Creation of non-human primate neurogenetic disease models by gene targeting and nuclear transfer.
Creation of non-human primate neurogenetic disease models by gene targeting and nuclear transfer.
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DOI：
10.1186/1477-7827-2-40
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发表时间：
2004-06-16
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影响因子：
--
通讯作者：
Norgren RB Jr
中科院分区：
文献类型：
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作者：
Norgren RB Jr
转基因恒河猴是必要的，因为小鼠模型不适用于许多重要的神经遗传性疾病;例如，卡尔曼综合征，莱施-尼汉病和共济失调-毛细血管扩张症。小鼠模型可能不适合，因为可能没有人感兴趣基因的小鼠直系同源物，如卡尔曼综合征的情况，或者因为突变小鼠不表现出在人类中观察到的相同表型，如莱-尼二氏病和共济失调-毛细血管扩张症的情况。神经遗传性疾病的非人灵长类动物模型预计比现有的小鼠模型更接近人类疾病。转基因恒河猴可以通过修改体细胞的基因组，然后将细胞核从该细胞转移到去核卵母细胞来产生。转基因的随机整合足以创建功能获得性遗传疾病的模型。绿色荧光蛋白在恒河猴成纤维细胞中得到稳定表达。然而，基因靶向对于创建功能丧失型遗传病模型是必要的。在产生无效突变的非人灵长类动物之前，必须克服几个技术挑战。在我们的经验中，胎儿成纤维细胞经常在选择程序完成之前就衰老了。我们已经克服了这个问题，通过转染体细胞与人端粒酶逆转录酶。这种酶延长了体细胞的端粒和寿命。长而准确的聚合酶链反应可用于获得足够的同源区域的同基因恒河猴基因组DNA的靶向构建体。这将提高基因靶向效率。基因打靶实验目前正在进行中。突变恒河猴可能会导致对神经遗传性疾病的理解取得突破性进展，并证明对新疗法的临床前试验非常宝贵。
Genetically modified rhesus macaques are necessary because mouse models are not suitable for a number of important neurogenetic disorders; for example, Kallmann's syndrome, Lesch-Nyhan's disease and Ataxia-Telangiectasia. Mouse models may not be suitable because there may be no mouse ortholog of the human gene of interest, as is the case for Kallmann's syndrome, or because mutant mice do not exhibit the same phenotype observed in humans, as is the the case for Lesch-Nyhan's disease and Ataxia-Telangiectasia. Non-human primate models of neurogenetic diseases are expected to more closely resemble human diseases than existing mouse models. Genetically modified rhesus macaques can be created by modifying the genome of a somatic cell and then transferring the nucleus from this cell to an enucleated oocyte. Random integration of a transgene is sufficient to create models of gain-of-function genetic diseases. Stable expression of green fluorescent protein has been achieved in rhesus macaque fibroblasts. However, gene targeting is necessary to create models of loss-of-function genetic diseases. Several technical challenges must be overcome before null mutant non-human primates can be produced. In our experience, fetal fibroblasts frequently become senescent before selection procedures can be completed. We have overcome this problem by transfecting somatic cells with human telomerase reverse transcriptase. This enzyme extends the telomeres, and lifespan, of somatic cells. Long and accurate polymerase chain reaction can be used to obtain sufficient regions of homology of isogenic rhesus genomic DNA for targeting constructs. This should improve gene targeting efficiency. Gene targeting experiments are currently underway. Null mutant rhesus macaques will likely result in breakthrough advances in the understanding of neurogenetic disease and prove invaluable for preclinical trials of new therapies.