Retrovirus-Mediated Gene Transduction Into Canine Peripheral Blood Repopulating Cells
Retrovirus-Mediated Gene Transduction Into Canine Peripheral Blood Repopulating Cells
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逆转录病毒介导的基因转导到犬外周血再生细胞中
DOI：
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发表时间：
1994
期刊：
影响因子：
--
通讯作者：
F. Schuening
中科院分区：
文献类型：
--
作者：
H. Kiem;H. Kiem;B. Darovsky;C. Kalle;S. Goehle;D. Stewart;T. Graham;R. Hackman;F. Appelbaum;H. Deeg;A. Miller;R. Storb;F. Schuening
基因标记的外周血祖细胞被用来研究它们在自体骨髓和外周血细胞移植后长期造血重建中的作用。3只犬自体骨髓采集和冷冻保存后，用重组犬干细胞因子处理8天，动员犬外周血祖细胞。采集外周血单个核细胞，通过亲和素-生物素免疫吸附富集主要组织相容性复合体(MHC)II类抗原阳性细胞，从而为细胞再扩增做好准备。随后，将细胞在辐照的载体产生包装细胞(PA317/LN)上共培养24小时，然后在包含长期骨髓培养系统的载体中孵育11天。移植当天，动物接受9.2Gy全身照射，并在全身照射后2小时内输注已转导的外周血细胞和未转导的冻存骨髓细胞。三只狗都被植入了。两只狗长期存活，移植后至48周，骨髓来源的粒细胞-巨噬细胞集落形成中位数分别为1%和2%(范围从1%到10%)，呈间歇性的G418抗性。通过聚合酶链式反应在移植后65周的外周血粒细胞和75周的外周血淋巴细胞中检测到新的特异性序列。狗的外周血液样本中没有辅助病毒，也没有观察到转导的副作用。其中一只狗在第84天死于慢性犬瘟硬化性脑炎，而另外两只狗分别在15个月和17个月时存活。我们的数据显示，逆转录病毒成功地转导了犬的外周血细胞。移植后明显的髓样细胞和淋巴样细胞的长期存留表明，外周血中含有有助于致命性脑损伤后长期造血重建的再生细胞。
Genetically marked peripheral blood progenitor cells were used to investigate their contribution to long-term hematopoietic reconstitution after autologous marrow and peripheral blood cell transplantation. After autologous marrow harvest and cryopreservation, canine peripheral blood progenitor cells were mobilized in three dogs by treatment with recombinant canine stem cell factor for 8 days. Peripheral blood mononuclear cells were collected and enriched for major histocompatibility complex (MHC) class II antigen-positive cells by avidin-biotin immunoadsorption, thereby enriching for repopulating cells. Subsequently, the cells were cocultivated for 24 hours on irradiated vector-producing packaging cells (PA317/LN), followed by an 11-day incubation in a vector containing long-term marrow culture system. On the day of transplantation, the animals were irradiated with 9.2 Gy total body irradiation (TBI), and transduced peripheral blood cells and untransduced cryopreserved marrow cells were infused within 2 hours of TBI. All three dogs engrafted. Two dogs are long-term survivors showing intermittently G418-resistant marrow-derived colony-forming unit granulocyte-macrophage colonies at a median of 1% and 2%, respectively (range, 1% to 10%), for now up to 48 weeks after transplantation. Neo-specific sequences were detected by polymerase chain reaction in peripheral blood granulocytes for now up to 65 weeks and in peripheral blood lymphocytes for up to 75 weeks after transplantation. Peripheral blood samples of the dogs were free of helper virus and no side effects from the transduction were observed. One of the three dogs died from chronic canine distemper sclerosing encephalitis on day 84, whereas the other two dogs are alive at 15 and 17 months. Our data show successful retroviral transduction of canine peripheral blood repopulating cells. Long-term persistence of marked myeloid and lymphoid cells after transplantation suggests that peripheral blood contains repopulating cells that contribute to long-term hematopoietic reconstitution after otherwise lethal TBI.
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影响因子：
8
作者：
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通讯作者：
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DOI：
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发表时间：
1989
影响因子：
11.1
作者：
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通讯作者：
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使用聚合酶链式反应监测离体肿瘤细胞清除的有效性。
影响因子：
20.3
作者：
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影响因子：
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作者：
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影响因子：
2.6
作者：
Schuening,FG;Storb,R;Meyer,J;Goehle,S
通讯作者：
Goehle,S