Bioengineering Design of Artificial Blood

描述(由申请人提供)：在第一阶段，本BRP的工作表明，MP4(聚乙二醇化修饰的血红蛋白)克服了开发基于修饰的血红蛋白的血液替代品的最重要的障碍，即血管收缩。尽管MP4具有与直觉相反的特性，包括增加氧气亲和力、粘度和肿胀压力，但它通过氧运输和维持功能性毛细血管密度的组合来促进组织氧合。MP4在使动物从严重的、无法控制的出血中复苏的能力上优于血液，并且已经在人类临床试验中被证明是安全的。在这一应用中，我们将检验MP4是血红素配体O2、一氧化碳(CO)和一氧化氮(NO)的有效载体的假设，并将进行相关的生理学研究，以了解其作为血液替代品的有效性，并确定其新的临床应用。我们将测试这一假设，即聚乙二醇-血红蛋白制剂是血管扩张剂，以与血液循环相互作用的方式与血红蛋白清除NO无关。我们认为，MP4的S特性部分是由于亚硝酸还原酶活性的增加和NO的运输。我们将开发一种使用MP4输送一氧化碳的程序，并利用CO-MP4即使在高温下也非常稳定的特点，使其在创伤现场应用中具有价值。项目1有一个GMP设施，提供质量和属性一致的MP4。它将开发和生产新的聚乙二醇Hb化合物，目标是在不增加胶体渗透压的情况下优化浓度，并增加氧气输送能力，从而增加MP4在更广泛的临床应用中的适用性。将通过生化分析、数学建模和包括对心肌梗死影响在内的大鼠系统实验来筛选特性。项目2将研究聚乙二醇化血红蛋白对糖基化的完整性的影响，聚乙二醇化血红蛋白的存在如何影响活性氧物种(ROS)，并将研究聚乙二醇化血红蛋白和增强的血浆粘度的联合作用。MP4将作为一氧化碳的运输载体，在缺血和出血时提供细胞保护。微循环研究将使用醒着的仓鼠窗口室模型，直接测量O2和NO水平、血管中的流量和直径以及功能性毛细血管密度。这项研究结合了输血的生理分析、工程运输过程的基本原理和机械转导与两个实验室合作超过12年的专业知识。有效的血液替代品将显著提高民用和军用环境中输血的安全性和有效性，精简和简化输血医学。将开发一种新型血液替代品的应用，这种替代品提供一种根本不同的治疗缺血的方法，其基础是增强微血管流量、心脏和肾脏功能、修复内皮细胞和提供血红素配体(O2、NO和CO)。

DESCRIPTION (provided by applicant): In the first period, work in this BRP demonstrated that MP4 (PEG-modified hemoglobin) overcomes the most significant hurdle to the development of modified hemoglobin-based blood substitutes, namely vasoconstriction. MP4 promotes tissue oxygenation through a combination of O2 transport and maintenance of functional capillary density in spite of its counterintuitive properties, including increased O2 affinity, viscosity and oncotic pressure. MP4 is superior to blood in its ability to resuscitate animals from severe, uncontrolled hemorrhage, and it has been shown to be safe in human clinical trials. In this present application we will test the hypothesis that MP4 is an effective carrier of the heme ligands O2, carbon monoxide (CO) and nitric oxide (NO), and will carry out the related physiological studies in order to understand its effectiveness as a blood substitute and identify new clinical applications for its use. We will test the hypothesis that PEG-Hb formulations are vasodilators which interact with the circulation in ways not related to NO scavenging by Hb. We propose that MP4's properties are in part due to an increased nitrite reductase activity and NO transport. We will develop a procedure for using MP4 to deliver CO and exploit that CO-MP4 is exceptionally stable, even at elevated temperatures, making it valuable in field use for trauma. Project 1 has a GMP facility that provides MP4 of consistent quality and properties. It will develop and produce new PEG-Hb compounds with goals to optimize concentration without increasing colloid osmotic pressure and augment O2 delivery capacity so as to increase the applicability of MP4 to a wider range of clinical uses. Properties will be screened via biochemical analysis, mathematical modeling, and systemic experiments in rats including the effect on myocardial infarction. Project 2 will examine PEG-Hbs' effects on the glycocalyx integrity, how PEG-Hbs' presence influences reactive O2 species (ROS), and will investigate the combined effect of PEG-Hb and enhanced plasma viscosity. MP4 will be used as a delivery vehicle for CO to provide cellular protection during ischemia & hemorrhage. Microcirculation studies will use the awake hamster window chamber model, with direct measurements of O2 and NO levels, flow and diameter in blood vessels and functional capillary density. This research combines physiological analysis of transfusion, fundamentals of engineering transport processes and mechanotransduction with the expertise of two laboratories with more than 12 years of collaboration. Effective blood substitutes will significantly increase the safety and efficacy of blood transfusions in civilian and military settings, streamline and simplify transfusion medicine. Applications for a new type of blood substitute will be developed, one that provides a fundamentally different treatment of ischemia, based on enhancement of microvascular flow, cardiac and renal functions, repair of the endothelium and delivery of heme ligands (O2, NO and CO).