Pre-Clinical Trial of MSC-Based Therapy for CNS Disease

描述(申请人提供)：这项建议的目的是在相关动物模型中评估成人干细胞注射到中枢神经系统治疗与溶酶体储存疾病相关的神经后遗症的安全性、可行性和有效性。具体地说，来自骨髓的成年干细胞，称为间充质干细胞(MSCs)，将使用MRI确定的立体定位坐标，单侧注射到婴儿恒河猴(Macaca Mulatta)的尾壳核中。在移植前和移植后长达一年的不同时期，每个移植受者的健康、发育、行为以及认知和运动技能将通过一系列适合年龄的测试进行评估，并与假手术动物或标准对照数据进行比较。此外，植入女性移植受者中枢神经系统的雄性MSCs的水平和解剖分布将使用我们开发的针对猕猴序列的实时聚合酶链式反应方法进行评估。Y染色体。为了确认实时PCR数据，植入的MSCs将通过免疫荧光染色或FISH在脑切片上可视化，并使用现代体视学技术进行计数。将MSC植入脑内与对健康、发育、行为和运动功能的长期影响相关联，将直接评估MSC在脑内移植的安全性和可行性。随后，MSCs将被注射到患有球状细胞白质营养不良(Krabbe病)的婴儿猕猴的中枢神经系统。后者的特点是缺乏半乳糖脑苷酶(GALC)活性，导致少突胶质细胞内储存物质异常堆积，导致神经系统进行性脱髓鞘。在这些研究中，MSC的植入水平和分布将与整个中枢神经系统中GALC mRNA的表达水平和酶的活性直接相关。此外，将使用生化和组织学方法以及行为和运动能力测试来评估对疾病进展的影响。因此，这一阶段的研究将直接评估应用MSC的治疗效果。总之，这些拟议的研究将在相关动物模型中产生重要的临床前数据，这将是开发基于干细胞的疗法以治疗患有各种溶酶体储存疾病的婴儿和儿童的神经后遗症所必需的。

DESCRIPTION (provided by applicant): This proposal aims to assess in a relevant animal model the safety, feasibility and efficacy of adult stem cell administration to the CNS for treating neurological sequelae associated with lysosomal storage diseases. Specifically, adult stem cells derived from bone marrow, referred to as mesenchymal stem cells (MSCs) will be injected unilaterally into the caudate putamen of infant rhesus macaques (Macaca mulatta) using stereotaxic coordinates determined by MRI. Prior to and at various periods up to 1 year post-transplantation, the health, development, behavior as well as cognitive and motor skills of each transplant recipient will be evaluated using a battery of age appropriate tests and compared to sham-operated animals or normative control data. Additionally, the levels and anatomical distribution of male MSCs engrafted in the CNS of female transplant recipients will be evaluated using a real-time PCR assay we developed that targets sequences in the Macaca sp. Y chromosome. To confirm the real-time PCR data engrafted MSCs will be visualized in brain sections by immunofluorescent staining or FISH and counted using modern stereological techniques. Correlating MSC engraftment in brain with long-term affects on health, development, behavior and motor function will directly assess the safety and feasibility of intracranial MSC administration. Subsequently, MSCs will be injected into the CNS of infant macaques afflicted with globoid cell leukodystrophy (Krabbe's Disease). The latter is characterized by a lack of galactocerebrosidase (GALC) activity, which results in abnormal accumulation of storage material in oligodendrocytes causing progressive demyelination in the nervous system. In these studies MSC engraftment levels and distribution will be directly correlated with levels of expressed GALC mRNA and enzyme activity throughout the CNS. Furthermore, affects on disease progression will be evaluated using biochemical and histological methods as well as behavioral and motor performance tests. Consequently, this phase of the study will directly evaluate the therapeutic efficacy of MSC administration. Collectively, these proposed studies will yield important pre-clinical data in a relevant animal model that will be essential for developing stem cell-based therapies to treat neurological sequelae in human infants and children afflicted with various lysosomal storage diseases.