组蛋白去乙酰化酶在小鼠神经元发育中的作用与机制研究

Role and mechanism of histone deacetylases in mouse neuronal development

目的探讨组蛋白去乙酰化酶(HDAC)1和HDAC2在小鼠神经元发育中的作用及机制。方法将具有突触蛋白1启动子Cre重组酶(Synapsin1-Cre)的小鼠与HDAC1&2flox/flox小鼠杂交,得到在神经元中特异性敲除HDAC1&2的小鼠纳入敲除组,同窝出生的未敲除HDAC1&2的小鼠纳入对照组。观察小鼠一般状态,绘制生存曲线。收集生后14 d敲除组和对照组小鼠脑组织,采用Western blot法及免疫组化法检测神经元发育标记物神经元核抗原(NeuN)、轴突标记物非磷酸化神经丝蛋白重链(np-NF200)、磷酸化神经丝蛋白重链(p-NF200)及哺乳动物雷帕霉素靶蛋白(mTOR)信号通路下游磷酸化核糖体S6(p-S6)蛋白表达水平。结果HDAC1&2敲除小鼠通常于1个月内死亡,体型和脑组织明显小于对照组,表现出震颤及紧抱后肢等运动功能异常。敲除组小鼠NeuN、np-NF200、p-NF200、p-S6蛋白表达量均较对照组明显减少(P<0.05,n=3)。结论缺失HDAC1和HDAC2导致小鼠神经元成熟障碍,轴突发育不良。推测这种变化与mTOR信号通路有关。

Objective To study the role and mechanism of histone deacetylase 1(HDAC1)and histone deacetylase 2(HDAC2)in mouse neuronal development.Methods The mice with Synapsin1-Cre recombinase were bred with HDAC1&2flox/flox mice to obtain the mice with neuron-specific HDAC1&2 conditional knockout(knockout group),and their littermates without HDAC1&2 knockout were used as the control group.The general status of the mice was observed and survival curves were plotted.Brain tissue samples were collected from the knockout group and the control group.Western blot and immunohistochemistry were used to measure the protein expression of related neuronal and axonal markers,neuronal nuclear antigen(NeuN),non-phosphorylated neurofilament heavy chain(np-NF200),and phosphorylated neurofilament heavy chain(p-NF200),as well as the downstream effector of the mTOR signaling pathway,phosphorylated S6 ribosomal protein(p-S6).Results The mice with HDAC1&2 conditional knockout usually died within one month after birth and were significantly smaller than those in the control group,with motor function abnormalities such as tremor and clasping of hindlimbs.Compared with the control group,the knockout group had significant reductions in the protein expression levels of NeuN,np-NF200,p-NF200,and p-S6(P<0.05;n=3).Conclusions Deletion of HDAC1 and HDAC2 in mouse neurons results in reduced neuronal maturation and axonal dysplasia,which may be associated with the mTOR signaling pathway.