香煙中的尼古丁對於人類乳癌影響之分子機制研究

(1)

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先前有許多文獻指出吸煙是引發肺癌的重要因子之ㄧ (1,2) 。我們在先前 (Tox icology and Applied Pharmacology, 2004) 也發表過尼古丁引發致癌因子的是仰賴肺臟表皮細胞中尼古丁接受體將 NF-Κb 活化後會結合至 cyclin D1 的啟動子位置上，進而達到加速細胞生長且維持細胞存活的目的 (3,4) 。為了深入探討吸煙習慣和乳癌發生率的相關性 (5-7) ，因此我們認為在乳癌的發生部位應該可以找到特定尼古丁接受體 (nAchR) 。在本實驗中我們在臺灣女性乳癌病人的腫瘤中找到特定的尼古丁接受體 (nAchR) ，我們也是第一個發現在乳癌細胞株 MCF-7 和 MDA-MB-231 有共同的 α5 、 α9 尼古丁接受體。接著我們收集了四十位女性乳癌病人的正常及腫瘤部位做個別尼古丁在 mRNA 上的表現中發現 nAchR ubtype α9 及 α10 是在正常及腫瘤組織中最常發現的尼古丁接受體。利用即時性 PCR (RealTime-PCR) 定量 mRNA 的表現也發現腫瘤組織中 α 9 尼古丁接受體的表現量有明顯的高於正常組織中 α9 尼古丁接受體的表現量的情況。接著在西方墨點法中也發現經由尼古丁刺激細胞而調控的蛋白最主要是依靠 PI3K/AKT 訊息傳遞路徑。我們也利用了 Si RNA 的技術做出了抑制 α5 和 / 或 α9 尼古丁接受體蛋白表現的 MDA-MB-231 乳癌細胞株，結果發現抑制 α9 尼古丁接受體的細胞生長速率明顯的比未抑制 α9 尼古丁接受體的乳癌細胞慢，以上的結果可能顯示 α9 尼古丁接受體可能在尼古丁所引發的乳癌中扮演非常重要的角色。

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The Studies on the Carcinogen Effect of Nicotine on Human Breast Cancer and its Molecular Mechanism

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Previous study indicated that tobacco-smoking is a well nderstanding carcinogenic factors that promote the

lung cancer formation(1,2). The mechanisms of nicotine-induced carcinogenesis were demonstrated in our

recent report (Toxicology and Applied Pharmacology, 2004, in press) indicated as specific binding of nicot

ine to the nicotinic acetylcholine receptor (nAchRs) in lung epithelial cells by activation of the cyclin D1 p

romoter through the NFkB signaling proteins to induced cell proliferation(3,4). To further investigate the t

obacco smoking habits and its correlations of breast tumor formation(5-7), therefore, we suggested that spe

香煙中的尼古丁對於人類乳癌影響之分子機制研究



Previous study indicated that tobacco-smoking is a well nderstanding carcinogenic factors that promote the

lung cancer formation(1,2). The mechanisms of nicotine-induced carcinogenesis were demonstrated in our

recent report (Toxicology and Applied Pharmacology, 2004, in press) indicated as specific binding of nicot

ine to the nicotinic acetylcholine receptor (nAchRs) in lung epithelial cells by activation of the cyclin D1 p

romoter through the NFkB signaling proteins to induced cell proliferation(3,4). To further investigate the t

obacco smoking habits and its correlations of breast tumor formation(5-7), therefore, we suggested that spe

cific nAchR subunits will be identified in the breast tumor. In this study, our results demonstrated that (nA

chR) was detected in breast tumor tissue in Taiwanese patients. We first demonstrated that the a5 and a9 n

AchRs were detected in both the MCF-7 and MDA-MB-231 breast cancer cell lines. We further study the e

xpression of the nAchR mRNA levels in breast tumor tissues collected from forty cases of breast cancer pa

tients in Taiwan and found that the α9 and α10 subunits of the nAchR was the most prevalence in both nor

mal and tumor tissue. Quantitative assays of the mRNA levels of the nAchRs was measured by Real-time P

CR analysis technique and revealed that expression of the a9-nAchR was higher in the tumor tissue when c

ompared to the normal tissue which dissected form the tumor margin. Western blotting analysis was then p

erformed and demonstrated that the PI3K/Akt-regulated proteins were the major signaling pathway that inv

olved in cell proliferation stimulated by nicotine treatment. Different clones of cell lines that inhibited the a

9 and a5 receptor expression by SiRNA technique was established in the MDAMB 231 cells. We found th 

at cell growth curve was significant inhibited in the a9-siRAN-nAchR cells. Such results implied that the a

9-nAchR may play some important role involved in nicotine-mediated breast tumor carcinogenesis.