Monitoring Red Cell Metabolism using a Lab on a Chip

使用芯片实验室监测红细胞代谢

• 批准号: 7850211
• 负责人: Dana M Spence
• 金额: $ 10.49万
• 依托单位: MICHIGAN STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-01-15 至 2011-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7850211
• 关键词: Adenosine Triphosphate; Antioxidants; Biological Assay; Blood Circulation; Blood Pressure; Cell Line; Chemicals; Complications of Diabetes Mellitus; Detection; Development; Devices; Diabetes Mellitus; Endothelial Cells; Endothelium; Erythrocyte Deformability; Erythrocytes; Health; Hypertension; Literature; Measurement; Measures; Metabolic Pathway; Metabolism; Methods; Microcirculation; Microfluidic Microchips; Monitor; NADP; Nitric Oxide; Non-Insulin-Dependent Diabetes Mellitus; Oryctolagus cuniculus; Outcome; Oxygen; Patients; Pentosephosphate Pathway; Physiological; Production; Relaxation; Reporting; Resistance; Role; Scheme; Smooth Muscle Myocytes; System; Technology; Time; Tissues; base; diabetic; improved; micro-total analysis system

DESCRIPTION (provided by applicant): In this proposal, we describe the development of an analytical system that will enable the elucidation of the role of red blood cells (RBCs) in complications resulting from type II diabetes. Specifically, using a lab on a chip approach, we propose to examine the ability of erythrocytes to maintain inherent deformability via their antioxidant defense system. Recently, it has been demonstrated that erythrocyte deformability is a determinant of deformation-induced release of adenosine triphosphate (ATP). Moreover, this ATP is a known stimulant of nitric oxide production in endothelial cells that line resistance vessels in the microcirculation. When released, this endothelium-derived NO results in the eventual relaxation of smooth muscle cells surrounding the resistance vessels and subsequent dilation of the circulatory vessel. This dilation allows for an increase in erythrocyte flow delivering oxygen to required tissues and maintaining proper blood pressure. It has been reported in the literature that the erythrocytes of patients with type II diabetes have erythrocytes that are less deformable than erythrocytes of healthy patients. Thus, it is possible that the decreased deformability of erythrocytes from type II diabetics may be related to a decrease in erythrocyte-derived ATP (which in turn may result in diminished NO production in endothelial cells) and a subsequent increase in hypertension or circulation problems (both of which are complications suffered by most type II diabetics). Here, we propose 1)To develop an amperometric detection scheme on a microfluidic device to determine the ratio of GSH:GSSG in the RBCs of rabbits, 2) To develop an amperometric assay on a microfluidic device to determine the levels of NADPH in the RBCs of rabbits, 3) To develop a chip-based method for the determination of RBC deformability, 4) To employ the measurement schemes developed in 1-3 to determine the levels of those molecules while at the same time quantitatively measuring the amount of ATP released from rabbit RBCs and the RBCs from patients with Type II diabetes mellitus. Importantly, this device will not be employed solely for monitoring endpoint metabolites; rather, it will enable, for the first time, the ability to monitor a crucial metabolic pathway (specifically, the pentose phosphate pathway) in real time while simultaneously measuring the physical outcome (ATP release). Such determinations will help identify the role of RBCs in diabetic complications, thus improving the health of patients with diabetes.

描述（由申请人提供）：在本提案中，我们描述了一种分析系统的开发，该系统将能够阐明红细胞（RBC）在II型糖尿病并发症中的作用。具体而言，使用芯片上的实验室的方法，我们建议检查红细胞通过其抗氧化防御系统保持固有的变形能力。最近，研究表明红细胞变形能力是变形诱导的三磷酸腺苷（ATP）释放的决定因素。此外，这种ATP是一种已知的刺激物，可以刺激微循环中的阻力血管内皮细胞产生一氧化氮。当释放时，这种内皮衍生的NO导致阻力血管周围的平滑肌细胞的最终松弛和随后的循环血管的扩张。这种扩张允许增加红细胞流量，将氧气输送到所需的组织并保持适当的血压。在文献中已经报道，患有II型糖尿病的患者的红细胞具有比健康患者的红细胞更不易变形的红细胞。因此，II型糖尿病患者的红细胞变形能力降低可能与红细胞来源的ATP减少（这反过来可能导致内皮细胞中NO产生减少）以及随后高血压或循环问题增加（这两者都是大多数II型糖尿病患者所遭受的并发症）有关。在这里，我们提出1）在微流控装置上开发安培检测方案以确定兔RBC中GSH：GSSG的比率，2）在微流控装置上开发安培测定以确定兔RBC中NADPH的水平，3）开发基于芯片的方法以确定RBC变形性，4）采用1-3中开发的测量方案来确定那些分子的水平，同时定量测量从兔RBC和来自II型糖尿病患者的RBC释放的ATP的量。重要的是，该设备将不仅仅用于监测终点代谢物;相反，它将首次能够在真实的时间内监测关键代谢途径（特别是戊糖磷酸途径），同时测量物理结果（ATP释放）。此类测定将有助于确定红细胞在糖尿病并发症中的作用，从而改善糖尿病患者的健康状况。