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目的 了解白血病细胞系 HL- 60中 ,肿瘤坏死因子 - α( TNF- α)诱导的核因子 - κB( NF- κB)活化与细胞凋亡之间的关系。方法 采用脂质体基因转染技术 ,将突变的核因子抑制蛋白 ( IκBα)质粒 ( p CMV4- IκBαS32 / 36 A)转染 HL-60细胞 ,于 48h后测其瞬时表达效应。用间接免疫荧光和 Western blot技术检测转染组和非转染组 HL- 60细胞的 NF-κB活化状态 ,同时用流式细胞术和 MTT法分别检测 TNF- α对两组 HL- 60细胞的凋亡诱导及生长抑制效应。结果 TNF- α可诱导非转染组 HL- 60细胞的 NF- κB活化 ,不能诱导转染组细胞的 NF- κB活化 ,即突变型 IκBα可特异性抑制HL- 60细胞的 NF- κB活化。结论 用突变型 IκBα特异性抑制 HL- 60细胞的 NF- κB活化 ,能明显增加其对 TNF- α的敏感性
Objective To investigate the relationship between the activation of nuclear factor - κB (NF - κB) induced by tumor necrosis factor - α (TNF - α) and apoptosis in leukemia cell line HL - 60. Methods The transfected HL-60 cells were transfected with the mutant plasmid pCMV4-IκBαS32 / 36 A by using lipofection method. The transient expression of HL-60 cells was detected after 48 hours. The activation of NF-κB in HL-60 cells was detected by indirect immunofluorescence and Western blot. The expressions of NF-κB in HL-60 cells were detected by flow cytometry and MTT assay Apoptosis induction and growth inhibitory effect. Results TNF-α induced NF-κB activation in non-transfected HL-60 cells and failed to induce NF-κB activation in transfected cells. That is, mutant IκBα specifically inhibited NF-κB activation in HL-60 cells. Conclusion The specific inhibition of NF-κB activation by mutant IκBα in HL-60 cells can significantly increase its sensitivity to TNF-α