Genetic manipulation and transplantation of cynomolgus embryonic stem cells

食蟹猴胚胎干细胞的基因操作与移植

• 批准号: 13671080
• 负责人: HANAZONO Yutaka
• 金额: $ 2.62万
• 依托单位: Jichi Medical School
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2001
• 资助国家: 日本
• 起止时间: 2001 至 2002
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-13671080/
• 关键词: embryonic stem cells; lentiviral vector; cynomolgus monkeys; primate model; NOD; SCID; 胎仔移植; カニクイザル; ヒツジ; GFP; OP9; BMP-4; ヒツジ胎仔

The ability to stably introduce genetic material into primate embryonic stem (ES) cells could allow broader application. In this study, we examined lentiviral gene transfer into cynomolgus ES cells. When cynomolgus ES cells were transduced once with a simian immunodeficiency virus (SIV)-based lentivirus vector encoding the green fluorescent protein (GFP) gene, most cells (around 90%) fluoresced and high levels of GFP expression persisted for 5 months without selection procedures. In addition, high levels of GFP expression were observed during embryoid body formation. On the other hand, transduction of mouse ES cells with the SIV based vector resulted in lower gene transfer rates, implying that primate viral vectors can transduce primate ES cells more efficiently than mouse ES cells. The use of GFP as a reporter gene allows direct and simple detection of successfully transduced ES cells and facilitates monitoring of ES cell proliferation and differentiation both in vitro and potentially in vivo. Furthermore, this highly efficient gene transfer method allows faithful gene delivery to primate ES cells with potential for both research and therapeutic application.

稳定地将遗传物质引入灵长类动物胚胎（ES）细胞的能力可以更广泛地应用。在这项研究中，我们检查了慢病毒基因转移到cynomolgus ES细胞中。当用Simian免疫缺陷病毒（SIV）基于基于绿色荧光蛋白（GFP）基因的基于SIV的慢病毒病毒（SIV）的慢病毒载体转导CYNOMOLGUS ES细胞时，大多数细胞（约90％）荧光，GFP表达的高水平持续了5个月。另外，在胚胎体形成期间观察到高水平的GFP表达。另一方面，用基于SIV的载体转导小鼠ES细胞导致基因转移率较低，这表明灵长类动物病毒载体比小鼠ES细胞更有效地传递灵长类动物ES细胞。 GFP用作报告基因的基因可以直接和简单检测成功转导的ES细胞，并促进在体外和可能在体内监测ES细胞增殖和分化。此外，这种高效的基因转移方法允许忠实的基因传递到具有研究和治疗应用潜力的灵长类动物ES细胞。

项目成果

Hanazono,Y., et al.: "Introduction of the GFP gene into hematopoietic stem cells results in prolonged discrepancy of in vivo transduction levels betweem bone marrow progenitors and peripheral blood cells in nonhuman primates"J.Gene Med.. 4. 470-477 (2002)

Hanazono,Y. 等人：“将 GFP 基因引入造血干细胞会导致非人灵长类动物的骨髓祖细胞和外周血细胞之间的体内转导水平存在长期差异”J.Gene Med.. 4. 470-

Asano,T., et al.: "Highly efficient gene transfer into primate embryonic stem cells with a simian lentivirus vector"Mol.Ther.. 6. 162-168 (2002)

Asano,T., et al.：“使用猿猴慢病毒载体将基因高效转移到灵长类胚胎干细胞中”Mol.Ther.. 6. 162-168 (2002)

Wang, L. et al.: "Delayed delivery of AAV-GDNF prevents nigral neurodegeneration and promotes functional recovery in a rat model of Parkinson's disease"Gene Ther.. 9巻. 381-389 (2002)

Wang, L. 等人：“延迟递送 AAV-GDNF 可预防帕金森病大鼠模型中的黑质神经变性并促进功能恢复”Gene Ther.. 9. 381-389 (2002)

Hanazono Y, et al.: "Genetic manipulation of primate embryonic and hematopoietic stem cells with simian lentivirus vectors"Trends in Cardiovascular Medicine. 13. 106-110 (2003)

Hanazono Y 等人：“用猿猴慢病毒载体对灵长类胚胎和造血干细胞进行基因操作”心血管医学趋势。

Hanazono, Y. et al.: "Introduction of the GFP gene into hematopoietic stem cells results in prolonged discrepancy of in vivo transduction levels between bone marrow progenitors and peripheral blood cells in nonhuman primates"J. Gene Med.. 4巻. 470-477 (200

Hanazono, Y. 等人：“将 GFP 基因引入造血干细胞会导致非人灵长类动物的骨髓祖细胞和外周血细胞之间的体内转导水平存在长期差异”J. Gene Med. 470-。 477 (200