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研究生:黃怡甄
研究生(外文):HUANG,YI-AHEN
論文名稱:透過 No.7 化合物抑制 PI3K/AKT /mTOR 訊息傳遞路徑誘導前列腺癌細胞死亡
論文名稱(外文):Induction of prostate cancer cell death by No. 7 compound was through inhibition of PI3K/AKT/mTOR signaling pathway
指導教授:洪瑞祥
指導教授(外文):HUNG,JUI-HSIANG
口試委員:蔡婉琪陳品晟
口試委員(外文):TSAI,WAN-CHICHEN,PIN-SHERN
口試日期:2020-07-23
學位類別:碩士
校院名稱:嘉南藥理大學
系所名稱:生物科技系
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:2020
畢業學年度:108
語文別:中文
論文頁數:65
中文關鍵詞:前列腺癌LNCaPPC-3MAP KinasePI3K/AKT/mTOR
外文關鍵詞:Prostate cancerLNCaPPC-3MAP KinasePI3K/AKT/mTOR
相關次數:
  • 被引用被引用:0
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  • 下載下載:15
  • 收藏至我的研究室書目清單書目收藏:0
前列腺癌是目前最常見的癌症之一,也是男性相關癌症死亡率之第二大原因。現今全球前列腺癌的發生率也迅速的在增長當中。因此尋求更有效的新治療藥物是急需的。首先利用5株不同細胞株來篩選 27 個化合物,結果顯示 No.7 的化合物可以專一性的誘導前列腺癌細胞死亡,進一步用 colony formation實驗的結果觀察,結果顯示No.7 化合物可以明顯抑制 LNCaP 及 PC-3 細胞群落的能力,另一方面利用 DAPI 染色實驗的結果觀察經過 No.7 化合物處理後細胞核的情形,觀察到 No.7 化合物可以明顯誘導 LNCaP 及 PC-3 細胞核皺縮的情形。進一步分析 No.7 化合物對 LNCaP 及 PC-3 細胞週期的影響,結果顯示No.7 化合物可以明顯增加G2/M 期的停滯。此外 No.7 化合物可以修飾 LNCaP 及 PC-3 細胞中p38、ERK、JNK、AKT、PTEN、mTOR 及 p70 S6K 蛋白磷酸化的情形。未來會再更進一步探討 No.7 化合物對前列腺癌細胞株 LNCaP 及PC-3的死亡機制,希望這些研究成果將有助於開發治療前列腺癌的潛力藥物。
Prostate cancer is one of the most common cancers and the second leading cause of death in men. The incidence of prostate cancer is also growing rapidly worldwide today. Therefore, the search for new and more effective treatments is urgently needed. First of all, we used 5 different cell lines to screen 27 compounds, the results showed that No.7 compound can be specifically induced the death in LNCaP and PC-3 cell prostate cancer cells. Furthermore, No.7 compound can significantly inhibit the colony formation ability of LNCaP and PC-3 cells. On the other hand, the nucleus of the cell begins to shrink after No.7 compound treatment in LNCaP and PC-3 cells. In cell cycle analysis, No.7 compound significantly increased G2/M phase arrest. In addition, No.7 compound can modification of the phosphorylation status of p38, ERK, JNK, AKT, PTEN, mTOR and p70 S6K proteins in LNCaP and PC-3 cells during No.7 compound treatment. Furthermore, the death mechanisms of No.7 compound to prostate cancer cell lines LNCaP and PC-3 will be further investigated in the future, and these findings will help develop potential drugs for the treatment of prostate cancer.
摘要 I
Abstract II
目錄 IV
第一章 緒論 1
1.1前列腺 1
1.2雄性激素 2
1.3前列腺癌 2
1.4前列腺癌之成因 3
1.5前列腺癌治療 4
1.6 PI3K/AKT/mTOR 路徑對前列腺癌的影響 5
1.7 MAPK Kinase 訊號對前列腺癌的影響 7
1.8細胞週期對前列腺癌的影響 8
1.9研究動機 11
第二章 材料與方法 13
2.1細胞培養 13
2.1.1實驗備製 13
2.1.2細胞培養條件 14
2.1.3細胞繼代培養 14
2.1.4細胞計算 15
2.1.5冷凍細胞 16
2.1.6解凍細胞 16
2.2 西方墨點法 (Western blot) 17
2.2.1 實驗備製 17
2.2.2 細胞內蛋白質萃取 18
2.2.3蛋白質定量 18
2.2.4膠體備製 19
2.2.5蛋白質電泳 20
2.2.6蛋白質轉漬 21
2.2.7免疫墨點法 22
2.3 Colony Formation 23
2.3.1實驗備製 23
2.3.2 Crystal violet 23
2.4 螢光染色 24
2.4.1實驗備製 24
2.4.2 DAPI染色 24
2.5 Flow cytometry 25
2.5.1實驗備製 25
2.5.2細胞週期分析 25
第三章 結果 27
3.1 No.7 化合物對於 LNCaP 與 PC-3 細胞增殖或集落形成能力的影響 27
3.2 No.7 化合物對於 LNCaP 與 PC-3 細胞外型的變化 27
3.3 No.7 化合物對於 LNCaP 與 PC-3 細胞凋亡的影響 28
3.4 No.7 化合物對於 LNCaP 與 PC-3 細胞週期的影響 28
3.5 No.7 化合物對於 LNCaP 與 PC-3 細胞 MAP Kinase 路徑的影響表現 29
3.6 No.7 化合物對於 LNCaP 與 PC-3 細胞 PI3K/AKT/mTOR 路徑的影響表現 30
第四章 討論 31
第五章 結論 35
第六章 參考文獻 36
圖表 47
圖 1. Colony formation 分析 LNCaP 細胞的增殖和集落形成能力。 47
圖 2.Colony formation 分析 PC-3 細胞的增殖和集落形成能力。 48
49
圖 3. LNCaP 細胞外型的變化。 49
圖 4. PC-3細胞外型的變化。 50
圖 5. LNCaP 螢光染色 ( DAPI ) 觀察細胞凋亡。 51
圖 6. PC-3 螢光染色 ( DAPI ) 觀察細胞凋亡。 52
圖7a. 以 Flow cytometry 分析 LNCaP 細胞在 No.7 化合物介導下以時間點方式處理對細胞週期的影響表現。 53
圖7b. LNCaP 細胞經 No.7 化合物處理 0、3、6、12、24hr 的細胞週期統計表。 54
圖8a. 以 Flow cytometry 分析 PC-3 細胞在 No.7 化合物介導下以時間點方式處理對細胞週期的影響表現。 55
圖8b.PC-3 細胞經 No.7 化合物處理 0、3、6、12、24hr 的細胞週期統計表。 56
圖 9. 以 Western blot 分析 LNCaP 細胞在 No.7 化合物誘導下以時間點方式處理對 MAP Kinase 路徑的影響表現。 57
圖 10. 以 Western blot 分析 PC-3 細胞在 No.7 化合物誘導下以時間點方式處理對 MAP Kinase 路徑的影響表現。 58
圖 11. 以 Western blot 分析 LNCaP 細胞在 No.7 化合物誘導下以時間點方式處理對 PI3K/AKT/mTOR 路徑的影響表現。 59
圖 12. 以 Western blot 分析 PC-3 細胞在 No.7 化合物誘導下以時間點方式處理對 PI3K/AKT/mTOR 路徑的影響表現。 60
附錄 61
附錄 1. 61
附錄 2. 62
附錄 3 63
附錄 4 64
附錄 5 65

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