| Abstract: | 13
中文摘要：根據行政院衛生署民國100年統計指出，子宮頸癌在台灣女性癌症死亡率位居第七名。經研究指出99.7 % 的子宮頸癌，都是感染人類乳突病毒 (Human Papillomavirus，HPV) 所導致。而主要造成子宮頸癌的HPV病毒又以16、18型最多，佔70％。零期的子宮頸癌在臨床上沒有特殊的自覺症狀，絕大多數都是在子宮頸抹片檢查意外地被發現。子宮頸癌最常見的症狀是不正常的陰道出血。而造成子宮頸癌病患死亡的主因在於癌症轉移 (metastasis) 。在癌轉移中，腫瘤會釋放出趨化因子、生長因子及基質金屬蛋白水解? (Matrix metalloproteinase, MMP) 來分解細胞外基質 (extracellular matrix, ECM) 而形成血管新生，腫瘤因此能藉由血液、淋巴液的流動進轉移到其他組織。欖仁樹葉 (Terminalia catappa leaves extracts)，有研究指出其主要的成分為 punicalagin 及 punicalin，此?種成分被發現具有很強的藥?作用，包括預防及治?肝炎、?死肝癌細胞、清除自由基、抗氧化、用於預防及治?皮膚發炎、抗細胞突變、?血?活性等功用。但對於欖仁樹葉抑制子宮頸癌其轉移與侵襲的詳細機轉尚未釐清。因此本篇研究探討欖仁樹葉在子宮頸癌中所扮演的角色及詳細機制為何。12- O-tetradecanoyl-phorbol-13-acetate (TPA)，又稱為PMA，是腫瘤促進因子。有研究指出TPA會誘發腫瘤細胞中的MMP-9，使細胞轉移更為明顯。因此在本實驗中，我們使用TPA當作誘發因子，並同時以50%酒精萃取的欖仁樹葉萃取物來處理HeLa細胞株，首先利用MTT assay分析方法發現欖仁樹葉萃取物並不會影響HeLa細胞株的存活率。接著利用gelatin zymography assay，發現欖仁樹葉萃取物會抑制HeLa細胞中TPA所誘發的MMP-9蛋白活性。接著我們在Boyden chamber assay中觀察到欖仁樹葉萃取物明顯抑制由TPA所誘發HeLa細胞的轉移及侵襲能力。因此我們推測，欖仁樹葉萃取物可能藉由調控MMP-9的基因活性來抑制子宮頸癌細胞的轉移及侵襲。我們進一步以 Western blot assay發現TPA所誘發的HeLa細胞，其MMP-9活性的調控是經由磷酸化ERK路徑。總結以上結果，我們發現在子宮頸癌細胞HeLa中，欖仁樹葉萃取物透過抑制TPA所誘發的MMP-9表現，使癌細胞轉移能力降低，但其上游訊息傳遞路徑有待進一步實驗確認。
According to Department of Health, Executive Yuan statistics, cervical cancer ranked seventh in cancer mortality of women. Previous studies showed that 99.7% of cervical cancers are caused by infection with Human Papillomavirus ( HPV). The subtype of 16 and 18 are the major cause of cervical cancer. HPV 16 and 18 lead to about 70% cervical cancer. Cervical cancer patients of clinical stage 0 have no particular symptoms and were almost found by the Pap smear accidentally. The most common symptom of cervical cancer is abnormal vaginal bleeding. Cervical cancer patients died primarily because of cancer metastasis. During cancer metastasis, the extracellular matrix (ECM) was decomposed by cancer-released chemokines, growth factors and matrix metalloproteinases (MMPs), and further promote cancer angiogenesis and distal metastasis through the blood flow. Studies have showed that the main compositions of Terminalia catappa leaves extracts were punicalagin and punicalin. The two compositions were discovered that have strong pharmacological effect, including prevention and treatment of hepatitis, killing liver cancer cells, removing free radical, antioxidant, treating inflammation of the skin, anti-cell mutation, antihyperglycemic activity and other functions. However, the anti-cancer effects and underlying mechanisms of this medicine on cervical cancer cell were still unclear. 12-O-tetradecanoyl -phorbol-13-acetate (TPA) is also called PMA, it is a tumor promoter. Some studies showed that TPA may induce MMP-9 expression in tumor cell and then increasing the metastatic ability. In this study, HeLa cells were treated with TPA and combined with or without Terminalia catappa leaves extracts extracted by 50% alcohol. At first, we found that the concentration of Terminalia catappa leaves extracts we used here did not affect cell viability of HeLa cells. The non-toxic Terminalia catappa leaves extracts also inhibits TPA-induced metastasis and invasion, as shown by results of the Boyden chamber assay. Therefore, we speculated that Terminalia catappa leaves extracts may inhibit metastasis and invasion of cervical cancer by regulation of MMP-9 enzyme activity. Results from western blot showed that Terminalia catappa leaves extracts also inhibits TPA-induced phosphorylation of ERK. In conclusion, We found that TCE might inhibit TPA-induced invasion of HeLa cells through modulating MMP-9 activity ,but the detail underlying mechanisms remains to be confirmed in the future work. |
