【摘要】 核转录因子-3B(NF-4B)通过调节其靶基因的表达,在机体免疫应答、炎症与肿瘤的发生发展过程中发挥着重要功能,因此也被认为是预防和治疗肿瘤与炎症的理想靶点.pNiFty-SEAP质粒被成功地转入HEK293细胞,因此该转染细胞含有5×NF-5B基序的增强子、SEAP报告基因和Zeocin抗生素抗性筛选基因.通过检测报告基因表达水平可反映NF-6B的活化状态.在以pNiFty-SEAP/HEK293细胞为基础建立的筛选体系中,TNF7和PMA等已知的NF-8B激活剂能剂量与时间依赖性地促进NF-9B的活化,而PDTC和NAC等NF-:B抑制剂的作用则相反.该筛选体系不仅具有稳定、特异、高通量等特点,而且可同时检测NF-;B的激活剂与抑制剂,将为发现作用于NF-<B信号传导通路的抗肿瘤与免疫调节药物提供有效的实验方法.更多还原

【Abstract】 NF-κB has been reported to play an important role in immunity, inflammation and carcinogenesis byregulating expression of its target genes. Thus, NF-"B was recognized as a promising target for the prevention andtreatment drugs against cancer and inflammation. NF-#B sequence, reporter gene SEAP and selection gene forZeocine resistance in plasmid pNiFty-SEAP were transfected into HEK293 cells. Therefore, NF-$B activity wasable to be evaluated by SEAP activity in the transfected cells. Based on pNiFtySEAP/HEK293 cells, a screeningassay was established, by which known NF-%B stimulators TNF& and PMA were demonstrated to activate theNF-’B in dose and time dependent manners. In contrast, NF-(B inhibitors PDTC and NAC showed inhibitoryeffects on the NF-)B activity in indicated doses and times. Except several characteristics such as stability,sensitivity, sepecifity, the assay was capable to screen both agents inducing and inhibiting NF-*B activity at thesame experiment, which will be useful for the discovery of compounds targeting NF-+B signal pathway.更多还原