Heparanase Expression And Regulation Of Human Liver Cancer

Objective: To detect the heparanase (HPSE) cDNA expression level of different humanliver cancer cell.We constructed green fluorescent protein vector ofpEGFP-HPSE,human HPSE gene miRNA vector and Eukaryotic expressionvector,screening and identification and analysis of its activity, regulate HPSE expressionof liver cancer cells.Methods:To detect the heparanase (HPSE) cDNA expression level of human livercancer cells in Bel-7402,HepG-2,HCCLM3by qPCR.According HSPE gene sequences,for which four interference targets were designed and synthesized four RNAiinterference fragment linearized miRNA vector and insertion sites, construct HPSE geneRNAi vector (pEGFP-1-RNAi).Polymerase chain reaction amplification HPSE genecoding region of the human genome DNA; then inserted into the multiple cloning sitesdigested pEGFP-1, building human HPSE eukaryotic expression vector(pIRES2-EGFP-HPSE,hereinafter to be referred as pEGFP-1-HPSE);Groupingtransfected HepG2cells and HCCLM3, fluorescence observation and analysis ofreal-time PCR and electrophoresis were detected pEGFP-1-HPSE promotingtranscriptional activity and four kinds of pEGFP-1-RNAi inhibition of transcriptionactivity.Results:In human hepatoma cells Bel-7402,HepG-2,HCCLM3with HPSE gene werehighly expressed, HCCLM3with HPSE highest in three groups.The pEGFP-1-RNAiand pEGFP-1-HPSE has been constructed,and meet the test requirements identified.All vectors can be specifically expressed in cancer cells, and the ability to filter outinterference HPSE expression strongest pEGFP-1-RNAi1, the maximum inhibitoryeffect on the expression of HPSE.Conclusions: Normal human liver cells (LO-2) HPSE cDNA expression was negative,HCCLM3, HepG-2, Bel-7402, three groups of human hepatoma cells, HCCLM3highest expression。Successfully constructed pEGFP-RNAi and pEGFP-1-HPSE andscreened effective target.