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

小胶质细胞，大脑的常驻免疫细胞，与脑缺血的病理生理学有关。 慢性缺血性脑小血管病（CSVD）和血管性认知功能障碍（VCI）是临床上 痴呆的重要原因，优先影响白色物质（WM）。集落刺激因子1受体 CSF 1 R是髓系细胞的关键调节因子。CSF 1 R的遗传缺失阻断了正常人群 常驻小胶质细胞和用CSF 1 R拮抗剂如PLX 5622的全身治疗导致显著的 CNS中小胶质细胞的耗竭。在这里，我们建议评估小胶质细胞和CSF 1 R信号转导的作用， 在我们的同事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University）过度表达转化生长因子β 1（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 类似于人类CSVD/VCI我们将首先评估TGFOE和HCD对区域和细胞类型的影响， 使用单细胞RNA测序在小胶质细胞和其他髓样细胞中特异性基因表达。接下来我们将 确定PLX 5622治疗是否影响神经系统、神经影像学和/或神经血管 TGFOE小鼠（+/-HCD）的结果。我们将评估纵向WM完整性的结果， 最先进的14特斯拉磁共振成像与扩散张量成像（DTI）和定量纤维束成像。我们还将使用 静息状态功能连接的复杂评估（内在信号的光学成像）， 神经认知此外，我们还将进行纵向双光子脑成像研究，以跟踪 在CSVD/ VCI疾病发生和进展的关键时期，我们将 通过交叉验证，我们使用正交遗传方法证实了我们研究结果的小胶质细胞特异性 具有Csf 1 rflox/flox：HexbCreERT 2双基因系的TGFOE小鼠表现出稳定的小胶质细胞特异性敲低， Csf1r。然后，我们将进料三基因TGFOE：Csf 1 rflox/flox：HexbCreERT 2系HCD，并进行相同的 上述纵向参数的综合集合。我们预计， 小胶质细胞的耗竭和小胶质细胞中特异性CSF 1 R信号转导的基因缺失将减弱WM 并导致多个结果参数改善。在Aim 3中，我们将合作 我们与放射科的同事一起使用一种新的CSF 1 R放射性配体进行纵向正电子扫描， 在疾病开始期间的多个时间点对TGFOE小鼠（+/-HCD）进行发射断层扫描（PET）， 进展这将提供关键信息的时间和空间的体内表达模式， CSF 1 R在CSVD/VCI的背景下，并为一种新的生物标志物生成基础数据， 神经炎症可以很容易地从小鼠扩大到人类。总的来说，这种小胶质细胞和CSF 1 R 信号靶向，WM为中心的方法将提供重要的机制信息的发病机制， CSVD/VCI中缺血性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.