NEW APPROACH TO ENDOTHELIAL CLEFT STRUCTURE

我们研究的总体目标是开发一种组合工程， 超微结构和生物物理方法的机制， 内皮细胞和细胞之间的裂缝调节微血管 磁导率 冷冻断裂研究和连续切片具有 表明内皮细胞通过一系列连接蛋白连接在一起， 以允许水通过的间隔中断的股线， 溶质。 细胞化学和渗透性研究也表明，所有或 部分裂缝可能填充有纤维基质。 我们描述小说 实验由一个新的概念理论框架指导， 单独灌注的微血管片段的渗透性特性， 具有已知的渗透性，对超微结构的 相邻内皮细胞和纤维基质之间的连接股 在裂缝中。 该提案的具体目标是：（1）评估 建议微血管壁对水的渗透性， 小溶质是由小的不连续性的频率， 连接链;和2）评估以下的相对贡献： 与连接股和纤维基质相关的结构， 微血管壁上的分子过滤器。 我们的新的主要重点是 方法是三维对流扩散的分析 示踪剂分子在中断的蛋白质侧的扩散， 连接股阵列。 这种蔓延类似于一个 在很低的雷诺数下，小物体下游侧的尾流。 亚当森博士连续切片的初步结果表明， 方法是必要的，以调查的几何形状和分布 假设的小孔或狭缝在连接股，不能 通过常规的30- 40 nm切片进行解析。 结合理论和 本文采用的实验方法利用了最近的 由Weinbaum博士开发的高度精确的分析解决方案 包含连接股的裂隙中的三维粘性流 具有离散的孔和细长的交叉桥接纤维组分。 模型 需要解释示踪剂的三维分布 具体目标1中提出的尾流实验，并分析结果， 具体目标2中提出的较大溶质分子的实验。 在两种实验设计中，将对个体进行测量。 灌注的微血管具有精确已知的渗透性， 咖喱博士的实验室已经广泛开发的技术。 我们相信这种理论和实验相结合的方法可以走得很远 除了过去和现在试图描绘连接和纤维基质 调节微血管通透性的结构。 这些研究是 最直接的方法来重新理解的性质， 决定正常组织中微血管通透性的阻力， 在受伤后的恢复期。

The overall aim of our research is to develop a combined engineering, ultrastructural and biophysical approach to the mechanisms whereby endothelial cells and the clefts between the cells modulate microvessel permeability. Freeze fracture studies and ultrathin serial sections have demonstrated that endothelial cells are joined by an array of junctional strands which are interrupted at intervals allowing passage of water and solutes. Cytochemical and permeability studies also indicate that all or part of the cleft may be filled with a fibrous matrix. We describe novel experiments guided by a new conceptual theoretical framework to relate permeability properties of segments of individually perfused microvessels, having known permeability properties, to the ultrastructure of the junctional strands between adjacent endothelial cells and a fiber matrix within the cleft. The specific aims of the proposal are 1) to evaluate the proposal that the permeability properties of microvessel walls to water and small solutes are determined by the frequency of small discontinuities in the junctional strand; and 2) to evaluate the relative contribution of structures associated with the junctional strand and the fiber matrix to the molecular filter at the microvessel wall. The primary focus of our new approach is the analysis of the three-dimensional convective-diffusive spread of tracer molecules on the albuminal side of the interruptions in the junction strand arrays. This spread is analogous to the growth of a wake on the downstream side of a small object at very low Reynolds numbers. Preliminary results from serial sections by Dr. Adamson indicate that this approach is necessary to investigate the geometry and distribution of hypothetical small pores or slits in the junction strands which cannot be resolved by conventional 30-40nm sections. The combined theoretical and experimental approach adopted herein takes advantage of a recently developed highly accurate analytic solution by Dr. Weinbaum for the three-dimensional viscous flow in a cleft, which contains junction strands with discrete pores and slender cross-bridging fiber components. The model is needed to interpret the three-dimensional distribution of tracer in the wake experiments proposed in Specific Aim 1, and to analyze the results of the experiments with larger solute molecules proposed in Specific Aim 2. In both experimental designs measurements will be performed on individual perfused microvessels of precisely known permeability properties using techniques that have been extensively developed in Dr. Curry's laboratory. We believe this combined theoretical and experimental approach goes far beyond past and current attempts to delineate the junction and fiber matrix structures which modulate microvessel permeability. These studies are the most direct approach to a new understanding of the nature of the resistances which determine microvessel permeability in normal tissue and during recovery from injury.