Reconstituting the oligogenic trait 'Eosinophilic myocarditis to heart disease' in Collaborative Cross lines to understand its genetic architecture

摘要/项目摘要 从一个自发的创始人突变，我们已经建立了几个品系的小鼠， 心脏病（HD），其迅速发展为心力衰竭和过早死亡。受影响者的特征 突变体在多个器官中显示嗜酸性粒细胞增多，包括心脏、肺和脾。受影响的小鼠发育 嗜酸性粒细胞性心肌炎（EM），导致广泛的纤维化和右心室扩张型心肌病， 大多数小鼠在约15周时死亡。临床上，突变小鼠的许多病理与人们认识到的一致。 诊断为嗜酸性粒细胞增多综合征（HES）或嗜酸性肉芽肿病和多血管炎的患者 (EGPA，又名Churg-Strauss综合征），这是两种罕见的嗜酸性粒细胞相关疾病， 伴随着HD。我们在近交A/J品系背景上保持了EM/HD性状（即，A/JHD）， 许多代，所以我们怀疑突变性状的遗传性是由一小部分基因控制的。 EM/HD疾病也可以通过过继转移来自小鼠的脾细胞转移到辐射A/J小鼠。 感染的小鼠，暗示了免疫系统的作用。因为我们对其中的基因知之甚少 在大多数嗜酸性粒细胞性疾病中，我们试图在该突变体中定位EM/HD。对于关联研究，我们系统地 经证明的A/JHD突变雄性（即先前父系受影响的后代）与4个近交系回交和杂交 菌株，并确定SJL/J（SJ）小鼠作为最佳品系杂交组合，以复制EM/HD的映射。 利用QTL分析SJ-F2和N2群体，将HD定位到3个高度显著的位点（命名为 emhd 1 -3表示“嗜酸性心肌炎至心脏病”）。连锁区包括Chr 5上的隐性变体 （Emhd 1），这是必要的，但不是足够的疾病，和2个独立的显性基因座Chr 17（Emhd 2和 Emhd3）。最近，我们发现使用A/JHD和SJ育种者的3代N2/F2交配方案可以 进一步提高产生HD的重组子的产生率。有了这些经验和知识 通过在几个近交系中重建EM/HD获得，沿着更有效的N2/F2育种策略， 我们现在建议概括选择的CC菌株中的寡基因特征。我们的假设是 CC系的遗传多样性将有助于细化连锁区间，确定任何额外的QTL，并确定 杂合区域的重要性（即，它们是否存在显性变异？）to the trait特质.的主要目标 这项资助是为了验证QTL，更好地了解整体遗传力。具体来说，我们将重建 从选择的CC系产生的重组体中的EM/HD，所述选择的CC系在关键位点处携带所需的菌株等位基因， 兴趣所有N2/F2 CC-重组体将被分型为EM/HD，并且相同数量的受影响和 对未受影响的CC-重组体进行基因分型，并进行QTL和单倍型分析。我们希望CC线 将灌输必要的遗传多样性，以明确识别和确认所有重要区域， 性状，并传授对EM/HD的整体遗传结构的理解。这些知识将提供一个 假设驱动研究的基础，旨在描述因果变量及其相互作用。

Abstract / Project Summary From a spontaneous founder mutant, we have established several lines of mice that naturally develop heart disease (HD), which rapidly progresses to heart failure and premature death. Characterization of affected mutants revealed eosinophilia in multiple organs, including the heart, lungs, and spleen. Affected mice develop eosinophilic myocarditis (EM) that leads to extensive fibrosis and right ventricular dilated cardiomyopathy, with most mice dying by ~15-weeks. Clinically, many of the pathologies in mutant mice coincide with those realized in patients diagnosed with hypereosinophilic syndrome (HES) or eosinophilic granulomatosis and polyangiitis (EGPA, aka Churg-Strauss Syndrome), two rare eosinophil-associated diseases with a worse prognosis when accompanied by HD. We have maintained the EM/HD trait on an inbred A/J strain background (i.e., A/JHD) for many generations, so we suspected that heritability of the mutant trait is controlled by a small set of genes. The EM/HD disease could also be transferred to irradiated A/J mice by adoptive transfer of splenocytes from affected mice, implicating a role for the immune system. Because little is known regarding the genes involved in most eosinophilic diseases, we sought to map EM/HD in this mutant. For linkage studies, we systematically backcrossed and intercrossed proven A/JHD mutant males (i.e. previously sired affected offspring) with 4 inbred strains and identified SJL/J (SJ) mice as the best strain–cross combination to reproduce EM/HD for mapping. Using QTL analysis of SJ-derived F2 and N2 populations, HD was mapped to 3 highly significant loci (named Emhd1-3 for ‘eosinophilic myocarditis to heart disease’). Linkage regions included a recessive variant on Chr5 (Emhd1) that is necessary but not sufficient for disease, and 2 separate dominant loci on Chr17 (Emhd2 and Emhd3). Recently, we found that using a 3-generation N2/F2 mating scheme with A/JHD and SJ breeders could further improve the rate of producing recombinants that developed HD. With this experience and knowledge gained from reconstituting EM/HD in several inbred strains, along with a more efficient N2/F2 breeding strategy, we now propose to recapitulate the oligogenic trait in select CC strains. Our working hypothesis is that the genetic diversity of CC lines will help refine linkage intervals, identify any additional QTLs and determine the importance of heterozygous regions (i.e., do they house a dominant variant?) to the trait. The primary goals of this grant are to validate QTLs and better understand the overall heritability. Specifically, we will reestablish EM/HD in recombinants generated from select CC lines that carry the desired strain alleles at critical sites of interest. All N2/F2 CC-recombinants will be phenotyped for EM/HD and an equal number of affected and unaffected CC-recombinants genotyped, and QTL and haplotype analyses performed. We expect the CC lines will instill the genetic diversity necessary to unambiguously identify and confirm all regions of importance to the trait and impart an understanding of the overall genetic architecture of EM/HD. This knowledge will provide a basis for hypothesis-driven studies designed to delineate the causal variants and their interactions.