SAA Protein Expression In CNE2Cells And Its Effect On Cell Growth And Apoptosis

Objective：In order to detecting its effects in Human Nasopharyngeal carcinom CNE-2cell,we use shRNA technology to estabish silence serum amyloid A (SAA).it willprovides an important experimental baisis for the association between theexpression of SAA and the development of nasopharyngeal carcinoma.Method：According to the principle of the short hairpin RNA (shRNA) design,weconstructed three shRNAs expression vector to target human SAA gene,then weconducted DNA sequencing and blasting.The eukaryotic expression vectors ofshRNA were transfected CNE2cells,we filtered out the effective siRNA sequence byRT-PCR and Western-blotting. By the technique of small interfering RNA and highexpression vectors,We detected its effects in Nasopharyngeal carcinom CNE2cell.By MTT and Hoechst33258fluorescent staining technique, We observed andanalyzed that how the expression of SAA influence the growth and apoptosis ofCNE2.Result：We successfully built disturbance eukaryotic expression vector group in SAAgene shRNA.By sequencing identification, blast, PT-PCR and Western blotting-detection, We determined that pGPU6/GFP/Neo-SAA-homo-482for SAAdesign is the most effective interference RNA genes.The recombinant plasmid vectorspGPU6/GFP/Neo-SAA-homo-482and pCD3.1(+)-SAA were successfullytransfected into CNE2nasopharyngeal carcinoma (NPC) cells.The result revealedthat pGPU6/GFP/Neo-SAA-homo-482and pCDNA3.1(+)-SAA effectively silenced and overexpressed the mRNA and protein of SAA by PT-PCR andWestern blotting, this result was the were statistically significant(P<0.05).By MTTand Hoechst33258fluorescence staining,we found that silencing of SAA genecould promote CNE2cell apoptosis and overexpression of SAA gene could accelerateCNE2cell growth.Conclusions：1、 pGPU6/GFP/Neo-SAA-homo-482could effectively silence the gene inCNE2cell.2、Silencing of SAA gene could promote CNE2cell apoptosis and overexpression ofSAA gene could accelerate CNE2cell growth.