INTRODUCTION Relative oxidation of different metabolic substrates in the heart varies both physiologically and pathologically, in order to meet metabolic demands under different circumstances. 13C labelled substrates have become a key tool for studying substrate use-yet an accurate model is required to analyse the complex data produced as these substrates become incorporated into the Krebs cycle. OBJECTIVES We aimed to generate a network model for the quantitative analysis of Krebs cycle intermediate isotopologue distributions measured by mass spectrometry, to determine the 13C labelled proportion of acetyl-CoA entering the Krebs cycle. METHODS A model was generated, and validated ex vivo using isotopic distributions measured from isolated hearts perfused with buffer containing 11 mM glucose in total, with varying fractions of universally labelled with 13C. The model was then employed to determine the relative oxidation of glucose and triacylglycerol by hearts perfused with 11 mM glucose and 0.4 mM equivalent Intralipid (a triacylglycerol mixture). RESULTS The contribution of glucose to Krebs cycle oxidation was measured to be 79.1 ± 0.9%, independent of the fraction of buffer glucose which was U-13C labelled, or of which Krebs cycle intermediate was assessed. In the presence of Intralipid, glucose and triglyceride were determined to contribute 58 ± 3.6% and 35.6 ± 0.8% of acetyl-CoA entering the Krebs cycle, respectively. CONCLUSION These results demonstrate the accuracy of a functional model of Krebs cycle metabolism, which can allow quantitative determination of the effects of therapeutics and pathology on cardiac substrate metabolism.

中文翻译：

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使用稳定的13C标记代谢物确定心脏线粒体底物利用率的模型。

简介心脏中不同代谢底物的相对氧化在生理和病理上均会发生变化，以便满足不同情况下的代谢需求。13C标记的底物已成为研究底物用途的关键工具，但需要精确的模型来分析随着这些底物被纳入克雷布斯循环而产生的复杂数据。目的我们旨在生成一个网络模型，用于对通过质谱法测量的克雷布斯循环中间同位素同位素分布进行定量分析，以确定进入克雷布斯循环的13 C标记的乙酰辅酶A比例。方法生成了一个模型，并使用同位素分布从离体心脏进行了验证，该同位素分布是从孤立的心脏中注入总含11 mM葡萄糖的缓冲液测得的，普遍使用13C标记的分数各不相同。然后使用该模型确定灌注11 mM葡萄糖和0.4 mM当量内脂（三酰甘油混合物）的心脏的葡萄糖和三酰甘油的相对氧化。结果测得葡萄糖对Krebs循环氧化的贡献为79.1±0.9％，与U-13C标记的缓冲液葡萄糖分数或评估的Krebs循环中间体无关。在存在内脂的情况下，确定葡萄糖和甘油三酸酯分别贡献了58±3.6％和35.6±0.8％的乙酰辅酶A进入Krebs循环。结论这些结果证明了克雷布斯循环代谢功能模型的准确性，该模型可以定量确定治疗方法和病理学对心脏底物代谢的影响。