臺灣博碩士論文加值系統

Meclizine (以下簡稱Mec)是一種用來治療頭暈和暈眩的用藥，為抗組織胺藥物的一種。在藥物與細胞的交互作用之中，我們發現Mec會讓人類直腸腫瘤細胞COLO 205誘發細胞凋亡現象。在人類直腸腫瘤細胞COLO 205中，以不同濃度的Mec處理細胞24小時後，觀察到細胞在50mM以上濃度有讓細胞走向細胞凋亡dose-dependent的情形。同時，在顯微鏡之下觀察可以看到隨著Mec藥物濃度增加，細胞數目隨之減少。在流式細胞儀的分析之中，也可以看到處於G0/G1 phase的細胞比例會因著Mec的作用而增加。在以西方墨點法分析各種相關蛋白質的變化之後，關於細胞週期停滯方面我們觀察到p53與p21蛋白在Mec作用之後都會升高，而CDK2和CDK4的活性則受到抑制，這就是Mec造成細胞週期停滯的機轉。在引起細胞凋亡方面，我們看到Mec可以dose-dependently造成以下結果: p53蛋白表現量增加，Bcl-2蛋白表現量減少，Bad蛋白表現量不變，cytochrome C由粒線體釋放到細胞質中，AIF由粒腺體進入細胞核中，Apaf-1蛋白表現量不變，Caspase 9，Caspase 8，Caspase 3活化，PARP被degrade。根據這些結果和文獻的探討，我們提出一個模式圖來解釋Meclizine引起COLO 205細胞走向細胞凋亡和細胞週期停滯的分子機轉。

Meclizine, a kind of histamine H1 antagonist, has been used in the treatment of motion sickness and vertigo. In studying the interaction of drugs and cancer cell lines, we have found that meclizine dose-dependently induced apoptosis in COLO 205 cells. By DNA ladder assay, we demonstrated that DNA ladder appeared with meclizine treatment in COLO 205 cells if dosage larger than 50 μM. Besides, we observed that cell numbers decreased dose-dependently after treatment with meclizine in COLO 205 cells. By flow cytometry, we noticed that the percentage of COLO 205 cells in G0/G1 phase increased dose-dependently. We analyzed the change of associated protein by Western blot. About cell cycle arrest, p53 and p21 were upregulated after treatment with meclizine and resulted in decreasing CDK2 and CDK4 kinase activity. About apoptosis, meclizine induces upregulation of p53, downregulation of Bcl-2, release of cytochrome C into cytosol from mitochondria, translocation of AIF to the nucleus from mitochondria, and activation of caspase 3, caspase 8, and caspase 9. According to these data and concepts from references, we propose a flowchart to explain the possible mechanism of meclizine-induced apoptosis and cell cycle arrest in COLO 205 cell.

目錄 (Contents)
縮寫表 (Abbreviations)…………………………………………………4
中文摘要 (Abstract in Chinese)………………………………………...5
英文摘要 (Abstract in English)…………………………………………6
第一章 背景資料 (Background)……………………………………….7
一、大腸直腸癌在台灣的現狀..…………………………………………..7
二、細胞週期………………………………………………………………..7
(一) 細胞週期的簡介………………………………………….……………7
(二) Cyclins與CDKs的調控…………………………………..……………8
三、細胞凋亡…………………………………………………………………11
(一) 細胞凋亡的簡介………………………………………………………11
(二) 細胞凋亡的特性………………………………………………………11
(三) 細胞凋亡的路徑和調控………………………………………………12
四、Meclizine的簡介…………………………………………………21
第二章 實驗材料與方法 (Materials and Methods)…………….…….23
一、實驗材料…………………………………………………………….…23
(一) 藥品試劑………………………………………………….……………23
(二) 常用溶液………………………………………………………….……26
二、實驗方法……………………………………………………………..…29
(一) 細胞培養………………………………………………………..……29
(二) 細胞生長曲線………………………………………………………..30
(三) 細胞週期同步化與細胞週期分析……………………………………30
(四) DNA裂片分析……………………………………..……………….31
(五) 西方墨點法…………………………………………………………..32
(六) CDK2和CDK4激脢活性試驗……………………………..……….…..35
第三章 結果 (Results)…………………………………..………………37
一. Meclizine可以誘發人類直腸腫瘤細胞(COLO 205)凋亡作用.37
二. Meclizine可以抑制人類直腸腫瘤細胞(COLO 205)生長，但不會抑制人類正常細胞(fibroblast)的生長…………………………………37
三. Meclizine會使人類直腸腫瘤細胞(COLO 205)停滯在細胞週期的G0/G1時期…………………………………………………………………38
四. Meclizine使人類直腸腫瘤細胞(COLO 205)產生細胞週期停滯是經由P53蛋白和P21蛋白升高所造成CDK2和CDK4活性被抑制……………39
五. Meclizine使人類直腸腫瘤細胞(COLO 205)產生細胞凋亡是經由cytochrome C釋放出來而活化Caspase 3………………………………40
第四章 結果討論 (Discussions)……………………………………….42
一、 Meclizine會抑制COLO 205 細胞生長，但對人類fibroblast影響不大…………………………………………………………..…………42
二、 Meclizine造成細胞週期停滯之探討………………………………43
三、 Meclizine造成細胞凋亡之探討……………………………………44
第五章 參考文獻 (References)…………………..……………………..49
第六章 圖表 (Figures)…………………………….………...………….55

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