Intrastriatal rAAV-mediated delivery of anti-huntingtin shRNAs induces partial reversal of disease progression in R6/1 Huntington's disease transgenic mice
Intrastriatal rAAV-mediated delivery of anti-huntingtin shRNAs induces partial reversal of disease progression in R6/1 Huntington's disease transgenic mice
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DOI：
10.1016/j.ymthe.2005.05.006
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发表时间：
2005-10-01
影响因子：
12.4
通讯作者：
Mandel, RJ
中科院分区：
文献类型：
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作者：
Rodriguez-Lebron, E;Denovan-Wright, EM;Mandel, RJ
亨廷顿病（Huntington's disease，HD）是一种由亨廷顿蛋白N端多聚谷氨酰胺结构域异常扩增引起的致死性神经退行性疾病。我们开发了一种重组腺相关病毒血清5型（rAAV5）基因转移策略，通过RNA干扰抑制R6/1 HD转基因小鼠纹状体突变型亨廷顿蛋白（mHtt）的水平。用表达源自R6/1转基因HD小鼠的人mHtt的一部分和针对mHtt mRNA的5 'UTR的短发夹RNA（siHUNT-1）的质粒瞬时共转染HEK293细胞导致mHtt mRNA（~75%）和蛋白质（~60%）水平的降低。在R6/1小鼠的纹状体中siHUNT-1的长期体内rAAV 5介导的表达降低了mHtt mRNA（~78%）和蛋白质（~28%）的水平，如分别通过定量RT-PCR和Western印迹分析所确定的。mHtt的减少伴随着神经元核内包涵体的大小和数量的减少，以及前脑啡肽原和多巴胺-和cAMP-反应性磷蛋白32 kDa mRNA的稳态水平的小但显著的正常化。最后，rAAV5-siHUNT-1的双侧表达导致R6/1小鼠表现出的后爪紧握表型的延迟发作。这些结果表明，纹状体mHtt水平的降低可以改善R6/1小鼠的HD表型。
Huntington's disease (HD) is a fatal neurodegenerative disorder caused by the presence of an abnormally expanded polyglutamine domain in the N-terminus of huntingtin. We developed a recombinant adeno-associated viral serotype 5 (rAAV5) gene transfer strategy to posttranscriptionally suppress the levels of striatal mutant huntingtin (mHtt) in the R6/1 HD transgenic mouse via RNA interference. Transient cotransfection of HEK293 cells with plasmids expressing a portion of human mHtt derived from R6/1 transgenic HD mice and a short-hairpin RNA directed against the 5' UTR of the mHtt mRNA (siHUNT-1) resulted in reduction in the levels of mHtt mRNA (-75%) and protein (-60%). Long-term in vivo rAAV5-mediated expression of siHUNT-1 in the striatum of R6/1 mice reduced the levels of mHtt mRNA (-78%) and protein (-28%) as determined by quantitative RT-PCR and Western blot analysis, respectively. The reduction in mHtt was concomitant with a reduction in the size and number of neuronal intranuclear inclusions and a small but significant normalization of the steady-state levels of preproenkephalin and dopamine- and cAMP-responsive phosphoprotein 32 kDa mRNA. Finally, bilateral expression of rAAV5-siHUNT-1 resulted in delayed onset of the rear paw clasping phenotype exhibited by the R6/1 mice. These results suggest that a reduction in the levels of striatal mHtt can ameliorate the HD phenotype of R6/1 mice.