A TRANSGENIC PIG MODEL FOR SICKLE CELL DISEASE

镰状细胞病转基因猪模型

• 批准号: 3754103
• 负责人: S G SHAPIRO
• 金额: --
• 依托单位: NATIONAL INSTITUTE OF DIABETES AND DIGESTIVE AND KIDNEY DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/3754103
• 关键词: animal breeding; disease /disorder model; follicle stimulating hormone; fusion gene; genetic regulatory element; genetically modified animals; globin; human genetic material tag; microinjections; miniature swine; model design /development; sickle cell anemia; swine

We have begun to make a transgenic pig model for sickle cell disease (SS disease) to facilitate studies for sickle cell pathology and evaluations of new drug and gene therapies. Previous attempts to make transgenic models using the mouse have failed to produce the clinical symptomatology of SS disease, in spite of the production, in some cases, of significant amounts of human sickle hemoglobin (HbS) and evidence for intracellular polymerization of hemoglobin. The failure of the transgenic mice to develop significant sickle cell disease pathology appears to be due to differences in the vascular anatomy and physiology between mice and humans. On the other hand, the pig provides an excellent model for human circulatory physiology, significantly, for our purposes, evincing cognate responses to vaso-active compounds, similarities in capillary bed density, cerebrovascular diseases and certain red cell structural properties. The technology to produce transgenic farm (full-size) pigs is also well established. To produce transgenic animals with high level globin transgene expression, a genetic regulatory region called by beta-globin locus control region (LCR), usually located upstream of the beta-globin gene cluster, has been shown to be required in cis. We have prepared DNA constructions containing the functional portions of the human beta-globin LCR (mini-LCR) linked to the human alpha-2-globin gene or a modified human sickle beta-globin gene with additional mutations to promote polymerization (beta SAD, containing additional mutations Antilles and D Punjab) for microinjection into the pigs. At the same time, to facilitate the maintenance and clinical analysis of the transgenic pigs we have begun to develop the NIH--minipig as a transgenic system. Using a regimen of purified FSH, significant superovulation has been obtained (40) eggs/pig). Following breeding, fertilize eggs have been recovered and procedures for microinjection, surgical reimplantation and maintenance of pregnancy are being developed to extend the transgenic technology to this more conveniently-sized pig variety. Once established, the transgenic pig model that produces high levels of human sickle hemoglobin and demonstrates SS disease pathology, would greatly accelerate our efforts to understand and treat the disease.

我们已经开始制作镰刀的转基因猪模型 细胞病（SS病），以促进镰状细胞病的研究 细胞病理学及新药和基因的评价 治疗 以前尝试制作转基因模型 使用老鼠的实验未能产生临床效果， SS疾病的病理学，尽管生产， 在某些情况下，大量的人类镰刀 血红蛋白（HbS）和细胞内 血红蛋白的聚合。 的失败 转基因小鼠患上严重的镰状细胞病 病理学似乎是由于血管的差异， 在解剖学和生理学上的差异。 上 另一方面，猪为人类提供了一个很好的模型， 循环生理学，对于我们的目的来说， 表明对血管活性化合物的同源反应， 毛细血管床密度、脑血管 疾病和某些红细胞结构特性。 的 生产转基因农场（全尺寸）猪技术， 也很好地建立了。 生产高水平球蛋白的转基因动物 转基因表达，一个遗传调控区，称为 β-珠蛋白基因座控制区（LCR），通常位于 β-珠蛋白基因簇的上游，已被证明 在CIS中需要。 我们准备了DNA结构 含有人β-珠蛋白的功能部分 与人α-2-珠蛋白基因连接的LCR（mini-LCR），或 一种修饰的人镰状β-珠蛋白基因， 促进聚合的突变（β SAD，含有 另外的突变安的列斯和D旁遮普）， 对猪进行显微注射。 同时为了 便于维护和临床分析 转基因猪我们已经开始开发NIH--迷你猪 转基因系统。 使用纯化FSH的方案， 已获得显著的超数排卵（40） 鸡蛋/猪）。 在繁殖之后，受精卵被 显微注射手术 再植入和维持妊娠正在 开发了将转基因技术扩展到这一点， 体型巨大的猪。 一旦建立， 转基因猪模型产生高水平的人类 镰状血红蛋白并表现出SS疾病病理， 将大大加快我们理解和 治疗疾病。