核因子-κB活化对肿瘤坏死因子表达的调控

EFFECTS OF THE ACTIVATION OF NF κ B ON THE EXPRESSION OF TUMOR NECROSIS FACTORα

为观察脂多糖（ＬＰＳ）处理单核细胞（Ｍ）后对核因子－κＢ抑制蛋白（ＩκＢα）降解，ＮＦ－κＢ活化及α肿瘤坏死因子（ＴＮＦα）表达的影响，分离、培养Ｍ，分为正常对照组和ＬＰＳ刺激组（ＬＰＳ１０μｇ／ｍ ｌ）；分别在刺激前（０）、刺激后１５、３０ｍ ｉｎ 和１、２、４ｈ，留取Ｍ 和细胞培养上清。检测Ｍ ＩκＢ水平和核提取物中ＮＦ－κＢ的活性和细胞培养上清中ＴＮＦα的含量。结果发现，ＬＰＳ刺激组ＩκＢ水平１５ｍ ｉｎ 开始降低，３０ｍ ｉｎ 降到最低值；ＮＦ－κＢ活性在３０ｍ ｉｎ～４ｈ 显著增高，峰值在刺激后１～２ｈ；ＬＰＳ刺激后１～２ｈ，ＴＮＦα显著升高（Ｐ＜ ０．０１），峰值在刺激后１ｈ。ＮＦ－κＢ活性与ＴＮＦα含量在刺激后１ｈ 峰值呈显著正相关（ｒ＝ ０．９０１，Ｐ＜ ０．０１）。结果提示， ＬＰＳ可诱导Ｍ 的ＩκＢ降解和ＮＦ－κＢ激活。ＮＦ－κＢ活化后可导致ＴＮＦα的基因转录和表达增加。

To investigate effects of the level of IκBα and the activation of NF κB on the expression of tumor necrosis factorα ( TNFα) in monocytes(M) stimulated with lipopolysaccharide(LPS). Monocytes from volenteers were cultured and divided into two groups: Control group:M were not stimulated with LPS; LPS stimulation group: M were stimulated with LPS(10μg/ml). Supernatant and M were harvested at 0、15、30min,and 1、2、4 h after LPS stimulation. The level of IκBα, the activation of NF κ B and the concentration of TNF were measured by ELISA and electrophoretic mobility shift assay(EMSA), respectively. Results (1) The level of IκBα significantly decreased from 15min to 60min after LPS stimulation. The trough value was obtained at 30min. (2) The activity of NF κB significantly increased from 30min to 4h after LPS stimulation( P <0.01); The peak value was seen at 1 to 2h. (3) The level of TNFα in the supernatant elevated markedly from 1 to 2h after LPS stimultion ( P <0.01); peaking at 1h. Positive correlation was found between the NF κB activity and the TNFα content in the supernatant( r =0.901, P <0.01). Conclusions LPS might induce downregulation of IκB, activation of NF κB and upregulation of transcription and expression of TNFα gene in M.