肝癌為排名第六名全世界最常見之惡性腫瘤，且根據2012年行政院衛生署統計，肝癌在國人十大癌症死因位居第二位，因此肝癌之治療為不可忽視之重要議題。過去研究中發現銀杏萃取物 EGb761有抗氧化、抑制血小板凝集、抑制癌細胞增殖的特性，其中有效成份銀杏內酯 B 約佔銀杏萃取物 EGb 761 之 0.8%，為 EGb761 中生物活性最高之類萜烯，其有保護心血管、抗癌、抗發炎及抑制血管新生之特性。雖已知 EGb761 有抗癌活性，但鮮少研究探討銀杏萃取物中何種成份具有抗癌作用。血管新生為從原有之血管發展出新血管之過程，研究發現阻礙癌細胞之血流可能有抑制肝癌進展之作用，故本實驗將37隻Sprague-Dawley (SD) 大鼠隨機分為四組：正常組 (給予基本飲食)、負對照組 (誘發肝癌，給予基本飲食)、EGb761組 (基本飲食含100 mg/kg body weight/day EGb761)、銀杏內酯B組 (基本飲食含 1 mg/kg body weight/day 銀杏內酯 B)，給予粉末飼料一週後，以 SD 大鼠肝癌細胞N1-S1誘發其肝癌，實驗期五週後犧牲取其血液、肝臟進行分析。結果顯示，給予銀杏內酯 B 之組別，其肝中介白素-6 顯著降低，並且顯著降低因肝癌血管新生而增加之肝中血管新生相關因子：血管內皮生長因子 (vascular endothelial growth factor, VEGF) 與血小板衍生性生長因子 (platelet derived growth factor, PDGF) 之濃度，除此之外，亦降低血管內皮生長因子接受器 2 (vascular endothelial growth factor receptor 2, VEGFR 2) 蛋白質表現量，推測銀杏內酯 B 可藉由抑制介白素-6 延緩肝癌進展，並且抑制促血管新生因子 VEGF 與 PDGF 及 VEGFR 2蛋白質表現來抑制血管新生，因此，銀杏內酯 B 可能為 EGb761 中具有抗癌活性之成份。

Hepatocellular carcinoma (HCC) is the sixth most common cancers in the world. HCC was the second leading cause of death among cancers for both genders in Taiwan in 2012 reported by the Department of Health. Therefore, the treatment for HCC is one of the important issues in Taiwan. Ginkgo biloba extract (EGb 761) has been reported to possess antioxidant, anti-platelet activation, and anti-proliferation properties. Ginkgolide B (0.8 % of EGb 761) is the most biological active terpene lactone in EGb761 extract, and exhibits the activities of cardiovascular protection, anti-cancer, anti-inflammatory, and anti-angiogenesis. However, very few studies have further determined which component in EGb 761 has anti-HCC activity. Angiogenesis is a process of vascular growth by sprouting of preexisting vessels. Blocking blood supply for tumor cells might be a novel therapeutic strategy to inhibit tumor growth. The study investigated the effects of EGb 761 and ginkgolide B on tumor growth and angiogenesis in rats with orthotopic induced hepatocellular carcinoma. Thirty-seven male Sprague-Dawley rats were randomly divided into four groups: normal (basal diet), negative control (HCC with basal diet), Ginkgo biloba extract (HCC with 100 mg/kg bw/day EGb761), and ginkgolide B (HCC with 1 mg/kg bw/day ginkgolide B) groups. Ginkgo biloba extract and ginkgolide B were orally administered 1 week before orthotopic tumor implantation. Hepatocellular carcinoma in the rats was induced using an orthotopic tumor implantation model after one-week feeding with or without supplementation. After 5-week supplementation, rats were killed and the liver was excised for the histopathological assessment of tissue damage. The results showed that supplementation with ginkgolide B (1 mg/kg bw/day) for 5 weeks significantly reduced elevated interleukin-6 caused by HCC, and significantly decreased hepatic vascular endothelial growth factor, platelet derived growth factor levels , and VEGFR 2 protein expression increased by angiogenesis. In conclusion, ginkgolide B can be considered the anti-angiogenesis agent of EGb761 and delayed the progression of HCC.

目錄
致謝 I
摘要 IV
Abstract VI
縮寫表 VII
圖目次 X
表目次 XII
第一章 緒論 1
第二章 文獻回顧 5
第一節 肝癌介紹及治療方式 5
一、 肝癌介紹 5
二、 肝癌治療 5
第二節 與肝癌進展相關之指標 6
一、 甲型胎兒蛋白 (α-fetoprotein, AFP) 6
二、 介白素-6 (interleukin-6, IL-6) 7
第三節 血管新生與肝癌相關性 10
一、 血管新生形成過程 10
二、 血管新生在腫瘤扮演的角色 14
三、 血管內皮生長因子 (vascular endothelial growth factor, VEGF)與血管內皮生長因子接受器 (vascular endothelial growth factor receptor, VEGFR) 16
四、 血小板衍生性生長因子 ( platelet-derived growth factor, PDGF) 17
第五節 銀杏 18
一、 銀杏 18
二、 銀杏萃取物 (EGb 761) 19
三、 銀杏內酯 B (ginkgolide B) 24
第三章 研究動機與目的 26
第四章 實驗材料與方法 27
第一節 細胞株與細胞培養 27
一、 細胞株與來源 27
二、 細胞培養 27
三、 細胞收集 27
第二節 動物實驗 28
一、 實驗動物 28
二、 實驗分組及飲食 28
第三節 實驗設計 29
第五章 分析項目 34
第一節 肝臟病理切片檢驗 34
第二節 肝功能指數測定 34
第三節 血清中α-feto-protein (AFP)測定 34
第四節 肝中 IL-6 測定 35
第五節 血管新生相關調節因子之蛋白質表現測定 35
一、 肝中 VEGF 與 PDGF 含量分析 35
二、 肝中 VEGFR 2 蛋白質表現量分析 36
第六節 統計分析 39
第六章 結果 40
第一節 體重 40
第二節 肝重與相對肝重 42
第三節 肝功能測定 44
第四節 誘發率 46
第五節 肝臟外觀 48
第六節 肝臟組織染色切片 50
第七節 肝癌指標 52
第八節 血管新生相關因子 56
第七章 討論 62
第一節 以注射肝癌細胞 N1-S1 作為誘發模式 62
第二節 銀杏內酯 B 劑量之討論 64
第三節 IL-6 與肝癌之相關性 65
第四節 銀杏對血管新生之影響 66
第五節 發炎反應與血管新生之相關性 68
第八章 總結 69
第九章 參考文獻 70

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