Xu Yang, Wang Wenling, Gou Aihong, Li Haitao, Tian Yanli, Yao Meihua, Yang Rongya
Chinese PLA Medical School, Beijing 100853, P.R. China.
Department of Dermatology, The General Hospital of Beijing Military Command, Beijing 100700, P.R. China.
Mol Med Rep. 2015 Jan;11(1):583-8. doi: 10.3892/mmr.2014.2674. Epub 2014 Oct 16.
The aim of the current study was to observe the effects of suppressor of cytokine signaling 1 (SOCS1) silencing in human melanoma cells on cell biological behavior and interferon-γ (IFN-γ) sensitivity, and to investigate the use of SOCS1 as a therapeutic target in the treatment of melanoma. Western blot analysis and reverse transcription-quantitative polymerase chain reaction (RT-qPCR) were used to verify that SOCS1 interference effectively silenced the expression of SOCS1 in the Mel526 human melanoma cell line. For IFN-γ stimulation, western blot analysis was used to observe changes in expression levels of signal transduction and transcription activator (STAT) 1 and phosphorylated STAT (pSTAT) 1. Changes in the expression levels of IFN-γ regulatory factor 1 (IRF-1) were measured with RT-qPCR. Changes in the sensitivity of melanoma cells to IFN-γ were detected using an MTT assay. The cell proliferation rate was observed by cell counting and changes in the cell cycle were detected with flow cytometry. The results revealed that SOCS1 interference effectively silences SOCS1 expression in Mel526 cells. However, the S stage of the cell cycle was markedly extended. Following the inhibition of SOCS1 expression, the proliferation experiment demonstrated that the proliferation ability of Mel526 cells was decreased. Following IFN-γ stimulation, the expression levels of pSTAT and IRF-1 increased significantly compared with those in the controls. The MTT experiment showed that SOCS1 interference caused the median inhibitory concentration (IC50) of oxaliplatin in Mel526 cells to decrease significantly. In conclusion, SOCS1 interference reduced the proliferation ability of Mel526 human melanoma cells and increased their sensitivity to IFN-γ.
本研究的目的是观察细胞因子信号转导抑制因子1(SOCS1)沉默对人黑色素瘤细胞生物学行为及干扰素-γ(IFN-γ)敏感性的影响,并探讨将SOCS1作为黑色素瘤治疗靶点的应用价值。采用蛋白质免疫印迹分析和逆转录-定量聚合酶链反应(RT-qPCR)验证SOCS1干扰可有效沉默人黑色素瘤细胞系Mel526中SOCS1的表达。对于IFN-γ刺激,采用蛋白质免疫印迹分析观察信号转导及转录激活因子(STAT)1和磷酸化STAT(pSTAT)1表达水平的变化。用RT-qPCR检测IFN-γ调节因子1(IRF-1)表达水平的变化。采用MTT法检测黑色素瘤细胞对IFN-γ敏感性的变化。通过细胞计数观察细胞增殖率,并用流式细胞术检测细胞周期的变化。结果显示,SOCS1干扰可有效沉默Mel526细胞中SOCS1的表达。然而,细胞周期的S期明显延长。抑制SOCS1表达后,增殖实验表明Mel526细胞的增殖能力下降。IFN-γ刺激后,与对照组相比,pSTAT和IRF-1的表达水平显著升高。MTT实验表明,SOCS1干扰使奥沙利铂对Mel526细胞的半数抑制浓度(IC50)显著降低。综上所述,SOCS1干扰降低了人黑色素瘤细胞Mel526的增殖能力,并增加了其对IFN-γ的敏感性。