Molecular Genetics of Lyme Arthritis

描述（由申请人提供）：莱姆病是由蜱传播的伯氏疏螺旋体感染引起的，是美国最常见的病媒传播疾病。感染与一系列疾病症状和严重程度相关，高达60%的感染患者发展为关节炎。关节炎严重程度的差异被发现， 近交系小鼠品系，其中C3 H小鼠显示严重疾病，C57 BL/6（B6）小鼠发展轻度疾病。使用正向遗传方法，我们已经确定了数量性状基因座（QTL），调节响应B。burgdorferi，其中6个QTL调节莱姆关节炎的严重程度（Bbaa’s 1，2，3，4，6，12）。先进的重组同源系的开发允许在Chr 5上的Bbaa 2内定位克隆β-葡糖醛酸酶（Gusb），作为关节炎严重程度的主要调节剂。Gusb的C3 H等位基因是亚型的，导致未消化的糖胺聚糖（GAG）在关节组织中的积累，其似乎起到导致B恶化的第二次打击的作用。引发关节炎和类风湿性关节炎。本申请结合了在莱姆病关节炎的调节中鉴定的基因的机制表征。在第一个目标中，GAG将被生物合成标记，并通过HPLC和质谱法从同源和转基因小鼠中进行表征，以鉴定那些在炎性关节炎中富集的GAG。然后将使用点击木糖酶来引发GAG合成，从而允许回收和评估具有炎症潜力的GAG。在突变小鼠中的GAG谱的表征将允许记录可能参与莱姆关节炎发展的外切糖苷酶的宽度。通过定量来自不同B阶段患者样本中涉及GAG降解的几种外切糖苷酶，将这些发现转化为莱姆病患者。感染和对抗生素治疗显示不同临床反应的患者。生物信息学方法已经开始鉴定GUSB功能的修饰剂，并与先进的重组体相结合，提供了令人信服的证据，证明Chr 5上连接的其他基因修饰了GUSB的关节炎作用。进一步发展先进的重组同源基因和双同源基因，目的是确定相互作用，调节莱姆关节炎的严重程度。第二个QTL的C3 H等位基因，Bbaa 1的Chr 4，也显示出强烈的渗透性关节炎表型。在这种情况下，关节炎依赖于I型IFN的产生。Bbaa 1也编码I型IFN基因簇。因此，先前在C3 H小鼠中观察到的I型IFN的过度活跃表达是由Bbaa 1内的基因直接调节的，并且可能是IFN基因座固有的。本实验拟研究I型干扰素的遗传调控和I型干扰素的过度产生导致莱姆病关节炎严重程度的机制。本申请利用了先前对莱姆关节炎的前瞻性遗传研究，并应提供对炎症病理的新颖且广泛适用的见解。

DESCRIPTION (provided by applicant): Lyme disease is caused by infection with the tick borne spirochete Borreliai burgdorferi and is the most common vector borne disease in the United States. Infection is associated with a spectrum of disease symptoms and severity, with up to 60% of infected patients developing arthritis. Differences in arthritis severity are found in inbred mouse strains, with C3H mice displaying severe disease and C57BL/6 (B6) mice developing mild disease. Using a forward genetic approach we have identified quantitative trait loci (QTL) that regulate the response to B. burgdorferi, with six of these QTL regulating the severity of Lyme arthritis (Bbaa's 1,2,3,4,6,12). Development of advanced recombinant congenic lines allowed the positional cloning of beta- Glucuronidase (Gusb) within Bbaa2 on Chr5, as a major regulator of arthritis severity. The C3H allele of Gusb is hypormorphic, resulting in the accumulation of undigested glycosaminoglycans (GAGs) in joint tissue, which appear to function as a second hit leading to the exacerbation of B. burgdorferi triggered arthriti and rheumatoid arthritis. The current application incorporates mechanistic characterization of the genes identified in the regulation of Lyme arthritis. In the first Aim, GAGs will be biosynthetically labeled and characterized by HPLC and Mass Spectrometry from congenic and transgenic mice in order to identify those enriched in inflammatory arthritis. Click-xylosidses wil then be used to prime GAG synthesis, allowing the recovery and assessment of GAGs with inflammatory potential. Characterization of the GAG-profile in mutant mice will allow documentation of the breadth of exoglycosidases that could be involved in Lyme arthritis development. These findings will be translated to patients with Lyme disease, by quantifying several exoglycosidases involved in GAG degradation in samples from patients at different stages of B. burgdorferi infection and from patients displaying different clinical responses to antibiotic treatment. A bioinformatics approach has been initiated to identify modifiers of GUSB function, and in conjunction with advanced recombinants provides convincing evidence for additional genes linked on Chr5 that modify the arthritis effect of Gusb. Further development of advance recombinant congenics and double congenics are proposed, with the goal of identifying interactions that modulate Lyme arthritis severity. The C3H allele of a second QTL, Bbaa1 on Chr4, also displays a strongly penetrant arthritis phenotype. In this case, arthritis is dependent on the production of Type I IFN. Bbaa1 also encodes the Type I IFN gene cluster. Therefore, the hyperactive expression of Type I IFN previously observed in C3H mice is directly regulated by genes within Bbaa1 and possibly inherent to the IFN locus. Experiments are proposed to study the genetic regulation of Type I IFN and the mechanism by which hyper-production of Type I IFN directs the severity of Lyme arthritis. This application capitalizes on ou prior forward genetic studies of Lyme arthritis and should provide novel and broadly applicable insight into inflammatory pathologies.