【摘要】 目的A20基因是一种肿瘤坏死因子(TNF-α)诱导下的原始反应基因。其编码的锌指结构蛋白在抑制NF-κB和TNF-α介导的细胞凋亡方面得到了广泛的研究。但对于其在神经系统中的作用研究较少。该实验采用脂质体转染及克隆筛选将A20基因导入神经母细胞瘤(SY-SH-5Y细胞系)中,通过体外实验观察A20-SY-SH-5Y细胞在抗凋亡方面的表现。方法SY-SH-5Y细胞系转染A20基因后,应用肿瘤坏死因子(TNF-α)刺激,采用流式细胞仪(FACS)测定细胞的死亡率,并进行DNA片断化分析(DNA Ladder)。结果实验表明:转染A20的SY-SH-5Y细胞系在TNF-α的诱导下死亡率小于对照组转染pCDNA3基因的SY-SH-5Y细胞系(P<0.05)。结论A20作为一种在细胞抗凋亡过程中发挥关键作用的基因,能够在神经母细胞瘤SY-SH-5Y细胞系中表现出较强抗凋亡能力,为临床上许多以凋亡为主要表现的神经系统疾病的治疗提供了新思路。更多还原

【Abstract】 [Objective]A20 is a TNF-induced primary response gene which encodes a novel zinc finger protein which can inhibits tumor necrosis factor (TNF)-mediated apoptosis in many cell lines. But rare work was done in central nervous system. [Methods] A20 was also delivered into the cell line SY-SH-5Y by liposome,then the positive clone was picked by G418. The expression of A20 protein was identified with anti-A20 IgG by immunocytochemical staining. Then the cells overexpressing A20 were exposed to TNF-α stimuli, the mortality was measured by fluorescence-activated cell sorting (FACS). DNA fragmentation were also used to evaluate the degree of cell death. [Results] The immunocytochemical staining showed the SY-SH-5Y cell line which had been picked by clone filtration proved to be mostly A20 positive. The test in vitro showed that the apoptosis rates of A20-SY-SH-5Y was 9.59%±3.88%, much lower than the control group which was 41.52%±3.29% under the stimuli of TNF-α(P <0.05). The results of DNA fragmentation was also consistent with the FACS. [Conclusion] For SY-SH-5Y cell line, A20 gene can offer the mean of achieving an anti-apoptosis effect, and may play a key role for the treatment of many diseases of nervous system which apoptosis is their main manifestation.更多还原