METABOLIC FLUX PATTERNS IN HEPATOMA CELLS

这个项目的重点是代谢流量的定量评估。 肝癌细胞株AS-30D中TCA周期相关通路的研究 细胞。该项目的两个长期目标是：1.进一步 建立三氯乙烷分析的定量模型方法 利用碳示踪剂分析的自行车路径。2.发展一种 对代谢异常有更全面的认识 截短TCA中所描述的肿瘤细胞的特征 循环论。该项目有五个具体目标。1.评估 乙酰乙酸酯作为胆固醇前体和作为一种 氧化能源。这个项目评估了可能的 乙酰辅酶A合成酶在胆固醇合成中的意义 肿瘤的症状。提供定量估计的关系 通过两条途径向胆固醇的通量将被引入。2.至 评价丙酮酸羧化的生理学意义 AS-30D细胞。这种通量是利用二氧化碳比率发现的。 技术。3.检验TCA循环苹果酸是 优先由苹果酸脱氢酶代谢，而在外部 供应的苹果酸由线粒体苹果酸酶代谢。 4.谷氨酰胺还原成脂的通量分析 α-酮戊二酸的羧化反应。程序旨在 评估这种通量是发生在细胞质中还是发生在细胞质中 线粒体。5.考察理论和实验两个方面 三氯乙烷循环中的稳态前示踪动力学 系统处于代谢稳定状态。这是它的一个关键特征 分析是示踪剂通量的时间常数之和 通过一条途径从14C02的生产剖面中确定。 对于所有项目，实验方法都使用相同的烧瓶 分离的细胞，每个细胞都有不同的14C标记化合物。全 生化分析，并基于已发表的技术。理论上的 这项工作是使用代数和简单的微分方程式来完成的。 这种方法的优势在于结合了传统的 生化方法和建模方法。

This project focuses on a quantitative assessment of metabolic flux through TCA cycle related pathways in a hepatoma cell line, AS-30D cells. The two long term goals of the project are: 1. To further develop a quantitative modeling approach for the analysis of TCA cycle pathways utilizing carbon tracer analysis. 2. To develop a more complete understanding of the metabolic abnormalities characteristic of tumor cells as described in the truncated TCA cycle theory. The project has five specific aims. 1. To evaluate the role of acetoacetate as a precursor for cholesterol and as an oxidative energy source. This project assess the possible significance of acetoacetyl CoA synthetase in cholesterol synthesis of tumors. A relationship providing a quantitative estimate of flux to cholesterol via two pathways will be introduced. 2. To assess the physiological significance of pyruvate carboxylation in AS-30D cells. This flux was discovered using the C02 ratios technique. 3. To examine the hypothesis that TCA cycle malate is preferentially metabolized by malate dehydrogenase while externally supplied malate is metabolized by the mitochondrial malic enzyme. 4. To analyze the flux of glutamine to lipid via reductive carboxylation of a-ketoglutarate. Procedures are designed to evaluate whether this flux occurs in the cytoplasm or in the mitochondria. 5. To examine theoretical and experimental aspects of pre-steady state tracer kinetics in the TCA cycle when the system is in metabolic steady state. A key feature of this analysis is that the sum of the time constants for tracer flux through a pathway are determined from the 14C02 production profile. For all projects experimental methods utilize identical flasks of isolated cells, each with a different 14C labeled compound. All biochemical assays and based on published techniques. Theoretical work is performed using algebra and simple differential equations. The strength of this approach is in the combination of conventional biochemical methods and modeling methodology.