12C对人肝癌SMMC-7721细胞中survivin表达的影响

目的 通过采用RNA干扰技术下调survivin基因在人肝癌SMMC-7721细胞中的表达,观察survivin表达下调对细胞G₂/M期阻滞、细胞凋亡和重离子辐射敏感性的影响。方法 针对survivin mRNA体外化学合成小干扰RNA(siRNA),转染细胞,以实时 PCR方法检测转染后24和48 h细胞survivin mRNA的表达。Annexin-FITC检测细胞的凋亡情况;流式细胞仪检测周期阻滞情况;克隆存活法确定细胞的辐射敏感性。结果 转染24和48 h后,细胞中survivin的表达量分别为未转染组的59%和39%;转染survivin siRNA后24 h,引起细胞G₂/M期阻滞,48 h阻滞明显。转染survivin siRNA 48 h后的细胞凋亡率为21.41%,明显高于未转染组细胞(t=9.13,P<0.01)。siRNA 转染组细胞受辐照后的存活率明显降低。结论 以siRNA下调survivin的表达可有效诱导细胞凋亡,引起G₂/M期阻滞,提高细胞对重离子的辐射敏感性。

Survivin as a factor of radioresistance to high-LET carbon ions in human hepatoma SMMC-7721 cells

Objective To investigate the influences of survivin down-regulation on cell G₂/M phase arrest,apoptosis and sensitivity to carbon ion irradiation.Methods Small interfering RNA (siRNA) targeting survivin mRNA was designed, in vitro chemo-synthesized and transfected into SMMC-7721 cells. Survivin mRNA expression in SMMC-7721 cells was measured by real-time PCR, and the apoptotic rates by Annexin-FITC at 24 and 48 h after transfection. Cell G₂/M phase arrest after transfection was assessed with flow cytometry as well. Cellular sensitivity to high-LET carbon ions was determined by means of colony-forming assay.Results The expressions of survivin at mRNA level were down-regulated to be 59% and 39% in relation to the non-treated cells at 24 and 48 h after siRNA transfection, respectively. G₂/M phase arrest in SMMC-7721 cells at 24 h after transfection was observed while much more obvious at 48 h. The apoptotic rate of SMMC-7721 cells was 21.41% at 48 h after survivin siRNA transfection, which was significantly higher than that of the cells transfected with negative siRNA. Moreover, a decreased clonogenic survival in siRNA treated group was shown.Conclusion Down-regulation of survivin gene expression in SMMC-7721 cells by siRNA could effectively induce cell apoptosis and G₂/M phase arrest, and enhance the cellular radiosensitivity to high-LET heavy ions.