Efficient lentiviral gene transfer to canine repopulating cells using an overnight transduction protocol
Efficient lentiviral gene transfer to canine repopulating cells using an overnight transduction protocol
复制标题
DOI：
10.1182/blood-2003-07-2414
复制
发表时间：
2004-05-15
期刊：
影响因子：
20.3
通讯作者：
Kiem, HP
中科院分区：
文献类型：
--
作者：
Horn, PA;Keyser, KA;Kiem, HP
使用慢病毒载体转导造血干细胞引起了人们的极大兴趣，因为它们能够稳定地整合到非分裂细胞的基因组中。然而，已发表的大型动物研究报告了高度可变的基因转移率，通常低于1%。在这里，我们报告使用慢病毒载体转导犬CD 34（+）造血再生细胞使用一个非常短的，18小时的转导协议。我们比较了慢病毒转导的造血再生细胞从干细胞因子（SCF）和粒细胞集落刺激因子（G-CSF）引发的骨髓或动员外周血中的竞争性再生试验在3只狗。所有犬均在9天内迅速植入。在所有谱系（B细胞、T细胞、粒细胞和红细胞以及血小板）中检测到转基因表达，表明转导细胞的多谱系植入，总体长期标记水平高达12%。动员的外周血细胞中的基因转移水平略高于引发的骨髓细胞。总之，我们显示了使用过夜转导方案的犬再生细胞的有效慢病毒转导。这些结果对干细胞基因治疗方案的设计具有重要意义，特别是对于那些在培养中维持干细胞是主要限制的疾病。(C)2004年，美国血液学会。
The use of lentiviral vectors for the transduction of hematopoietic stem cells has evoked much interest owing to their ability to stably integrate into the genome of nondividing cells. However, published large animal studies have reported highly variable gene transfer rates of typically less than 1%. Here we report the use of lentiviral vectors for the transduction of canine CD34(+) hematopoietic repopulating cells using a very short, 18-hour transduction protocol. We compared lentiviral transduction of hematopoietic repopulating cells from either stem cell factor (SCF)- and granulocyte-colony stimulating factor (G-CSF)-primed marrow or mobilized peripheral blood in a competitive repopulation assay in 3 dogs. All dogs engrafted rapidly within 9 days. Transgene expression was detected in all lineages (B cells, T cells, granulocytes, and red blood cells as well as platelets) indicating multilineage engraftment of transduced cells, with overall long-term marking levels of up to 12%. Gene transfer levels in mobilized peripheral blood cells were slightly higher than in primed marrow cells. In conclusion, we show efficient lentiviral transduction of canine repopulating cells using an overnight transduction protocol. These results have important implications for the design of stem cell gene therapy protocols, especially for those diseases in which the maintenance of stem cells in culture is a major limitation. (C) 2004 by The American Society of Hematology.