Ion Gradients And Metabolic Energy In Animal Tissue

总结：工作灌注心脏中九种主要无机离子的梯度能量彼此接近平衡，胞外和胞内相之间的电势和ATP水解的Δ G接近平衡[增田T等，生物化学杂志1990;265：20321-34]。乙醇代谢使肝细胞的静息电位从-28 mV增加至-40 mV [Veech RL et al，Alch Clin Exp Res 1994;18：1040-56]。以前，我们表明，仅仅改变可用的底物就改变了心脏中ATP水解的Δ G，并将心脏的液压功效率提高了28% [Kashiwaya Y等人，Am J Cardiol 1997;80：50 A-64 A]。由于任何类型的损伤都会诱导细胞离子分布的定型变化，其中细胞获得Na+，失去K+并肿胀，因此这些损伤的定型变化可能通过简单地改变给予损伤或烧伤受害者的液体组成而逆转。我们继续表明，细胞的细胞内和细胞外相之间的所有九种常见无机离子的梯度形成使相之间的电势与ATP水解的能量相关的唐南近平衡系统[Veech RL等人，IUBMB Life 2002;54：241-52]。由于这些研究和我们早期的研究批评了现有的胃肠外液体的组成[Veech RL，Am J Clin Nutr 1986;44：519-51]，由医学院召集的专家组建议开始研究制备新的复苏液体的可行性[Fluid Resuscitation，Natl Acad Press，Wash DC，1999]。 从那时起，我们就参加了国防部赞助的这些研究。目标是改善出血和烧伤的标准治疗，这在过去50年中没有改变。我们早先已经证明乳酸的d-异构体（林格氏乳酸盐中的一种组分）在经历烧伤复苏的儿童中诱导心律失常，并且可以在实验动物中诱导致死性心律失常[Chan L等人，Physiol Behav Sci 1994;29：383-94]。我们与海军血液研究实验室合作，设计并制造了替代品林格氏溶液，用于实验动物出血复苏。在USUHS进行的这些新生产的液体的测试表明，复苏出血与标准的高容量林格氏乳酸盐协议导致细胞凋亡在肺出血后24小时，这种不良影响可以避免，如果钠D-？O-羟基丁酸酯取代此类液体中的D，L-乳酸酯[Alam HB等人，J Am Col Surg 2001;193：255-63]。复苏后的肺损伤，所谓的岘港肺是越南战争期间手术后死亡的主要原因。虽然没有被承认，但在2003年期间，美国液体制造商将林格氏乳酸盐的组成从D，L改为L乳酸盐。这一变更不是通过向FDA提交新数据完成的，而是通过将美国药典中乳酸盐的定义从无旋光度物质变更为具有旋光度物质的权宜之计完成的。 在胃肠外液体中包含L-乳酸盐，虽然比含有D，L-乳酸盐的液体有所改进，但不是最佳的。需要复苏的情况几乎不可避免地与缺氧和乳酸酸中毒有关。L-乳酸盐的额外输注克服了这种状况，并增加了糖酵解的阻断，糖酵解是缺氧期间能量产生的主要来源，如合作者的研究所示[Cross HR等，J Mol Cell Cardiol 1995;27：1369-81]。我们认为含有D-？O-羟基丁酸酯代替L-乳酸酯是优选的。在与海军血液实验室的合作中，我们正试图设计和制备在cGMP条件下制备的用于人类受试者测试的此类流体。一篇题为“与血红蛋白氧载体相关的严重不良事件：复苏液和液体保存红细胞的作用”的手稿详细介绍了当前复苏实践的毒性作用以及如何避免这些毒性作用，已被创伤杂志接受发表。 这项工作对本研究所方案的意义 摄入酒精可降低肝细胞质中游离[NAD+]/[NADH]比值，静脉注射l-乳酸盐也可降低肝细胞质中游离[NAD +]/[NADH]比值。因此，本文的工作对该研究所的工作具有一定的指导意义。然而，这项工作对一般的医疗和外科治疗，特别是对那些参加战斗或在大规模灾难中受伤的人的治疗具有更大的意义。

Summary: The energy of the gradients of the nine major inorganic ions in working perfused heart are in near equilibrium with each other, the electrical potential between extra- and intracellular phase and the delta G of ATP hydrolysis [Masuda T et al, J Biol Chem 1990;265:20321-34]. The metabolism of ethanol increases the resting electrical potential of hepatocytes from -28 to -40 mV [Veech RL et al, Alch Clin Exp Res 1994;18:1040-56]. Previously, we showed that merely changing the substrate available altered the delta G of ATP hydrolysis in heart and improved the efficiency of hydraulic work of heart by 28% [Kashiwaya Y et al, Am J Cardiol 1997;80:50A-64A]. Since injuries of any sort induce a stereotypic change in cellular ionic distributions wherein the cell gains Na+, loses K+ and swells, these stereotypic changes of injury can possibly be reversed by simple changes in the compositions of fluids administered to victims of injury or burns. We went on to show that the gradients of all nine common inorganic ions between intra and extracellular phases of a cell form a Donnan near equilibrium system relating the electrical potential between the phases to the energy of ATP hydrolysis [Veech RL et al, IUBMB Life 2002;54:241-52]. As a result of these studies and our earlier studies criticizing the composition of existing parenteral fluids [Veech RL, Am J Clin Nutr 1986;44:519-51] a panel convened by the Academy of Medicine, recommended that investigation of the feasibility of making new resuscitation fluids be initiated [Fluid Resuscitation, Natl Acad Press, Wash DC, 1999]. We have taken part in these studies sponsored by the Department of Defense since that time. The goal is to improve the standard treatment of hemorrhage and burns, which has not changed over the past 50 years. We had earlier shown that the d isomer of lactic acid, a component in Ringers Lactate, induced cardiac arrythmias in children undergoing resuscitation from burn and could induce fatal arrythmias in experimental animals [Chan L et al, Physiol Behav Sci 1994;29:383-94]. In collaboration with the Naval Blood Research Laboratory, we designed and caused to be manufactured alternatives Ringer's solutions for use in resuscitation from hemorrhage in experimental animals. Tests of these newly manufactured fluids performed at USUHS have shown that resuscitation from hemorrhage with the standard high volume Ringer's lactate protocol leads to apoptosis in the lung 24 hours after hemorrhage and that this untoward effect can be avoided if Na D-?O-hydroxybutyrate is substituted for the D,L-lactate in such fluids [Alam HB et al, J Am Col Surg 2001;193:255-63]. Lung damage after resuscitation, so called DaNang lung was a leading cause of post surgical death during the Viet Nam war. Although not admittedly, during 2003, the US fluid manufacturers changed the composition of Ringer's lactate from D,L to L lactate. This change was not done through submission of new data to FDA, but rather through the expedient of changing the definition of lactate in the US Pharmacopeia from a substance without optical rotation, to one with optical rotation. The inclusion of L-lactate in parenteral fluids, while an improvement over fluids containing D,L-lactate, is not optimum. Conditions requiring resuscitation are almost inevitably associated with hypoxia and lactic acidosis. Additional infusion of L-lactate worsens this condition and adds to the blockade of glycolysis, the major source of energy generation during periods of hypoxia as has been shown in studies by collaborators [Cross HR et al, J Mol Cell Cardiol 1995;27:1369-81]. It is our opinion that resuscitation fluids containing D-?O-hydroxybutyrate in place of L-lactate are preferable. In collaboration with the Naval Blood Laboratory, we are attempting to design and prepare such fluids made under cGMP conditions for testing in human subjects. A manuscript entitled "Severe adverse events associated with hemoglobin based oxygen carriers: role of resuscitative fluids and liquid preserved RBC" detailing the toxic effects of current resuscitation practices and how they can be avoided has been accepted for publication in the Journal of Trauma. SIGNIFICANCE OF THIS WORK TO THE PROGRAMS OF THIS INSTITUTE Ingestion of alcohol reduces the free [NAD+]/[NADH] ratio in cytoplasm of liver as does intravenous administration of l-lactate. Therefore this work has some significance to the work of this institute. However, this work is of greater significance to medical and surgical treatments in general particularly to the treatment of those engaged in combat or subject to injury from mass disasters.