基因體規模肝細胞代謝網路之通量耦合與沃伯格效應分析

肝臟是身體內以代謝功能為主的器官，其中肝細胞 (Hepatocyte) 在身體裡面扮演著去毒素、儲存醣原（肝醣）和分泌蛋白質合成等重要生理機制，為人體重要的代謝器官。大部分的肝臟疾病都會有黃疸症狀，這是由於肝臟無法持續將膽紅素排出，所以就在體內累積而造成疾病。另外，粒線體 (Ｍitochondria) 為肝細胞經三羧酸循環 (Tricarboxylic acid cycle) 與呼吸鏈生成三磷酸腺苷 (Adenosine triphosphate，ATP) 之主要場所，提供細胞所需之能量，以維持肝細胞正常生理運作，但在異常細胞或癌細胞中，糖酵解 (glycolysis) 路徑通量會增強而粒線體裡面的氧化磷酸化（Oxidative phosphorylation）與三羧酸循環通量會減少。符合這種現象的狀態稱做沃伯格效應 (Warburg Effect)。本研究利用人類肝細胞代謝網路模型- Recon2 liver hepatocytes model，藉由觀察以合成ATP反應之間的耦合反應關係，模擬肝細胞在攝取不同營養基質 (氧氣、胺基酸、脂肪酸、維生素) 的情況下，如何執行膽紅素 (Bilirubin) 排出等生理功能。並且假設人類肝細胞在受到環境干擾或基因突變等狀態下，會傾向於合成ATP通量最大化，以提供應變所需的能量，並且使用通量均衡分析方法 (Flux balance analysis, FBA) 求得在穩定狀態下的通量分佈。另外，我們以腫瘤抑制基因 Mir122a 為例，模擬 Mir122a 基因異常肝細胞的代謝變化，以突變通量均衡分析方法 (Mutant Flux balance analysis, mFBA) 求得在穩定狀態下的通量分佈，進行通量均衡分析與突變通量均衡分析的比較，最後再利用通量變異性分析 (Flux Variability Analysis, FVA) 發現其結果符合沃伯格效應並將模擬結果與實驗數據做交叉驗證，發現實驗組的22個重要物質裡面有14個物質模擬出來的結果與實驗相符。

關鍵字

Mir122a 基因；肝細胞；肝臟；最適化模擬；代謝網路；膽紅素

並列摘要

Liver is a vital organ with the main metabolic functions in human body. The liver cells (Hepatocyte) inside the human body play a role of important physiological mechanism, including getting rid of the toxin, storage of glycogen and synthesis of secreted protein. Most of liver diseases occurred with jaundice symptoms, because bilirubin was unable discharged continuously by liver and accumulated in human body. Mitochondrion is the main organelle in which ATP (Adenosine Triphosphate) was generated through TCA cycle (Tricarboxylic Acid Cycle) and respiratory chain and provided hepatocyte cells enough energy to maintain their normal physiological operation. Unlike in normal cells, glycolysis is enhanced and both tricarboxylic acid cycle and oxidative phosphorylation capacity are reduced in various cancer cells. This phenomenon is corresponding to the Warburg Effect. In this study, we used a mathematical hepatocyte-specific model downloaded from Recon2 to simulate bilirubin metabolic functions of liver under different environment conditions by observing the relationship of coupling between the objective function, e.g., different uptakes of oxygen, amino acids, lipids, and vitamins. We assumed mammalian hepatocyte cells will produce more ATP as possible as they can to meet the energy requirement for responding to genetic perturbations and environmental changes. The internal flux distribution of mutants at steady states can be obtained by flux balance analysis (FBA) under the assumption of maximization of internal fluxes. Mir122a is a tumor suppressor gene. We computed the flux distribution of a Mir122a mutant at steady states by mutant flux balance analysis (mFBA) and compare flux balance analysis and mutation flux balance analysis, found that the results is corresponding to the Warburg effect. Therefore, the simulation results and experimental data do cross-validation, found that 22 important substances which has 14 substances in agreement with experiment.

並列關鍵字

metabolic network ； Mir-122a ； Hepatocyte ； Liver ； Optimization ； Bilirubin

參考文獻

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基因體規模肝細胞代謝網路之通量耦合與沃伯格效應分析

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