Targeted conversion of the transthyretin gene in vitro and in vivo
Targeted conversion of the transthyretin gene in vitro and in vivo
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DOI：
10.1038/sj.gt.3302228
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发表时间：
2004-05-01
期刊：
影响因子：
5.1
通讯作者：
Yamamura, K
中科院分区：
文献类型：
--
作者：
Nakamura, M;Ando, Y;Yamamura, K
家族性淀粉样多发性神经病（FAP）是遗传性泛发性淀粉样变性的常见形式，其特征是淀粉样纤维在周围神经和其他器官中的积聚。肝移植已被用作FAP的治疗，因为变体甲状腺素运载蛋白（TTR）主要由肝脏合成，但这种治疗与几个问题有关。因此，我们需要开发一种新的治疗方法，以防止在肝脏中产生TTR变体。在这项研究中，我们使用HepG2细胞显示在体外转换TTR基因的单链寡核苷酸（SSO），嵌在去端胶原，旨在促进内源性修复的基因组DNA。对于研究的体内部分，我们使用来自转基因小鼠的肝脏，所述转基因小鼠的固有野生型TTR基因被鼠TTR Val 30Met基因替换。基因转换水平通过实时RCR结合多等位基因特异性扩增来确定。我们的研究结果表明，基因转换的水平是约11%和9%的总TTR基因在HepG2细胞和肝脏转基因小鼠，分别。因此，通过这种方法的基因治疗可能是一个有前途的替代肝移植治疗FAP。
Familial amyloidotic polyneuropathy (FAP) is the common form of hereditary generalized amyloidosis and is characterized by the accumulation of amyloid fibrils in the peripheral nerves and other organs. Liver transplantation has been utilized as a therapy for FAP, because the variant transthyretin (TTR) is predominantly synthesized by the liver, but this therapy is associated with several problems. Thus, we need to develop a new treatment that prevents the production of the variant TTR in the liver. In this study, we used HepG2 cells to show in vitro conversion of the TTR gene by single-stranded oligonucleotides (SSOs), embedded in atelocollagen, designed to promote endogenous repair of genomic DNA. For the in vivo portion of the study, we used liver from transgenic mice whose intrinsic wild-type TTR gene was replaced by the murine TTR Val30Met gene. The level of gene conversion was determined by real-time RCR combined with mutant-allele-specific amplification. Our results indicated that the level of gene conversion was approximately 11 and 9% of the total TTR gene in HepG2 cells and liver from transgenic mice, respectively. Gene therapy via this method may therefore be a promising alternative to liver transplantation for treatment of FAP.