Towards a Mouse Model of Complement-Mediated Diseases

描述(由申请人提供)：广泛的临床和实验证据有力地表明，补体系统在包括一些溶血性贫血和男性不育在内的多种人类疾病的发病机制中起着关键作用。鉴于这些疾病的复杂和慢性性质，补体的致病作用仍需确定，我们建议在本申请中使用的分子工程小鼠模型提供了强大的工具来进行不可能在人类身上进行的实验。补体系统通过三种不同的激活途径和膜攻击复合体(MAC)的形成，介导获得性和先天反应，以防御细菌感染，并处理免疫复合体或凋亡细胞。激活的补体和MAC也可以损伤自身细胞，从而介导导致慢性病的病理反应。一系列补体调节蛋白，包括MAC形成的关键抑制因子CD59，已经进化成保护自身细胞免受补体损伤，补体损伤可能是由于MAC形成导致补体激活增加，也可能是调节减少所致。由于小鼠基因组中意外存在两个CD59基因(称为mCD59a和mCD59b)，阻碍了研究MAC的相对作用及其在人类疾病发病机制中的调控的适当动物模型的发展。对mCD59a和mCD59b基因敲除小鼠的初步研究表明，我们通过靶向缺失这两个基因成功地获得了mCD59ab双基因敲除小鼠(mCDSgab-/-)，需要研究MAC调节在人类疾病发病中的作用。我们的假设是，CD59功能的缺失将导致MAC介导的组织损伤和不同器官的局灶性病理。基于在mCdSgb-/-‘和mCdSgab-/-小鼠中观察到的表型特征，我们将对mCdSgab’-/-小鼠进行研究，重点研究补体在人类疾病中的两个表型特征：1)补体介导的溶血性贫血伴血小板激活；2)男性不育。利用现有的C3或C4缺陷小鼠，我们将区分补体及其激活途径的特定作用与CD59的补体非依赖功能。利用只在红细胞中表达hCD59的转基因小鼠，并将其与mCDSgab-/-杂交，我们将验证溶血性贫血与mCD59ab-/-中CD59缺失之间的因果关系。我们的目标的成功实现将确定MAC及其在人类三种典型疾病中的调控作用，并将为研究界提供一个亟需的模型，以继续研究补体在人类病理学中的作用。

DESCRIPTION (provided by applicant): Extensive clinical and experimental evidence strongly indicates that the complement system plays a critical role in the pathogenesis of a diverse group of human diseases that includes some of the hemolytic anemias and male infertility. Given the complex and chronic nature of these diseases in which a pathogenic role of complement still needs to be defined, molecular engineered mouse models that we propose to use in this application provide powerful tools to conduct experiments that would be impossible to perform in humans. Through three different activation pathways and formation of the membrane attack complex (MAC), the complement system mediates both acquired and innate responses to defend against bacterial infection, and to dispose of immune complexes or apoptotic cells. Activated complement and MAC can also damage self cells and thereby mediate pathological responses leading to chronic diseases. An array of complement regulatory proteins including CD59, a key inhibitor of MAC formation, have evolved to protect self cells from complement damage, which can result from either increased complement activation with MAC formation or decreased regulation. Development of adequate animal models to study the relative role of the MAC and its regulation in the pathogenesis of human diseases has been hampered because of the unexpected presence of two Cd59 genes (termed mCd59a and mCd59b) in the mouse genome. Preliminary studies on mCd59a and mCd59b knockout mice demonstrated that the mCd59ab double knockout mouse (mCdSgab-/-), which we have successfully generated by targeted deletion of both genes, was required to investigate the role of MAC regulation in the pathogenesis of human diseases. Our hypothesis is that absence of CD59 function will result in MAC-mediated tissue damage and focal pathology in different organs. Based on the phenotypic features observed in mCdSgb-/-' and mCdSgab-/- mice, we will study mCdSgab'-/- mice focusing on two phenotypic traits representative of human diseases in which a role of complement has long been suspected: 1) complement-mediated hemolytic anemia with platelet activation; and 2) male infertility. Using available C3 or C4 deficient mice, we will differentiate specific roles of complement and its activation pathways from complement-independent functions of CD59. Using transgenic mice that express hCD59 only in RBC and crossing them with mCdSgab-/-, we will verify the causal relationships between the hemolytic anemia and the loss of CD59 in mCd59ab-/-. Successful accomplishment of our goals will define the role of MAC and its regulation in three paradigmatic human diseases, and will provide the research community with a much-needed model to continue studying the role of complement in human pathology.