Neuronal Expression of Hemoglobin in Multiple Sclerosis Cortex

描述(申请人提供)：神经元和轴突的损伤和丢失与多发性硬化症(MS)的残疾相关，因此迫切需要神经保护性治疗。在以前的研究中，我们已经发现参与呼吸的线粒体基因和蛋白的表达改变，导致死后MS皮质中线粒体活性异常。最近，我们发现血红蛋白在MS皮质中表达上调，并定位于MS脑内的神经元。我们还发现从大鼠脑中分离的神经元的原代培养中有血红蛋白的表达。这是一个令人惊讶的新发现，并为我们的假设提供了支持，即血红蛋白表达可能涉及神经元呼吸。本研究的目的是了解血红蛋白在MS神经元中的表达功能及其在神经病理学中的意义。在本研究中，我们将通过原位杂交、RT-PCR、Western blotting、共聚焦成像和电子显微镜观察MS和对照组织的血红蛋白在MS脑不同区域的表达范围和定位。我们将通过对哺乳动物表达载体中表达血红蛋白亚单位的神经母细胞瘤细胞和经促红细胞生成素处理的小鼠脑片进行高分辨率呼吸测量，来研究在原代神经元培养中调节血红蛋白表达的机制，并通过对表达血红蛋白亚单位的神经母细胞瘤细胞进行高分辨率呼吸测量来评估血红蛋白在神经元呼吸和氧化磷酸化中的作用。我们还将研究血红蛋白作为自由基清除剂在大脑中发挥神经保护作用的可能性。 公共卫生相关性：多发性硬化症(MS)是一种衰弱的神经系统疾病，患者在壮年时期发作。我们之前已经证明，线粒体参与了这种疾病。这项拟议的研究与公共健康相关，因为它将有助于更好地理解导致多发性硬化症线粒体活动改变的机制，并可能确定新的神经保护性治疗靶点。

DESCRIPTION (provided by applicant): Damage to and loss of neurons and axons is correlated with disability in multiple sclerosis (MS), thus neuroprotective therapies are urgently needed. In previous studies we have found altered expression of mitochondrial genes and proteins involved in respiration resulting in dysfunctional mitochondrial activity in postmortem MS cortex. Recently, we have shown that hemoglobin is upregulated in MS cortex and have localized hemoglobin expression to neurons in the MS brain. We have also found hemoglobin expression in primary cultures of neurons isolated from rat brain. This is a surprising and novel finding and provides support for our hypothesis that hemoglobin expression may be involved in neuronal respiration. The goal of this study is to understand the function of hemoglobin expression in neurons and its significance to neuropathology in MS. In the proposed study we will investigate the extent and localization of hemoglobin expression in different regions of the MS brain by in situ hybridization, RT-PCR, western blotting, confocal imaging, and electron microscopy of postmortem MS and control tissue. We will investigate the mechanisms which regulate hemoglobin expression in primary neuronal cultures and will assess the role of hemoglobin in neuronal respiration and oxidative phosphorylation by performing high resolution respirometry on neuroblastoma cells expressing hemoglobin subunits from a mammalian expression vector, and on brain slices from mice treated with erythropoietin which increases hemoglobin expression in the brain. We will also investigate the possibility that hemoglobin serves a neuroprotective role in the brain as a scavenger of free radicals. PUBLIC HEALTH RELEVANCE: Multiple sclerosis (MS) is a debilitating neurological disease which strikes patients in the prime of their life. We have previously shown that there is mitochondrial involvement in this disease. The proposed study is relevant to public health in that it will lead to a better understanding of mechanisms leading to altered mitochondrial activity in MS and will potentially identify new neuroprotective therapeutic targets.