TRANSCRIPTIONAL BASIS OF CARDIAC ALLOGRAFT REMODELING

心脏同种异体移植重构的转录基础

• 批准号: 6638499
• 负责人: ARTHUR ROGER STRAUCH
• 金额: $ 28.76万
• 依托单位: OHIO STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-07-01 至 2005-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6638499
• 关键词: actins; cardiac myocytes; cell type; confocal scanning microscopy; gel mobility shift assay; genetic promoter element; genetically modified animals; heart transplantation; histopathology; homologous transplantation; immunocytochemistry; in situ hybridization; laboratory mouse; muscle proteins; reporter genes; transcription factor; transplant rejection; vascular smooth muscle

DESCRIPTION (adapted from applicant's abstract): The major objective of the research plan is to investigate relationships between transcriptional modulation of the vascular smooth muscle (VSM) alpha-actin gene, cardiac and aortic allograft remodeling, and chronic rejection. Inappropriate expression of this essential smooth muscle actin isoform is known to impair contractility and adversely affect cardiovascular function. Loss of VSM alpha-actin expression is a characteristic feature of transplant vascular sclerosis that appears to be mediated by transcriptional regulatory proteins that bind and repress the VSM alpha-actin gene promoter in neointimal smooth muscle cells. Examination of mouse heterotopic cardiac allografts has revealed additional tissue remodeling processes during chronic rejection that are associated with elevated VSM alpha-actin gene transcription in graft stromal (cardiac fibroblast) and parenchymal (cardiomyocyte) cell beds. The proposed research focuses on the control of the mouse VSM alpha-actin gene with an emphasis on characterizing the properties of its cognate transcriptional regulatory (VAC) proteins in isolated VSM cells, cardiomyocytes, and cardiac fibroblasts as well as their genetically reprogrammed counterparts that arise following transplant. Chronic rejection conditions will be simulated by exposing isolated cells to pro-inflammatory cytokines to determine how immune cell products influence VAC protein activity and actin gene reprogramming. Regulatory elements from the VSM alpha-actin promoter will be identified, mutated, linked to a reporter gene encoding a histologically detectable product, and used to prepare transgenic mice for supplying donor hearts and aortae for in situ assay of transcription in the graft stroma, parenchyma, and arterial beds following transplant. Spatial aspects of cardiac allograft remodeling will be revealed by using histologic probes for VSM alpha-actin and VAC proteins coupled with serial microtomy, confocal microscopy, and in situ hybridization to map the distribution of cellular phenotypes in allografts. These studies are expected to provide a global view of gene reprogramming following transplant and permit early identification of anatomical regions and cellular beds in heart grafts that are most at risk for structural remodeling. The proposed research will extend our understanding of gene transcriptional control mechanisms that mediate very early cellular maladaptations to heart transplantation. Diagnostic assays and therapeutic protocols based on aspects of VSM alpha-actin transcription may represent new tools for recognizing and managing early-stage chronic rejection well before a functional decline in graft performance becomes clinically detectable.

描述（改编自申请人的摘要）： 研究计划是调查转录之间的关系， 调节血管平滑肌（VSM）α-肌动蛋白基因，心脏和 主动脉同种异体移植物重塑和慢性排斥。的不适当表达 已知这种必需的平滑肌肌动蛋白同种型损害收缩性， 对心血管功能有不利影响。VSM α-肌动蛋白表达的缺失是 这是移植血管硬化的一个特征， 由结合并抑制VSM的转录调节蛋白介导 α-肌动蛋白基因启动子在新生内膜平滑肌细胞。审查 小鼠异位心脏同种异体移植显示了额外的组织重塑 与VSM升高相关的慢性排斥反应过程 α-肌动蛋白基因在移植物基质（心脏成纤维细胞）中的转录， 实质（心肌细胞）细胞床。拟议的研究重点是 控制小鼠VSM α-肌动蛋白基因，重点是表征 其同源转录调节（VAC）蛋白的性质， 分离的VSM细胞、心肌细胞和心脏成纤维细胞以及它们的 在移植后产生的基因重组的对应物。慢性 排斥条件将通过将分离的细胞暴露于 促炎细胞因子，以确定免疫细胞产物如何影响VAC 蛋白质活性和肌动蛋白基因重编程。VSM的监管要素 α-肌动蛋白启动子将被鉴定、突变、连接到报告基因 编码组织学上可检测的产物，并用于制备转基因 用于原位转录测定的供体心脏和供体骨髓的小鼠 在移植后的移植物基质、实质和动脉床中。 心脏移植物重塑的空间方面将通过使用 VSM α-肌动蛋白和VAC蛋白的组织学探针偶联了串联 显微镜，共聚焦显微镜，原位杂交，以映射 同种异体移植物中细胞表型的分布。这些研究预计 提供移植后基因重编程的全局视图，并允许 心脏移植物解剖区域和细胞床的早期识别 最容易发生结构重塑的区域拟议的研究将 扩展我们对基因转录控制机制的理解， 介导对心脏移植的非常早期的细胞适应不良。诊断 基于VSM α-肌动蛋白方面的测定和治疗方案 转录可能代表了识别和管理早期阶段的新工具， 在移植物性能出现功能性下降之前， 临床检测。