Genetics of arthritis suscuptibility in outbred HS mice

描述（由申请人提供）：关节炎疾病的致病链尚不清楚，但人们认为不同的机制聚集在一个共同的效应阶段，涉及先天免疫系统的几个关键细胞和分子。在小鼠和人类中，相当程度的遗传变异导致了这种炎症过程。虽然已经确定了一些位点（IL-1, C5, Ncf1），但大多数这种遗传变异仍然未知。为了在种群基础上最好地了解易感性，需要一个综合的观点，尽可能完整地反映物种的遗传变异。该建议建立在一个成功的R21上，通过“异质库存”（HS）小鼠提供的潜力来解剖K/BxN模型中的关节炎数量性状位点（qtl）。这些动物是由8个创始自交系经50代左右杂交而成的；累积的重组事件带来了高精度QTL定位的潜力，包括物种等位基因库的广泛代表。通过K/BxN血清转移对600只HS小鼠进行筛选，鉴定出33个高分辨率qtl (3-8 Mb)，且具有较高的统计学显著性（LOD评分bbb10）。这个高数字反映了参与这一过程的先天细胞和分子途径的数量。鉴定12个最突出的qtl背后的基因将被解决。在Aim1中，通过在qtl周围进行更密集的基因分型和测试来自定向杂交的其他小鼠，它们的位置将被精确到Mb范围。致病基因将通过序列分析和基因表达谱的组合来寻找，这些序列分析和基因表达谱来自于创始近交系和已经筛选的HS小鼠子集的RNA。在Aim2中，将使用慢病毒载体通过RNA干扰创建表达亚型，并测试结果系对关节炎的易感性，从而评估鉴定出的强定量性状候选基因（QTG）。在Aim3中，将在RA患者中评估HS小鼠中鉴定的QTGs的人类同源物：将对来自24例RA患者的基因的编码和启动子区域进行测序，以确定潜在的疾病特异性变异，并对三组RA患者（2700例+细胞）进行基因分型，并对这些单倍型标记snp进行基因分型，寻找与疾病或严重程度指标的关联。除了确定与关节炎相关的qtl外，该项目还将为先天免疫系统的遗传变异提供有价值的信息。这个提议的项目试图确定控制关节炎炎症的严重程度和破坏性后果的基因。为此，我们正在绘制一种小鼠品系中导致强烈疾病的基因图谱，这对遗传分析尤其有用。一旦确定，我们将测试这些基因是否也可能与人类关节炎患者的易感性有关。这些决定将导致更好地了解关节炎的原因，并提出新的治疗途径。

DESCRIPTION (provided by applicant): The pathogenic chain of events in arthritic diseases is still poorly understood, but it is thought that different mechanisms converge on a common effector phase, which involves several key cells and molecules of the innate immune system. A considerable degree of genetic variation, in mice and human, conditions this inflammatory process. While some loci have been identified (IL-1, C5, Ncf1), most of this genetic variation remains uncharted. In order to best understand susceptibility on a population basis, there is a need for an integrated perspective that incorporates a representation, as complete as possible, of the species' genetic variation. This proposal builds on a successful R21 to dissect arthritis Quantitative Trait Loci (QTLs) in the K/BxN model via the potential offered by "Heterogenous Stock" (HS) mice. These animals were generated from 8 founder inbred strains by >50 generations' intercrossing; the accumulated recombination events bring the potential for high-precision QTL mapping, encompassing a broad representation of the species' allele pool. A screen involving 600 HS mice tested by transfer of K/BxN serum identified 33 QTLs at high resolution (3-8 Mb) and a high level of statistical significance (LOD score >10). This high number reflects the number of innate cellular and molecular pathways involved in the process. Identification of the genes underlying the 12 most prominent QTLs will be tackled. In Aim1, their position will be refined to the Mb range by denser genotyping around the QTLs and by testing additional mice from directed crosses. The causal genes will be pursued by a combination of sequence analysis and gene-expression profiling in founder inbreds and in RNA from a subset of the HS mice already screened. In Aim2, the strong Quantitative Trait Genes candidates (QTG) thus identified will be evaluated by using lentiviral vectors to create expression hypomorphs through RNA interference, and testing the resulting lines for susceptibility to arthritis. In Aim3, the human orthologs of the QTGs identified in the HS mice will be evaluated in RA patients: coding and promoter regions of the genes from 24 RA patients will be sequenced in order to identify potential disease-specific variants, and three cohorts of RA patients (2700 cases +ctls) will be genotyped for these and haplotype-tagging SNPs, searching for association to disease or to severity indices. Beyond identifying QTLs of relevance to arthritis, this project should provide valuable information on genetic variation in the innate immune system in general. This proposed project attempts to identify genes that control the severity and destructive consequences of inflammation in arthritis. To this end, we are mapping the genes that lead to strong disease in a strain of mice particularly informative for genetic analyses. Once identified, we will test whether these genes may also be involved in setting susceptibility to arthritis in human patients. These determinations should lead to a better understanding of the causes of arthritis, and suggest novel avenues for treatments.