摘要
目的:了解SC58125与TNF-α是否具有协同诱导HT29细胞凋亡的作用,同时探讨其可能的分子机制。方法:用MTT、Hoechst 33342染色及琼脂糖凝胶电泳,观察SC58125/TNF-α对HT29结肠癌细胞增殖与凋亡的作用;用EMSA及Western blotting检测转录因子NF-κB的结合活性、IκBα以及磷酸化IκBα的表达。结果:SC58125与TNF-α在抑制HT29细胞增殖及诱导其凋亡方面具有明显的协同作用;表现为细胞核浓缩、核碎裂及"DNA梯带"形成;凋亡过程中伴随caspase活性的激活。经TNF-α刺激后的HT29细胞,IκBα,磷酸化IκBα迅速降解,NF-κB的结合活性大大提高,而加入SC58125后,IκBα降解及NF-κB的结合活性明显受抑制。结论:SC58125与TNF-α在诱导HT29细胞凋亡方面具有明显的协同作用,这可能与激活caspase-3及抑制IκBα降解有关。
AIM: To investigate whether SC58125 synergizes with TNF-α to induce HT- 29 cell apoptosis and study the possible molecular mechanism. METHODS: By using MTT, Hoechst 33342 staining and agarose gel electrophoresis, the effect of SC58125/TNF -α on the proliferation and apoptosis of HT -29 cell line was examined. The activity of caspase - 3, the expression of IκBα, the phosphorylation level of IκBα, and the activation of NF -κB were measured after treatment with SC58125 by electrophoretic mobility shift assay and Western blotting. RESULTS: Both SC58125 and TNF- α exhibited cytotoxicity. The combination of the two reagents significantly reduced HT -29 cell viability in a dose - dependent manner. SC58125 and TNF - α co - treated cells showed induced nuclear condensation and fragmentation, and led to oligonucleosomal cleavage of genomic DNA, which was accompanied by the induction of caspase activity. IκBα levels were substantially decreased by the treatment of TNF - α. The degradation of IκBα was almost completely inhibited when SC58125 was added. NF -κB was activated in HT -29 cells after treatment with TNF -α, whereas pretreatment of HT- 29 cells with SC58125 for 2 h, TNF -α induced NF -κB DNA binding was profoundly suppressed. CONCLUSION: SC58125 synergizes with TNF -α to inhibit cell growth and induces apoptosis in HT- 29 cells, which may be mediated by activating caspase -3 and preventing degradation of IκBα .
出处
《中国病理生理杂志》
CAS
CSCD
北大核心
2007年第8期1499-1502,共4页
Chinese Journal of Pathophysiology
基金
国家科技部重大基础研究前期研究专项资助项目(No.2002ccc0400)
2003年广东省科技计划资助项目(No.2003B30101)
2004年教育部博士点科研基金资助项目(No.20040559008)
2004年广东省卫生厅青年基金资助项目(No.B2004072)