目的:研究复方利肝冲剂对肝星状细胞NF-κB结合活性的影响,探讨复方利肝冲剂抗肝纤维化的主要作用机制。 方法:复方利肝冲剂药物血清与鼠肝星状细胞共孵育48h后,NF-κB结合活性的检测采用凝胶电泳移动抑制法,细胞培养液中细胞因子的检...目的:研究复方利肝冲剂对肝星状细胞NF-κB结合活性的影响,探讨复方利肝冲剂抗肝纤维化的主要作用机制。 方法:复方利肝冲剂药物血清与鼠肝星状细胞共孵育48h后,NF-κB结合活性的检测采用凝胶电泳移动抑制法,细胞培养液中细胞因子的检测采用ELISA法,细胞凋亡和细胞增殖分别采用流式细胞术和~3H-TdR掺入法检测。 结果:复方利肝冲剂药物血清使体外培养的鼠肝星状细胞NF-κB结合活性比无药血清对照组显著减弱(图1),细胞培养液中的IL-6和sICAM水平降低(1.56±0.42ng·L^(-1) vs 3.64±0.58ng·L^(-1),P<0.05;3.82±0.98ng·L^(-1) vs 12.64±1.22ng·L^(-1),P<0.01).星状细胞凋亡增加24.8％±3.60％ vs 6.6％±1.2％,P<0.01,增殖减少2623±654/孔 vs 5061±346/孔,P<0.01. 结论:复方利肝冲剂显著抑制体外培养的肝星状细胞NF-κB结合活性,可能为其抗纤维化的重要机制之一。展开更多
The developmental control of the human e-globin gene expression is mediated by transcription regulatory elements in the 5’ flanking DNA of this gene. Sequence analysis has revealed a DNA motif (GGGGAATTTGCT) similar ...The developmental control of the human e-globin gene expression is mediated by transcription regulatory elements in the 5’ flanking DNA of this gene. Sequence analysis has revealed a DNA motif (GGGGAATTTGCT) similar to NF-кB consensus sequence resides in the negative regulatory element (-3028bp～ -2902bp, termed ε-NRAII) 5’ to the cap site of this gene. NRF DNA fragment (-3010bp～ -2986bp) containing the NF-кB motif similar sequence was synthesized and used in electrophoresis mobility shift assay (EMSA) and competitive analysis. Data showed that a protein factor from nuclear extracts of K562 cells specifically interacted with NRF DNA fragment. The synthetic NF DNA fragment (containing NF-кB consensus sequence) could competed for the protein binding, but MNF DNA fragment (mutated NF-кB motif) could not, suggesting that the binding protein is a member of NF-кB/Rel family. Western blot assay demonstrated that the molecular weight of NF-кB protein in the nuclei of K562 cells is 50ku. We suggested that NF-кB p50 may play an important role in the regulation of human c-globin gene expression.展开更多
文摘目的:研究复方利肝冲剂对肝星状细胞NF-κB结合活性的影响,探讨复方利肝冲剂抗肝纤维化的主要作用机制。 方法:复方利肝冲剂药物血清与鼠肝星状细胞共孵育48h后,NF-κB结合活性的检测采用凝胶电泳移动抑制法,细胞培养液中细胞因子的检测采用ELISA法,细胞凋亡和细胞增殖分别采用流式细胞术和~3H-TdR掺入法检测。 结果:复方利肝冲剂药物血清使体外培养的鼠肝星状细胞NF-κB结合活性比无药血清对照组显著减弱(图1),细胞培养液中的IL-6和sICAM水平降低(1.56±0.42ng·L^(-1) vs 3.64±0.58ng·L^(-1),P<0.05;3.82±0.98ng·L^(-1) vs 12.64±1.22ng·L^(-1),P<0.01).星状细胞凋亡增加24.8％±3.60％ vs 6.6％±1.2％,P<0.01,增殖减少2623±654/孔 vs 5061±346/孔,P<0.01. 结论:复方利肝冲剂显著抑制体外培养的肝星状细胞NF-κB结合活性,可能为其抗纤维化的重要机制之一。
基金This work was supported by National Natural Sciences Foun-dation of China, No. 39893320 and No. 39870378.
文摘The developmental control of the human e-globin gene expression is mediated by transcription regulatory elements in the 5’ flanking DNA of this gene. Sequence analysis has revealed a DNA motif (GGGGAATTTGCT) similar to NF-кB consensus sequence resides in the negative regulatory element (-3028bp～ -2902bp, termed ε-NRAII) 5’ to the cap site of this gene. NRF DNA fragment (-3010bp～ -2986bp) containing the NF-кB motif similar sequence was synthesized and used in electrophoresis mobility shift assay (EMSA) and competitive analysis. Data showed that a protein factor from nuclear extracts of K562 cells specifically interacted with NRF DNA fragment. The synthetic NF DNA fragment (containing NF-кB consensus sequence) could competed for the protein binding, but MNF DNA fragment (mutated NF-кB motif) could not, suggesting that the binding protein is a member of NF-кB/Rel family. Western blot assay demonstrated that the molecular weight of NF-кB protein in the nuclei of K562 cells is 50ku. We suggested that NF-кB p50 may play an important role in the regulation of human c-globin gene expression.
