槲皮素調控小鼠C2C12肌纖維細胞粒線體生合成及功能之研究

Quercetin regulates mitochondrial biogenesis and function in C2C12 muscle cells

周映君(Ying-Chun Chou)

指導教授 ： 謝榮鴻

臺北醫學大學/公共衛生暨營養學院/保健營養學研究所/碩士(2013年)

摘要

隨著世界人口老化的增加，如何延緩慢性疾病的發生為重要的議題。老化伴隨著肌肉的減少及功能的衰退，影響粒線體失調，與許多慢性疾病息息相關。槲皮素 (quercetin)為天然存在的類黃酮物質，廣泛存在食物中，臨床研究顯示槲皮素調控粒線體生合成相關蛋白與促進粒線體生合成呈現相關性且可改善肌肉耐力。本篇論文主要的研究方向為利用分化的C2C12肌纖維細胞，探討槲皮素的介入對粒線體生合成的影響及與AMPK (AMP-activated protein kinase)-SIRT1 (sirtuin 1)-PGC1α (peroxisome proliferator-activated receptor γ coactivator-1α)調控路徑的相關性。C2C12肌纖維細胞介入槲皮素24小時後觀察其細胞存活率、ATP 生成量、細胞氧消耗率、粒線體膜電位、粒線體酵素活性。在測定基因表現方面，以同步定量聚合酶反應、西方墨點法，測定目標基因之表現。實驗結果顯示，在分化的C2C12肌肉細胞，槲皮素可增加粒線體DNA (mitochondrial DNA, mtDNA) 的套數、增加PGC-1α及Tfam (mitochondrial transcription factor A)之mRNA表現，且投以24小時之25 µM槲皮素可提高粒線體電子傳遞鏈酵素SCCR (succinate- cytochrome c reductase)活性、粒線體膜電位、粒線體產生ATP之氧消耗率，且增加細胞內ATP的產生。由上述實驗數據推論，榭皮素可經由促進粒線體生合成且增加粒線體電子傳遞鏈SCCR活性，進而增加細胞內ATP產生。隨著槲皮素投以的劑量增加，Sirt1蛋白表現量有增加的趨勢，然而槲皮素影響粒線體生合成是否藉由AMPK-SIRT1-PGC-1α路徑來調控，仍需再進一步研究。

關鍵字

槲皮素 ； C2C12肌纖維細胞 ； 粒線體

並列摘要

The increasing number of aging population has implications for the occurrence of chronic diseases in the elderly. Studies have shown that people with Type 2 DM have impaired mitochondrial function, primary because of the alteration in muscle aging and functional decline. Quercetin, the flavonoid molecules, are widely present in foods. Previous studies has shown that quercetin have benefits on muscle fitness and mitochondrial biogenesis. The main goal of this work was to investigate effects of quercetin on mitochondrial biogenesis and the regulation mechanism of AMPK (AMP-activated protein kinas) -SIRT1 (sirtuin 1) -PGC1α (peroxisome proliferator-activated receptor γ coactivator-1α) pathway in C2C12 myotubes.We analyzed cell viability after treatment with quercetin for 24 hr using neutral red assay. We also evaluated intracellular ATP production, celluar oxygen respiration, mtochondrial membrane potential and mitochondrial enyzyme activity. Expression of mitochondrial biogenesis related genes were determined using real-time RT-PCR and western blot.The results showed that quercetin significantly increase mtDNA copy number, mRNA expression of PGC-1α and Tfam (mitochondrial transcription factor A). After 25 µM quercetin treatment for 24hr would enhanced mitochondrial succinate- cytochrome c reductase activity, mitochondrial membrane potential, cellular xygen respiration and ATP production.In conclusion, quercetin increases mitochondrial biogenesis and succinate- cytochrome c reductase activity, thereby increasing the production of ATP by increasing the number of electron entering the electron transport chain.

並列關鍵字

quercetin ； C2C12 myotube ； mitochondria

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