中文摘要：

目的巨噬细胞凋亡在动脉粥样斑块的发展、病灶的坏死、斑块的不稳定以及急性血管事件中起着重要的作用,因此研究死亡结构相关蛋白（Daxx）对巨噬细胞凋亡的调控作用有利于揭示新的药物靶点,为动脉粥样硬化的发生与发展提供理论依据。方法应用脂质体2000将p CDNA3.1-Daxx、p CDNA3.1转入RAW264.7细胞中,G418（500 mg/L）筛选15天后,建立过表达Daxx稳定细胞株,RT-PCR和Western blot检测稳定细胞株内Daxx的转染效率。模型组和转染的细胞用胆固醇β环糊精（Chol∶MβCD）孵育48 h,建立RAW264.7荷脂和细胞凋亡模型,运用酶荧光法检测细胞内胆固醇蓄积情况,MTT检测细胞活性,流式细胞术观察细胞凋亡。Western blot测定细胞内Caspase3蛋白的表达。结果 RT-PCR和Western blot检测转染细胞内Daxx mRNA和蛋白明显高表达,表明稳定转染成功。在Chol∶MβCD的诱导下,模型组细胞内胆固醇的含量升高,细胞活性下降,凋亡数目增加。而Daxx高表达组可以加剧Chol∶MβCD诱导的胆固醇蓄积,细胞活性显著下降,凋亡细胞数目明显增加;同时,Daxx转染细胞组Caspase3蛋白表达明显增高。结论过表达Daxx可以加剧胆固醇β环糊精诱导的巨噬细胞凋亡。

英文摘要：

Aim Macrophage apoptosis plays an important role in development of atherosclerotic plaques,necrotic lesions,plaque instability and acute vascular events,so the study of the regulation of Daxx macrophage apoptosis is favorable to reveal new drug targets,providing a theoretical basis for the occurrence of atherosclerosis. Methods The eukaryotic vector of Daxx and p CDNA3. 1 were constructed to be stably transfected into RAW264. 7 cells,the mRNA and protein expressions of Daxx were used by RT- PCR and Western blot respectively. The intracellular lipid droplets and lipid levels was assayed by enzyme fluorescent analysis. The growth inhibition and apoptotic effect of RAW264. 7 cells induced by Chol∶ MβCD were analyzed by MTT and flow cytometric analysis. The protein expressions of caspase3 was assayed by western blotting. Results RT-PCR and Western blot analysis showed that the expression of Daxx was signifi-cantly increased in gene-transfected RAW264. 7 cells. Chol ∶ MβCD induced cholesterol accumulation and apoptosis in RAW264. 7 cells was increased by Daxx. Caspase3 protein levels were significantly elevated in Daxx transfected cells.Conclusion Daxx overexpression could increase Chol ∶ MβCD induced apoptosis in macrophages.