多酚天然物 resveratrol 和 quercetin 增強 cisplatin 毒殺肺癌細胞之能力

Synergisms of naturally-occuring products resveratrol and quercetin on cisplatin -induced cytotoxicity in lung cancer cells

李易親(Yi Chin Lee)

指導教授 ： 李輝

中山醫學大學/醫學院/醫學分子毒理學研究所/碩士(2011年)

https://doi.org/10.6834/CSMU.2011.00052

摘要

Cisplatin 是治療肺癌之主要化療藥物，但其毒性和副作用是造成患者之預後較差的可能原因。因此提高腫瘤對 cisplatin 之敏感性，減少藥物毒性和副作用，是當前重要之研究課題。腫瘤細胞對化療藥物具有不同之敏感性，主要與細胞凋亡相關基因之表現有關。本研究擬比較最常被研究之多酚類－resveratrol (RSV) 和 quercetin (QUE) 是否會增強 cisplatin 毒殺肺癌細胞能力？兩者能力是否不同？RSV 和 QUE 促進 cisplatin 毒殺能力，是否是主要經由改變凋亡或抗凋亡基因之表現所致？本研究選取 A549、H358、TL1 和 TL4 肺癌細胞處理不同濃度之 cisplatin，以 MTT assay 分析 cisplatin 毒殺 50% 肺癌細胞之濃度 (IC50) 分別為 11.1、17.9、21.6 和 26.6 μM。RSV 和 QUE 分別在 20 μM 以及 100 μM 之濃度範圍內，對這四種細胞都僅能低於 20% 的細胞毒性。為了解在此濃度範圍內 RSV 和 QUE 是否會增強 cisplatin 毒殺細胞之能力? 結果發現 20 μM RSV 能增強 A549、TL1 和 TL4 細胞對 cisplatin 之敏感性增強 1.5~2.4 倍。但是 RSV 對 H358 細胞則沒有影響。另外，QUE 僅增強 A549 細胞對 cisplatin 之敏感性 1.6 倍，而對其他三種細胞都沒有增強。因此 RSV 協同增強 cisplatin 毒殺這些肺癌細胞之能力高於 QUE 之能力。西方點墨法的結果發現 QUE 會抑制 A549 細胞之抗凋亡基因－Mcl-1、XIAP、Bcl-2 之表現，同時又會促進凋亡基因－Bak 之表現，但不會改變 H358 細胞這些基因的表現。若以 Bcl-2 類之已知抑制劑－HA14-1 則會顯著促進 cisplatin 毒殺 A549 細胞之能力。這結果顯示 QUE 經由抑制 Mcl-1、XIAP 和 Bcl-2 之表現，是促進 cisplatin 毒殺 A549 細胞之能力之原因。本研究又發現 QUE 會在轉錄層次活化 Bak 基因之表現，對 QUE 增強 cisplatin 毒殺 A549 肺癌細胞可能有些貢獻，但此推測仍須進一步之研究。

關鍵字

肺炎 ； 白藜蘆醇 ； 榭黃素 ； 毒殺能力

並列摘要

Cisplatin is frequently used for lung cancer chemotherapy. However, a high toxicity and side effects of cisplatin result in patients with poor outcome. How to reduce the toxicity and side effects of cisplatin is a very important medical issue. It is well documented that apoptostic and antiapoptotic gene expression are related with the resistance to cisplatin in cancer cells. In the present study, two polyphenols, resveratrol (RSV) and quercetin (QUE) were used to test 1) whether both compounds could synergistically increase cisplatin cytotoxicity to lung cancer cells; 2) which compound could have more effective; and 3) whether the changes of Bcl-2 family and apoptic gene expression by both compounds could be linked with increased cisplatin sensitivity? Four lung cancer cells (A549, H358, TL-1, and TL-4) were enrolled to evaluate their 50% inhibition concentration of cisplatin (IC50) by MTT assay. Data showed that the IC50 value of these four lung cancer cells was 11.1, 17.9, 21.6, and 26.6 μM, respectively. The IC50 was decreased to 8.7, 8.5 and 15.5 μM in A549, TL-1, and TL-4 cells when these cells were treated with RSV, but the change of IC50 was not observed in H358 cells. However, cisplatin sensitivity increased by QUE was only observed in A549 cells (17.7 vs. 11.3 μM), not in the other three cells. These results showed that cisplatin sensitivity increased by RSV is more effective than QUE. To verify which apoptosis-related genes could be linked with the cisplatin sensitivity, A549 and H358 cells were treated with RSV and QUE. Expressions of Mcl-1, XIAP, and Bcl-2 in A549 cells were markedly decreased by QUE in a dose-dependent manner; conversely, QUE elevated Bak expression. The gene expressions in H358 cells were not changed by QUE. As expected, the cisplatin sensitivity in A549 cells was signicantly increased by HA14-1, an inhibitor of Bcl-2 family, suggesting that the decrease of Mcl-1, XIAP, and Bcl-2 expressions by QUE could contribute to increased cisplatin sensitivity. In addition, Bak mRNA and protein expression markedly increased by QUE could partially contribute to cisplatin sensivity in A549 cells. This speculation should be further investigated.

並列關鍵字

lung cancer ； cisplatin ； resveratrol ； quercetin ； cytotoxicity

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