The Role of Microglia in Models of ALS

描述（由申请人提供）：肌萎缩性侧索硬化症（ALS）是一种快速进展和致命的神经肌肉疾病；大约10%的ALS患者有这种疾病的家族史。这些家族性ALS病例中有20-25%是由Cu2+/Zn2+超氧化物歧化酶基因（mSOD1）突变引起的。mSOD1在转基因小鼠中过表达时也会引起疾病。然而，有证据表明，除了运动神经元外，可能还需要其他细胞来产生ALS表型，即非细胞自主表型。首先，mSOD1在仅神经元和仅星形胶质细胞中的表达不会产生疾病；然而，mSOD1的表达水平和时间与原mSOD1G93A小鼠不同。其次，嵌合小鼠的数据表明，被正常胶质细胞包围的表达msod1的神经元似乎没有受到损伤，而被表达msod1的胶质细胞包围的正常神经元则出现异常。第三，病理存在于中枢神经系统的几种不同细胞类型，包括神经元、星形胶质细胞和小胶质细胞。为了确定mSOD1在小胶质细胞和免疫细胞中的表达对ALS的影响，我们选择了PU.1-/-小鼠。出生时，这些小鼠缺乏巨噬细胞、中性粒细胞、T细胞和B细胞，重要的是缺乏小胶质细胞；这类小鼠只能在骨髓移植后存活，产生供体来源的中枢神经小胶质细胞。具体而言，我们的目的是：特异性目的1：确定mSOD1在小胶质细胞和免疫细胞中的表达是否可以诱导als样疾病和/或启动亚临床病理。我们将PU.1 -/-小鼠与mSOD1-、正常人(h)SOD1和野生型骨髓进行移植。移植mSOD1骨髓的PU.1 -/-小鼠未表现出任何明显的运动神经元疾病迹象；尽管移植的小胶质细胞在形态上可能与对照小鼠的小胶质细胞不同。我们将检查这些小鼠的亚临床病理。特异性目的2：确定mSOD1在小胶质细胞和免疫细胞中的表达是否需要mSOD1小鼠als样疾病的启动和/或扩增。我们正在移植与PU.1 -/-小鼠杂交的mSOD1小鼠（mSOD1/ PU.1 -/-）。mSOD1 / PU。移植野生型骨髓的1 -/-小鼠表现出als样表型。有趣的是，这些小鼠的疾病持续时间可能长于mSOD1/PU的持续时间。1 -/-小鼠移植mSOD1骨髓。特异性目的3：明确小胶质细胞介导的运动神经元损伤的潜在机制。我们正在培养被激活的野生型小胶质细胞损伤的初级运动神经元，mSOD1初级小胶质细胞似乎更活跃，我们将评估它们损伤野生型和mSOD1运动神经元的能力。

DESCRIPTION (provided by applicant): Amyotrophic lateral sclerosis (ALS) is a rapidly progressive and fatal neuromuscular disease; approximately 10% of ALS patients have a family history of the disease. 20-25% of these familial ALS cases are caused by mutations in the Cu2+/Zn2+ superoxide dismutase gene (mSOD1). Overexpression of mSOD1 in transgenic mice also induces disease when ubiquitously expressed. However, evidence indicates that other cells, in addition to motor neurons, may be required to produce an ALS phenotype, i.e. non-cell autonomous. First, expression of mSOD1 in neurons-only and in astroglia-only did not produce disease; however, the level and timing of mSOD1 expression were not the same as the original mSOD1G93A mouse. Second, data from chimeric mice suggest that mSOD1-expressing neurons surrounded by normal glia appear uninjured, whereas normal neurons surrounded by mSOD-expressing glia appear abnormal. Third, pathology is present in several different cell types in the CNS, including neurons, astrocytes, and microglia. To determine the effects of mSOD1 expression in microglia and immune cells in ALS, we chose to use the PU.1-/- mouse. At birth, these mice lack macrophages, neutrophils, T and B cells, and importantly microglia; such mice survive only after bone marrow transplantation, resulting in donor-derived CNS microglia. Specifically, our aims are: Specific Aim 1: To determine if mSOD1 expression in microglia and immune cells alone can induce an ALS-like disease and/or initiate subclinical pathology. We are transplanting PU.1 -/- mice with mSOD1-, normal human (h)SOD1, and wildtype-derived bone marrow. PU.1 -/- mice transplanted with mSOD1 bone marrow did not appear to show any overt signs of motoneuron disease; even though the transplanted microglia may be morphologically different than microglia in control mice. We will examine these mice for subclinical pathology. Specific Aim 2: To determine if mSOD1 expression in microglia and immune cells is required for the initiation and/or amplification of an ALS-like disease in the mSOD1 mouse. We are transplanting the mSOD1 mice crossed with the PU.1 -/- mice (mSOD1/PU.I-/-). mSOD1/PU.1 -/- mice transplanted with wildtype bone marrow exhibit an ALS-like phenotype. Interestingly, the disease duration with these few mice may be longer than the duration of the mSOD1/PU.1 -/- mice transplanted with mSOD1 bone marrow. Specific Aim 3: To define potential mechanisms of microglia-mediated motor neuron injury. We are culturing primary motor neurons, which are injured by activated wildtype microglia, mSOD1 primary microglia appear more activated and we will assess their ability to injure wildtype and mSOD1 motor neurons.