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

研究总结 多发性硬化症(MS)是一种炎症性自身免疫性中枢神经系统疾病，导致 脱髓鞘所致的神经功能缺陷和慢性神经炎所致的神经元丢失。多发性硬化症更为普遍 然而，在女性中，临床症状的改善发生在怀孕期间，随后疾病恶化 产后。众所周知，性激素具有免疫调节特性，并可能保护 怀孕期间患有多发性硬化症的个体。我们之前的工作表明，用低剂量雌激素(17β- 雌二醇对实验性自身免疫性脑脊髓炎(EAE)小鼠的保护作用 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 KO小鼠免受EAE的发展(表明IL-10 不需要保护)，这些小鼠上调PD-1/PD-L1/2途径中的蛋白质以及 增加不同的调节性B细胞亚群。因此，在这项建议中，我们将详细研究这两大 为了解决我们的假说，E2在EAE期间保护IL-10 KO小鼠的途径 IL-10基因敲除小鼠通过代偿机制保护EAE免受EAE的侵袭。目标1，意志 明确PD-1/PD-L通路在保护IL-10基因敲除小鼠和野生型小鼠中的作用。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处理的IL-10KO小鼠中的每一种蛋白；目标2，将确定调控B的哪些亚群 在E2介导的IL-10保护中，细胞对分泌IL-10的调节性B细胞的丧失进行补偿 基因敲除老鼠。来自脾和淋巴结的调节性B细胞将在E2预处理的野生型中进行检测 和IL-10 KO EAE小鼠，以确定在每个品系的小鼠中产生哪些调节性B细胞；Aim 3，将 确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 免疫后天数：野生型+E2组、野生型+假手术组、IL-10KO+E2组和IL-10+组 假的。总体而言，这项提案将提供对免疫调节的更全面的了解 E2的特性，并可能导致识别可被激活或扩展的优势通路 治疗多发性硬化症

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.