【摘要】 目的研究佛手柑内酯是否能抑制肺癌细胞分泌可溶性白细胞介素2受体(s IL-2R)及探讨其可能机制。方法将浓度梯度(1μmol/L、10μmol/L和100μmol/L)的佛手柑内酯处理肺癌细胞(Calu-3和NCI-H292)后,利用酶联免疫吸附反应检测肺癌细胞上清s IL-2R的含量,利用实时定量聚合酶链反应检测肺癌细胞s IL-2R编码基因IL2RA的表达。利用生物信息学分析IL2RA是否受转录因子STAT3的调控,并预测IL2RA启动子区域内STAT3的结合位点。利用染色体免疫共沉淀和双荧光素酶报告基因技术,观察佛手柑内酯处理后,STAT3对IL2RA启动子结合及转录激活能力的变化。每次实验设3个复孔,每个实验重复3次以上,两组间数据比较采用t检验,多组间数据比较采用单因素方差分析,检验水平α=0.05。结果在排除细胞活性的影响后,佛手柑内酯能呈浓度梯度抑制肺癌细胞分泌s IL-2R,并抑制其编码基因IL2RA的表达。公共数据库分析显示,IL2RA受转录因子STAT3的调控,其启动子区域内有两个STAT3的结合位点。佛手柑内酯处理肺癌细胞后,能呈浓度梯度抑制STAT3对IL2RA启动子区域的结合,并能降低STAT3对IL2RA的转录激活。结论佛手柑内酯能在体外模型中,通过抑制STAT3对IL2RA启动子区域的结合及转录激活,阻抑肺癌细胞分泌s IL-2R。更多还原

【Abstract】 Objective To investigate the effect of bergapten suppressing the expression of soluble interleukin-2 receptor(s IL-2R) in lung cancer cell. Methods The lung cancer cells(Calu-3 and NCI-H292) were treated by bergapten on different concentration(1 μmol/L, 10 μmol/L, and 100 μmol/L), and the s IL-2R in cultural supernatant was detected by Enzyme linked immunosorbent assay(ELISA). The IL2 RA m RNA level in lung cancer cells were measured by Real-time quantitive polymerase chain reaction(real-time PCR), and the binding sites that STAT3 recognizing on the promoter of IL2 RA were predicted by bioinformatics software. The Chromatin immunoprecipitation was using to detect the binding between STAT3 and promoter of IL2 RA, and the dual-luciferase reporter gene assay was present to detect the transcriptional activity of STAT3 on promoter of IL2 RA. All values were presented as means±standard deviation(SD). Student’s t-test was used to determine statistical differences between two groups, and One-Way ANOVA analysis was used to determine statistical differences among more than two groups. Results The concentration of s IL-2R and the IL2 RA m RNA level were decreased on lung cancer cell by treating bergapten in dose-dependent. The bioinformatics software and publication array datasets were showed that IL2 RA could be transcriptional activated by STAT3, and there were two predicted binding sites of STAT3 on the promoter of IL2 RA. The binding and the transcriptional activity of STAT3 on the promoter of IL2 RA was suppressed by treating bergapten on lung cancer cell in dose-dependent. Conclusions Bergapten could suppresses lung cancer cells secreting soluble interleukin-2 receptor by inhibiting the binding and the transcriptional activity of STAT3 on the promoter of IL2 RA.更多还原