Role of Microglia in Cerebral Small Vessel Disease (CSVD)/Vascular Cognitive Impairment (VCI)

项目摘要：小胶质细胞是大脑的常驻免疫细胞，与脑血管疾病的病理生理学有关。 慢性缺血性脑小血管病(CSVD)和血管认知功能障碍(VCI)的临床研究 痴呆的重要原因，优先影响脑白质(WM)。集落刺激因子1受体 (CSF1R)是髓系细胞的关键调节因子。CSF1R的遗传缺失阻碍了正常人群的 驻留的小胶质细胞和全身应用CSF1R拮抗剂，如PLX5622，结果显著 中枢神经系统小胶质细胞耗竭。在这里，我们建议评估小胶质细胞和CSF1R信号的作用 在我们同事Edith Hamel博士的实验室开发的一种新的CSVD/VCI啮齿动物模型中 (麦吉尔大学)。过度表达转化生长因子-β1(TGFOE)并喂食高胆固醇的小鼠 饮食(HCD)显示脑血管反应性受损、弥漫性白质脑病和认知功能障碍 这与人类的CSVD/VCI相似。我们将首先评估TGFOE和HCD对区域和细胞类型的影响 用单细胞RNA测序法研究小胶质细胞和其他髓系细胞中特异性基因的表达。接下来我们将 确定PLX5622治疗是否影响神经、神经影像和/或神经血管 TGFOE小鼠(+/-HCD)的结果。我们将使用以下状态评估纵向WM完整性结果 -ART-14T磁共振弥散张量成像(DTI)和定量纤维束成像。我们还将使用 静息状态功能连通性的复杂评估(固有信号的光学成像)和 神经认知。此外，我们将进行纵向双光子脑成像研究，以跟踪 CSVD/VCI疾病发生发展关键时期的脑血管系统我们会 通过杂交，使用正交遗传方法确认我们的研究结果的小胶质细胞特异性 具有Csf1rflx/Flox：HexbCreERT2双基因系的TGFOE小鼠表现出稳定的小胶质细胞特异性基因敲除 CSF1R.然后，我们将供给TGFOE：Csf1rflx/Flox：HexbCreERT2线HCD并执行相同的操作 如上所述的全套纵向参数。我们预计这两种药物 小胶质细胞的耗尽和CSF1R信号的基因缺失将使WM减弱 炎症和损伤，并导致多个结局参数的改善。在Aim 3中，我们将合作 与我们放射学的同事一起使用一种新型的CSF1R放射性配基进行纵向正电子 TGFOE小鼠(+/-HCD)发病和发病过程中多个时间点的发射断层扫描(PET) 进步。这将提供有关在体时间和空间表达模式的关键信息 CSF1R在CSVD/VCI的背景下，并为一种新的生物标记物生成基础数据 神经炎症可以很容易地从老鼠扩大到人类。总体而言，这种小胶质细胞-和CSF1R 以信号转导为靶点，以西医为中心的方法将为急性髓细胞白血病的发病机制提供重要的机制信息 CSVD/VCI中的缺血性WM损伤，并阐明新的潜在诊断和治疗方法。

PROJECT SUMMARY: Microglia, the brain’s resident immune cells, are implicated in the pathophysiology of chronic ischemic cerebral small vessel disease (CSVD) and vascular cognitive impairment (VCI), a clinically important cause of dementia that preferentially affects white matter (WM). Colony stimulating factor 1 receptor (CSF1R) is a key regulator of myeloid lineage cells. Genetic loss of CSF1R blocks the normal population of resident microglia and systemic treatment with CSF1R antagonists such as PLX5622 results in a marked depletion of microglia in the CNS. Here we propose to evaluate the role of microglia and CSF1R signaling in a novel rodent model of CSVD/VCI developed in the laboratory of our colleague Dr. Edith Hamel (McGill University). Mice overexpressing transforming growth factor-beta1 (TGFOE) and fed a high cholesterol diet (HCD) display impaired cerebrovascular reactivity, diffuse leukoencephalopathy, and cognitive dysfunction that resembles human CSVD/VCI. We will first assess effects of TGFOE and HCD on regional and cell type specific gene expression in microglia and other myeloid cells using single cell RNA sequencing. Next we will determine if treatment with PLX5622 influences neurological, neuroimaging, and/or neurovascular outcomes in TGFOE mice (+/-HCD). We will assess longitudinal WM integrity outcomes using state-of- the-art 14 Tesla MRI with diffusion tensor imaging (DTI) and quantitative tractography. We will also use sophisticated assessments of resting-state functional connectivity (optical imaging of intrinsic signals) and neurocognition. Furthermore, we will perform longitudinal 2-photon brain imaging studies to track changes in the cerebrovasculature during critical periods of CSVD/ VCI disease initiation and progression. We will confirm the microglia-specific nature of our findings using an orthogonal genetic approach by crossing TGFOE mice with a Csf1rflox/flox : HexbCreERT2 bigenic line that exhibits stable, microglia-specific knockdown of Csf1r. We will then feed the trigenic TGFOE : Csf1rflox/flox : HexbCreERT2 line HCD and carry out the same comprehensive set of longitudinal parameters described above. We anticipate that both pharmacologic depletion of microglia and genetic deletion of CSF1R signaling specifically in microglia will attenuate WM inflammation and injury and result in improvement in multiple outcome parameters. In aim 3 we will collaborate with our colleagues in Radiology to use a novel CSF1R radioligand to carry out longitudinal positron emission tomography (PET) on TGFOE mice (+/-HCD) at multiple time points during disease initiation and progression. This will provide critical information on temporal and spatial in vivo expression patterns of CSF1R in the context of CSVD/VCI and generate foundational data for a novel biomarker for neuroinflammation that can be readily scaled up from mouse to humans. Overall, this microglia- and CSF1R signaling-targeted, WM-centric approach will provide important mechanistic information on the pathogenesis of ischemic WM injury in CSVD/VCI and elucidate new potential diagnostic and therapeutic approaches.