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AIM:To construct the retroviral vector of p~(125FAK)specificribozyme genes and to explore the feasibility of ribozymein BGC-823 gene therapy in vitro.METHODS:A hammerhead ribozyme DNA targetingp~(125FAK)mRNA from nt 1010 to nt 1032 was synthesizedand recombinated into the retroviral vector pLXSN form-ing pLRZXSN recon.Using the lipofectin-mediated DNAtransfection technique,pLRZXSN was introduced intoBGC-823 cells.The effects of ribozyme on the growth ofBGC-823 cells and apoptosis were studied by cell colonyassay,flow cytometry(FCM),reverse transcriptase-polymerase chain reaction(RT-PCR),detection of DNAfragmentation and electron microscopy.RESULTS:The number of BGC-823 cell colonies wasinhibited by 56% after the cells were treated for 48 h.The cell proliferation was inhibited effectively by p~(125FAK)ribozyme and the inhibitory effect depended on the con-centration and the time of incubation.The expressionof p~(125FAK)mRNA and protein p~(125) decreased sharply inBGC-823 cells treated with p~(125FAK)ribozyme.The charac-teristics of apoptosis,namely sub-G1 peak,DNA frag-mentation and morphological changes,were revealed inBGC-823 cells treated with p~(125FAK) ribozyme.CONCLUSION:p~(125FAK) ribozyme decreases p~(125FAK)geneexpression and induces apoptosis of human gastric can-cer cells in vitro.
AIM: To construct the retroviral vector of p ~ (125FAK) specific ribozyme genes and to explore the feasibility of ribozymein BGC-823 gene therapy in vitro. METHODS: A hammerhead ribozyme DNA targeting p ~ (125FAK) mRNA from nt 1010 to nt 1032 was synthesizedand recombinated into the retroviral vector pLXSN form-ing pLRZXSN recon. Using the lipofectin-mediated DNA transfection technique, pLRZXSN was introduced into BGC-823 cells. The effects of ribozyme on the growth of BGC-823 cells and apoptosis were studied by cell colony assay, flow cytometry FCM), reverse transcriptase-polymerase chain reaction (RT-PCR), detection of DNAfragmentation and electron microscopy .RESULTS: The number of BGC-823 cell colonies was inhibited by 56% after the cells were treated for 48 h. effectively by p ~ (125FAK) ribozyme and the inhibitory effect depended on the con-centration and the time of incubation. The expression of p ~ (125FAK) mRNA and protein p ~ 125 decreased sharply inBGC-823 cells treated w ith p ~ (125FAK) ribozyme.The charac-teristics of apoptosis, that sub-G1 peak, DNA frag mentation and morphological changes, were revealed in BGC-823 cells treated with p ~ (125FAK) ribozyme.CONCLUSION: p ~ ) ribozyme decreases p ~ (125FAK) geneexpression and induces apoptosis of human gastric can-cer cells in vitro.