Weiss R G, Gloth S T, Kalil-Filho R, Chacko V P, Stern M D, Gerstenblith G
Peter Belfer Laboratory of the Cardiology Division, Department of Medicine, Johns Hopkins Hospital, Baltimore, Md. 21205.
Circ Res. 1992 Feb;70(2):392-408. doi: 10.1161/01.res.70.2.392.
Although the tricarboxylic acid (TCA) cycle is the prime means of carbon metabolism for energy generation in normal myocardium, the noninvasive quantification of TCA cycle flux in intact cardiac tissues is difficult. A novel approach for estimating citric acid cycle flux using 13C nuclear magnetic resonance (NMR) is presented and evaluated experimentally by comparison with measured myocardial oxygen consumption over a wide range of cardiac contractile function in intact, beating rat hearts. Continuous series of 13C NMR spectra, obtained after the introduction of [2-13C]acetate as substrate, quantified the time course of 13C appearance in the carbon positions of myocardial glutamate, which are sequentially enriched via citric acid cycle metabolism. A TCA cycle flux parameter was calculated using the premise that TCA cycle flux is inversely proportional to the time difference between 13C appearance in the C-4 and C-2 positions of glutamate (glutamate delta t50 [minutes]), which are enriched in subsequent "turns" of the TCA cycle. This TCA cycle flux parameter, termed KT, correlated strongly with myocardial oxygen consumption over a range of developed pressures in hearts perfused with 5 mM acetate (r = 0.98, p less than 0.001), as well as in separate studies in hearts perfused with 5 mM glucose and 0.5-0.8 mM acetate (r = 0.94, p less than 0.001). Results of numerical modeling of 13C glutamate kinetics suggest that this TCA cycle flux parameter, KT, is relatively insensitive to changes in metabolite pool sizes that could occur during metabolism of other substrates or during conditions of altered oxygen availability. Additional studies in separate hearts indicated that the time course of 13C appearance in citrate, which is predominantly mitochondrial in the rat heart, is similar to that in glutamate, further supporting the premise that the described 13C NMR parameters reflect mitochondrial citric acid cycle activity in intact cardiac tissues.
虽然三羧酸（TCA）循环是正常心肌中碳代谢产生能量的主要方式，但在完整心脏组织中对TCA循环通量进行无创定量却很困难。本文提出了一种使用13C核磁共振（NMR）估算柠檬酸循环通量的新方法，并通过与完整跳动大鼠心脏在广泛心脏收缩功能范围内测得的心肌耗氧量进行比较，进行了实验评估。在引入[2-13C]乙酸盐作为底物后获得的连续13C NMR光谱系列，对心肌谷氨酸碳位置上13C出现的时间进程进行了定量，这些位置通过柠檬酸循环代谢依次富集。使用TCA循环通量与谷氨酸C-4和C-2位置（谷氨酸δt50[分钟]）上13C出现的时间差成反比这一前提来计算TCA循环通量参数，这两个位置在TCA循环的后续“轮次”中富集。这个称为KT的TCA循环通量参数，在灌注5 mM乙酸盐的心脏中，在一系列发育压力范围内与心肌耗氧量密切相关（r = 0.98，p小于0.001），在灌注5 mM葡萄糖和0.5 - 0.8 mM乙酸盐的心脏的单独研究中也是如此（r = 0.94，p小于0.001）。13C谷氨酸动力学的数值模拟结果表明，这个TCA循环通量参数KT对其他底物代谢过程中或氧可用性改变条件下可能发生的代谢物池大小变化相对不敏感。在单独心脏中的额外研究表明，柠檬酸中13C出现的时间进程（在大鼠心脏中主要存在于线粒体中）与谷氨酸中的相似，进一步支持了所描述的13C NMR参数反映完整心脏组织中线粒体柠檬酸循环活性这一前提。
