南方医科大学学报

南方医科大学学报 ›› 2013, Vol. 33 ›› Issue (02): 172-.

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反式激活蛋白-NEMO结合域抑制核因子-κB活化在胆红素诱导大鼠海马神经元凋亡中的作用

宋思捷，朱叶芳，党姗姗，王少雯，华子瑜   

  • 出版日期:2013-02-20 发布日期:2013-02-20

Role of nuclear factor-κB activation in bilirubin-induced rat hippocampal neuronal apoptosis and the effect of TAT-NBD intervention

  • Online:2013-02-20 Published:2013-02-20

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摘要： 目的明确核因子-κB（NF-κB）活化在胆红素诱导大鼠海马神经元凋亡中的作用，及反式激活蛋白-NEMO 结合域
（TAT-NBD）对胆红素神经毒性的干预作用。方法原代培养海马神经元分为对照组、胆红素组、TAT-NBD早期干预组、持续干
预组及晚期干预组。免疫细胞化学法检测NF-κB p65蛋白表达，改良MTT法、Annexin V-FITC/PI双染法及TUNEL法检测细胞
相对存活率及凋亡率，ELISA法检测原代培养基上清IL-1β水平。结果胆红素组海马神经元NF-κB p65 蛋白平均光密度值
（MOD）明显高于对照组（P<0.01），6、24 h达高峰。TAT-NBD早期干预组细胞相对存活率为（80.784±9.767）%低于对照组（P<
0.01）、高于胆红素组（P<0.01），细胞凋亡率为（14.100±2.252）%（Annexin V-FITC/PI双染法）、（12.883±1.629）%（TUNEL法）高
于对照组（P<0.01）、低于胆红素组（P<0.01），IL-1β水平为15.348±0.812 pg/ml低于胆红素组（P<0.05）。TAT-NBD持续干预及晚
期干预组细胞存活率、凋亡率、IL-1β水平与胆红素组无显著性差异（P>0.05）。结论NF-κB双向调控胆红素诱导的海马神经元
凋亡。TAT-NBD抑制NF-κB早期高峰有神经保护作用，有可能用于胆红素脑损伤的预防。

Abstract: Objective To investigate the role of nuclear factor-κB (NF-κB) activation in bilirubin-induced apoptosis of rat
hippocampal neurons and the effect of TAT-NBD intervention on bilirubin neurotoxicity. Methods Primary-cultured rat
hippocampal neurons were treated with TAT-NBD in the initial 6 or 24 h or in the latter 6 h during a 24-h bilirubin exposure of
the cells (early, continuous and late intervention groups, respectively). Immunocytochemistry was performed to detect NF-κB
p65 protein expression, and the cell survival and apoptosis were assessed with a modified MTT assay, Annexin V-FITC/PI and
TUNEL assay. IL-1β concentration in the supernatant was determined with ELISA. Results Compared with the control cells,
bilirurin-treated cells showed a significantly increased NF-κB p65 protein expression (P<0.01), which reached the peak level at
6 and 24 h (P<0.01). The cell survival rate in early TAT-NBD intervention group was (80.784±9.767)%, significantly lower than
that of the control group (P<0.01) but higher than that of bilirubin group (P<0.01); the apoptotic rate in early TAT-NBD
intervention group was significantly higher than that of control group (P<0.01) but lower than that of bilirubin group (P<0.01).
IL-1β concentration was significantly lower in early TAT-NBD intervention group (15.348±0.812 pg/ml) than in bilirubin group
(P<0.05). The continuous and late TAT-NBD intervention groups showed comparable cell survival rate, apoptotic rate and
IL-1β concentration with bilirubin group (P>0.05). Conclusion NF-κB bidirectionally regulates bilirubin-induced apoptosis of
rat hippocampal neurons. Selective inhibition of the early peak of NF-κB by TAT-NBD offers neuroprotective effect. TAT-NBD
can be potentially used for prophylaxis of bilirubin-induced brain injury.