Core--Pathology

病理学核心的目的是为所有三个项目的免疫病理学研究提供通用设施和专业知识。在这些同种异体移植方案中，病理分析的主要作用是记录和表征移植物的状态（浸润的性质、急性或慢性排斥反应的存在，包括体液成分）以及方案的全身效应（毒性、并发症）。此外，免疫病理学和分子研究将深入了解移植物排斥和接受方案活检的机制。将使用的技术包括常规组织学、免疫组织化学、免疫荧光、电子显微镜、PCR、ELISPOT/细胞培养和流式细胞术。使用一组单克隆抗体的免疫过氧化物酶技术将区分浸润细胞类型、粘附和细胞因子分子和受体以及激活标记。免疫荧光将用于定位免疫球蛋白的沉积和 补体，以及使用数字成像对细胞标记物进行双重/三重染色。 C4d 染色将用于检测体液排斥。 RT-PCR 将用于检测和定量关键细胞因子的合成。大多数项目中的一些研究将使用激光捕获显微切割来选择合适的细胞。细胞凋亡/DNA 片段化 (TUNEL) 标记物将评估细胞损伤。移植细胞的培养将与 ELISPOT 一起使用，以与血液中的功能研究相关联。嵌合现象将通过流式细胞术技术检测。每个单独的项目中都详细给出了协议。所有研究的病理学特征将由同一组病理学家进行评估和分级，我们认为这是该计划项目的重要资产。抗体和细胞因子探针组将 如有可能，也应具有可比性。这应该通过扩展新的发现来促进项目之间的交叉受精，并通过比较和对比不同物种和不同器官之间的移植物来深入了解一个项目中结果的一般意义。

The purpose of the Pathology Core is to provide common facilities and expertise for the immunopathological studies in all three projects. The primary role of the pathological analyses in these allograft protocols is to document and characterize the status of the graft (nature of infiltrate, presence of acute or chronic rejection, including a humoral component) and the systemic effects of the protocols (toxicity, complications). In addition, the immunopathologic and molecular studies will give insights into the mechanisms of graft rejection and acceptance with protocol biopsies The techniques that will be utilized include routine histology, immunohistochemistry, immunofluorescence, electron microscopy, PCR, ELISPOT/cell culture, and flow cytometry. Immunoperoxidase techniques with a panel of mAbs will distinguish the infiltrating cell types, adhesion and cytokine molecules and receptors, and activation markers. Immunofluorescence will be used to localize the deposition of immunoglobulin and complement, as well as double/triple staining for cell markers using digital imaging. C4d stains will be used to detect humoral rejection. RT-PCR will be used to detect and quantify synthesis of key cytokines. Laser capture microdissection will be used for some of these studies in most of the projects to select the appropriate cells. Markers of apoptosis/DNA fragmentation (TUNEL) will assess cell injury. Culture of graft cells will be used with ELISPOT to correlate with the functional studies in the blood. Chimerism will be detected by flow cytometry techniques. The protocols are given in detail in each of the individual Projects. The pathology features will be assessed and graded by the same group pathologists for all studies, which we consider an important asset of the Program Project. The panel of antibodies and cytokine probes will also be comparable, whenever possible. This should promote cross-fertilization among the projects by extension of novel findings and provide insights into the general significance of the results in one project by comparing and contrasting grafts in different species and between different organs.