Histopathology core

组织病理学核心将在Paul D.Weilstone MDCRC中发挥至关重要的作用。CORE的负责人是一位经验丰富的肌肉病理学家(Z.Sahenk，医学博士，国立儿童医院组织病理学实验室主任)。项目1将要求对直接从DMD患者身上提取的肌肉组织进行分析。在项目1的目标1中，T细胞的渗透将被检测为对Dstrophin的预先存在的免疫反应的一种表达。此外，还将进行DMD mRNA的测序，以评估剪接改变和抗原性Dstrophin表位之间的关系。在项目1的目标2中，将研究皮质类固醇治疗前后的特异性T细胞免疫。项目1的目标3密切观察AAV-微量肌营养不良蛋白注射到肌肉后的基因表达和免疫反应。在项目2中，重点切换到使用恒河猴的实验范式。这对这个实验室来说也是熟悉的领域，因为这个动物模型的组织在过去几年里一直在这个实验室处理。在AAV1项目的目标1中，将在耗尽中和抗体后检测微肌营养不良蛋白基因的传递，以研究T细胞免疫对病毒衣壳的作用。在项目2的目标2中，组织切片将被用来帮助确定针对外来转基因产品的CD4和CD8 T细胞的作用。 在项目2的最终目标3中，将有必要进行类似的组织研究，以了解可能的免疫抑制剂的作用。在人类和非人类灵长类动物组织的研究过程中，组织切片将采用标准染色和免疫组织化学方法检测T细胞亚群的基因表达和免疫标记，以及与本实验室日常研究类似的蛋白质印迹法。这一提议的成功取决于该实验室的经验和卓越表现，原因如下： 1)每个项目的任务将由一个实验室的专业知识提供便利，该实验室具有内置的质量控制措施，以一致的方式处理所有组织。 2)通过将项目1和2中的PI从结果度量中分离出来，将提高结果的客观性。 3)所有组织切片都由相同的工作人员处理，每个项目之间将有更大的一致性。

The Histopathology Core will perform a vital function in Paul D. Weilstone MDCRC. The director of the core is an experienced muscle pathologist (Z. Sahenk, MD, Ph.D, Director of the Histopathology Laboratory at Nationwide Children's Hospital). Project 1 will require analysis of muscle tissue taken directly from DMD patients. In Aim 1 of Project 1, T cell infiltration will be examined as an expression of a pre-existing immune response to dystrophin. In addition, sequencing of the DMD mRNA will be performed, in order to assess the relationship between altered splicing and antigenic dystrophin epitopes. In Aim 2 of Project 1, specific T cell immunity will be studied before and after corticosteroid treatment. Aim 3 of Project 1 closely scrutinizes gene expression and immune response following AAV-micro-dystrophin delivery to muscle. In Project 2 emphasis switches to an experimental paradigm using the rhesus macaque. This too is familiar territory to this laboratory because tissue from this animal model has been processed in this laboratory for the past several years. In Aim 1 of project 1 AAV.micro-dystrophin gene delivery will be examined after depleting neutralizing antibodies to study the role of T cell immunity to viral capsid. In Aim 2 of project 2 tissue sections will be used to help define the role of CD4+ and CD8+ T cells against a foreign transgene product. In the final Aim 3 of Project 2 similar tissue studies will be necessary to understand the role of possible immunosuppressant agents. In the course of study of both human and non-human primate tissue, histologic sections will employ standard stains and immunohistochemistry for gene expression and Immunological markers for T cell subsets, and western blots similar to every day studies in this laboratory. The success of this proposal is dependent on the experience and excellence ofthis laboratory for the following reasons: 1) The mission of each Project will be facilitated by the expertise of a single laboratory with built in quality control measures to process all tissues in a consistent manner. 2) The objectivity of the results will be enhanced by separating the Pi's in Projects 1 and 2 from the outcome measures. 3) There will be greater consistency between each of the Projects by having all tissue sections processed by the same staff.