To Explore The Signal Transduction Of Hepatoma HepG2 Cells Transfected With NOR₁ Gene

Objective:To investigate the function of NOR1 in hepatoma,a NOR₁ over-expressed HepG2 cell line was established by gene transfection.We screen the differentially expressed genes in NOR₁ over-expressed HepG2 cells by cDNA microarrays and validation of the differentially expressed genes by RT-PCR,real-time quantitative PCR and western blot.Based on our experimental data and the data of signal transduction studio, hypothesize the signal transduction network.Methods:1 Screening for the differentially expressed genes in NOR₁ over-expressed HepG2 cells by cDNA microarrays.2 Validation of the differentially expressed genes by RT-PCR,real-time quantitative PCR and western blot.3 Based on our experimental data and the data of signal transduction studio,hypothesize the signal transduction network.Results:1 Using cDNA microarray,we showed that 59 genes up-regulated, such as Grb2,HBP17,TNFRSF11B.In addition,103 genes down-regulated,which included Bik,MAP2K6,ZFP95 et al.2 The cDNA microarray data had been validated by real-time fluorogenetic quantitative PCR.The real-time fluorogenetic quantitative PCR results showed that Grb2,HBP17,TNFRSF11B in pcDNA3.1(+)NOR₁/HepG2 were all up-regulated compared with pcDNA3.1(+)/HepG2.The relative quantification were 2.34,3.14,6.79 respectively.The three genes were also up-regulated compared pcDNA3.1(+)NOR₁/HepG2 with HepG2,the relative quantification were 4.87,5.23,6.33 respectively.The real-time fluorogenetic quantitative PCR results agreed with the microarray data.The result of RT-PCR of Grb2 and the western blot result of Grb2 also agreed with the cDNA microarray results.3 Based on our experimental data and the data of signal transduction studio,hypothesize the signal transduction network.Conclusions:1 The results of cDNA microarray showed 59 genes up-regulated and 103 genes down-regulated.2 The results of cDNA microarray,RT-PCR,Real-time fluorogenetic quantitative PCR and western blot uniformly showed NOR₁ may influence the HepG2's biological behaviors by its effect on the expression of cell signal transduction,cell cycle regulation,transcription regulation and translation control related genes.3 Based on our experimental data and the data of signal transduction studio,we hypothesize that during the entire procedure of tumorigenesis and tumor development,NOR₁ gene may play a regulatory role in the signal transduction pathway.