Evaluation of primitive murine hematopoietic stem and progenitor cell transduction in vitro and in vivo by recombinant adeno-associated virus vector serotypes 1 through 5.
Evaluation of primitive murine hematopoietic stem and progenitor cell transduction in vitro and in vivo by recombinant adeno-associated virus vector serotypes 1 through 5.
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重组腺相关病毒载体血清型 1 至 5 对原始小鼠造血干细胞和祖细胞转导的体外和体内评价。
DOI：
10.1089/hum.2006.17.321
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发表时间：
2006
影响因子：
4.2
通讯作者：
Srivastav
中科院分区：
文献类型：
--
作者：
Zhong,Li;Li,Weiming;Li,Yanjun;Zhao,Weihong;Wu,Jianqing;Li,Baozheng;Maina,Njeri;Bischof,Daniela;Qing,Keyun;Weigel-Kelley,KirstenA;Zolotukhin,Irene;WarringtonJr,KennethH;Li,Xiaomiao;Slayton,WilliamB;Yoder,MervinC;Srivastav
关于 AAV 血清型 2 (AAV2) 载体的造血细胞转导存在相互矛盾的数据，并且尚未评估其他 AAV 血清型载体在造血干/祖细胞转导中的功效。我们在体外和体内评估了传统单链 AAV 血清型载体 1 至 5 在原始小鼠造血干/祖细胞中的功效。在使用 Sca1+c-kit+Lin−造血细胞进行的祖细胞测定中，感染 AAV1 的培养物中 9% 的集落表达转基因。 AAV1 与携带 T 细胞蛋白酪氨酸磷酸酶基因 (scAAV-TC-PTP) 的自我互补 AAV 载体的共感染将转导效率提高到 24%，表明病毒第二链 DNA 合成是一个限速步骤。 scAAV 载体的使用进一步证实了这一点，该载体绕过了这一要求。在涉及经致死辐射的同基因小鼠的骨髓移植研究中，与 AAV1 ± scAAV-TC-PTP 载体共感染的 Sca1+c-kit+Lin− 细胞在移植后 6 个月分别导致 2% 和 7.5% 的外周血 (PB) 细胞出现转基因表达。在二次移植实验中，移植后 6 个月，7% 的 PB 细胞和 3% 的骨髓 (BM) 细胞表达转基因。大约 21% 的 BM 衍生菌落含有整合形式的原病毒 DNA 序列。这些结果证明AAV1是迄今为止转导原始鼠造血干/祖细胞最有效的载体。涉及 scAAV 基因组和造血细胞特异性启动子的进一步研究应进一步提高 AAV1 载体的转导效率，这对这些载体在造血干细胞基因治疗中的最佳使用具有影响。
Conflicting data exist on hematopoietic cell transduction by AAV serotype 2 (AAV2) vectors, and additional AAV serotype vectors have not been evaluated for their efficacy in hematopoietic stem/progenitor cell transduction. We evaluated the efficacy of conventional, single-stranded AAV serotype vectors 1 through 5 in primitive murine hematopoietic stem/progenitor cellsin vitroas well asin vivo. In progenitor cell assays using Sca1+c-kit+Lin−hematopoietic cells, 9% of the colonies in cultures infected with AAV1 expressed the transgene. Coinfection of AAV1 with self-complementary AAV vectors carrying the gene for T cell protein tyrosine phosphatase (scAAV-TC-PTP) increased the transduction efficiency to 24%, indicating that viral secondstrand DNA synthesis is a rate-limiting step. This was further corroborated by the use of scAAV vectors, which bypass this requirement. In bone marrow transplantation studies involving lethally irradiated syngeneic mice, Sca1+c-kit+Lin−cells coinfected with AAV1 ± scAAV-TC-PTP vectors led to transgene expression in 2 and 7.5% of peripheral blood (PB) cells, respectively, 6 months posttransplantation. In secondary transplantation experiments, 7% of PB cells and 3% of bone marrow (BM) cells expressed the transgene 6 months posttransplantation. Approximately 21% of BM-derived colonies harbored the proviral DNA sequences in integrated forms. These results document that AAV1 is thus far the most efficient vector in transducing primitive murine hematopoietic stem/progenitor cells. Further studies involving scAAV genomes and hematopoietic cell-specific promoters should further augment the transduction efficiency of AAV1 vectors, which should have implications in the optimal use of these vectors in hematopoietic stem cell gene therapy.