AIM： To construct the retroviral vector of p^125FAK specific ribozyme genes and to explore the feasibility of ribozyme in BGC-823 gene therapy in vitro. METHODS： A hammerhead ribozyme DNA targeting p^125FAK mRNA fro...AIM： To construct the retroviral vector of p^125FAK specific ribozyme genes and to explore the feasibility of ribozyme in BGC-823 gene therapy in vitro. METHODS： A hammerhead ribozyme DNA targeting p^125FAK mRNA from nt 1010 to nt 1032 was synthesized and recombinated into the retroviral vector pLXSN forming 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 transcriptasepolymerase chain reaction （RT-PCR）, detection of DNA fragmentation and electron microscopy. RESULTS： The number of BGC-823 cell colonies was inhibited 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 concentration and the time of incubation. The expression of p^125FAK mRNA and protein p^125 decreased sharply in BGC-823 cells treated with p^125FAK ribozyme. The characteristics of apoptosis, namely sub-G1 peak, DNA fragmentation and morphological changes, were revealed in BGC-823 cells treated with p^125FAK ribozyme. CONCLUSION： p^125FAK ribozyme decreases p^125FAK gene expression and induces apoptosis of human gastric cancer cells in vitro.展开更多
基金Supported by the Natural Science Foundation of Fujian Province,No.C0010015
文摘AIM： To construct the retroviral vector of p^125FAK specific ribozyme genes and to explore the feasibility of ribozyme in BGC-823 gene therapy in vitro. METHODS： A hammerhead ribozyme DNA targeting p^125FAK mRNA from nt 1010 to nt 1032 was synthesized and recombinated into the retroviral vector pLXSN forming 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 transcriptasepolymerase chain reaction （RT-PCR）, detection of DNA fragmentation and electron microscopy. RESULTS： The number of BGC-823 cell colonies was inhibited 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 concentration and the time of incubation. The expression of p^125FAK mRNA and protein p^125 decreased sharply in BGC-823 cells treated with p^125FAK ribozyme. The characteristics of apoptosis, namely sub-G1 peak, DNA fragmentation and morphological changes, were revealed in BGC-823 cells treated with p^125FAK ribozyme. CONCLUSION： p^125FAK ribozyme decreases p^125FAK gene expression and induces apoptosis of human gastric cancer cells in vitro.
