TRANSGENIC MODLES AND GENETIC MODIFIERS OF SICKLE CELL DISEASE

这项计划的第一个目标是培育出改良的转基因小鼠。 镰状细胞病(SCD)模型。通向血管的过程- SCD的闭塞、疼痛危象和其他病理生理方面是 很复杂，也很难理解。动物模型可能对活体有价值 急性和慢性血管闭塞过程的研究和测试 药物或基因疗法。一种表达人HBs的转基因株将是 与阿尔法和贝塔珠蛋白基因缺失的小鼠杂交， 产生高达100%人类HBS表达的后代。这些老鼠将会是 表型特征为新的SCD模型。第二个目标是 使用技术识别镰状细胞病的新基因修饰物 用来绘制老鼠的基因图谱。而主要的遗传决定因素是 SCD是Beta6Val突变，在 SS纯合子中疾病的表型表达，这可能是 这在一定程度上是由于遗传修饰者。这种基因的鉴定 改良剂可以为治疗提出新的策略，并提供 单项SCD患者的预后指标。虽然它会是 最近，通过人类基因研究很难识别这样的基因 老鼠遗传学的发展使得修饰基因的快速定位成为可能。 首先将转SAD-1基因的SCD模型培育成不同的自交系 背景鉴定两株具有相同转基因的菌株 数量上不同的表型效应。修改器轨迹数 将通过对亲本、F1和N2回交的分析来估计 几代人。为了绘制修饰基因座图，一大群n2动物将 利用多态聚合酶链式反应标记进行表型分析和基因分型， 横跨整个基因组。候选基因定位在预测修饰物附近 将检查两个菌株之间的等位基因差异，以 确定潜在的修饰基因。除了这种全球方法之外， 预测的红细胞-内皮相互作用调节剂von的作用 威勒布兰德因子(VWF)将通过SAD-1小鼠之间的杂交进行测试 以及近交系RIIIS/J小鼠，表达较低水平的vWF。

The first goal of this proposal is to produce improved transgenic mouse models of sickle cell disease (SCD). The processes leading to vaso- occlusion, painful crisis and other pathophysiological aspects of SCD are complex and poorly understood. Animal models may be valuable for in vivo studies of acute and chronic vaso-occlusive processes, and for testing drug or gene therapies. A transgenic line expressing human HbS will be crossed with mice whose alpha and beta globin genes are deleted, to generate offspring expressing up to 100% human HbS. These mice will be phenotypically characterized as new SCD models. The second goal is to identify novel genetic modifiers of sickle cell disease, using techniques for genetic mapping the mouse. While the primary genetic determinant of SCD is the beta6Val mutation, there is considerable variability in the phenotypic expression of the disease among SS homozygotes, which may be due in part to genetic modifiers. The identification of such genetic modifiers could suggest new strategies for treatment, and provide prognostic indicators for individual SCD patients. While it would be difficult to identify such genes through human genetic studies, recent developments in mouse genetics permit the rapid mapping of modifier genes. The SAD-1 transgenic SCD model first will be bred into different inbred backgrounds to identify two strains in which the same transgene has quantitatively different phenotypic effects. The number of modifier loci will be estimated from an analysis of parental, F1 and N2 backcross generations. To map the modifier loci, a large pool of N2 animals will phenotypically characterized, and genotyped using polymorphic PCR markers, spanning the genome. Candidate genes mapping near the predicted modifier loci will be examined for allelic differences between the two strains, to identify potential modifier genes. In addition to this global approach, the role of a predicted modifier of red cell-endothelial interactions, von Willebrand factor (vWF), will be tested through crosses between SAD-1 mice and inbred RIIIS/J mice, which express lowered levels of vWF.