Quantitative Vasodilation Studies

描述(申请人提供)：这项研究方案描述了一系列实验，旨在帮助确定循环系统中阻力血管(即小动脉和毛细血管)控制血管张力的确切机制。具体地说，这项提议将有助于确定红细胞(RBC)在控制肺血管阻力中的作用。当红细胞穿过微血管床时，例如在肺中，红细胞会受到机械变形。以前的发现表明，红细胞是合成一氧化氮(NO)所必需的，并且红细胞通过微米大小的毛孔或管子会导致ATP的释放，这是一种已知的刺激内皮细胞合成NO的物质。在这项建议中，我们描述了一系列旨在量化红细胞穿过聚二甲基硅氧烷(PDMS)芯片制成的微孔通道时的ATP释放速率和持续时间的研究，这些芯片的内径与完整循环中的阻力血管的内径相当。具体地说，我们将研究微孔通道内径和长度的变化以及流速对兔红细胞释放ATP的影响。此外，我们还将监测在存在和不存在红细胞来源的ATP的情况下产生的NO的量。因此，在这个方案中，我们提出的假设是：红细胞在机械变形时释放的ATP是内源性NO合成的刺激因素，因此是肺循环中血管阻力的重要决定因素。在这里，我们打算1)证明在制造的微芯片中，通道直径的减小以及通道长度和流速的增加可以刺激这些细胞释放ATP；2)证明红细胞固有的某些特性，即细胞的变形性和细胞年龄，可以影响红细胞释放ATP；3)证明固定在微芯片通道管腔上的内皮细胞可以被用来在体内模拟真实阻力血管的内皮，并且在RBC衍生的ATP刺激下，固定化内皮细胞产生和释放的NO可以在芯片上以电流计测量。这些研究的成功完成将使人们更全面地了解那些负责控制肺循环中血管阻力的机制。这些信息将允许开发关于红细胞对控制健康和疾病中的血管口径的贡献的新假说。

DESCRIPTION (provided by applicant): This research proposal describes a series of experiments that are designed help to determine the exact mechanism by which resistance vessels in the circulatory system, namely arterioles and capillaries, control vascular tone. Specifically, this proposal will help define the role of the red blood cell (RBC) in the control of pulmonary vascular resistance. When traversing microvascular beds, such as in the lung, RBCs are subjected to mechanical deformation. Previous findings indicate the RBCs are required for nitric oxide (NO) synthesis in the lung, and that passage of RBCs through micrometer-sized pores or tubing results in the release of ATP, a known stimulus for endothelial cell NO synthesis. In this proposal, we describe a series of studies designed to quantify both the rate and duration of ATP release from RBCs as they traverse microbore channels fabricated in polydimethylsiloxane (PDMS) chips with internal diameters comparable to those of resistance vessels in the intact circulation. Specifically, we will examine the effect of alterations in the internal diameter and the length of the microbore channel, as well as the velocity of flow on ATP release from RBCs of rabbits. In addition, we will also monitor the amount of NO produced in the presence and absence of RBC derived ATP. Thus, in this proposal we address the hypothesis that: ATP, released from RBCs in response to mechanical deformation, is a stimulus for endogenous NO synthesis and, thereby, is an important determinant of vascular resistance in the pulmonary circulation. Here, we intend to 1) demonstrate that decreases in channel diameter and increases in channel length and flow velocity stimulate ATP release from these cells in a fabricated microchip, 2) demonstrate that certain properties intrinsic to the RBC, namely cell deformability and cell age, can affect ATP release from RBCs and 3) demonstrate that endothelial cells immobilized to the lumen of a microchip channel can be employed to mimic the endothelium of a real resistance vessel in vivo, and that the NO production and release from immobilized endothelial cells, stimulated by RBC-derived ATP, can be measured amperometrically on-chip. The successful completion of these studies will lead to a more comprehensive understanding of those mechanisms that are responsible for he control of vascular resistance in the pulmonary circulation. This information will permit the development of new hypotheses regarding the contribution of RBCs to the control of vascular caliber in health and disease.