Mechanisms of Transmigration of HIV Infected Cells Across the Blood Brain Barrier

HIV感染细胞穿越血脑屏障的迁移机制

• 批准号: 7321940
• 负责人: TONI KAY ROBERTS
• 金额: $ 4.6万
• 依托单位: ALBERT EINSTEIN COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-07-01 至 2011-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7321940
• 关键词: AIDS Dementia Complex; Adaptor Signaling Protein; Address; Antibodies; Autopsy; Autoradiography; Binding; Biological Assay; Blood - brain barrier anatomy; Brain; CCL2 gene; Candidate Disease Gene; Cell Adhesion; Cell Adhesion Molecules; Cell Communication; Cell Line; Cell surface; Cells; Chemotactic Factors; Clinical; Coculture Techniques; Collection; Complex; Consequences of HIV; Cytoskeleton; Data; Dementia; Disruption; Elements; Encephalitis; Endothelial Cells; Environment; Exposure to; Gene Expression; Genes; Genetic; Goals; HIV; HIV Infections; HIV encephalitis; HIV-1; Highly Active Antiretroviral Therapy; Human; Image; Immunoprecipitation; Impaired cognition; Impairment; Incubated; Individual; Infection; Infiltration; Intercellular Junctions; Laboratories; Leukocytes; Life; Maintenance; Measures; Mediating; Microarray Analysis; Microglia; Modeling; Molecular; Monkeys; Neurocognitive; Neuropathogenesis; Patients; Phosphorylation; Phosphotransferases; Population; Post-Translational Protein Processing; Process; Property; Proteins; Radioactivity; Radiolabeled; Relative (related person); Research; Reverse Transcriptase Polymerase Chain Reaction; SIV; Scaffolding Protein; Source; Specimen; Stimulus; Tight Junctions; Tissue Stains; Transcriptional Activation; Tyrosine; Up-Regulation; Urticaria; Viral; Viral Load result; Virus; Western Blotting; Work; cell motility; cell type; chemokine; interest; intracellular protein transport; macrophage; migration; monocyte; neuroinflammation; phosphatase inhibitor; prevent; protein expression; protein localization location; radiotracer; response; success

DESCRIPTION (provided by applicant): With the success of HAART, HIV infected individuals are living longer and HIV mediated damage to the CNS, including HIV Associated Dementia (HAD) and other forms of neurocognitive impairment, are increasingly presenting as a devastating consequence of HIV infection. Although a late clinical finding, compromise of the blood brain barrier (BBB) and infiltration into the CNS parenchyma by HIV infected monocytes have been shown to occur early in infection. In fact, cognitive impairment correlates more directly with neuroinflammation than with the presence of virus within the CNS. CCL2, the predominate monocyte chemoattractant, has been shown to be substantially elevated in patients with HAD and HIV Encephalitis. Thus, understanding the mechanisms of HIV and CCL2 mediated monocyte transmigration into the CNS and concomitant disruption of the BBB will enable the identification of potential targets of therapy, with the ultimate goal of preventing HIV related neurocognitive decline. We hypothesize that HIV infection of monocytes alters their expression of CCR2, cell adhesion molecules, and adherens and tight junction proteins, and that transmigration of these infected monocytes in response to CCL2 in the CNS results in aberrant monocyte-BBB endothelial cell interactions, promoting enhanced migration and BBB disruption. The goal of this research is to define the molecular interactions responsible for the enhanced diapedesis by HIV infected monocytes into the CNS in response to the CCL2 chemokine and the subsequent BBB breakdown. The specific goals of this research are: (1) to identify differentially expressed genes or changes to protein localization in monocytes as a result of HIV infection and CCL2 chemotactic stimulation that may cofitribute to their enhanced diapedesis using microarray, RT-PCR, Western blot, and confocal imaging; (2) to characterize the effects of CCL2 on brain microvascular endothelial cell junction proteins that may potentiate monocyte diapedesis and disrupt BBB integrity using microarray, RT-PCR, Western blot, confocal imaging, and coimmunoprecipitation (ColP) studies; (3) to characterize the dynamic interactions between junctional proteins, adaptor proteins, and the cytoskeleton in HIV infected monocytes and in human brain microvascular endothelial cells that contribute to the enhanced transmigration properties of monocytes and disruption of BBB integrity in response to CCL2 using radiolabeling, ColP, Western blot, autoradiography, confocal imaging, and tissue staining; and (4) to determine the phosphorylation changes in cell juction proteins during transmigration of HIV infected monocytes across the BBB in response to CCL2 using radiolabeling, immunoprecipitation, autoradiography, and kinase and phosphatase inhibitors.

描述（由申请人提供）：随着HAART的成功，HIV感染的个体的寿命更长，HIV对中枢神经系统的损害，包括HIV相关的痴呆症（HAT）和其他形式的神经认知障碍，越来越多地表现为HIV感染的毁灭性后果。尽管是临床后期的发现，但血液脑屏障（BBB）和HIV感染的单核细胞的血液屏障（BBB）和渗透到CNS实质中，已证明在感染的早期发生。实际上，认知障碍与神经炎症更直接相关，而不是与中枢神经系统内的病毒存在。 CCL2是主要的单核细胞趋化剂，已证明在HAT和HIV脑炎患者中显着升高。因此，了解HIV和CCL2的机制介导的单核细胞转移到中枢神经系统中，并伴随着BBB的破坏，将使能够鉴定出潜在的治疗靶标，其最终目标是防止HIV相关的神经认知下降。我们假设单核细胞的HIV感染改变了它们对CCR2的表达，细胞粘附分子，粘附剂和紧密连接蛋白的表达，并且这些感染感染的单核细胞在CNS中响应CCL2的迁移导致异常的单核细胞单细胞BB BB胚胎相互作用，从而增强了CCL2。这项研究的目的是定义导致HIV感染单核细胞增强的分子相互作用，以响应CCL2趋化因子和随后的BBB分解，以响应CCL2趋化因子响应CNS。这项研究的具体目标是：（1）由于HIV感染和CCL2趋化性刺激，鉴定单核细胞中蛋白质定位的差异基因或变化，可以使用微阵列，RT-PCR，Western Blot和共焦成像来辅助其增强的尿症。 （2）表征CCL2对使用微阵列，RT-PCR，Western Blot，Condocal Imaging和copomunoprecipitation（COLP）的研究（COLP）研究（COLP）研究（COLP）研究的CCL2对脑微血管内皮细胞连接蛋白的影响，并破坏BBB完整性。 （3）表征连接蛋白，衔接蛋白和HIV感染的细胞骨架之间的动态相互作用，以及人类脑脑微血管内皮细胞中的动态相互作用，这些细胞有助于使用radiolabeling，ccli ccl2构造，从而有助于单核和对BBB完整性响应的单核和破坏的迁移，并促进迁移的迁移特性。和组织染色； （4）使用放射性标记，免疫沉淀，放射自显影和激酶和磷酸酶抑制剂，在艾滋病毒感染的单核细胞跨BBB的迁移过程中，细胞Juction蛋白的磷酸化变化。