檳榔生物鹼對於前列腺癌細胞生長和轉移 的影響

Effect of BNAs on growth and migration of human prostate cancer cells

鄭靖耀(Ching-Yao Cheng)

指導教授 ： 高永旭

國立中央大學/理學院/生命科學系/碩士(2014年)

摘要

檳榔在亞洲是常見的食品，包括台灣，其中包含四個主要的生物鹼，如
arecoline，arecaidine，guvacoline 和guvacine 。雖然檳榔生物鹼，尤其是arecoline，在大鼠中發現會增加血睾酮水和前列腺生長，但並未報告檳榔生物鹼和人類前列腺癌細胞之間的確切關係。使用正常人類前列腺 (RWPE -1）和癌化（LNCaP- FGC，- 104 -S，- 104 -R 1，- 104 -R 2，和PC-3 ）細胞中，我們觀察到arecoline會顯著降低這些前列腺細胞存活率，呈劑量依賴性和時間依賴性，但arecaidine或guvacine 並不會。流式細胞儀分析顯示，arecoline 會將正常RWPE -1 細胞和PC-3 腫瘤細胞停留在G2 / M 期，並且在G1 期降低細胞週期。然而在LNCaP -FGC ，- 104 -R 1 和- 104 -R 2 前列腺癌細胞的細胞週期，arecoline 誘導增加了G1 期和Sub-G1 期，並在S 期下降細胞週期。在LNCaP-104 -S 前列腺癌細胞的細胞週期arecoline 誘導增加Sub-G1 期的細胞週期。此外，在正常和癌化人類前列腺細胞中arecoline 會誘導生產自由基氧物種（ROS），但guvacine 不會。有趣的是，我們發現arecoline 和guvacine 會呈劑量依賴性顯著降低前列腺癌細胞的細胞轉移，但arecaidine 不會， RWPE-1 細胞不會轉移。雖然在其他細胞的研究中arecoline 被報告為毒蕈鹼性乙酰膽鹼受體（mAChR）的促進劑，所以我們預
先處理mAChR的拮抗劑atropine，但並沒有改變arecoline 所誘導的細胞存活率、ROS 的產生。Atropine 也無法改變PC- 3 和LNCaP 腫瘤細胞的轉移和在RWPE1前列腺上皮細胞中，arecoline 誘導下降細胞存活率和ROS 的產生。然而，N-乙酰半胱氨酸（NAC），穀胱甘肽的活化劑，會阻擋arecoline 所改變的細胞生存率，以及產生ROS 和在正常和癌化前列腺細胞的細胞週期。在人類前列腺癌細胞，NAC 還可以防止的arecoline 抑制的癌細胞的轉移，但不能改變guvacine 抑制的癌細胞的轉移。這些數據表明，arecoline 抑制前列腺癌細胞的存活率會經由穀胱甘肽和ROS 路徑，但和mAChR 的路徑無關。結果推測guvacine 對前列腺癌細胞轉移的作用與穀胱甘肽、ROS 和mAChR 的途徑無關。我們得出結論，檳榔生物鹼對人類前列腺上皮細胞和癌細胞會去調節細胞生長和轉移，其是通過不同的信號轉導分子、結構種類和細胞種類相關。

關鍵字

前列腺癌 ； 檳榔生物鹼 ； 檳榔鹼 ； 氧化自由基 ； 轉移

並列摘要

Betel nut, a common substance consumed in Asia, including Taiwan, contains four major alkaloids, such as arecoline, arecaidine, guvacoline, and guvacine. Although betel nut alkaloids (BNAs), particularly arecoline, were found to increase blood testosterone level and prostate gland growth in rats, a clear relationship between BNAs and human prostate cancer cells was not reported. Using normal(i.e., RWPE-1) and cancerous human prostate(i.e., LNCaP-FGC、-104-S、-104-R1、-104-R2, and PC-3) cells, we observed that arecoline, but not arecaidine or guvacine, significantly reduced cell viability of these prostate cells, in dose-dependent and time-dependent manners. Flow cytometric analysis indicated that arecoline arrest normal RWPE-1 cells and PC-3 cancer cells at G2/M phase of their cell cycle, and it reduced G1 phase of their cell cycle. However, arecoline induced increase of the G1 and Sub-G1 phase and decrease of the S phase in the cell cycle of LNCaP-FGC、-104-R1, and -104-R2 prostate cancer cells. And, arecoline induced increase of the Sub-G1 phase in the cell cycle of LNCaP-104-S prostate cancer cells. In addition, arecoline but not guvacine induced the production of radical oxygen species (ROS) from normal and cancerous human prostate cells. Interestingly, we found that arecoline and guvacine, but not arecaidine, significantly reduced cell migration of prostate cancer cells in dose-dependent manners. RWPE1 cells did not migrate. Although arecoline was reported as a muscarinic acetylcholine receptor (mAChR) agonist from other cell studies, pretreatment with a mAChR antagonist atropine did not alter the arecoline-induced cell viability, ROS production, or migration of PC-3 and LNCaP cancer cells. Atropine neither altered the arecoline induced cell viability and ROS production of RWPE1 prostate cell nor changed the guvacine-induced cell migration of LNCaP and PC-3 cancer cells. However, N-acetyl-cysteine (NAC), a glutathione activator, blocked the arecoline-induced changes in the cell viability, ROS production, and diffirent phase of the cell cycle of normal and cancerous human prostate cells. NAC also prevent the arecoline-inhibited but not guvacine-inhibited migration of human prostate cencer cells. These data suggest that arecoline inhibits cell viability and migration of prostate cancer cells through the glutathione-, and ROS-dependent and mAChR-independent pathways. The results also suggest the glutathione-, ROS-, and mAChR-independent effect of guvacine on the migration of prostate cancer cells. We conclude that BNAs mediate growth and cell migration of human prostate cancer cells through the distinct signaling molecule, structure type-dependent, and cell type-dependent pathway.

並列關鍵字

prostate cancer ； Betel nut alkaloids ； Arecoline ； ROS ； migration

參考文獻

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