NMR STUDIES OF THE MECHANISMS OF CELL INJURY

细胞损伤机制的核磁共振研究

• 批准号: 3840980
• 负责人: R E LONDON
• 金额: --
• 依托单位: NATIONAL INSTITUTE OF ENVIRONMENTAL HEALTH SCIENCES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/3840980
• 关键词: adenosine; adenosine triphosphate; calcium flux; cytotoxicity; heart; ion transport; laboratory rat; membrane channels; membrane transport proteins; nuclear magnetic resonance spectroscopy; physiologic stressor; stress proteins

The goal of this research is to elucidate the critical cellular changes that lead to irreversible cell injury. These mechanisms must be understood before rational interventions can be proposed. Previously, we have shown that an increase in cytosolic free calcium is associated with cell injury. In many systems, it has been observed that exposure of cells or animals to a brief stress provides considerable protection against a subsequent more severe stress. This brief period of stress is known to result in the synthesis of stress proteins; however, in spite of the identification of these proteins, there is little understanding of the mechanisms of protection. Exposing perfused rat heart to several brief (5 minute) periods of anoxic stress (referred to as preconditioning) affords protection when these preconditioned hearts are subjected to a longer 30-60 minute period of anoxic stress such that the preconditioned hearts have less necrosis and better recovery of function compared to untreated controls. Using 31P nuclear magnetic resonance(NMR) we have determined that preconditioning decreased the rate of ATP utilization; preconditioned hearts have a slower rate of decline in ATP and a reduced rate of anaerobic glycolysis. In addition, using a combination of 31P NMR and 19F NMR (to measure intracellular calcium) we have determined that the reduced metabolism results in less H+ production, and therefore less Na/H and Na/Ca exchange and ultimately less of an increase in cytosolic free calcium. We conclude that at least part of the protective effect of preconditioning is due to reduced calcium overload. We were interested in addressing the mechanism(s) responsible for the reduced rate of ATP utilization. We have investigated the hypothesis that adenosine is the mediator of preconditioning; however, in perfused rat heart adenosine does not appear to play such a role since we were unable to block the effect with an adenosine antagonist. In addition, we were not able to block preconditioning by the addition of glibenclamide, thus ruling out a role for the ATP sensitive K channels as part of this process.

这项研究的目的是阐明关键的细胞变化 从而导致不可逆的细胞损伤。 这些机制必须了解 在提出合理干预措施之前。 之前我们已经展示过 细胞质游离钙的增加与细胞损伤有关。 在许多系统中，已经观察到细胞或动物暴露于 短暂的压力可以提供相当大的保护，防止随后的更多压力 严重的压力。 众所周知，这段短暂的压力会导致 应激蛋白的合成；然而，尽管鉴定 人们对这些蛋白质的作用机制知之甚少 保护。 将灌注的大鼠心脏暴露于几个简短的时间（5 分钟） 缺氧应激期（称为预处理）提供 当这些经过预处理的心脏受到更长时间的 30-60 小时的保护时 短暂的缺氧应激，使预先适应的心脏 与未治疗相比，坏死更少，功能恢复更好 控制。 使用 31P 核磁共振 (NMR) 我们确定 预处理降低了 ATP 利用率；预条件化的 心脏 ATP 下降速度较慢，无氧代谢率降低 糖酵解。 此外，结合使用 31P NMR 和 19F NMR（以 测量细胞内钙）我们已经确定减少 新陈代谢导致 H+ 产生减少，因此 Na/H 和 Na/Ca 减少 交换并最终减少胞质游离钙的增加。 我们 得出的结论是，预处理的至少部分保护作用是 由于钙超载减少。 我们有兴趣解决 导致 ATP 利用率降低的机制。 我们有 研究了腺苷是介导的假设 预处理；然而，在灌注的大鼠心脏中不出现腺苷 发挥这样的作用，因为我们无法用 腺苷拮抗剂。 此外，我们无法阻止 通过添加格列本脲进行预处理，从而排除了作用 ATP 敏感 K 通道作为此过程的一部分。