香煙中的尼古丁對於人類乳癌影響之分子機制研究

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

李嘉華(Chia-Hwa Lee)

指導教授 ： 何元順

臺北醫學大學/醫學院/醫學檢驗生物技術學研究所/碩士(2005年)

摘要

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(Toxicology 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α9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。

關鍵字

即時性 ； 表現量 ； 細胞株 ； 墨點法 ； 發生率

並列摘要

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 nicotine to the nicotinic acetylcholine receptor (nAchRs) in lung epithelial cells by activation of the cyclin D1 promoter through the NFkB signaling proteins to induced cell proliferation(3,4). To further investigate the tobacco smoking habits and its correlations of breast tumor formation(5-7), therefore, we suggested that specific nAchR subunits will be identified in the breast tumor. In this study, our results demonstrated that (nAchR) was detected in breast tumor tissue in Taiwanese patients. We first demonstrated that the a5 and a9 nAchRs were detected in both the MCF-7 and MDA-MB-231 breast cancer cell lines. We further study the expression of the nAchR mRNA levels in breast tumor tissues collected from forty cases of breast cancer patients in Taiwan and found that the α9 and α10 subunits of the nAchR was the most prevalence in both normal and tumor tissue. Quantitative assays of the mRNA levels of the nAchRs was measured by Real-time PCR analysis technique and revealed that expression of the a9-nAchR was higher in the tumor tissue when compared to the normal tissue which dissected form the tumor margin. Western blotting analysis was then performed and demonstrated that the PI3K/Akt-regulated proteins were the major signaling pathway that involved in cell proliferation stimulated by nicotine treatment. Different clones of cell lines that inhibited the a9 and?a5 receptor expression by SiRNA technique was established in the MDAMB 231 cells. We found that cell growth curve was significant inhibited in the a9-siRAN-nAchR cells. Such results implied that the a9-nAchR may play some important role involved in nicotine-mediated breast tumor carcinogenesis.

並列關鍵字

nAchR

參考文獻

1.Proctor, R. N. (2001) Nat Rev Cancer 1(1), 82-86

2.Shields, P. G. (2002) Oncogene 21(45), 6870-6876

5.Schumacher, H. H. (2005) Ann Plast Surg 54(2), 117-119

10.Giovino, G. A. (2002) Oncogene 21(48), 7326-7340

11.Hecht, S. S. (2003) Nat Rev Cancer 3(10), 733-744