Conditional Deletion of Dnaic1 as a Model of Primary Ciliary Dyskinesia

描述（由申请人提供）：原发性纤毛运动障碍（PCD）是一种罕见的肺部疾病，通常以常染色体隐性遗传方式遗传。PCD是由损害纤毛功能特性的蛋白质突变引起的。在气道中，这导致粘膜纤毛清除缺失或减少，进而导致呼吸系统的慢性和复发性感染。PCD患者通常表现为慢性鼻炎/鼻窦炎，复发性中耳炎和支气管炎。最近的研究工作已经确定了两种纤毛蛋白DNAI1和DNAH5的突变，这两种突变共同导致了约20%的被分析患者患上这种疾病。然而，尽管这些最近的研究已经开始揭示这种多等位基因疾病的遗传学，但仍有许多关于疾病发病机制的问题尚未得到解答。其中最主要的是在PCD患者中观察到的极低水平呼出的鼻一氧化氮（NO）的机制，以及这种急剧降低的NO水平在疾病发病机制中的作用。在某种程度上，缺乏关于PCD的可用信息是由于缺乏合适的模型系统。这主要是三个主要限制的结果。首先，PCD是纤毛细胞的疾病，由于纤毛细胞是终末分化的，因此没有可用的细胞系来研究PCD。其次，PCD从定义上讲是一种罕见的疾病，可以获得的人体组织的频率和数量都非常有限。最后，在现有的PCD动物模型中，动物会出现脑积水，使它们不适合研究肺部疾病。本研究的主要目标是利用可诱导的CreER/loxP系统，在人类基因DNAI1的小鼠同源物中引入缺失，建立可行的人类PCD小鼠模型。CreER/loxP系统将允许在脑积水发生的一段时间后诱导小鼠PCD表型。为了建立人类PCD疾病的小鼠模型，提出了以下具体目标：目的：通过条件缺失Dnaid建立原发性纤毛畸形模型。具体目标2a。测定dna缺失对纤毛结构、纤毛搏动频率的影响。dna缺失小鼠纤毛粘膜清除率、呼出一氧化氮和对细菌感染的易感性。具体目标2b。确定小鼠杂合dna缺失是否减少了外动力蛋白臂的数量，纤毛功能和清除受损，呼出一氧化氮水平降低，以及对细菌感染的易感性增加。通过建立PCD疾病的小鼠模型，将有可能深入研究关于PCD发病机制和可能治疗的许多关键问题。如果成功，该模型也将为研究有关肺部健康和疾病的其他问题提供一个新的系统。

DESCRIPTION (provided by applicant): Primary ciliary dyskinesia (PCD) is a rare disorder that causes pulmonary disease and is usually inherited in an autosomal recessive manner. PCD is caused by mutations in proteins that impair the ability of cilia to function property. In the airway, this results in absent or reduced mucociliary clearance, that in turn leads to chronic and recurrent infections of the respiratory system. Patients with PCD typically exhibit chronic rhinitis/sinusitis, recurrent otitis media, and bronchitis. Recent work has identified mutations in two ciliary proteins, DNAI1 and DNAH5, which together account for the disease in about 20% of the patients analyzed. However, while these recent studies have begun to unravel the genetics of this multi-allelic disease, there are still many questions regarding disease pathogenesis that remain unanswered. Primary among these is the mechanism responsible for the extremely low levels of exhaled nasal nitric oxide (NO) observed in PCD patients, and the role this drastically reduced level of NO plays in disease pathogenesis. In part, the lack of information available concerning PCD is due to the lack of a suitable model system. This is largely the result of three major limitations. First, PCD is a disease of ciliated cells, and as these cells are terminally differentiated, there are no cell lines available to study PCD. Second, PCD is by definition a rare disease, and the frequency and quantity of human tissue that may be obtained is extremely limited. And finally, in the available animal models of PCD, the animals develop hydrocephalus, making them unsuitable for studying pulmonary disease. The primary goal of this proposal is to develop a viable murine model of human PCD by using an inducible CreER/loxP system to introduce a deletion into the murine homologue of the human gene DNAI1. The CreER/loxP system will allow the induction of the PCD phenotype in mice after the time period during which hydrocephalus develops. To develop a mouse model for the human disease PCD, the following specific aims are proposed: Specific aim 1. To generate a model of Primary Ciliary Dvskinesia by conditional deletion of Dnaid. Specific aim 2a. To measure the effect of Dnaid deletion on ciliary structure, ciliary beat frequency. mucociliary clearance, exhaled nitric oxide, and susceptibility to bacterial infection in Dnaid null mice. Specific aim 2b. To determine if mice heterozygous for Dnaid deletion have reduced numbers of outer dynein arms, impaired ciliary function and clearance, reduced levels of exhaled nitric oxide, and increased susceptibility to bacterial infection. By creating a mouse model for the disease PCD, it will be possible to study in depth many key questions regarding the pathogenesis and possible treatments of PCD. If successful, this model will also provide a novel system to investigate other questions concerning pulmonary health and disease.