Interaction of GCSF with the Endocannabinoid System in Promoting Brain Repair

GCSF 与内源性大麻素系统的相互作用促进大脑修复

• 批准号: 10292953
• 负责人: Shijie Song
• 金额: --
• 依托单位: JAMES A. HALEY VA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-10-01 至 2023-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10292953
• 关键词: 2-arachidonylglycerol; Agonist; Amides; Animals; Anti-Inflammatory Agents; Apoptosis; Astrocytosis; Basic Science; Behavioral; Blood Cells; Bone Marrow; Bone Marrow Transplantation; Brain; Brain Injuries; Brain region; Brain-Derived Neurotrophic Factor; CB2 receptor antagonist; CNR1 gene; CNR2 gene; CSF3 gene; Cannabinoids; Cell Differentiation process; Cells; Clinic; Colony-Stimulating Factor Therapy; Contralateral; Corpus striatum structure; Disease; Endocannabinoids; Ethanolamines; Fatty Acids; Fostering; Goals; Granulocyte Colony-Stimulating Factor; Hippocampus (Brain); Human; IL3 Gene; Injury; Knockout Mice; Lesion; Leukocytes; Ligands; Mediating; Messenger RNA; Microglia; Mus; Neuraxis; Neurodegenerative Disorders; Neurons; Performance; Phenotype; Population; Process; Proteins; Protocols documentation; Radial; Recovery; Recovery of Function; Research; Role; Site; Stroke; System; Testing; Therapeutic Agents; Time; Transgenic Organisms; Traumatic Brain Injury; Traumatic Brain Injury recovery; Veterans; anandamide; antagonist; arm; bone circulation; brain repair; chemokine; controlled cortical impact; cytokine; design; endogenous cannabinoid system; frontal lobe; glial cell-line derived neurotrophic factor; improved; inhibitor; military veteran; monocyte; mouse model; neurogenesis; neuroinflammation; neurotrophic factor; programs; receptor; receptor expression; recruit; regenerative; relating to nervous system; repaired; response; stem; stem cells; translational goal; treatment guidelines; water maze

Background: G-CSF has been identified as a potential therapeutic agent for TBI. Administration of G- CSF soon after injury results in proliferation and release into the circulation of bone marrow-derived cells (BMDC). Monocytes from blood are recruited into the brain to the site of the lesion where they differentiate into microglia. These, in turn generate a number of neurotrophic factors and cytokines involved in repair and regenerative processes. Numerous studies indicate that TBI impacts the endogenous cannabinoid system (eCBs), altering expression of eCB receptors CB1 and CB2, and changing levels of the endocannabinoids anandamide (N-arachidonoyl-ethanolamine; AEA) and 2-AG (2- arachidonoylglycerol). Moreover, administration of cannabinoid agents have been shown to enhance recovery from TBI mediated by anti-inflammatory cytokines and regenerative processes that parallel those triggered by G-CSF. Specific Aims of this research program are designed to test the hypotheses a) that G-CSF interacts with the endocannabinoid system (eCS) to promote brain repair and b) that blockade of CB1 and/or CB2 receptors will diminish or enhance the brain’s repair response to TBI. Aim 1: To investigate the effects of G-CSF on the expression of CB1 and CB2 and their natural ligands in mouse brain (cortex, striatum and hippocampus) following TBI. Dependent variables: CB1 and CB2 receptor expression (mRNA and protein), levels of the eCB ligands AEA and 2-AG, extent of apoptosis, microgliosis, astrocytosis, neuro-inflammation, levels of neurotrophic factors (BDNF, GDNF); changes in hippocampal neurogenesis. Aim 2a: To determine if stimulation of the brain endocannabinoid system with a) an inhibitor of fatty acid amide (FAAH) to increase levels of AEA and 2AG, or b) administration of selective CB1 and CB2 receptor antagonists will potentiate (or diminish) the known beneficial effects of G- CSF on brain repair and recovery. Dependent variables are the same as in Aim 1, with the additional parameter of recovery of performance in the radial arm water maze (RAWM). Aim 2b: To determine the extent to which the CB1-R or the CB2-R is responsible for enhanced recovery from TBI, the CB1-R and CB2-R knockout mice will be studied using the protocol detailed in Aim1. Dependent variables: CB1 and CB2 receptor expression (mRNA and protein), levels of AEA, 2AG in cortex, striatum and hippocampus; extent of microgliosis and astrocytosis in these 3 brain regions; levels of BDNF and GDNF, hippocampal neurogenesis and recovery of performance in RAWM. Aim 3: To determine the role of the CB2 receptor in the mobilization of BMDC in mediating the beneficial effects of G-CSF, we will administer a CB2 antagonist (or utilize CB2-R knockout mice) to animals treated with G-CSF or vehicle. Tracking of BMDC will utilize chimeric mice that have had bone marrow transplants from transgenic “green mice”. Dependent variables: percentage of total circulating white blood cells that co-express GFP and markers of monocytes, quantitative estimates of the distribution GFP+ cells in the lesioned and contralateral hemisphere, proportion of GFP+ cells that differentiate into microglial phenotypes (M1, M2), profile of cytokines/chemokines, hippocampal neurogenesis, and performance in RAWM. Aim 4: Effects of delayed treatment (1, 2, 4 wks after “severe” TBI) with G-CSF on behavioral recovery and repair mediated by eCBs. Dependent variables: are the same as in Aim 1, with the additional parameter of rate of recovery of performance in RAWM. Impact on the Field of Brain Repair: The consequences of TBI, stroke and neurodegenerative diseases are highly prevalent in the veteran population. There is a need for effective, and safe therapies for these disorders. The optimal timing of G-CSF therapy after TBI, with or without cannabinoid agonists or antagonists as determined from this project, will be helpful in developing guidelines for treatment of humans who have suffered brain injury.

背景：G-CSF已被确定为TBI的潜在治疗剂。 g- 受伤后不久，CSF导致增殖并释放到骨髓衍生细胞的循环中 （BMDC）。血液中的单核细胞被招募到大脑到病变的部位 分化为小胶质细胞。这些反过来产生许多神经营养因素和细胞因子 参与维修和再生过程。大量研究表明TBI影响 内源性大麻素系统（ECB），改变了欧洲央行受体CB1和CB2的表达，以及 内源性大麻素的水平变化（N-芳基烯酰基 - 乙醇胺； AEA）和2-AG（2- 芳基烯丙基甘油）。此外，已经证明了大麻素剂的给药可增强 从抗炎细胞因子和再生过程中介导的TBI恢复，这些过程平行 由G-CSF触发。该研究计划的具体目的旨在测试假设a） G-CSF与内源性大麻素系统（EC）相互作用，以促进脑修复和b） CB1和/或CB2受体将减少或增强大脑对TBI的修复反应。目标1：到 研究G-CSF对CB1和CB2表达及其天然配体在小鼠中的影响 TBI之后，大脑（皮层，纹状体和海马）。因变量：CB1和CB2受体 表达（mRNA和蛋白质），欧洲央行配体AEA的水平和2-ag，凋亡的程度， 小胶质细胞增多，星形细胞增多症，神经炎症，神经营养因子水平（BDNF，GDNF）；变更 海马神经发生。 AIM 2A：确定是否刺激脑内源性大麻素系统 a）一种脂肪酸酰胺抑制剂（FAAH）增加AEA和2AG的水平，或b）给药 选择性CB1和CB2受体拮抗剂将潜在（或减少）G-的已知有益作用 CSF关于大脑修复和恢复。因变量与AIM 1相同，附加 径向臂水迷宫（RAWM）中性能恢复的参数。目标2b：确定 CB1-R或CB2-R负责从TBI，CB1-R和 CB2-R基因敲除小鼠将使用AIM1中详细介绍的协议进行研究。因变量：CB1和 CB2受体表达（mRNA和蛋白质），AEA水平，2ag皮层，纹状体和海马； 这三个大脑区域的小胶质细胞增多症和星形细胞增多症的程度； BDNF和GDNF的水平，海马 RAWM中的神经发生和性能的恢复。目标3：确定CB2受体的作用 在动员BMDC在介导G-CSF的有益作用中，我们将管理CB2 拮抗剂（或利用CB2-R基因敲除小鼠）对用G-CSF或媒介物治疗的动物。跟踪BMDC 将利用具有转基因“绿色小鼠”的骨髓移植的嵌合小鼠。依赖 变量：共表达GFP和标记的总循环白细胞的百分比 单核细胞，病变和对侧分布GFP+细胞的定量估计值 半球，分化为小胶质表型（M1，M2）的GFP+细胞的比例， RAWM中的细胞因子/趋化因子，海马神经发生和性能。目标4：效果 延迟治疗（“严重” TBI之后的1、2、4周），G-CSF进行行为恢复和修复介导的 由ECB。因变量：与AIM 1相同，速率的附加参数 RAWM的性能恢复。对大脑修复领域的影响：TBI的后果， 在退伍军人人群中，中风和神经退行性疾病非常普遍。有需要 为这些疾病有效且安全的疗法。 TBI后G-CSF治疗的最佳时机， 或没有该项目确定的大麻素激动剂或对手，将有帮助 制定治疗脑损伤的人的准则。