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目的优化不同长度聚肌苷酸胞苷酸[polyinosinic acid:polycytidylic acid,poly(I∶C)]在HEK293T和BEAS-2B细胞中激活Ⅰ型干扰素(IFNβ)荧光素酶报告基因系统的实验条件。方法将质粒IFN-β-luc分别转染HEK293T及BEAS-2B细胞后,HEK293T细胞转染不同浓度(0.25、0.5、1.0、1.5和2.0μg/孔)的高分子量(HMW)poly(I∶C)和低分子量(LMW)poly(I∶C),检测其对HEK293T细胞中IFNβ启动子激活的影响;BEAS-2B细胞直接加入不同浓度(0.1、1.0、10.0、20.0和50.0μg/孔)及转染不同浓度(同HEK293T细胞)的HMW poly(I∶C)和LMW poly(I∶C),检测其对BEAS-2B细胞中IFNβ启动子激活的影响。结果 poly(I∶C)转染HEK293T细胞激活IFNβ启动子的最佳实验条件为:0.5μg/孔的HMW poly(I∶C)和LMW poly(I∶C)转染24 h。poly(I∶C)直接加入BEAS-2B细胞激活IFNβ启动子的最佳实验条件为:50μg/孔的HMW poly(I∶C)和LMW poly(I∶C)作用6 h;poly(I∶C)转染BEAS-2B细胞激活IFNβ启动子的最佳实验条件为:0.25μg/孔的HMW poly(I∶C)转染12 h,0.5μg/孔的LMW poly(I∶C)转染12 h。结论成功优化了poly(I∶C)在HEK293T和BEAS-2B细胞中激活IFNβ启动子的实验条件,为poly(I∶C)激活免疫反应机制的研究提供了实验依据。
Objective To optimize the experimental conditions for activation of IFNβ luciferase reporter system in polycytidylic acid (poly (I: C)] cells of different lengths in HEK293T and BEAS-2B cells . Methods HEK293T cells were transfected with high molecular weight (HMW) poly (I: C ) And low molecular weight (LMW) poly (I: C) were detected in HEK293T cells. BEAS-2B cells were treated with different concentrations of 0.1,1.0,10.0,20.0 and 50.0μg / well and The effect of IFNβ promoter activation in BEAS-2B cells was examined by transfecting HMW poly (I: C) and LMW poly (I: C) at different concentrations (same as HEK293T cells). Results The optimal conditions of HEK293T cells transfected with poly (I: C) were as follows: 0.5μg / well HMW poly (I: C) and LMW poly (I: C) were transfected for 24 hours. The optimal conditions of poly (I: C) BEAS-2B cells activating IFNβ promoter were as follows: 50μg / well HMW poly (I: C) and LMW poly C) Transfection of BEAS-2B cells activated IFNβ promoter with the best experimental conditions: 0.25μg / well of HMW poly (I: C) transfected 12h, 0.5μg / well of LMW poly 12 h. Conclusion The experimental conditions of poly (I: C) activating IFNβ promoter in HEK293T and BEAS-2B cells were successfully optimized, which provided an experimental basis for the study of the mechanism of poly (I: C) activating immune response.