Blood-Brain Barrier Ion Transport in Cerebral Ischemia

脑缺血中的血脑屏障离子转运

基本信息

批准号：
9059450
负责人：
Martha E O'Donnell
金额：
$ 41.68万
依托单位：
UNIVERSITY OF CALIFORNIA AT DAVIS
依托单位国家：
美国
项目类别：
财政年份：
2002
资助国家：
美国
起止时间：
2002-08-01 至 2018-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9059450
关键词：
Animals Attenuated Bicarbonates Biological Assay Blood Blood - brain barrier anatomy Blood Glucose Brain Brain Edema Brain Injuries Bumetanide Cause of Death Cells Cerebral Edema Cerebral Infarction Cerebral Ischemia Comorbidity Diabetes Mellitus Edema Endothelial Cells Evaluation Goals Grant Health Hour Human Hyperglycemia Hypoxia In Situ In Vitro Individual Infarction Intravenous Ion Transport Ions Ischemia Ischemic Stroke Magnetic Resonance Imaging Mediating Membrane Methods Middle Cerebral Artery Occlusion Modeling Nuclear Magnetic Resonance Outcome Patients Perfusion Population Process Proteins Rattus Sodium Sodium-Potassium-Chloride Symporters Streptozocin Stroke Swelling Testing Therapeutic Translating Vasopressins Water Western Blotting Work brain endothelial cell diabetic effective therapy in vivo inhibitor/antagonist intravenous administration research study response targeted treatment therapeutic target uptake

项目摘要

DESCRIPTION (provided by applicant): The long term goal of this project is to identify blood-brain barrier (BBB) ion transporters that mediate ischemia-induced brain edema. During the early hours of ischemic stroke, edema forms in the presence of an intact BBB by a process involving BBB transport of Na and Cl from blood into brain. Our previous studies have shown that Na-K-Cl cotransport (NKCC) and Na/H exchange (NHE), present in the luminal BBB membrane, are stimulated by ischemic factors, including hypoxia, aglycemia and vasopressin (AVP) and that inhibiting BBB NKCC and/or NHE activity reduces brain edema in the rat middle cerebral artery occlusion (MCAO) model of stroke. In preliminary studies we have now found evidence that a BBB Na-HCO3 cotransporter (NBC) constitutes a 3rd prominent BBB Na transporter that is also stimulated during ischemia to substantially participate in cerebral edema formation. This gives us an important additional therapeutic target for reducing the highly injurious early edema formation in ischemic stroke. While our studies indicate that NKCC, NHE and NBC appear to be promising therapeutic targets for reducing edema in otherwise healthy stroke patients, it must be recognized that hyperglycemia, a common co-morbidity in stroke present in ~30% of patients, causes greater cerebral edema and worsened outcome. The underlying mechanisms for this are poorly understood. However, in recent studies we have found that hyperglycemia increases expression and activity of BBB NKCC, NHE and NBC and augments their stimulation by ischemic factors. Further, we have also found evidence that the hyperglycemia exacerbation of brain edema in MCAO is reduced by inhibition of BBB NKCC, NHE and NBC. Our hypothesis is that BBB NKCC, NHE and/or NBC participate in the hyperglycemia-induced augmentation of cerebral edema formation and constitute effective therapeutic targets in hyperglycemic stroke. The first aim is to determine whether NBC protein is present at the luminal BBB membrane and is stimulated by ischemic factors. We will use Western blot and immunoEM to evaluate BBB NBC protein in CMEC and in BBB in situ, and microspectrofluorometry to assess ischemic factor effects on CMEC NBC activity. The second aim is to determine whether hyperglycemic conditions increase expression and activity of BBB NKCC, NHE and/or NBC and augment stimulation of these Na transporters by ischemic factors. We will use Western blot, microspectrofluorometry and radioisotopic flux assays to test the effects of hyperglycemia on CMEC NKCC, NHE and NBC protein and activity. We will also use immunoEM to evaluate NKCC, NHE and NBC in BBB in situ of hyperglycemic rats. The third aim is to determine whether inhibition of BBB NKCC, NHE and/or NBC attenuates ischemia-induced edema in hyperglycemic animals. Here, we will examine the effects of inhibiting BBB NKCC, NHE and NBC on ischemia-induced changes in rat brain Na and water, using nuclear magnetic resonance methods and rats with STZ-induced hyperglycemia. We will also assess the efficacy of NKCC, NHE and NBC inhibitors for reduction of edema when administered after the onset of ischemia.

描述（由申请人提供）：该项目的长期目标是识别介导缺血引起的脑水肿的血脑屏障（BBB）离子转运蛋白。在缺血性中风的早期，通过涉及Na和Cl从血液转移到大脑的BBB转运的过程中，在完整的BBB存在下形成了水肿。 Our previous studies have shown that Na-K-Cl cotransport (NKCC) and Na/H exchange (NHE), present in the luminal BBB membrane, are stimulated by ischemic factors, including hypoxia, aglycemia and vasopressin (AVP) and that inhibiting BBB NKCC and/or NHE activity reduces brain edema in the rat middle cerebral artery occlusion (MCAO) model of 中风。在初步研究中，我们现在发现了证据表明，BBB Na-HCO3共转运蛋白（NBC）构成了第三名突出的BBB NA转运蛋白，在缺血期间也刺激了大量参与大脑肿瘤的形成。这为我们提供了一个重要的额外治疗靶标，可以减少缺血性中风中高度伤害的早期水肿形成。虽然我们的研究表明，NKCC，NHE和NBC似乎是减少其他中风患者水肿的有前途的治疗靶标，但必须认识到，高血糖症是约30％的患者中风中常见的共生性，导致大脑水肿和恶化的结果。对此的基本机制知之甚少。但是，在最近的研究中，我们发现高血糖会增加BBB NKCC，NHE和NBC的表达和活性，并通过缺血因素增强其刺激。此外，我们还发现了证据表明，通过抑制BBB NKCC，NHE和NBC，MCAO中脑水肿的高血糖恶化会降低。我们的假设是BBB NKCC，NHE和/或NBC参与了高血糖引起的大脑水肿形成的增强，并构成了高血糖中风的有效治疗靶标。第一个目的是确定在腔内BBB膜上是否存在NBC蛋白，并被缺血因素刺激。我们将使用Western印迹和免疫系统评估CMEC和BBB原位中的BBB NBC蛋白，以及微光谱荧光测定法来评估缺血因素对CMEC NBC活性的影响。第二个目的是确定高血糖条件是否会增加BBB NKCC，NHE和/或NBC的表达和活性，以及通过缺血因素对这些NA转运蛋白的增强刺激。我们将使用Western印迹，微光谱荧光法和放射性药物通量测定法来测试高血糖对CMEC NKCC，NHE和NBC蛋白质以及活性的影响。我们还将使用免疫原子来评估BBB的NKCC，NHE和NBC的原位高血糖大鼠。第三个目的是确定抑制BBB NKCC，NHE和/或NBC衰减是否抑制高血糖动物中缺血诱导的水肿。在这里，我们将使用核磁共振方法和具有STZ诱导的高血糖的大鼠来研究抑制BBB NKCC，NHE和NBC对缺血诱导的大鼠脑Na和水变化的影响。我们还将评估NKCC，NHE和NBC抑制剂在缺血发作后施用时降低水肿的功效。