Study On Effects Of Gene Expression Profiles On HepG2 Cell Line Of NOR₁ Gene

Objective:1 To establish the stable transfection of hepatoma cell line pcDNA3.1(+)/NOR₁-HepG2.2 To study the effects of the gene expression profiles of human hepatocarcinoma HepG2 cells after transfection of NOR₁ gene with microarray assay and Real-time PCR.3 To discuss the possible mechanism between NOR₁ gene and up-regulated/down-regulated in the occurrence and progression of human hepatocarcinoma.Methods:1 The mammal expression vector of NOR₁(pcDNA3.1(+) /NOR₁)and eukaryotic expressing plasmid pcDNA3.1(+)were introduced into human hepatocarcinoma cell line HepG2 mediated by lipofectamin transfection.The stable G418-resistant clones were isolated. After extracting total RNA,the overexpression of NOR₁ gene were detected by RT-PCR.2 The differential expression profiles genes from NOR₁-transfected cells(HepG2-NOR₁)and empty vector cells(HepG2-PC)were determined using the human 17K cDNA expression chip with 17101 cDNA probes.3 The expression of p15 and MAGE-A6 genes were verified by Real-time PCR. Results:1 Recombinant plasmid pcDNA3.1(+)/NOR₁ can express in HepG2 cells.Stable expression of NOR₁ gene in HepG2 cells was confirmed by RT-PCR.The lever of NOR₁ mRNA increased apparently in HepG2-NOR₁ cells compared with HepG2-PC cells.2 From the scanning results of gene chips,162 genes were differentialiy expressed in the two cells lines,among which 59 genes(or EST)expression were up-regulated and 103 genes(or EST)expression were down-regulated in the HepG2-NOR₁ cells.The genes were correlated with cell apoptosis or tumor,cell cycle,cell transcription regulator activity,cell signal transducer activity,cell translation regulator activity,and so on.3 The expression of P15 gene in HepG2-NOR₁ cells increased to 2.692 compared with HepG2-PC cells,and MAGE -A6 increased to 2.533 by Real-time PCR.Conclusion:1 Over-expression of NOR₁ gene by transfecting NOR₁ can change the gene expression profiles in HepG2 cells,leading to change of cell growth,differentiation and apoptosis in HepG2-NOR₁ cells.cDNA chip is a useful method in understanding the mechanism of cancer with its high throughput characteristics.2 The expression of P15 gene in HepG2-NOR₁ cells increased to 2.692 compared with HepG2-PC cells,and MAGE-A6 increased to 2.533 by Real-time PCR.P15 is a very important factor in the occurrence and progression of human cancer,one of the mechanisms of NOR₁ gene inhibited the growth of HepG2-NOR₁ cells was NOR₁ gene induced p15 gene up-regulatation.The study laid a solid foundation for the further research of mechanism of NOR₁ gene during the occurrence and progression of hepatocarcinoma.