Neurovascular Mechanisns of Brain Function and Disease

脑功能和疾病的神经血管机制

• 批准号: 7448456
• 负责人: Philip K Liu
• 金额: $ 19.08万
• 依托单位: MASSACHUSETTS GENERAL HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-06-16 至 2010-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7448456
• 关键词: A Mouse; Actins; Affect; Animals; Applications Grants; Autopsy; Bilateral; Binding; Biological Assay; Biological Markers; Biopsy; Blood Circulation; Blood Platelets; Blood Vessels; Brain; Brain Edema; Brain Injuries; Cardiac Edema; Cell membrane; Cells; Cerebral Edema; Cerebral Ischemia; Cerebrum; Condition; Contrast Media; Contrast Sensitivity; Control Groups; DNA; DNA Probes; Detection; Development; Disease; Doctor of Philosophy; Dose; Edema; Elevation; Endothelium; Evaluation; Extravasation; Functional disorder; Future; Gelatinase B; Gene Expression; Genes; Genetic Transcription; Goals; Heart Arrest; Histology; Human; Image; Imaging technology; In Situ; Infusion procedures; Injury; Intraperitoneal Injections; Invasive; Investigation; Knock-out; Label; Life; Link; Magnetic Resonance; Magnetic Resonance Imaging; Maps; Matrix Metalloproteinases; Messenger RNA; Metalloproteases; Methods; Modeling; Modification; Mus; Myocardial Infarction; Nervous System Trauma; Neurological outcome; Oligonucleotides; Operative Surgical Procedures; Organ; Oxidative Stress; Physiological reperfusion; Polymerase Chain Reaction; RNA; Radioactive; Randomized; Recovery; Reperfusion Therapy; Reporter; Reporting; Resolution; Reverse Transcription; Running; Sampling; Signal Transduction; Simulate; Stress; Stroke; Techniques; Testing; Therapeutic Agents; Tissue Sample; Tissues; Toxic effect; Transcript; Transcriptional Activation; Up-Regulation; Validation; Work; beta Actin; brain cell; design; dextran-coated SPIO; gene correction; gene therapy; human subject; improved; iron oxide; mRNA Expression; male; nanoparticle; nervous system disorder; neurogenesis; neutrophil; novel; phosphorothioate; preclinical study; repaired; restoration; uptake; A Mouse; Actins; Affect; Animals; Applications Grants; Autopsy; Bilateral; Binding; Biological Assay; Biological Markers; Biopsy; Blood Circulation; Blood Platelets; Blood Vessels; Brain; Brain Edema; Brain Injuries; Cardiac Edema; Cell membrane; Cells; Cerebral Edema; Cerebral Ischemia; Cerebrum; Condition; Contrast Media; Contrast Sensitivity; Control Groups; DNA; DNA Probes; Detection; Development; Disease; Doctor of Philosophy; Dose; Edema; Elevation; Endothelium; Evaluation; Extravasation; Functional disorder; Future; Gelatinase B; Gene Expression; Genes; Genetic Transcription; Goals; Heart Arrest; Histology; Human; Image; Imaging technology; In Situ; Infusion procedures; Injury; Intraperitoneal Injections; Invasive; Investigation; Knock-out; Label; Life; Link; Magnetic Resonance; Magnetic Resonance Imaging; Maps; Matrix Metalloproteinases; Messenger RNA; Metalloproteases; Methods; Modeling; Modification; Mus; Myocardial Infarction; Nervous System Trauma; Neurological outcome; Oligonucleotides; Operative Surgical Procedures; Organ; Oxidative Stress; Physiological reperfusion; Polymerase Chain Reaction; RNA; Radioactive; Randomized; Recovery; Reperfusion Therapy; Reporter; Reporting; Resolution; Reverse Transcription; Running; Sampling; Signal Transduction; Simulate; Stress; Stroke; Techniques; Testing; Therapeutic Agents; Tissue Sample; Tissues; Toxic effect; Transcript; Transcriptional Activation; Up-Regulation; Validation; Work; beta Actin; brain cell; design; dextran-coated SPIO; gene correction; gene therapy; human subject; improved; iron oxide; mRNA Expression; male; nanoparticle; nervous system disorder; neurogenesis; neutrophil; novel; phosphorothioate; preclinical study; repaired; restoration; uptake

DESCRIPTION (provided by applicant): Altered expression of endogenous genes in the brain often accompanies neurological disorders. Genes or cells have been used to correct gene trancription so that brain can repair itself. Currently, most detection techniques of gene transcription require biopsy or autopsy samples. Invasive surgical procedures for removing tissue samples severely limit the benefits, especially to the cells we try to cure. Our goals are to develop methods to image and quantitatively compare endogenous gene expression at the transcript level using magnetic resonance (MR) in live animal or human subjects. We made a novel MR probe using short phosphorothioate-modified oligodeoxynucleotides (sODN) with SuperParamagnetic Iron Oxide Nanoparticles (SPION, an MR T2 agent). The work outlined in this application investigates the utility of this contrast probe as an biomarker for mRNA transcripts in live animals. We designed three probes for MRI: two are with sequence complementary to matrix metalloprotease-9 (sODN-mmp9) or beta- actin (sODN-bactin) mRNA and a randomized s-ODN (sODN-Ran) with no sequence complementary to mRNA. The SPION-Ran probes will serve as controls. We will evaluate conditions that allow optimal imaging endogenous gene expression using MR.The specific anims are to demonstrate: Aim 1: Maximize MR Contrast Enhancement using SPION-bactin in High-resolution MRI in Mouse Brains. The hypothesis is that sODN-linked SPION will be retained by brain cells for detection using MRI in live animals, and for validation using histology (iron oxide) and binding assay (SPION-bactin) in postmortem samples. We select beta-actin mRNA as a target because beta-actin mRNA is constant and is inert to stress. To support this hypothesis, we will: (a) select an optimal SPION-retention using MRI in live animals after infusion with various doses of SPION-bactin; We will demonstrate that the uptake of SPION in the brain is sODN-linkage dependent; (b) demonstrate the presence of intracellular iron oxide after infusion of SPION-bactin at the optimal dose, and (c) show that the internalized SPION-bactin binds to its target mRNA. Aim 2: Retention of cerebral SPION-mmp9 in live C57black6 mice predicts brain edema after stroke inducted by 60 or 90 minutes bilateral carotid occlusion. The hypothesis is that cerebral mmp-9 mRNA transcript reports MMP-9 expression. To support this hypothesis, we will demonstrate:(a) elevation of mmp-9 mRNA expression is positively correlated with cerebral edema after stroke, (b) retention of SPION-mmp9 is higher in the stroke-treated than in the sham-operated mice. Specifically, we will identify the hotspot of SPION-mmp9 retention in stroke-treated animals using subtraction of R2* map between stroke-treated and sham-operated animals. In addition, we will compare the hotspots of SPION-mmp9 retention to stroke-induced damage in the brain of wild type and mmp-9 knockout strains. R21NS05755601-A1 PI: Liu, Philip K. PhD Project Narratives: Brain edema is one neurological disorder that complicates the recovery of several diseases in the humans. Cardiac arrest in humans can induce brain edema despite restoration of circulation using various means (Roine et al. 1993; Fujioka et al. 1994; Xiao 2002); brain damage is predictive of a poor neurological outcome. Currently few models simulate brain edema from cardiac arrest for us to test therapeutic agents that can reduce neurological damage. To improve our understanding of cerebral injury caused by heart attack, we proposed this grant application a global cerebral ischemia-reperfusion model to investigate the pathophysiology in the brain of male C57black/6 mice simulating cardiac arrest (Liu et al. 1996). Our long-term goal is to establish new methods using MRI contrast agent to gain understanding on the pathological change in gene expression in live subjects, which can be saved for further evaluation on efficacy and toxicity of the therapy. The immediate goal is to show our probe can report the elevation of gene transcripts after cerebral ischemia.

描述（申请人提供）：内源基因在大脑中的表达改变通常伴随神经系统疾病。基因或细胞已用于纠正基因trancriptim，以便大脑可以自身修复。当前，大多数基因转录技术都需要活检或尸检样品。去除组织样品的侵入性手术程序严重限制了益处，尤其是我们试图治愈的细胞。我们的目标是开发使用活动物或人类受试者中的磁共振（MR）在转录水平上图像和定量比较内源基因表达的方法。我们使用短磷酸硫酸盐修饰的寡脱氧核苷酸（SODN）与超帕磁铁氧化铁纳米颗粒（SPION，MR T2剂）进行了新的MR探针。该应用程序中概述的工作调查了该对比探针作为活动物中mRNA转录物的生物标志物的实用性。我们设计了三个用于MRI的探针：两个探针与基质金属蛋白酶9（SODN-MMP9）或β-肌动蛋白（SODN-BACTIN）mRNA和随机的S-ODN（SODN-RAN）互补，没有序列互补。 SPION-RAN探针将用作对照。我们将评估使用特定动画的最佳成像内源基因表达的条件：AIM 1：在小鼠大脑中使用SPION-BACTIN使用SPION-BACTIN在小鼠大脑中最大化MR对比度增强。假设是，将SODN连接的SPION保留在活动物中使用MRI检测，并使用组织学（铁氧化铁）和结合分析（SPION-BACTIN（SPION-BACTIN））在后验尸样品中进行验证。我们选择β-肌动蛋白mRNA作为靶标，因为β-肌动蛋白mRNA是恒定的，并且是压力的。为了支持这一假设，我们将：（a）在输注各种剂量的SPION-BACTIN后，在活体动物中选择最佳的SPION-SPION-使用MRI；我们将证明大脑中SPION的摄取取决于sodn-linkage。 （b）证明在最佳剂量下输注Spion-Bactin后，存在细胞内氧化铁，并且（c）表明内部化的SPION-BACTIN与其靶mRNA结合。 AIM 2：在活的C57BLACK6小鼠中保留脑SPION-MMP9，可预测脑湿气在感应60或90分钟双侧颈动脉闭塞后的中风后脑水肿。假设是脑MMP-9 mRNA转录本报告MMP-9表达。为了支持这一假设，我们将证明：（a）MMP-9 mRNA表达的升高与中风后的脑水肿呈正相关，（b）中风治疗中SPION-MMP9的保留比在假手术小鼠中更高。具体而言，我们将使用中风处理和假手术动物之间的R2*缩写在中风处理的动物中确定SPION-MMP9保留的热点。此外，我们将比较SPION-MMP9保留的热点与野生型和MMP-9敲除菌株的中风诱导的损伤。 R21NS05755601-A1 PI：Liu，Philip K. Phd项目叙事：脑水肿是一种神经系统疾病，使人类中几种疾病的恢复复杂化。尽管使用各种手段恢复了循环，但人类心脏骤停仍可以诱导大脑浮肿（Roine等，1993； Fujioka等，1994； Xiao 2002）。脑损伤可以预测神经系统差的结果。目前，很少有模型模拟我们从心脏骤停的大脑水肿来测试可以减少神经系统损害的治疗剂。为了提高我们对由心脏病发作引起的脑损伤的理解，我们提出了这项赠款应用一种全球脑缺血 - 再灌注模型，以研究雄性C57Black/6小鼠模拟心脏骤停大脑中的病理生理学（Liu等人，1996年）。我们的长期目标是使用MRI对比剂建立新方法，以了解现场受试者基因表达的病理变化，这些方法可以保存，以进一步评估治疗的疗效和毒性。直接目标是表明我们的探测器可以报告脑缺血后基因转录物的升高。