Mechanisms Underlying the Suppression of Transcytosis at the Blood Brain Barrier

抑制血脑屏障转胞吞作用的机制

• 批准号: 8830634
• 负责人: Benjamin Joseph Andreone
• 金额: $ 3.48万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-09-30 至 2017-09-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8830634
• 关键词: Ablation; Affect; Alzheimer' s Disease; Amyotrophic Lateral Sclerosis; Biochemical; Biological Assay; Blood - brain barrier anatomy; Blood Vessels; Blood capillaries; Brain; Brain Neoplasms; Caveolae; Cell Culture Techniques; Cell Line; Cell membrane; Cells; Central Nervous System Diseases; Cerebrum; Chemicals; Clathrin; Clathrin-Coated Vesicles; Co-Immunoprecipitations; Complex; Cytoplasm; Data; Drug Delivery Systems; Endothelial Cells; Endothelium; Ensure; Environment; Event; Extravasation; Family; Functional disorder; Genetic; Goals; Homeostasis; In Vitro; Ions; Label; Mediating; Molecular; Multiple Sclerosis; Mus; Mutate; Nerve Degeneration; Neuraxis; Neurodegenerative Disorders; Nutrient; Pathway interactions; Peripheral; Permeability; Pharmaceutical Preparations; Phenotype; Physiological; Process; Property; Proteins; Research; Solid; Structure; Synaptic Transmission; System; Testing; Therapeutic; Therapeutic Agents; Tight Junctions; Tracer; Vesicle; Work; base; capillary; density; immunoreactivity; in vitro Assay; in vivo; insight; member; monolayer; mutant; nervous system disorder; novel; overexpression; polarized cell; prevent; public health relevance; repaired; research study; seal; tool; trafficking; transcytosis

DESCRIPTION (provided by applicant): The tightly controlled chemical environment of the CNS required for proper synaptic transmission is maintained by the 'blood brain barrier' (BBB), which is composed of highly specialized blood vessels whose endothelial cells act as a physiological barrier to seal the CNS and control substance influx and efflux. Two unique features of CNS endothelial cells determine BBB integrity, namely specialized tight junctions between the endothelial cells lining the CNS capillaries and extremely low rates of vesicular trafficking between the luminal and abluminal plasma membranes, a process termed transcytosis. A solid understanding of how BBB function is regulated to ensure brain homeostasis is missing in the field, as there are little to no molecular insights into how CNS endothelial cells acquire and maintain their specialized properties. Furthermore, the most promising strategies for delivering therapeutic agents across the BBB involve the manipulation of transcytotic pathways, an avenue of research which requires mechanistic understanding of how this process is normally regulated. Previous work in our lab has identified MFSD2A as the first molecule to maintain BBB integrity specifically by suppressing transcytosis. MFSD2A is exclusively expressed in CNS endothelial cells, and genetic ablation of Mfsd2a results in both extravasation of exogenous tracer from the vessel lumen to brain parenchyma and an increased density of vesicles within CNS endothelial cells, while intercellular tight junctions remain normal The goal of this study is to understand the molecular mechanism whereby MFSD2A suppresses transcytosis. My preliminary data show that MFSD2A is localized to the luminal plasma membrane of CNS endothelial cells and that loss of Mfsd2a results in increased immunoreactivity of Cav-1. Therefore, I hypothesize that MFSD2A acts as a suppressor of caveolae-mediated transcytosis in CNS endothelial cells, possibly by interacting with transcytotic machinery to inhibit caveolae dynamics at the luminal plasma membrane. To this end, I have developed a rigorous experimental plan with three aims: I will (1) use in vivo tools and an in vitro cell-based system I have developed to determine which specific transcytotic pathway is suppressed by MFSD2A in CNS endothelial cells, (2) perform structure-function experiments to dissect how structural domains of MFSD2A contribute to its function in suppressing transcytosis, and (3) test the hypothesis that MFSD2A interacts with well-known molecular determinants of caveolae-mediated transcytosis to elicit its function. I expect that these experiments will provide novel insight into how BBB function is regulated, leading to new pathways for BBB manipulation for therapeutic purposes.

描述（由申请人提供）：适当突触传播所需的中枢神经系统的紧密控制的化学环境由“血脑屏障”（BBB）维持，该环境由高度专业的血管组成，该血管的内皮细胞充当密封CNS和控制物质的生理障碍，并控制了CNS和控制物质的膨胀和溢出。中枢神经系统内皮细胞的两个独特特征决定了BBB完整性，即中枢神经系统毛细管内皮细胞之间的专业紧密连接，以及腔内和空白质膜之间的囊泡运输速率极低，这是一种称为转胞胞菌的过程。对BBB功能如何受到调节，以确保在现场缺少大脑稳态，因为几乎没有分子见解对CNS内皮细胞如何获得和维持其专业特性。此外，在BBB上提供治疗剂的最有前途的策略涉及对跨性神经途径的操纵，这是一种研究途径，这是一种需要机械理解该过程通常受到调节的理解。我们实验室中的先前工作已将MFSD2A确定为通过抑制转胞细胞增多而专门维持BBB完整性的第一个分子。 MFSD2A仅在中枢神经系统内皮细胞中表达，MFSD2A的遗传消融都导致外源示踪剂从血管腔到脑实质的外源示踪剂渗入，同进的囊泡密度增加，并且在CNS内的囊泡密度增加，而MOFTRISE的opriention otcression却保持了抑制的正常状态，这是分子间的分子，是分子间的分子，是分子的分子。转胞病。我的初步数据表明，MFSD2A位于中枢神经系统内皮细胞的腔质膜上，MFSD2A的丧失导致CAV-1的免疫反应性升高。因此，我假设MFSD2A充当CNS内皮细胞中小窝介导的跨胞菌病的抑制剂，这可能是通过与跨经细胞性机制相互作用以抑制腔质质膜膜上的小窝动力学的相互作用。 To this end, I have developed a rigorous experimental plan with three aims: I will (1) use in vivo tools and an in vitro cell-based system I have developed to determine which specific transcytotic pathway is suppressed by MFSD2A in CNS endothelial cells, (2) perform structure-function experiments to dissect how structural domains of MFSD2A contribute to its function in suppressing transcytosis, and (3) test the假设MFSD2A与小窝介导的跨介症的众所周知的分子决定因素相互作用，从而引起其功能。我希望这些实验将提供有关如何调节BBB功能的新见解，从而导致用于治疗目的的BBB操作的新途径。