Role of HLA Class II Genes in Demyelination

描述(由申请人提供)：多发性硬化症(MS)，一种炎症性和脱髓鞘的自身免疫性疾病，具有遗传和环境易感性。在所有与MS易感性相关的遗传因素中，DR2/DQ6、DR3/DQ2、DR4/DQ8等HLAII类单倍型与MS的相关性最强。虽然人类白细胞抗原-DR等位基因在多发性硬化发病中的直接作用已被证实，但由于强烈的连锁不平衡，人类白细胞抗原-DQ等位基因在疾病发病机制中的作用一直难以理解。人群研究表明，DQ等位基因可能在MS的进展中起着调节作用。为了更好地了解人类白细胞抗原-DR和-DQ基因在MS易感和抵抗中的作用机制，我们建立了表达HLAII类基因但缺乏内源性II类基因的单、双转基因小鼠。已有研究表明，人类白细胞抗原DR3转基因小鼠对PLP91-110诱导的EAE易感，而DQ6(DQB1*0601)和DQ8(DQB1*0302)转基因小鼠对PLP91-110诱导的EAE具有抗性。令人惊讶的是，DQ6/DR3双转基因小鼠具有抵抗力，而DQ8/DR3小鼠的发病率和严重程度更高。DQ6对DQ6/DR3小鼠的保护作用是通过抗炎因子IFN3介导的，而DQ8分子的加重作用是通过IL17介导的。基于这些观察结果，我们推测，人类白细胞抗原-DR和-DQ基因之间的上位性相互作用在MS的易感性中起着重要作用。这一建议旨在加深对人类白细胞抗原-DQ和-DR分子之间的上位性相互作用决定疾病易感性和抵抗力的机制的理解。我们提出了两个目的来了解在人类白细胞抗原DR/DQ双转基因小鼠中，人类白细胞抗原-DQ分子调节疾病转归的机制。在第一个目标中，我们将通过以下分析来研究人类白细胞抗原-DQ8分子如何增加DQ8/DR3双转基因小鼠的发病率和严重程度：i)人类白细胞抗原多态和多肽-MHC亲和力在DQ8限制性CD4T细胞产生促炎性IL17中的作用；ii)IL17导致DR3DQ8小鼠脑原性增加的机制；以及iii)DQ8限制性CD4T细胞产生的GM-CSF在DR3DQ8双转基因小鼠EAE加重中的作用。在第二个目标中，我们将研究人类白细胞抗原-DQ6限制性免疫反应如何导致产生高分泌IFN3的调节性CD4+T细胞，特别是MHC-肽亲和力/功能亲和力的作用。接下来，我们将分析DQ6限制性CD4+T细胞抑制HLADR3/DQ6双转基因小鼠EAE的机制。我们还将产生一个表达疾病易感基因-DR3、疾病保护基因-DQ6和疾病增强基因-DQ8的三重转基因小鼠，以模拟人类杂合条件。这项提案中概述的全面研究应该能深入了解人类白细胞抗原II类分子塑造T细胞库和调节促炎和抗炎细胞因子的机制。这将促进治疗MS等炎症性疾病的新疗法的翻译研究开发。 公共卫生相关性：尽管人类白细胞抗原单倍型与多发性硬化症的易感性和发病有关，但过去很难明确人类白细胞抗原分子(S)在多发性硬化症中的作用。我们已经成功地利用人类白细胞抗原转基因小鼠，通过实验性自身免疫性脑脊髓炎(EAE)模型，了解了它们在多发性硬化症等中枢神经系统炎症和脱髓鞘疾病中的作用。这些人源化的II类EAE模型可用于评估未来适用于MS患者的潜在治疗方案和多肽疫苗。

DESCRIPTION (provided by applicant): Multiple sclerosis (MS), an inflammatory and demyelinating autoimmune disease has both a genetic and an environmental predisposition. Among all the genetic factors associated with MS susceptibility, HLA-class II haplotypes such as DR2/DQ6, DR3/DQ2, DR4/DQ8, show the strongest association. Although a direct role of HLA-DR alleles in MS have been confirmed, it has been difficult to understand the contribution of HLA-DQ alleles in disease pathogenesis, due to strong linkage disequilibrium. Population studies have indicated that DQ alleles may play a modulatory role in progression of MS. To better understand the mechanism by which HLA-DR and -DQ genes contribute to susceptibility and resistance to MS, we generated single and double transgenic mice expressing HLA class II genes and lacking endogenous mouse class II genes. Previously, we have shown that HLA-DR3 transgenic mice were susceptible to PLP91-110 induced EAE, while DQ6 (DQB1*0601) and DQ8 (DQB1*0302) transgenic mice were resistant. Surprisingly DQ6/DR3 double transgenic mice were resistant while DQ8/DR3 mice showed higher disease incidence and severity. Protective effect of DQ6 in DQ6/DR3 mice was mediated by anti-inflammatory IFN3, while disease exacerbating effect of DQ8 molecule was mediated by IL17. Based on these observations, we hypothesize that epistatic interaction between HLA-DR and -DQ genes play an important role in predisposition to MS. This proposal is aimed to enhance understanding of the mechanism by which epistatic interactions between HLA-DQ and -DR molecules determine the susceptibility vs. resistance to disease. We are proposing two aims to understand the mechanism by which HLA-DQ molecule modulate the disease outcome in HLA-DR/DQ double transgenic mice. In the first aim, we will examine how HLA-DQ8 molecule increases disease incidence and severity in DQ8/DR3 double transgenic mice by analyzing- i) role of HLA polymorphism and peptide-MHC affinity/avidity in generation of pro-inflammatory IL17 production from DQ8 restricted CD4 T cells; ii) mechanism by which IL17 cause increased encephalitogenicity in DR3DQ8 mice; and iii) role of GM-CSF produced by DQ8 restricted CD4 T cells in exacerbation of EAE in DR3DQ8 double transgenic mice. In the second aim, we will investigate how HLA-DQ6 restricted immune response leads to generation of high IFN3 producing regulatory CD4+ T cells, especially the role of MHC-peptide affinity/functional avidity. Next we will analyze mechanisms by which DQ6 restricted CD4+ T cells suppress EAE in HLA-DR3/DQ6 double transgenic mice. We will also generate a triple transgenic mice expressing disease susceptible HLA-DR3, disease protective -DQ6, and disease enhancing -DQ8 gene to simulate human heterozygous condition. The comprehensive studies outlined in this proposal should yield an insight into mechanism by which HLA class II molecules shape the T cell repertoire and regulate the pro-inflammatory and anti-inflammatory cytokine profile. This will facilitate the translational research development of novel therapeutic to treat inflammatory disease such as MS. PUBLIC HEALTH RELEVANCE: Although HLA haplotypes are linked to predisposition and onset of multiple sclerosis, it has been difficult in past to define a clear role of HLA molecule(s) in MS. We have used HLA transgenic mice successfully to understand their role in inflammatory and demyelinating disease of CNS such as MS using an experimental model experimental autoimmune encephalomyelitis (EAE). These humanized class II model of EAE can be used to evaluate potential therapeutic protocols and peptide vaccines applicable in the future to MS patients.