BLADDER SMOOTH MUSCLE RESPONSE TO MECHANICAL FORCE

描述(改编自申请人的摘要)：本报告的目标 建议确定膀胱平滑的分子机制 肌肉和固有层成纤维细胞对纤维状胶原基因的调控 表情。假设仅靠机械刺激就能 调节其编码蛋白的转录因子基因的表达 调节和触发其他负责基因的表达 膀胱细胞有明显的表型变化。通过鉴定这些基因 并鉴定了它们在膀胱平滑肌和成纤维细胞中的激活情况 细胞，调查人员希望获得关于 膀胱的结构和功能特性的机制 都是受监管的。他们已经开发出一种设备，可以让 对细胞施加精确的机械载荷，即机械拉伸 在试管中。他们将使用这样的装置将等轴拉伸应用于 膀胱壁细胞与机械敏感性调节过程的研究 基因。他们建议：1)研究应变和应变率对 细胞生长和基质蛋白合成，2)表征 I型和III型胶原基因中的机械反应元件，3)到 识别和表征其表达受调控的基因 机械刺激，以及4)使用靶向引物筛选该家系 转录因子的表达受应用的调节 并对其基因的启动子活性进行表征 目标。基因表达的变化将在mRNA和 使用分子生物学和生化技术的蛋白质水平，即， Northern印迹、Western印迹和差异显示RT-PCR。转染法 实验和DNA-蛋白质相互作用分析将被用来定义 血管紧张素转换酶启动子中的拉伸调节序列(S)和反式作用因子(S) 机械敏感基因。通过这些方法，调查人员预计将 胶原蛋白表达的变化与表达和/或 反式作用因素的活动性，并定义物理力量的作用 在分子水平上，膀胱的正常功能。

DESCRIPTION (Adapted from the Applicant's Abstract): The goal of this proposal is to determine the molecular mechanism by which bladder smooth muscle and lamina propria fibroblast regulate fibrillar collagen gene expression. The hypothesis is that mechanical stimulation alone can modulate the expression of transcription factor genes whose encoded proteins regulate and trigger the expression of other genes that are responsible for substantial phenotypic changes of bladder cells. By identifying these genes and characterizing their activation in bladder smooth muscle and fibroblast cells, the investigators expect to gain valuable information on the mechanisms by which the structural and functional properties of the bladder are regulated. They have developed an apparatus that permits the application of precise mechanical load, i.e., mechanical stretch, to cells in vitro. They will use such a device to apply an equiaxial stretch to bladder wall cells and study the regulation process of mechano-sensitive genes. They propose: 1) to study the effects of strain and strain rate on cell growth and matrix protein synthesis, 2) to characterize the mechano-response element in the type I and III collagen genes, 3) to identify and characterize genes whose expression is modulated as a result of mechanical stimulation, and 4) to use targeted primers to screen the family of transcription factors whose expression is modulated by the application of mechanical forces and to characterize the promoter activity of their gene targets. Changes in gene expression will be evaluated at the mRNA and protein levels using molecular biological and biochemical techniques, i.e., Northern blot, Western blot and differential display RT-PCR. Transfection experiments and DNA-protein interaction analyses will be used to define stretch-regulatory sequence(s) and trans acting factor(s) in the promoter of mechano-sensitive genes. With these approaches, the investigators expect to correlate changes in collagen expression to changes in expression and/or activity of trans acting factors, and to define the role of physical forces at the molecular level in normal functioning of the bladder.