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研究生:王贈惟
研究生(外文):Zeng-Wei
論文名稱:松萃取物抑制人類前骨髓性白血病細胞HL-60細胞增生並促進細胞凋亡之研究
論文名稱(外文):Pine extracts inhibit cell proliferation and promote apoptosis of human promyelocytic HL-60 cells
指導教授:劉光耀劉光耀引用關係
指導教授(外文):Guang-Yaw Liu
學位類別:碩士
校院名稱:中山醫學大學
系所名稱:免疫學研究所
學門:醫藥衛生學門
學類:醫學學類
論文種類:學術論文
論文出版年:2010
畢業學年度:98
語文別:英文
論文頁數:46
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在中國的古典書籍中,「神農本草經」和「本草綱目」描述能維持生命有助於長壽者是「上藥」,像是松、木、草,在「上藥」中,最常被提到的就是天然植物藥草「松」。而台灣五葉松(Pinus morrisonicola Hayata)是臺灣唯有、本土特殊植物五葉松(白松)。在先前研究文獻指出松萃取物的許多生物活性且具有治療、預防疾病效應,其中也包括了癌症的治療。但其中台灣五葉松的相關研究卻不多,而對於台灣五葉松的生物活性效應至今也尚未被明確闡明。故本研究是想了解台灣五葉松萃取物的抑制癌細胞增生與促進細胞凋亡之影響,並找出台灣五葉松萃取物中有效的活性成份。研究結果顯示台灣五葉松萃取物、水相萃取物和其純化成份pinocembrin,tiliroside和chrysin皆有抑制癌細胞生長和促進細胞凋亡的作用。此外,我們的結果也指出台灣五葉松萃取物(500 μg/ml)對於正常人類周邊血液單核球細胞與斑馬魚胚胎並無毒性作用。總括來說,我們發現台灣五葉松萃取物對於抑制癌細胞生長具有顯著的效應,並且台灣五葉松萃取藉由調節細胞週期和細胞凋亡的調控蛋白來誘導白血病癌細胞走向細胞凋亡路徑。本研究結果建議,台灣五葉松萃取物是一種具有抑制血癌細胞生長並促使血癌細胞走向細胞凋亡的天然物,未來台灣五葉松萃取物也許可提供做為新的癌症預防或癌症治療的輔助劑。

In the ancient China’s books, [Shen Nong Ben Cao Jing] and [Bencao Gangmu], mention that plants such as pines, woods and grasses are the best source of drugs for lengthen human life and to be the health foods. People have all along been seeking to prolong life. Pinus morrisonicola Hayata is one of original coniferous species in Taiwan. Many previous studies indicate pine’s physiological activities and therapeutic effects, including as a remedy for carcinoma. But there are few reports regarding the biological effects of Pinus morrisonicola Hayata have been found so far. The objective of this study is to investigate the effects on inhibiting cell proliferation and promoting apoptosis of Pinus morrisonicola Hayata extracts and its active compounds. Results from this study indicated that Pinus morrisonicola Hayata extract, the partition of water phase and its compounds, pinocembrin, tiliroside, and chrysin, inhibiting cancer cell growth and promoting apoptosis. In addition, our data showed that Pinus morrisonicola Hayata extract at 500 μg/ml did not have significant cytotoxicity on peripheral blood mononuclear cells and zebrafish embryos. In conclusion, we have shown that Pinus morrisonicola Hayata extract exerts a significant effect on inhibition of cancer cell growth and induction of apoptosis in leukemia cancer cells mediated by cell cycle and apoptosis regulatory proteins. These data suggest that Pinus morrisonicola Hayata extract, as natural substances with powerful growth inhibition and induce-apoptosis effects on leukemia cells, will be a good candidate for chemoprevention or chemotherapeutic adjuvant in the future.

目錄(Index)
目錄……………………………………………………………………I
圖表目錄………………………………………………………………IV
中文摘要………………………………………………………………VI
Abstract ……………………………………………………………VII
前言(Introduction)…………………………………………………1
松的介紹…………………………………………………………………1
台灣五葉松(Pinus morrisonicola Hayata)的介紹………………1
松樹(屬)之植物純化成份……………………………………………2
松萃取物抑制腫瘤及其生物功能之研究………………………………3
台灣五葉松及其生物活性………………………………………………4
白血病簡介………………………………………………………………4
細胞週期…………………………………………………………………5
細胞凋亡…………………………………………………………………6
研究動機與目的…………………………………………………………6
研究流程圖(Research flow chart)…………………………………7
材料與方法(Materials and methods)………………………………8
Chemicals and cell culture…………………………………………8
The extract purification of Pinus morrisonicola Hayata……………………8
DNA fragmentation analysis…………………………………………8
Cell viability and acridine orange staining assay……………9
MTT assays………………………………………………………………9
Apoptotic assay by flow cytometer…………………………………9
Immunoblotting…………………………………………………………10
The analysis of high-performance liquid chromatography (HPLC) ……11
Zebrafish strains and collection of eggs………………………11
Statistical analysis…………………………………………………11
結果(Results) ………………………………………………………12
Effects of Pinus morrisonicola Hayata extract and its compounds on cell population growth……………………………12
The cell cycle effect of Pinus morrisonicola Hayata extract treated in HL-60 cells………………………………………………12
Pinus morrisonicola Hayata extract induces apoptotic cell death…………13
Pinus morrisonicola Hayata extract induces apoptosis via intrinsically mediated pathway……………………………………13
Effects of water-phase partition from Pinus morrisonicola Hayata extract induce apoptotic cell death……………………14
Effects of compounds from Pinus morrisonicola Hayata extract on cell population growth………………………………14
Chrysin induces apoptotic cell death……………………………15
Chrysin induces apoptosis via intrinsically mediated pathway……………15
The cell cycle effect of chrysin treated in HL-60 cells………………………15
Dihydromyricetin induces apoptotic cell death………………16
Dihydromyricetin induces apoptosis via intrinsically mediated pathway……………………………………………………16
The cell cycle effect of dihydromyricetin treated in HL-60 cells…………17
Effects of Pinus morrisonicola Hayata extract prevents ultraviolet (UV) radiation -induced zebrafish embryos death………………………………17
討論 (Discussion) ……………………………………………………18
圖表(Figures)………………………………………………………21
參考文獻 (References) ………………………………………………37
附錄(Appendixes)……………………………………………………40


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