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目的 探讨人核糖体蛋白S13(RPS13)在胃癌多药耐药 (MDR)机制中的作用。方法 采用RT PCR法扩增RPS13cDNA片段编码区序列全长 ,DNA重组技术构建正反义真核表达载体 ,经脂质体介导转染胃癌细胞SGC790 1及胃癌耐药细胞SGC790 1/VCR ,斑点杂交检测转染细胞mRNA水平的变化 ;MTT法测定细胞对化疗药物的敏感性 ,流式细胞仪检测细胞周期。结果 RT PCR法成功扩增出RPS13cDNA片段编码区序列全长 ,并构建正反义真核表达载体 ;斑点杂交试验证实 :正义转染细胞RPS13mRNA水平上调 ,反义转染细胞其mRNA水平下调。RPS13正义核酸转染SGC790 1细胞后 ,细胞对阿霉素、5 氟尿嘧啶和长春新碱的敏感性降低 ;转染反义核酸后 ,耐药细胞对丝裂霉素和长春新碱的敏感性增加。细胞周期测定表明高表达RPS13后 ,G1期、S期和G2期细胞的比例分别为 4 7.0 %、33.2 %和 19.8% ;低表达RPS13后 ,G1期、S期和G2期细胞的比例分别为 6 2 .9%、1.0 %和 36 .1%。结论 RPS13参与胃癌耐药细胞SGC790 1/VCR的多药耐药。
Objective To investigate the role of human ribosomal protein S13 (RPS13) in the multidrug resistance (MDR) mechanism of gastric cancer. Methods The full-length coding region of RPS13 cDNA was amplified by RT-PCR. The sense and antisense eukaryotic expression vector was constructed by DNA recombination technique. The plasmid was transfected into SGC790 1 cells and SGC790 1 / VCR cells by liposome. The changes of mRNA level of transfected cells were detected by hybridization. The sensitivity of cells to chemotherapeutic drugs was determined by MTT assay. The cell cycle was detected by flow cytometry. Results The full-length coding sequence of RPS13 cDNA was amplified by RT-PCR and the sense and antisense eukaryotic expression vectors were constructed. Dot blot hybridization showed that RPS13 mRNA was up-regulated in antisense transfected cells and down-regulated in antisense transfected cells. The sensitivity of cells to doxorubicin, 5-fluorouracil and vincristine was decreased after transfection of RPS13 sense nucleic acid into SGC7901 cells; the sensitivity of resistant cells to mitomycin and vincristine was increased after transfected with antisense nucleic acid . Cell cycle analysis showed that the percentage of cells in G1, S and G2 phase was 4 7.0%, 33.2% and 19.8% respectively after RPS13 was overexpressed. The percentage of cells in G1, S and G2 phase after RPS13 was low was 6 2 .9%, 1.0% and 36 .1%. Conclusion RPS13 is involved in the multidrug resistance of gastric cancer cell line SGC790 1 / VCR.