinoma cells from lncRNA level.Methods:1.The suspension prepared by Biejiajianwan was given to rats intragastric administration,and the control group was given the same amount of normal saline for 3 consecutive days to prepare the medicated serum and control serum of Biejiajianwan.After the intervention of human hepatocellular carcinoma cells line SNU182 with medicated serum,the inhibition effect of Biejiajianwan was observed by CCK8 test,migration test and apoptosis test.2.After the intervention of Biejiajianwan in hepatocellular carcinoma cells,the differentially expressed lncRNAs were screened by whole-cell transcriptional analysis,and the signaling pathway,biological function,cell composition and molecular function of differentially expressed lncRNAs were obtained by bioinformatics KEGG and GO enrichment analysis.Survival analysis was used to evaluate the relationship between differential lncRNAs and prognosis of hepatocellular carcinoma and its clinical application value,and lncRNA-miRNA-mRNA network was constructed based on ceRNA regulatory mechanism.The genes with high connectivity(Hub genes)and their corresponding lncRNAs were identified through the protein interaction network.Take intersection of lncRNAs relevant to survival analysis and the corresponding lncRNAs of Hub gene.3.After the intervention of Biejiajianwan,these Intersecting lncRNAs were verified by fluorescence quantitative PCR to find the lncRNAs consistent with the whole-cell transcriptome analysis results,compare their expression in hepatocellular carcinoma cells and normal hepatocyte cells,and analyze their target genes.Results:1.The serum containing Biejiajianwan was prepared successfully.After the intervention of Biejiajianwan,the proliferation of human hepatocellular carcinoma cells line SNU182 was inhibited,migration ability was weakened,and apoptosis increased.2.42 differentially expressed lncRNAs were screened after whole-cell transcriptome analysis,.KEGG enrichment analysis suggested that differentially expressed lncRNAs might be involved in the following signaling pathways:cell cycle,cell senescence,ubiquitin-mediated proteolysis,HIF-1 signaling pathway,insulin signaling pathway,endocytosis,amino acid biosynthesis,cysteine and methionine metabolism.GO enrichment analysis suggested that differentially expressed lncRNAs were involved in biological processes including cell cycle,protein transport,cell division,Ubl conjugate pathway,DNA damage,mitosis,mrna processing,apoptosis and DNA repair.In cell composition,differentially expressed lncRNAs were mainly involved in cytoskeleton,endoplasmic reticulum,mitochondria,Golgi bodies,chromosomes,endosomes,cellular microtubules,centromeres,spliceosomes and centromeres.In terms of molecular function,differentially expressed lncRNAs were mainly involved in transferases,hydrolases,kinases,inhibitors,catalysts,serine threonine protein kinases,chromatin regulation,ribonucleoproteins and glycosyltransferases.Four lncRNAs were identified by survival analysis,namely lncRNA SERTAD4-AS1,EGOT,LINC00243 and FAM182A,and the ceRNA networks of these four lncRNAs were constructed.Protein interaction network(PPI)analysis showed that the top 10 Hub genes were CDK1,RRM2 and CDCA8,etc.lncRNAs corresponding to 10 Hub genes were lncRNA SERTAD4-AS1,LINC00243,EGOT,MIR3142HG and FAM182A.4 IncRNAs including SERTAD4-AS1,EGOT,LINC00243 and FAM182A were obtained after intersection.3.Fluorescence quantitative PCR showed that SERTAD4-AS1 expression increased and LINC00243 expression decreased after intervented with Biejiajianwan,which was consistent with the results of whole-cell transcriptome analysis.Compared with normal hepatocyte cells,the expression level of SERTAD4-AS1 in hepatocellular carcinoma cells decreased,while the expression level of LINC00243 in hepatocellular carcinoma cells increased.SERTAD4-AS1 was negatively correlated with target gene RRM2,and LINC00243 might regulate target gene CDK1 through hsa-miR-31-5p.Conclusions:1.The serum containing Biejiajianwan can inhibit the proliferation and migration of human hepatocellular carcinoma cells line SNU182 and promote its apoptosis.2.The molecular mechanism of Biejiajianwan inhibiting hepatocellular carcinoma cells may involve the regulatory axis of SERTAD4-AS1/RRM2 and LINC00243/hsa-miR-31-5p/CDK1.