Compensatory mechanisms of estrogen mediated protection from EAE in IL-10 KO mice

研究综述 多发性硬化症（MS）是中枢神经系统的炎性自身免疫性疾病，其导致 脱髓鞘引起的神经缺陷和慢性神经炎症引起的神经元损失。MS更普遍 在妇女中，临床症状改善发生在怀孕期间，随后疾病加重 产后众所周知，性激素具有免疫调节特性， 怀孕期间患有MS的人我们以前的工作表明，预处理低剂量雌激素（17β- 雌二醇（E2）保护小鼠免于发生实验性自身免疫性脑脊髓炎（EAE），并促进 调节性B细胞。我们已经证明，E2通过诱导表达PD-1的调节性B细胞来保护小鼠。 L1和/或IL-10。最近我们发现E2可以保护IL-10基因敲除小鼠免于发生EAE（表明IL-10基因敲除小鼠的EAE发生率低于IL-10基因敲除小鼠）。 这些小鼠上调了PD-1/PD-L1/2通路中的蛋白质， 增加调节性B细胞的不同亚群。因此，在本建议中，我们将详细研究这两个主要问题。 E2在EAE期间保护IL-10 KO小鼠的途径，以解决我们的假设 IL-10基因敲除小鼠通过E2的代偿机制而免受EAE的影响。目标一，威尔 定义PD-1/PD-L通路在保护IL-10敲除小鼠中的作用，与野生型小鼠相比。PD-L1， PD-L2和PD-1将被中和抗体抑制，以确定提供的补偿程度 通过E2预处理的IL-10 KO EAE小鼠中的每种蛋白质;目的2，将确定哪些调节性B亚群 在E2介导的IL-10保护作用中，IL-10分泌调节性B细胞正在补偿IL-10的损失 敲除小鼠将在E2预处理的野生型中检查来自脾和淋巴结的调节性B细胞 和IL-10 KO EAE小鼠，以确定在每种小鼠品系中产生哪些调节性B细胞;目的3，将 鉴定哪些细胞因子和趋化因子途径在缺乏IL-10的E2预处理小鼠中上调/下调 与野生型小鼠相比。脊髓将在21岁时使用细胞因子阵列和RT-PCR分析进行评估。 在四组小鼠中免疫后10天：野生型+ E2、野生型+假手术、IL-10 KO + E2和IL-10 + E2。 假的总的来说，这项建议将提供一个更全面的了解免疫调节， E2的性质，并可能导致识别的主导途径，可以激活或扩大 用于治疗MS。

Research Summary Multiple sclerosis (MS) is an inflammatory autoimmune disease of the central nervous system that results in neurological deficits from demyelination and neuron loss from chronic neuroinflammation. MS is more prevalent in women yet improvement in clinical symptoms occurs during pregnancy followed by disease exacerbations post-partum. It is well accepted that sex hormones have immunoregulatory properties and may protect individuals with MS during pregnancy. Our previous work has shown pretreatment with low dose estrogen (17β- estradiol, E2) protects mice from developing experimental autoimmune encephalomyelitis (EAE) and promotes regulatory B cells. We have demonstrated that E2 protects mice by inducing regulatory B cells that express PD- L1 and/or IL-10. Recently we showed E2 can protect IL-10 KO mice from developing EAE (indicating that IL-10 is not required for protection) and that these mice up regulate proteins in the PD-1/PD-L1/2 pathway as well as increasing different subsets of regulatory B cells. In this proposal we thus will examine in detail these two major pathways through which E2 appears to protect IL-10 KO mice during EAE in order to address our hypothesis that IL-10 knockout mice are protected from EAE by E2 through compensatory mechanisms. Aim 1, will define the role of the PD-1/PD-L pathway in protecting IL-10 knockout mice compared to wild type mice. PD-L1, PD-L2, and PD-1 will be inhibited with neutralizing antibodies to determine the extent of compensation provided by each protein in E2 pretreated IL-10 KO mice with EAE; Aim 2, will determine which subsets of regulatory B cells are compensating for the loss of IL-10 secreting regulatory B cells in E2 mediated protection of IL-10 knockout mice. Regulatory B cells from the spleen and lymph nodes will be examined in E2 pretreated wild type and IL-10 KO EAE mice to determine which regulatory B cells are produced in each strain of mouse; Aim 3, will identify which cytokine and chemokine pathways are up/down-regulated in E2 pretreated mice lacking IL-10 compared to wild type mice. Spinal cords will be evaluated using a cytokine array and RT-PCR analysis at 21 days post immunization in four groups of mice: wild type + E2, wild type + sham, IL-10 KO + E2, and IL-10 + sham. Overall this proposal will provide a more comprehensive understanding of the immunoregulatory properties of E2 and could lead to the identification of dominant pathways that could be activated or expanded for the treatment of MS.