Cell-specific expression of AAV-Nurr1 in a model of Parkinson's Disease

AAV-Nurr1 在帕金森病模型中的细胞特异性表达

• 批准号: 9121963
• 负责人: Sean Lee Hammond
• 金额: $ 3.49万
• 依托单位: COLORADO STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-04-01 至 2018-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9121963
• 关键词: 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine; Adverse effects; Affect; Animals; Astrocytes; Attenuated; Brain; Candidate Disease Gene; Cell Culture Techniques; Cells; Corpus striatum structure; Dependovirus; Disease; Disease Progression; Dopamine; Gene Delivery; Gene Expression; Gene Targeting; Genes; Genetic; Genetic Engineering; Genetic Transcription; Glial Fibrillary Acidic Protein; Health; Homeostasis; Image; Image Analysis; Immunotherapeutic agent; In Situ; In Vitro; Injection of therapeutic agent; Knowledge; Laboratories; Lead; Ligands; Luciferases; Maintenance; Midbrain structure; Modeling; Modification; Molecular; Mus; NR4A1 gene; Neonatal; Neurodegenerative Disorders; Neuroglia; Neurologic; Neurons; Nuclear; Nuclear Orphan Receptor; Orphan; Oxidative Stress; Parkinson Disease; Pathogenesis; Pharmaceutical Preparations; Pharmacodynamics; Phenotype; Phytochemical; Predisposition; Primary Cell Cultures; Probenecid; Production; Recombinant adeno-associated virus (rAAV); Reporter; Research; Role; Serotyping; Substantia nigra structure; Symptoms; Techniques; Testing; Therapeutic; Time; Transgenes; Viral Vector; adeno-associated viral vector; aged; base; cell type; design; dopaminergic neuron; glial activation; in vivo; innovation; mature animal; motor symptom; mouse model; neurochemistry; neuroinflammation; neuron loss; neuronal survival; neuroprotection; neurotoxic; neurotoxicity; neurotropic; novel; overexpression; pars compacta; promoter; protein aggregation; public health relevance; receptor; red fluorescent protein; research study; response; tool; transcription factor; transgene expression

 DESCRIPTION (provided by applicant): To date, there is no treatment to slow the progression of Parkinson's disease (PD). PD symptoms are caused by the slow degeneration of dopaminergic (DA) neurons in the ventral midbrain, which originate in the substantia nigra pars compacta (SNpc). DA neurons are characterized by specific genes necessary for dopamine production, which are regulated by orphan nuclear transcription factor Nurr1 (NR4A1). Decreased expression of Nurr1 causes loss of the dopaminergic phenotype in vivo and in vitro, thus is required for normal neurologic health and DA maintenance. Neuroinflammation triggered by glial cell activation has been proven to be a major causing factor of DA degeneration in PD. Interestingly, Nurr1 also provides a cell-specific role by constitutive suppression of neuroinflammatory gene expression in glial cells. This suggests Nurr1 could provide DA phenotypic support and neuroprotection to neurons of the SNpc if forced expression was maintained in astrocytes and neurons throughout PD progression. Nurr1 activation is also necessary for nuclear transcription, yet no endogenous ligands have been identified. However, we have demonstrated small lipophilic molecular compound, C-DIM12, to be a potent activator of Nurr1 and possess neuroprotection in a MPTP mouse model of PD. Challenges remain on genetically modifying specific cell types in the brain without off- target side effects. Yet, we hae tested specific non-pathogenic adeno-associated virus (AAV) vectors capable of transmitting a Nurr1 transgene into the SNpc and infecting both DA neurons and astrocytes. Thus, it is the Central Hypothesis of this proposal that selective overexpression of Nurr1 in DA neurons and astrocytes using AAV-based gene delivery will provide neuroprotection from MPTP-induced neurotoxicity and enhance the neuroprotective efficacy of C-DIM12 both in vitro and in vivo. The following Specific Aims will test this hypothesis: Specific Aim 1- Optimize the expression of AAV-Nurr1 in astrocytes and neurons and determine the effect of cell-specific Nurr1 expression against neurotoxic insult in vitro; Specific Aim 2- Determine the neuroprotective efficacy of cell specific AAV-Nurr1 overexpression in a mouse model of PD. Completion of these aims will increase our understanding of mechanisms by which Nurr1 regulates neuronal survival and glial activation in models of PD. Additionally, we anticipate these studies will expand our knowledge of candidate genes targeted for PD therapeutics.

 描述（由申请人提供）：迄今为止，尚无治疗方法可以减缓帕金森病（PD）的进展。 PD 症状是由腹侧中脑多巴胺能 (DA) 神经元缓慢变性引起的，这些神经元起源于黑质致密部 (SNpc)。 DA 神经元的特征是产生多巴胺所需的特定基因，这些基因受孤儿核转录因子 Nurr1 (NR4A1) 调节。 Nurr1 表达减少会导致体内和体外多巴胺能表型的丧失，因此是正常神经系统健康和 DA 维持所必需的。胶质细胞激活引发的神经炎症已被证明是 PD 中 DA 变性的主要原因。有趣的是，Nurr1 还通过组成性抑制神经胶质细胞中的神经炎症基因表达来发挥细胞特异性作用。这表明，如果在整个 PD 进展过程中星形胶质细胞和神经元中维持强制表达，Nurr1 可以为 SNpc 神经元提供 DA 表型支持和神经保护。 Nurr1 激活对于核转录也是必需的，但尚未鉴定出内源配体。然而，我们已经证明小亲脂性分子化合物 C-DIM12 是 Nurr1 的有效激活剂，并且在 PD 的 MPTP 小鼠模型中具有神经保护作用。在不产生脱靶副作用的情况下对大脑中的特定细胞类型进行基因改造仍然面临挑战。然而，我们已经测试了特定的非致病性腺相关病毒 (AAV) 载体，该载体能够将 Nurr1 转基因传递到 SNpc 中并感染 DA 神经元和星形胶质细胞。因此，该提案的中心假设是，使用基于 AAV 的基因递送在 DA 神经元和星形胶质细胞中选择性过度表达 Nurr1 将提供神经保护，免受 MPTP 诱导的神经毒性，并增强 C-DIM12 的体外和体内神经保护功效。以下具体目标将检验这一假设： 具体目标 1- 优化星形胶质细胞和神经元中 AAV-Nurr1 的表达，并确定细胞特异性 Nurr1 表达对体外神经毒性损伤的作用；具体目标 2 - 确定细胞特异性 AAV-Nurr1 过度表达在 PD 小鼠模型中的神经保护功效。完成这些目标将增加我们对 Nurr1 在 PD 模型中调节神经元存活和神经胶质激活的机制的理解。此外，我们预计这些研究将扩大我们对 PD 治疗候选基因的了解。