IMMUNOREGULATORY EFFECTS OF ESTROGEN IN EAE

描述(申请人提供)：多发性硬化症患者 (Ms)，存在明显的性别偏见，大约是这个数字的两倍 受影响的雌性与雄性相同。性激素可能导致易感性或 通过影响潜在的发育和功能对MS的抗性 中枢神经系统(CNS)抗原特异的致病T细胞 作为调节性T细胞，可能会改变疾病的进程。此前，我们 据报道，低剂量的17β-雌二醇(E2)可通过以下方式减轻EAE的严重程度 抑制激活、细胞因子和趋化因子的产生，以及 小鼠髓鞘少突胶质细胞特异性T细胞的脑源性研究 糖蛋白(MOG)、蛋白脂蛋白(PLP)或髓鞘碱性蛋白(MBP)， 并通过抑制炎症细胞向中枢神经系统的募集。的 尤其重要的是，雌激素治疗大大减少了细胞内 在EAE和EAE中产生作为关键炎性细胞因子的肿瘤坏死因子-α MS最近使用微阵列分析评估了E2对EAE的影响 脾细胞基因表达证实了对肿瘤坏死因子-α的有效抑制，并且 进一步确定了几个以前未被怀疑与免疫相关的候选对象 在体内，似乎受到E2处理强烈影响的基因。这些 雌激素对致病细胞、募集细胞和调节细胞的调节作用 在EAE中可能通过与雌激素受体(ER)直接相互作用而介导， 其中包括经典的ER-α和ER-β，它们是内部的f受体 (IER)，以及可能与IERS不同的膜ERs(Mer)。这 该提案将检验雌激素对EAE的调节包括 受体介导的肿瘤坏死因子-α调节及其他几种新的免疫相关因子 基因，从而抑制巨噬细胞的炎症效应， 树突状细胞和T细胞，增强调节性NKT细胞活性。我们的 主要目标是确定E2对EAE的抑制作用是否可以 在缺乏调节性T和NKT细胞的自发性EAE中观察到，以及 如果E2对EAE的抑制作用是通过IER-α和/或IER-β介导的， 或者相反，这两个经典的IERS都不是。使用新开发的 ER-α和ER-β基因敲除小鼠我们将首次将E2- IERS中的一种或两种IERS的依赖调节，或MER IF E2的替代调节 这种影响在双KO小鼠身上持续存在。此外，对于每个新的受E2影响的基因 从微阵列分析中推断，我们将调查细胞 来源，E2对转录和蛋白质产生的影响，以及 对体内诱发EAE的贡献。发现在EAE中重要的基因 将在不同水平的女性的血细胞中进一步评估 E2。这项拟议的工作将确定关键的雌激素敏感基因，包括 肿瘤坏死因子α，有助于EAE的诱导和抗性。这些基因的变化 可在雌激素治疗期间作为替代标记物进行验证 有效剂量的雌激素。从这项研究中，我们将制定一个坚实的 使用雌激素替代疗法治疗MS的依据

DESCRIPTION (provided by applicant): In patients with multiple sclerosis (MS), there is a distinct gender bias, with approximately twice as many affected females as males. Sex hormones may contribute to susceptibility or resistance to MS by influencing development and function of potentially pathogenic T cells specific for central nervous system (CNS) antigens, as well as regulatory T cells that might modify the course of disease. Previously, we reported that low doses of 17beta-estradiol (E2) can reduce severity of EAE by inhibiting activation, cytokine and chemokine production, and encephalitogenicity of murine T cells specific for myelin oligodendrocyte glycoprotein (MOG), proteolipid protein (PLP), or myelin basic protein (MBP), and by inhibiting I recruitment of inflammatory cells into the CNS. Of particular importance, estrogen treatment profoundly reduced intracellular production of TNF-alpha, known as a critical inflammatory cytokine in EAE and MS. Recent evaluation of E2 effects on EAE using microarray analysis of splenocyte gene expression confirmed potent inhibition of TNF-alpha, and further identified several previously unsuspected immune-associated candidate genes appearing to be strongly affected by E2 treatment in vivo. These modulatory effects of estrogen on pathogenic, recruited, and regulatory cells in EAE are likely mediated y direct interaction with estrogen receptors (ER), which include the classical ER-alpha and ER-beta that are internal f receptors (iER), and possibly membrane ERs (mER) that may be distinct from iERs. This proposal will test the hypothesis that modulation of EAE by estrogen involves receptor-mediated regulation of TNF-alpha and several other novel immunerelated genes, thereby inhibiting inflammatory effects of macrophages, dendritic cells and T cells and enhancing regulatory NKT cell activity. Our primary goals are to determine if the inhibitory effects of E2 on EAE can be observed in spontaneous EAE in the absence of regulatory T and NKT cells, and if inhibitory effects of E2 on EAE are mediated through iER-alpha and /or iERbeta, or in contrast, by neither of the classical iERs. Using newly developed ER-alpha and ER-beta knockout mice we will for the first time associate E2- dependent regulation with either or both iERs, or alternatively with mER if E2 effects persist in double KO mice. Moreover, for each novel E2-affected gene implicated from the microarray analysis, we will investigate the cellular source, effects of E2 on transcription and protein production, and contribution to EAE induction in vivo. Genes found to be important in EAE will be further evaluated in blood cells from women with different levels of E2. The work proposed will identify key estrogen-sensitive genes, including TNFalpha, contributing to EAE induction and resistance. Changes in these genes can be followed during estrogen treatment as surrogate markers to verify effective doses of estrogen. From this research, we will develop a solid basis for using estrogen replacement therapy for MS.