- 學位論文
白尾蜈蚣草水萃物誘導人類鼻咽癌及咽癌細胞株細胞週期G2/M停滯及生長抑制之探討
Induction of G2/M Arrest and Growth Inhibition by Aqueous Extract of Ajuga bracteosa (AEAB) in Human Nasopharyngeal and Pharyngeal Carcinoma Cell Lines
摘要
惡性腫瘤是台灣十大死因的第一名，可見癌症的預防與治療重要性與日俱增。近年來許多研究發現天然植物或食療性素材中具有生物活性的成分同時也有較強的化學預防性(chemopreventive)功效，可能具有協助癌症化學治療的作用。 白尾蜈蚣草(Ajuga bracteosa)遍布於東南亞，可用來治療許多疾病包括:肝硬化、喉痛、鼻咽癌、關節炎等，早期在印度將其作為抗瘧疾用藥，但其抗腫瘤功效仍較少被科學研究證實。在本研究中利用細胞體外實驗探討白尾蜈蚣草水萃物(AEAB)對人類鼻咽和咽癌細胞株的抗腫瘤效果，分別使用人類鼻咽癌細胞(Hone-1)和咽癌細胞(Detroit 562)來進行分析。 首先利用MTT assay進行細胞存活率測試，經AEAB處理Hone-1和Detroit562細胞株24至72小時後，觀察到明顯的劑量與時間依賴性的細胞活性抑制作用;利用流式細胞儀配合Propidium Iodide 染色以及Annexin V 加上 Propidium Iodide 雙染色觀察經AEAB處理後細胞週期的變化和細胞是否進行細胞凋亡，實驗結果發現經AEAB處理後造成細胞週期G2/M phase停滯且AEAB不會造成Hone-1和Detroit562細胞株的細胞凋亡與壞死的作用。並以MPM-2-FITC偵測經AEAB處理Hone-1和Detroit562細胞株的有絲分裂期的變化量，發現隨著劑量的增加，有絲分裂期停滯而造成有絲分裂期的變化量有增加的趨勢，意謂著調控G2/M相關蛋白質有受到AEAB的影響。利用西方墨點法和Q-PCR分析G2/M期相關蛋白質”CyclinB1及Cdk1”表現量及其mRNA基因表現，發現在Hone-1及Detroit562細胞株都有下降的趨勢。 本研究利用白尾蜈蚣草作為鼻咽癌及咽癌治療(或輔助化學治療)用藥物，並且同時深入探討其對於人類鼻咽癌及咽癌之抑癌作用機轉，可以提供鼻咽癌及咽癌治療及預防一個新的治療契機。
並列摘要
Malignant tumor is leading mortality in Taiwan, therefore the prevention and therapy of cancer is more important now. In recent years, many studies have found that natural plant materials or biologically active ingredients has strong chemopreventive effects and may play an important role in chemotherapy . Ajuga bracteosa (Labiatae) is a common wild herb growing in open fields throughout Asia. The entire plant has been used to treat various diseases, including liver cirrhosis, sore throat, nasopharyngeal carcinoma, and arthritis. Traditionally, it was used to treat malaria in India. But the anti-tumor activities of Ajuga bracteosa were still unclear. In the study, we have investigated the anti-tumor effect of the aqueous extract of Ajuga bracteosa (AEAB) in human pharyngeal and nasopharyngeal carcinoma cell lines (Hone-1 and Detroit 562 cells) . Our results show that cell viability of Hone-1 and Detroit562 were decreased by treated with AEAB for 24 to 72 hours by the MTT assay. It’s in a dose-dependent and time course manner. We also used the flow cytometry with Propidium Iodide and Annexin V plus Propidium Iodide double staining assay to see whether the growth inhibition or apoptosis/necrosis induced by AEAB. The Hone-1 and Detroit562 cells treated with AEAB will go to cell cycle G2/M phase arrest but not apoptosis/necrosis. The mitotic index (MPM-2) detected by flow cytometry was elevated in both cell lines after treatment with AEAB. The elevated mitotic index meant that the cell cycle G2/M-related proteins were influenced by AEAB. We used Western blot and Q-PCR to detect the cell cycle G2/M-related proteins” Cyclin B1 and Cdk1” and its mRNA expressions. The results showed that the AEAB could induce Cyclin B1 and Cdk1 proteins and mRNA decrease in both cell lines. In this study, Ajuga bracteosa may be an effective treatment of nasopharyngeal carcinoma and pharyngeal carcinoma (or adjuvant chemotherapy), and simultaneously we discussed the tumor suppressor role of the human nasopharyngeal carcinoma and pharyngeal carcinoma to provide a new direction of cancer therapy and prevention.
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