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完整後設資料紀錄
DC 欄位 | 值 | 語言 |
---|---|---|
dc.contributor.advisor | 顧記華 | |
dc.contributor.author | Pin-Hsuan Lu | en |
dc.contributor.author | 盧品璇 | zh_TW |
dc.date.accessioned | 2021-06-15T06:00:37Z | - |
dc.date.available | 2012-09-13 | |
dc.date.copyright | 2010-09-13 | |
dc.date.issued | 2010 | |
dc.date.submitted | 2010-08-17 | |
dc.identifier.citation | 参考文獻:
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dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/47455 | - |
dc.description.abstract | 轉移性荷爾蒙不依賴型前列腺癌是此癌症的主要死因,所以藥物研發多著重於此。許多抗癌療法的目的,大多是要抑制癌細胞生長或引起細胞凋亡。因此本篇論文以轉移性荷爾蒙不依賴型前列腺癌細胞株PC-3細胞為主,探討了數種具有抑制細胞生長及細胞凋亡的化合物的機轉。
第一個部份我們探討了海洋海綿代謝物ilimaquinone的抑制生長作用。Ilimaquinone對於許多癌細胞株,如前列腺癌細胞PC-3及LNCaP 、非小細胞型肺癌細胞A549及肝癌細胞Hep3B有生長抑制的作用。從流式細胞儀分析細胞週期發現ilimaquinone增加G1期細胞的比例及cyclin E的表現與細胞核的分布,顯示細胞週期停滯於G1/S期。我們的研究排除了ilimaquinone對高基氏體囊泡化及抑制NF-κB的活性和生長抑制作用的關係。同時我們也發現ilimaquinone增加CHOP/GADD153 (growth arrest- and DNA damage-inducible gene 153) 的表現及細胞核的分布。進一步的研究發現CHOP/GADD153的antisense可以明顯減少ilimaquinone所引起的細胞生長抑制,顯示CHOP/GADD153對於生長抑制的重要性。總結以上結果,我們發現ilimaquinone使細胞週期停滯在G1期並且增加CHOP/GADD153的表現及核轉移來達到生長抑制的作用。 接下來,第二部份我們探討G2/M期停滯對於癌細胞生長抑制作用的影響。以小分子化合物K2154 [ethyl-2-[N-ρ-chlorobenzyl-(2′-methoxy)]-anilino-4-oxo-4,5-dihydro-furan-3-carboxylate]作為機轉探討的主角,並且比較其與taxol及vincristine的異同。K2154對於PC-3、Hep3B、A549、HT-29及HCT-116及對taxol與vincristine有抗藥性的NCI/ADR-RES細胞有生長抑制的作用。由流式細胞儀分析發現,K2154使細胞週期停滯在G2/M期,並且伴隨sub-G1期的增加,但是這個現象在正常的前列腺細胞並未被觀察到。在in vitro turbidity assay及in vivo microtubule spin-down assay顯示K2154可以抑制微小管的聚合,並且從免疫螢光染色的實驗可知,K2154會影響紡錘體的形成,並且對於betaII及betaIII微管蛋白的作用特別明顯。從西方墨點法得知K2154增加Cdk1的活性,並且其抑制劑roscovitine可以減少K2154所造成的sub-G1細胞增加,顯示Cdk1在K2154引起的細胞凋亡扮演重要的角色。另外K2154引起Bcl-2及Bcl-xL的磷酸化、Mcl-1的裂解、caspase-9和caspase-3的活化及AIF(apoptosis-inducing factor)的釋放作用,顯示粒線體參與細胞凋亡的過程。綜合以上結果,K2154可作用在微小管並且接著引起有絲分裂停滯及細胞凋亡的訊息來達到其抗癌活性。 雖然在第一部分我們證明了高基氏體在ilimaquinone的抗癌作用中並非扮演重要的角色,但是卻引起我們想要探討此胞器是否與粒線體及內質網同樣參與細胞凋亡過程的興趣。由於高基氏體的形態受到微小管動態平衡的影響以及在有絲分裂的過程中會產生碎裂,因此第三部份我們使用taxol來探討高基氏體在細胞凋亡過程中所扮演的角色。我們發現taxol會讓高基氏體產生碎裂,而這些碎片在細胞內的分布有靠近粒線體的情形。另外,taxol也會活化高基氏體上的caspase-8和caspase-3及增加δCF(catalytic fragment of PKCδ)的表現,顯示高基氏體參與細胞凋亡的過程。由西方墨點法結果得知taxol會增加高基氏體的Cdk1的活性,而且Cdk1抑制劑roscovitine可以抑制高基氏體上caspase-8和caspase-3的活化及δCF的增加,顯示高基氏體的Cdk1扮演細胞凋亡因子的角色。此外,我們也發現PKCδ抑制劑rottlerin可以抑制taxol所引起的細胞凋亡及高基氏體Cdk1的活性,顯示PKCδ為活化Cdk1的上游因子。然而PKCδ在高基氏體上的活性是減少的,進一步的結果顯示PKCδ在細胞質內並無活化的現象,在細胞核內短時間有顯著活化的情形,因此推測高基氏體Cdk1的活性增加可能來自於細胞核內PKCδ活化的結果。綜合以上結果,我們發現PKCδ及其下游高基氏體Cdk1的活化,參與taxol所造成的細胞凋亡。 | zh_TW |
dc.description.abstract | Metastatic hormone resistant prostate cancer is the major cause of death in patients with prostate cancers. Therefore, drug discovery against metastatic prostate cancers is highlighted currently. The goal of anticancer therapy is to inhibit growth of tumor cells as well as to induce cell apoptosis. Accordingly, human hormone-resistant prostate cancer PC-3 cells were used as the model to study the anticancer mechanisms of several compounds in this thesis.
In the first part, we discovered that ilimaquinone, a metabolite of sea sponge, inhibited proliferation of PC-3 cells. Ilimaquinone induced anti-proliferative effect in several types of cancer cell lines, including prostate cancer PC-3 and LNCaP, non-small cell lung cancer A549 and hepatocellular carcinoma Hep3B cells. From cell cycle analysis, the data showed that ilimaquinone increased the population of cells at G1 phase of the cell cycle and increased the expression of cyclin E as well as its nuclear localization, indicating the induction G1-phase arrest. Furthermore, the data ruled out the role of ilimaquinone-induced Golgi vesiculation on antiproliferative effect. Ilimaquinone also inhibited NF-κB binding to DNA. However, the inhibitory effect could not explain ilimaquinone-induced anticancer effect. Ilimaquinone induced the up-regulation and nuclear translocation of growth arrest and DNA damage inducible gene 153 (CHOP/GADD153). Ilimaquinone-mediated anti-proliferative effect was significantly reduced by the transfection of cells with antisense CHOP/GADD153. In summary, it is suggested that ilimaquinone induced the G1 arrest of the cell cycle and anti-proliferative effect through the up-regulation and nuclear translocation of CHOP/GADD153. In the second part, we used K2154 [ethyl-2-[N-ρ-chlorobenzyl-(2′-methoxy)]- anilino-4-oxo-4,5-dihydro-furan-3-carboxylate] as a tool to study the effect of G2/M arrest on anticancer mechanisms. As well, the anticancer effects of taxol and vincristine were compared in this part. K2154 induced anti-proliferative effect on several cancer cell lines, including PC-3, Hep3B, A549, HT-29, HCT-116 and NCI/ADR-RES, a taxol- and vincristine-resisitant cell line. The cytoflowmetric analysis showed that K2154 induced G2/M arrest of the cell cycle and the subsequent increase of sub-G1 population (apoptosis) in PC-3 cells but not normal prostate cells. K2154 inhibited tubulin polymerization in both in vitro turbidity assay and in vivo microtubule spin-down assay. Furthermore, the immunocytochemical examination showed that K2154 disturbed the formation of spindle, in particular, in tubulin isotypes betaII and betaIII. K2154 increased Cdk1 activity. However, roscovitine (a Cdk1 inhibitor) significantly rescued K2154-mediated cell death suggesting the apoptotic role of Cdk1. The Western blot data showed that K2154 increased the phosphorylation of Bcl-2 and Bcl-xL, cleavage of Mcl-1, activation of caspase-9 and -3 and release of apoptosis-inducing factor (AIF) to cytosol, indicating the involvement of mitochondrial apoptosis pathways. In summary, K2154 displayed an anticancer activity through targeting on tubulin/microtubule, leading to mitochondria-involved apoptotic signaling cascades. Although Golgi vesiculation was not involved in ilimaquinone-induced antiproliferative effect, it aroused our interest to investigate whether Golgi complex similar to mitochondria and ER was participated in apoptosis. Furthermore, Golgi complex was fragmented due to microtubule dynamics and during mitosis. Therefore, we used taxol as a tool to study the involvement of Golgi complex in taxol-mediated apoptosis. Taxol dispersed Golgi complex into fragments which translocated to proximity of mitochondria. Furthermore, taxol induced the activation of caspase-8 and -3 and increase of δCF (catalytic fragment of PKCδ) on Golgi complex, suggesting the critical role of Golgi complex. Taxol also increased the activity of Golgi complex-associated Cdk1. Roscovitine significantly inhibited the activation of caspase-8 and -3 and decreased the formation of δCF on Golgi complex, indicating the function involvement of Golgi complex-associated Cdk1 in apoptosis. In addition, rottlerin (a PKCδ inhibitor) was also able to inhibit taxol-induced Cdk1 activation on Golgi complex and apoptosis, suggesting that PKCδ served as an upstream regulator. However, activated PKCδ was not associated with Golgi complex. The activity of PKCδ was decreased on Golgi complex, undetected in cytoplasm and increased in nuclei. Therefore, the activation of Golgi complex-associated Cdk1 may result from nuclear activated PKCδ. In summary, the data suggest that the activation of PKCδ and Golgi complex-associated Cdk1 was involved in taxol-mediated apoptosis in PC-3 cells. | en |
dc.description.provenance | Made available in DSpace on 2021-06-15T06:00:37Z (GMT). No. of bitstreams: 1 ntu-99-F92423002-1.pdf: 3954570 bytes, checksum: 313b54f4a86382a1f4c3a58f02e897ac (MD5) Previous issue date: 2010 | en |
dc.description.tableofcontents | 縮寫表...............................................................1
中文摘要.............................................................2 英文摘要.............................................................4 研究動機與目的.......................................................8 文獻回顧............................................................10 實驗材料與方法 實驗材料........................................................... 18 實驗方法........................................................... 20 第一部份 海綿代謝物ilimaquinone透過CHOP/GADD153來展現其抗癌活性機轉探討 中文摘要...........................................................26 英文摘要...........................................................27 緒論...............................................................29 結果...............................................................30 討論...............................................................33 第二部份 K2154抗癌機轉探討:對於異型微管蛋白、有絲分裂停滯及細胞凋亡的影響 中文摘要...........................................................37 英文摘要...........................................................38 緒論...............................................................40 結果...............................................................41 討論...............................................................44 第三部份 高基氏體上Cdk1的活化在Taxol引起的細胞凋亡角色探討 中文摘要...........................................................48 英文摘要...........................................................49 緒論...............................................................51 結果...............................................................52 討論...............................................................55 總結與展望..........................................................59 參考文獻............................................................61 附圖................................................................75 圖..................................................................79 | |
dc.language.iso | zh-TW | |
dc.title | 探討數種抗癌藥物活性成分在前列腺癌細胞的作用機轉及高基氏體在紫杉醇所引起的凋亡作用的角色 | zh_TW |
dc.title | Study of the mechanisms of several anticancer agents in prostate cancer cells and the role of Golgi complex on taxol-mediated apoptosis | en |
dc.type | Thesis | |
dc.date.schoolyear | 98-2 | |
dc.description.degree | 博士 | |
dc.contributor.oralexamcommittee | 蕭哲志,黃聰龍,楊家榮,孔繁璐 | |
dc.subject.keyword | 前列腺癌,高基氏體,紫杉醇, | zh_TW |
dc.subject.keyword | prostate cancer,Golgi complex,taxol, | en |
dc.relation.page | 103 | |
dc.rights.note | 有償授權 | |
dc.date.accepted | 2010-08-17 | |
dc.contributor.author-college | 醫學院 | zh_TW |
dc.contributor.author-dept | 藥學研究所 | zh_TW |
顯示於系所單位: | 藥學系 |
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