METABOLIC FLUX PATTERNS IN HEPATOMA CELLS

这个项目的重点是代谢通量的定量评估 肝癌细胞系AS-30 D中TCA循环相关通路 细胞 该项目的两个长期目标是：1。进一步 为TCA分析开发定量建模方法 循环途径利用碳示踪分析。2.开发一个 对代谢异常有更全面的了解 肿瘤细胞的特征，如截短的TCA中所述 循环理论 该项目有五个具体目标。1.评价 乙酰乙酸作为胆固醇前体和作为 氧化能源 该项目评估了 乙酰乙酰辅酶A合成酶在胆固醇合成中意义 肿瘤。 一种关系，提供对 将引入通过两种途径流向胆固醇的流量。2. 到 评估丙酮酸羧化的生理意义 AS-30 D细胞。 使用CO2比率发现了该通量 法 3.为了检验假设TCA循环苹果酸是 优先由苹果酸脱氢酶代谢，而外部 供应的苹果酸通过线粒体苹果酸酶代谢。 4.通过还原法分析谷氨酰胺向脂质的通量， α-酮戊二酸的羧化。 程序旨在 评估这种通量是否发生在细胞质中或细胞质中， 线粒体5.从理论和实验两个方面进行研究 的前稳态示踪剂动力学在TCA循环时， 系统处于代谢稳定状态。 其中一个关键特征是， 分析是，示踪剂通量的时间常数之和 从14 C 02生产曲线确定通过途径的量。 对于所有项目，实验方法都使用相同的烧瓶， 分离的细胞，每个细胞具有不同的14 C标记的化合物。 所有 生物化学测定和基于公开的技术。 理论 使用代数和简单的微分方程进行工作。 这种方法的优势在于结合了传统的 生物化学方法和建模方法。

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.