Border-associated macrophages in an alpha-synuclein overabundance model of Parkinson Disease

项目摘要/摘要 这项建议旨在确定边界相关巨噬细胞(BAMS)的贡献，以及 α-突触核蛋白中单核细胞浸润和神经变性的炎性信号 建立帕金森病(PD)小鼠模型。我们的实验室一直在追寻一种想法，即天然免疫系统 对α-SYN的反应激活是帕金森病神经炎症和神经变性的触发因素，并已 与模型进展有关的外周血单核细胞和小胶质细胞。越来越多的尸检证据 组织和帕金森病血液和脑脊液强调了炎症在疾病中的作用，如小胶质细胞增多症和 病变周围可见淋巴细胞浸润和变性。此外，促炎作用 细胞因子在帕金森病死后脑和脑脊液中升高，而趋化因子如CCL2，这是重要的 在CCR2+中，单核细胞募集到组织中，在PD血液中显著升高。鼠标模型重述关键 帕金森病的先天免疫激活、外周细胞浸润和神经变性的特征。 然而，BAMS在外周免疫细胞募集和外渗到中枢神经系统中起着关键作用，但 在帕金森病的研究中基本上被忽视了。这一建议试图提高我们对中枢神经系统炎症的认识 通过研究α-SYN诱导的BAM激活和CCl_2分泌所需的假说的机制 用于炎性单核细胞募集和神经变性。 使用体内模型，其中使用腺相关病毒产生α-SYN过表达 (AAV)，小胶质细胞MHCII表达增加，BAM数量和MHCII表达增加，CCR2+数量增加 已观察到促炎症的单核细胞。此外，通过以下途径防止单核细胞招募 CCR2基因敲除具有神经保护作用，可阻止小胶质细胞MHCII的表达。拟议的项目将首先 应用流式细胞仪、流式细胞仪和一种定量聚合酶链式反应细胞因子研究α-SYN对骨髓间充质干细胞的激活状态 和AAV2-SYN模型中的趋化因子阵列。接下来，将专门耗尽BAMS以确定其 在α-SYN诱导的外周免疫细胞浸润中的作用。这将通过流式细胞仪进行测量 和免疫组织化学。最后，BAMS耗尽，并用CCL2敲除的外周细胞重新填充 将被用来确定它们在炎症和神经退化中的作用。这些研究使用的是骨髓 嵌合体、流式细胞术、免疫组织化学和体视学，以确定对炎症和 神经退行性变。 帕金森病是第二种最常见的神经退行性疾病，但仍然没有 足够的治疗。这项建议旨在阐明BAMS在大脑和大脑之间的交流中的作用 帕金森病进展过程中的外周状态。这项研究将为靶向趋化因子信号转导和 髓系细胞作为开发帕金森病治疗药物的创新靶点。

Project Summary/Abstract This proposal aims to determine the contribution of the border associated macrophages (BAMs), and inflammatory signaling from these cells, to monocyte infiltration and neurodegeneration in an alpha-synuclein based mouse model of Parkinson Disease (PD). Our lab has pursued the idea that innate immune system activation in response to α-syn is a trigger for such neuroinflammation and neurodegeneration in PD, and has implicated peripheral monocytes and microglia in model progression. Increasing evidence from both postmortem tissue and PD blood and CSF has highlighted the role of inflammation in the disease, as microgliosis and lymphocyte infiltration is found surrounding pathology and degeneration. Additionally, pro-inflammatory cytokines are elevated in PD post-mortem brain and CSF, while chemokines such as CCL2, which is important in CCR2+ monocyte recruitment to tissues, are robustly elevated in PD blood. Mouse models recapitulate key features of the innate immune activation, peripheral cell infiltration, and neurodegeneration found in PD. However, BAMs have key roles in peripheral immune cell recruitment and extravasation into the CNS, but have largely been ignored in PD research. This proposal attempts advance our knowledge of CNS inflammatory mechanisms by investigating the hypothesis that α-syn induced BAM activation and CCL2 secretion is required for inflammatory monocyte recruitment and neurodegeneration. Using an in vivo model in which α-syn overexpression is produced using an adeno-associated virus (AAV), increases in microglial MHCII expression, BAM number and MHCII expression, and number of CCR2+ pro-inflammatory monocytes have been observed. Furthermore, prevention of monocyte recruitment through CCR2 knock-out is neuroprotective and prevents microglial MHCII expression. The proposed project will first investigate the activation status of BAMs in response to α-syn, using flow cytometry, FACS, and a qPCR cytokine and chemokine array in the AAV2-SYN model of PD. Next, BAMs will be specifically depleted to determine their functional role in α-syn induced peripheral immune cell infiltration. This will be measured through flow cytometry and immunohistochemistry. Finally, depletion of BAMs and re-population with CCL2 knock-out peripheral cells will be used to determine their role in inflammation and neurodegeneration. These studies use bone marrow chimeras, flow cytometry, immunohistochemistry, and stereology to determine protection from inflammation and neurodegeneration. Parkinson disease is the second most common neurodegenerative disorder, but remains without sufficient treatments. This proposal aims to elucidate the role of BAMs in communication between the brain and periphery during PD progression. This study would provide rationale for targeting chemokine signaling and myeloid cells as an innovative target for the development of PD therapeutics.