綠茶唲茶素對前脂肪細胞內活性氧及榖胱甘?的影響; Effects of green tea catechins on reactive oxygen species and glutathione in 3T3-L1 preadipocytes

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題名:	綠茶唲茶素對前脂肪細胞內活性氧及榖胱甘?的影響;Effects of green tea catechins on reactive oxygen species and glutathione in 3T3-L1 preadipocytes
作者:	王姿婷;Chin-Ting Wang
貢獻者:	生命科學研究所
關鍵詞:	活性氧;氧化壓力;榖胱甘肽;1-Methyl-3-isobutylxanthine;N-acetyl-L-cysteine.Phosphate buffered saline;Reactive oxygen species;L-buthionine sulfoximine;Bovine serum albumin;Epigallocatechin;Epigallocatechin gallate;Fetal bovine serum;Glutathione;Insulin;Dexamethasone;Epicatechin;Epicatechin gallate
日期:	2007-07-12
上傳時間:	2009-09-22 10:20:18 (UTC+8)
出版者:	國立中央大學圖書館
摘要:	當活性氧 (ROS) 的生成和抗氧化防禦系統之間失去了平衡時，過多的活性氧會使細胞產生氧化壓力。有趣的是，多數多酚類植物同時擁有antioxidant以及prooxidant兩種特性。過去的研究指出綠茶茶多酚，例如EGCG，在細胞培養狀態下有能力使過氧化氫這種氧化物產生而改變自由基在細胞內的生成情況。然而，在3T3-L1前脂肪細胞裡，EGCG和氧化壓力之間的分子作用機制仍然不清楚。本研究是使用小白鼠3T3-L1 preadipocytes作為研究細胞株模型。我們發現，當3T3-L1 preadipocytes暴露於高濃度的EGCG，ROS的形成會隨著時間而增加。這結果顯示高劑量的EGCG在3T3-L1 preadipocytes擁有prooxidant的特性。細胞內的穀胱甘肽 (GSH) 被認為是可以用來評估細胞內氧化還原的狀態的一個重要參數。我們也發現EGCG可能減少細胞內穀胱甘肽，而減少的情況是跟時間及劑量有相關性的。為了研究GSH和ROS之間的關聯性，將3T3-L1 preadipocytes暴露在半胱胺酸　(NAC，GSH前驅物)，發現EGCG導致的氧化作用會緩化下來。另外，將3T3-L1 preadipocytes暴露在L-buthionine sulfoximine (BSO，γ-glutamylcysteine合成酶抑制劑)，發現EGCG導致的氧化壓力被強化。除此之外，進一步對綠茶中結構相似的唲茶素 (例如EC，ECG，EGC和EGCG) 作研究，發現其中仍以EGCG具有較明顯引起活性氧生成的能力，顯示綠茶具有特異性的作用效果。這些結果發現EGCG藉由改變細胞內穀胱甘肽的量來引起活性氧的生成。EGCG造成活性氧的生成及穀胱甘肽量的減少的現象同樣可以在3T3-L1 adipocytes中觀察的到。雖然證明了EGCG會造成活性氧的生成及穀胱甘肽量的減少，然而這種現象會被EGCG接受器（即67-kDa的laminin接受器）的抗體給限制住。這樣的結果顯示EGCG對活性氧和穀胱甘肽的影響是可經由67-kDa　laminin接受器的這條途徑。 Oxidative stress represents an imbalance between production of reactive oxygen species (ROS) and the antioxidant defense system. Interestingly, most plant polyphenols were found to possess the antioxidant and prooxidant properties. It is evident that green tea polyphenols, such as EGCG, have the ability to produce H2O2 and alter the free radical generation in the cell culture system. However, the molecular effects and mechanisms underlying EGCG and oxidative stress in the 3T3-L1 preadipocytes are still not clear. The present study was to use murine 3T3-L1 preadipocytes as the research cell model to examine whether EGCG regulates production of ROS and glutathione (GSH) of fat cells. We found that when 3T3-L1 preadipocytes were exposed to high concentrations (50~100 μM) of EGCG, but not to low doses (5~20 μM) of EGCG, in short-term period (8hrs), their ROS production increased with a time-dependent manner. This suggests that high dose of EGCG possesses prooxidant capacity in 3T3-L1 preadipocytes. Also, EGCG could reduce the amount of intracellular GSH in dose- and time- dependent manners. To further examine the relation of GSH to ROS altered by EGCG, exposure of 3T3-L1 preadipocytes to N-acetyl-L-cysteine (NAC, a GSH precursor) blocked the EGCG-induced increases in ROS level and decreases in GSH levels. Moreover, treatment with L-buthionine sulfoximine (BSO, a γ-glutamylcysteine synthetase inhibitor) enhanced the EGCG-induced increases in ROS level and decreases in GSH levels. EGCG was more effective than other structure-related EC, ECG and EGC in changing ROS and GSH levels, suggesting the catechin-specific effect of green tea. These observations suggest the GSH-dependent effect of EGCG on ROS production. Similar changes in the EGCG-induced increases in ROS levels and decreases in GSH levels were observed in 3T3-L1 adipocytes. While EGCG was demonstrated to alter levels of ROS and GSH, signaling was blocked by an EGCG receptor, 67-kDa laminin receptor (67LR) antibody. These results suggest that EGCG exerts its effects on GSH and ROS levels via the 67LR pathway.
顯示於類別:	[生命科學研究所 ] 博碩士論文

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