摘要
目的研究小鼠未编辑的谷氨酸受体2(GluR2)Q/R部位缺失RNA2的次黄嘌呤腺苷脱氨酶(ADAR2)是否引起缓慢的运动神经元死亡。方法利用Cre/loxP重组系统制作一个条件性基因敲除ADAR2小鼠品系(AR2),利用基因组PCR和反转录PCR技术,将AR2小鼠的运动神经元ADAR2基因条件性定位,观察及检测AR2小鼠、携带内源性GluR2等位基因的未编辑GluR2 Q/R小鼠(AR2res)和对照组小鼠的GluR2 Q/R部位编辑率、行为特征、电生理以及组织形态。结果 AR2小鼠GluR2 Q/R部位编辑率降低,脊髓与脑运动神经核细胞显示因ADAR2缺乏引起的运动神经元死亡,并且出现运动神经功能衰退症状,而GluR2 Q/R部位的编辑率在动眼神经核无明显减少。当失去ADAR2活性的AR2小鼠携带内源性的GluR2等位基因时,细胞和表型的变化可以被阻止。结论 ADAR2失活可以引起运动神经元α-氨基-3-羟基-5甲基-4异恶唑-丙酸受体介导的死亡,而在未编辑的GluR2 Q/R部位缺乏ADAR2可以引起运动神经元缓慢死亡。
Aim To study whether decreased adenosine deaminase acting on RNA2 （ADAR2） induces slow death of motor neuron from unedited GluR2 subunit of the α-amino-3-hydroxy-5-methl-4-isoxazolepropionic acid （AMPA） receptor. Methods Genetically modified mice （designated as AR2） were generated, in which theADAR2 gene was conditionally targeted in motor neurons using the Cre/loxP system. Chimeric mice were generated by injection of a targeted embryonic stem cell clone into C57BL/6-derived blastocysts, knockouting ADAR2 activity selectively in motor neurons, crossing ADAR2^flox/flox mice with VAChT-Cre. Fast mice the GluR2 Q/R side editing efficiency, behavioral characteristics, motor neurons were detected respectively. Results Among 116 specimens from AR2 mice were examined, 8 specimens showed 0% and 42 showed 100% Q/R site editing, with the remaining 66 specimens distributed between the ranges from 17% to 98%. AR2 mice were hypokinetic and abnormal in posture, but they displayed no overt paralysis or vesico-urinary disturbances and exhibited a normal withdrawal response to noxious stimuli. They showed a lower rotarod performance than their control littermates after 5 weeks of age when the Cre expression reached the maximum level. The kinetics of neuronal loss were consistent with the kinetics of progressive motor-selective behavioral deficits. The long survival with hypoactivity beyond 6 months of age indicated that the remaining ADAR2- expressing neurons functioned normally during the remainder of life. The numbers of large neurons in facial and hypoglossal nerve nuclei in AR2 mice were significantly smaller than those in the control mice at 12 months of age, whereas the numbers of neurons in nuclei of oculomotor nerves were not decreased. The anterior horn cells （AHCs）, including 30% AHCs lacking ADAR2 from Cre-mediated recombination, were viable in AR2 mice at 6 months of age, and the total number of AHCs was the same as in age-matched control mice. There was no detectable increase in GFAP or MAC2 immunore
出处
《中国临床神经科学》
2013年第1期1-11,共11页
Chinese Journal of Clinical Neurosciences
基金
Ministry of Education
Culture
Sports
Science
and Technology of Japan Grants-in-Aid for Scientific Research(编号:17390251
19390235与20023008
负责人:Shin K)
Ministry of Health
Labor
and Welfare of Japan Grant(编号:H18-Kokoro-017
负责人:Shin K)
日本肌萎缩侧索硬化症协会基金(编号:875 P.H.S.)
上海浦江人才计划资助项目(编号:09PJ1409300
负责人:潘卫东)
上海市中医药事业发展3年行动计划(重大研究)项目(ZYSNXD-CC-ZDYJ028
负责人:蔡定芳)
关键词
RNA2
次黄嘌呤腺苷脱氨酶
RNA编辑
Q
R部位
肌萎缩侧索硬化症
神经元
谷氨酸受体
RNA2
adenosine deaminase acting
RNA editing
Q/R site
amyotrophic lateralsclerosis
neuron
α-amino-3-hydroxy-5-methl-4-isoxazolepropionic acid
