天津医药

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MiR-204对癫痫神经元中BDNF-TrkB信号通路的影响

常伟,潘立平,吴秋静,宋毅军   

  1. 天津医科大学总医院
  • 收稿日期:2013-10-10 修回日期:2013-11-21 出版日期:2014-03-15 发布日期:2014-03-15
  • 通讯作者: 宋毅军

Study of miR-204 regulates BDNF/TrkB signaling expression in epileptic neurons

wei CHANG 2, yijun song   

  • Received:2013-10-10 Revised:2013-11-21 Published:2014-03-15 Online:2014-03-15
  • Contact: yijun song

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摘要： 【摘要】目的研究海马神经元癫痫模型中转染miR-204后对BDNF/TrkB通路调控作用的影响。方法取原代培养7天后的细胞，随机分为正常组、正常+BDNF组、癫痫组、癫痫+BDNF组、正常转染miR-204组、癫痫转染miR-204组、癫痫转染miR-204+BDNF组。癫痫组用无媄液处理3h制作癫痫模型。制备成功miR-204慢病毒表达载体。采用免疫荧光、膜片钳及免疫印迹技术鉴定及观察miR-204对BDNF/TrkB通路的表达的影响。结果外源性加入BDNF可激活BDNF/TrkB通路。在癫痫状态下BDNF/TrkB通路处于受抑制状态，并且TrkB.T表达上调，TrkB.FL表达下调，TrkB.T对TrkB.FL有显性抑制作用，从而抑制了BDNF/TrkB通路的激活。转染miR-204能解除TrkB.T对BDNF/TrkB通路的抑制作用。结论BDNF及mir-204可以改善BDNF/TRKB受抑制的状态，从而在癫痫疾病的缓解中可能发挥重要作用。

关键词: 癫痫, TrkB, 海马神经元, miR-204, 蛋白印迹

Abstract: 【Abstract】ObjectiveTo study the functions of miR-204 in BDNF/TrkB signaling and pathogenesis in the epilepsy model after the transfected with miR-204.Methods Primary hippocampal neurons were cultured in vitro after 7 days.Epilepsy model of hippocampal neurons were established by being exposed to Mg2+ free media for 3 hours.Construct miR-204 lentivirusvector.Detected by immunohistochemistry and patch clamp technique and Western Blot technique.Results BDNF/TrkB signaling can be activated by exogenous BDNF and is inhibited under epileptic condition. BDNF/TrkB signaling is inhibited by high expression of TrkB.T rather than TrkB.FL under epileptic condition.miR-204 can down-regulate expression of TrkB.T, so as to remove the inhibition of BDNF/TrkB signaling. Conclusion BDNF and mir-204 can improve the inhibiting condition of BDNF/TrkB signaling,and then playing an important role in alleviating epilepsy disease.

Key words: Epilepsy, Trkb, Hippocampal neurons, MiR-204, Western Blot