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研究生:郭曉平
研究生(外文):KUO, HSIAO-PING
論文名稱:異硫氰酸烯丙酯對HeLa細胞有絲分裂的干擾
論文名稱(外文):Disruption of mitosis by Allyl isothiocyanates in HeLa cells
指導教授:張芝瑞
指導教授(外文):CHANG, CHIH-JUI
口試委員:蘇淑惠陳紀雄
口試委員(外文):SU, SHU-HUICHEN, JI-HSHIUNG
口試日期:2022-03-18
學位類別:碩士
校院名稱:慈濟大學
系所名稱:分子生物暨人類遺傳學系碩士班
學門:生命科學學門
學類:生物科技學類
論文種類:學術論文
論文出版年:2022
畢業學年度:110
語文別:中文
論文頁數:42
中文關鍵詞:有絲分裂異硫氰酸烯丙酯
外文關鍵詞:mitosisAllyl isothiocyanates
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在許多人的觀念裡,一旦罹患癌症便等同患上絕症,過去三十九年來,惡性腫瘤位居台灣十大死因之首,隨著醫療技術的進步,治療手段日新月異,但化學療法的副作用問題依舊存在。治療的方向漸漸轉向飲食科學,例如十字花科植物被廣泛認為可以降低罹患癌症風險,其中含有的天然小分子異硫氰酸酯,經研究發現具有抗癌潛力。異硫氰酸酯的衍伸分子異硫氰酸烯丙酯存在於山葵、白蘿蔔、芥末等植物中,為無色、氣味刺鼻的油狀液體,在研究中展現出抑制癌細胞生長、抗發炎、抗血管新生、促進細胞凋亡的潛力。我們的研究發現異硫氰酸烯丙酯會抑制HeLa細胞的生長,並觀察到在細胞有絲分裂的過程中紡錘絲出現異常,產生multipolar和monopolar spindles的現象。中心體在有絲分裂的時候形成微管組織中心,負責調控微管的排列,呈現紡錘狀的微管便是紡錘絲,紡錘絲結構的異常可能觸發有絲分裂檢查點,誘導細胞週期停滯。極光激酶A是負責調控中心體成熟的蛋白質,需要磷酸化才具備活性。我們發現了一個可能導致M phase停滯的未發表途徑,經過異硫氰酸烯丙酯處理後,HeLa細胞的極光激酶A磷酸化程度有降低的趨勢。結合實驗的結果,我們確認異硫氰酸烯丙酯會減少極光激酶A的磷酸化程度,這導致紡錘絲的結構異常,最終因細胞停滯於有絲分裂檢查點,達到抑制HeLa細胞生長的效果。 
People always perceive cancer as a terminal illness. Over the past thirty-nine years, malignant tumors death is the first rank among the top ten causes of death in Taiwan. With the growth of scientific research, the treatments have developed rapidly, but the side effects of chemotherapy still exist. The direction of treatment is gradually turning to diet science, such as people generally believes that cruciferous plants can reduce the risk of cancer. Natural small molecule isothiocyanates contained in cruciferous plants have anticancer potential. The derivative molecule, allyl isothiocyanate, is a colorless, pungent oily liquid found in horseradish, white radish, and mustard. In studies, this molecule shows the potential of inhibiting cancer cell growth, anti-inflammation, anti-angiogenesis and pro-apoptosis. In our study, we discovered that allyl isothiocyanate restrained the growth of HeLa cells, multipolar and monopolar spindles were observed during mitosis. The abnormal spindle structure may initiate spindle assembly checkpoint, induced cell cycle arrest. Centrosome forms microtubule-organization center during mitosis that regulating the arrangement of microtubules. The spindle-shaped microtubules is called spindles. Aurora kinase A, a protein responsible for regulating centrosome maturation, requires phosphorylation to become active. Our research identified an unpublished pathway that may lead to M phase arrest. Phosphorylation of Aurora A kinase tended to decrease in HeLa cells following treatment with allyl isothiocyanate, which may account for the phenotype. In conclusion, we confirmed that allyl isothiocyanate reduced the phosphorylation of Aurora A kinase, the abnormal spindle structure initiates spindle assembly checkpoint, induced cell cycle arrest, and finally inhibiting the growth of HeLa cells.
1. 前言 6
1.1 現今的癌症治療 6
1.2異硫氰酸烯丙酯 (Allyl isothiocyanate, AITC) 7
1.3 細胞週期 (cell cycle) 10
1.4 有絲分裂檢查點 (spindle assembly checkpoint) 12
1.5 極光激酶 A (Aurora A kinase,Aurora A) 13
2. 材料與方法 15
2.1化學分子 (Chemical compound) 15
2.2 細胞株及培養 (Cell line and culture) 15
2.3 MTT試驗 (MTT assay) 16
2.4 免疫螢光染色 (Immunofluorescence) 16
2.5 有絲分裂指數 (Mitotic index) 17
2.6 西方墨點法 (Western blot) 18
3. 結果 22
3.1 AITC對細胞生長的抑制效果 22
3.2 AITC對有絲分裂的影響 23
3.3 AITC影響細胞有絲分裂的細胞型態 24
3.4 AITC誘使metaphase發生異常 25
3.5 AITC對細胞在有絲分裂過程中之蛋白質影響 26
4. 結論 28
5. 討論 29
參考文獻 40


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