SOCS1基因沉默对人胰腺癌细胞增殖和干扰素-γ敏感性的影响

摘要

目的：在人胰腺癌细胞中观察细胞因子信号转导抑制因子1（SOCS1）沉默对细胞增殖及干扰素-γ（IFN-γ）敏感性的影响，探讨SOCS1作为胰腺癌治疗靶点的可能。方法 Western blot及PCR验证SOCS1干扰序列沉默人胰腺癌细胞系PANC1中SOCS1的表达；给予IFN-γ刺激后，采用Western blotting方法观察转录激活因子（STAT）1及磷酸化STAT（pSTAT）1的变化；采用定量PCR的方法观察IFN-γ调节因子1（IRF-1）表达水平变化；MTT法检测胰腺癌细胞对IFN-γ敏感性的变化；细胞计数的方法观察细胞的增殖速度；采用流式细胞术的方法检测细胞周期的变化。结果将SOCS1干扰序列转染PANC1细胞后，SOCS1 mRNA及蛋白表达水平均明显下降。沉默SOCS1表达后，pshSOCS1-PANC1组细胞的IRF-1 mRNA水平及pSTAT1蛋白表达水平均显著升高（P<0.05），IFN-γ对PANC1细胞的半数抑制浓度（IC50）显著降低（P<0.01）；转染72 h后PANC1细胞数量较对照组显著减少（P<0.05）；SOCS1表达抑制后PANC1细胞G0/G1期细胞比例明显升高，而S期和G2/M期细胞比例明显减小，与对照组比较差异均有统计学意义。结论 SOCS1表达抑制后，人胰腺癌细胞株PANC1的增殖能力下降，并且对IFN-γ的敏感性增强。%Objective To detect the changes of cell proliferation and IFN-γsusceptibility of human pancreatic can-cer cells after suppressor of cytokine signaling-1 (SOCS1) gene silencing, and to explore the SOCS1 as the target of anti-tu-mor therapy through enhancing the function of IFN-γ. Methods Western blot assay, PCR and real-time PCR were used to verify the down regulation of SOCS1 in human pancreatic cancer cell (PANC1) after transfection;subsequently, PANC1 was stimulated with IFN-γ. Western blot assay was also used to detect the expression of signal transducers and activators of tran-scription (STAT)1 and phosphorylation STAT(pSTAT)1;and the change of IFN-γsusceptibility was detected by MTT assay. Real-time PCR was used to detect the mRNA of interferon regulatory factor-1(IRF-1). Flow cytometry was used to detect the cell cycle. Results The expression levels of SOCS1 mRNA and protein were significantly decreased in small hairpin SCOS1 (shSOCS1) transfected PANC1 cells. After the silence of SOCS1, the expression levels of IRF-1 and pSTAT1 in-creased significantly (P<0.05), and the median inhibitory concentration(IC50)of IFN-γfor PANC1 cells decreased signifi-cantly (P<0.01). The cell count of shSOCS1 cells dropped significantly compared with that of control group after the SOCS1 silencing for 72 hours (P<0.05). The cell cycle arrest was promoted at the G0/G1 phase, but the percentage of cells in S phase and G2/M decreased compared to that of control groups (P<0.05). Conclusion After the inhibition of SOCS1 gene expression, the proliferation ability of human pancreatic cancer cell line PANC1 decreased, and the sensitivity of PANC1 cells to IFN-γwas enhanced.