臺灣博碩士論文加值系統

膠質細胞瘤屬於腦瘤中病程惡化速度最快且癒後差的類型之一。其中的膠質母細胞瘤是最常見且致死率最高的。在台灣，腦部惡性腫瘤盛行率約每10萬人有5人。據統計，台灣每年新增近600例原發性惡性腦瘤，膠質母細胞瘤佔近40％，發現時多已晚期且存活率低，難以治癒。從天然柑橘類水果中分離得到的類黃酮已被證明可以抑制乳腺癌，結腸癌，胃癌和前列腺癌細胞的生長。然而，尚未有研究探討其是否影響人類膠質母細胞瘤細胞的侵襲和遷移。在本實驗中，我們發現利用不同濃度的柚皮素處理神經膠質母細胞瘤細胞株8901或是8401細胞，能夠抑制癌細胞的侵襲和遷移的作用。此外，藉由酵素活性測試和西方墨點法結果分析，柚皮素隨著濃度增加隨之抑制基質金屬蛋白酶 （MMP-2、MMP-9） 活性以及蛋白表現量。 Naringenin藉由調控 snail 及 slug 路徑誘發上皮間質轉換 （EMT） 。此外，在柚皮素作用下磷酸化 p38及Erk的表現下降。總之，實驗結果可知，柚皮素能夠通過下調MAPK訊號路徑來抑制膠質母細胞瘤細胞遷移。

Gliomas are one of the most malignant and treatment-resistant types of human brain cancer. Glioblastoma, the most common type of glioma, is associated with very poor survival. In Taiwan, the prevalence of brain cancer is about 10 per 100,000 people. In Taiwan, the annual increase of nearly 600 cases of primary malignant brain tumor and glioblastoma accounted for around 40% with more advanced and low survival rate. Naringenin, a flavanone isolated from natural citrus fruits, has been shown to inhibit the growth of breast, colon, gastric and prostate cancer cells. However, there are no reports describing its effects on the invasion and migration of human glioblastoma cell lines. In this report, we found that the treatment of glioblastoma cell 8901 and 8401 lines with different concentrations of naringenin inhibited the invasion and migration of these cells. Furthermore, zymography and Western blot analysis revealed that naringenin downregulated the levels of matrix metalloproteinase-2 and -9 (MMP-2 and MMP-9) expression in a concentration-dependent manner.Naringenin reversed epithelial–mesenchymal transition (EMT) as shown by decreased expression of snail/slug. Moreover, naringenin also decreased the phosphorylation p38, Erk(extracellular signal–regulated kinases). In conclusion, our study shows that naringenin is capable of inhibiting glioblastoma cancer cell migration and invasion through multiple mechanisms including downregulation of MMP-2 and MMP-9 activities, decreased MAPK signaling pathway and reverse of EMT phenomenon.

壹、中文摘要 i
貳、英文摘要 ii
參、縮寫表 iii
肆、緒論 1
一、 腦癌 1
二、 膠質母細胞瘤(GBM) 2
三、 癌症的侵襲與轉移 4
四、 基質金屬蛋白酶 5
五、 上皮細胞間質轉化機制 7
六、 柚皮素 8
七、 柚皮素與癌症關係 9
伍、實驗動機 12
陸、實驗材料與設備 13
一、 藥品試劑 13
二、 儀器設備 16
柒、實驗方法 18
一、 細胞培養 18
二、 細胞存活率分析 (MTT assay) 20
三、 水解蛋白酵素分析 (Zymography) 21
四、 細胞移動性分析 (migration assay) 22
五、 細胞侵襲性分析 23
六、 細胞蛋白質製備 23
七、 西方墨點法 25
捌、實驗結果 28
一、 柚皮素對 GBM 存活率的影響 28
二、 柚皮素對 GBM 細胞週期的影響 28
三、 柚皮素對 GBM 侵襲和移動能力之分析 29
四、 柚皮素對 MMP-2 及 MMP-9 的活性影響 31
五、 柚皮素對 Erk1/2、p38 訊號傳遞路徑之影響 31
六、 柚皮素對 EMT轉錄因子的影響 32
玖、討論 33
壹拾、參考文獻 36
壹拾壹、圖表與圖表說明 42

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