Dynamics of Regulating Oxygen Supply by Erythrocytes

脉管系统最重要的功能是向身体的每个细胞输送足够的氧气（O2）。由于氧气从血管中的红细胞（红细胞）扩散到在细胞中被消耗产生能量的地方的距离小于0.1 mm，因此血管系统必须适当地将氧气供应分配到单个末端小动脉（非常小的动脉）的水平，为支持这些细胞的少量毛细血管提供支持。这种分配氧气的准确度不能在发育过程中建立在血管结构中，它必须动态调节以响应被服务组织的不同需求（例如，根据身体活动水平在骨骼肌或心肌中）。我们是第一个提出红细胞，它是氧气的主要载体，也是传感器，它能精确地控制氧气的分布。红细胞通过红细胞中血红蛋白携带的氧气量的变化来感知氧气供应和氧气消耗之间的平衡。当氧与血红蛋白结合时，分子的形状发生变化，这既激活了红细胞内的酶产生三磷酸腺苷（ATP），又触发了红细胞膜上的信号通路，使ATP从红细胞中释放出来。ATP激活血管内皮细胞内表面的特化受体（P2y），通过释放一氧化氮或前列腺素导致局部小动脉直径增加，或者通过内皮细胞传导到小动脉的电信号从毛细血管和小静脉远端增加。关于ATP的O2依赖性释放有许多尚未解决的问题，现有技术无法解决。主要的问题是，当血红蛋白氧饱和度发生变化时，ATP需要多长时间才能从红细胞中释放出来？答案很重要，因为释放时间决定了向血管内皮报告氧饱和度水平的空间准确性。现有的技术只能提供在氧饱和度变化时释放多少ATP，而不能用于确定ATP的释放速度。我的博士生Richard sov<s:1>的最新研究成果，使我们有可能回答这个问题。获得对ATP释放调节机制的新见解。sov<s:1>开发并发表了一种微流体装置的计算模型，该装置可用于测量ATP释放的动力学。他在微流控模型中加入了ATP释放的动态信号通路模型，并证明了这种微流控方法能够揭示信号通路中每个阶段的速度。sov<s:1>的方法还可以回答这样一个问题：ATP从红细胞释放的速率是由氧饱和度水平决定的，还是由氧饱和度变化的速率决定的？另一种模型预测，如果ATP释放依赖于O2饱和度变化的速率，那么氧的分布只能由红细胞调节；因此，这个答案对于我们理解RBC在调节氧气供应中的作用至关重要。本研究的重点是开发由sov<s:1>提出的微流体系统，以充分表征红细胞中O2依赖性ATP释放的动力学。所有的研究都将在大鼠血液中进行，以保持与以往研究的一致性。这项技术为其他研究生项目提供了机会，研究红细胞在体内微血管中遇到的条件的影响，并为未来的合作健康研究项目奠定了基础，将这项研究转化为心血管疾病患者的筛查工具。

Arguably the most important function of the vasculature is the delivery of adequate oxygen (O2) to every cell of the body. Since the distance O2 can diffuse from red blood cells (RBCs) in a vessel to where it is consumed in the cell to produce energy is <0.1-mm, the vasculature must properly distribute O2 supply down to the level of a single terminal arteriole (very small artery) supplying a small number of capillaries supporting these cells. This level of accuracy in distributing O2 cannot be built into the vascular structure during development, it must be dynamically regulated to respond to varying needs of the tissue being served (e.g. in skeletal or cardiac muscle depending on the level of physical activity). We were the first to propose that the red blood cell, which is the primary carrier of O2, is also the sensor which enables this fine level of control of the distribution of O2. RBCs sense the balance between the O2 supply and O2 consumption by the change in the amount of O2 carried by hemoglobin in the RBC. As O2 binds to hemoglobin the shape of the molecule changes which both activates enzymes within the RBC to produce adenosine triphosphate (ATP) and triggers a signalling pathway in the RBC membrane for ATP to be released from the RBC. The ATP activates specialized receptors (P2y) on the inner surface of endothelial cells lining the vessel which result in an increase in arteriolar diameter, either locally in arterioles by the release of nitric oxide or prostaglandins or remotely from capillaries and venules via a electrical signal conducted through endothelial cells to the arterioles.There are unresolved issues about the O2 dependent release of ATP which existing technologies cannot answers. The primary question is how long does it take for ATP to be released from the RBC following a change in hemoglobin O2 saturation? The answer is important since the release time determines the spatial accuracy with which O2 saturation levels can be reported to the vascular endothelium. Existing techniques only provide the how much ATP is released for a change in O2 saturation and cannot be used to determine fast ATP is released. Recent advances by my Ph.D. student, Richard Sové, make it possible for us to answer this question and, in doing so; gain new insights into the mechanisms regulating ATP release. Sové developed and published a computational model for the potential design of a microfluidic device which could be used to measure the dynamics of ATP release. He added a dynamic signaling pathway model for ATP release into his microfluidic model and has demonstrated that this microfluidic approach is able to reveal how the speed of each stage in the signaling pathway. Sové’s approach can also answer the question is the release rate of ATP from the RBC governed by the level of O2 saturation or by the rate at which O2 saturation is changing? Another model predicts that O2 distribution can only be regulated by RBCs if ATP release depends on the rate of O2 saturation change; thus the answer is critical to our understanding of the RBC’s role in regulating O2 supply.This proposal focuses on developing the microfluidic system proposed by Sové for fully characterizing the dynamics of O2 dependent ATP release from RBCs. All research will be carried out on rat blood to maintain consistency with previous research. This technology creates the opportunity for other graduate student projects investigating the impact of conditions the RBC would encounter in the microvasculature in vivo and forms the basis for a future collaborative health research projects to translate this research into a screening to tool for patients with cardiovascular disease.