Neuronal Expression of Hemoglobin in Multiple Sclerosis Cortex

描述（由申请人提供）：神经元和轴突的损伤和损失与多发性硬化症（MS）的残疾相关，因此迫切需要神经保护治疗。在以前的研究中，我们发现线粒体基因和蛋白质的表达改变，参与呼吸导致功能障碍的线粒体活性在死后MS皮质。最近，我们发现血红蛋白在MS皮质中上调，并在MS脑中的神经元中定位血红蛋白表达。我们还发现从大鼠脑分离的神经元的原代培养物中有血红蛋白表达。这是一个令人惊讶的和新颖的发现，并为我们的假设，血红蛋白的表达可能参与神经元呼吸提供了支持。本研究的目的是了解血红蛋白在神经元中表达的功能及其对MS神经病理学的意义。在拟议的研究中，我们将通过原位杂交，RT-PCR，蛋白质印迹，共聚焦成像和电子显微镜尸检MS和对照组织的不同区域的MS脑中的血红蛋白表达的程度和定位。我们将调查机制，调节血红蛋白的表达在原代神经元文化，并将评估血红蛋白在神经元呼吸和氧化磷酸化的作用，通过执行高分辨率呼吸测定神经母细胞瘤细胞表达血红蛋白亚基从哺乳动物表达载体，并从小鼠脑切片与促红细胞生成素，增加血红蛋白在大脑中的表达。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。

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.