| Objective:To explore the effects of sh RNA-mediated down-regulation of the RACK1 gene on the chemotherapeutic sensitivities in human lung adenocarcinoma cell line A549 cells,and to investigate its possible mechanism.Methods :The sh RNA recombinant plasmid targeting to hunmn RACK1 gene was designed,transferred into A549 cells by lipofectin technique.Experiments were divided into three groups: RACK1-sh RNA group, Vector-sh RNA group and Control group.(1)Transfection efficiency of A549 cells was analysed by fluorescence microscopy.(2)The proteins expressions of transfected A549 cells RACK1 was detected by Western blot.(3)MTT assays was used to analyse drug sensitivities of A549 cells to cisplatin,gemcitabine,pemetrexed and paclitaxel.(4)The proteins expressions of LRP,MRP were detected by Western blot.Results:1 After 24 hours of transfection,the green fluorescence in the A549 cells could be seen under the fluorescence microscope,which suggests the success of the transfection;2 After 36 hours of transfection,The relative expression quantity of RACK1 pr otein in RACK1-sh RNA group was 0.267±0.47, which was significantly lower than that in Vector-sh RNA group(RQ= 0.821±0.109) and Control group(RQ=0.842±0.060),( F=54.438,p<0.05);3 MTT showed that the growth of A549 cells in the RACK1-shRNA group was markedly inhibited.The sensitivities of A549 cells to cisplatin and paclitaxel were significantly enhanced compared to that in the Vector-sh RNA group and Control group(p<0.05);4 Western-blot analysis indicated that the relative expression quantity of LRP and MRP protein in RACK1-sh RNA group were 0.163±0.056 and 0.246±0.050, which were lower than that in Vector-sh RNA group and Control group( FLRP=19.430,FMRP=61.548,p<0.05).Conclusion:sh RNA targeting to inhibit gene RACK1 expression in A549 cells significantly increased the sensitivities of A549 cells to the ascisplatin and paclitaxel medicines, which might be through down-regulation of the expressions of LRP and MRP. |
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