探討蛇莓草萃取物對降低非小細胞肺癌H1299遠端轉移能力之機轉

To investigate the effect of Duchesnea indica extracts on inhibition of metastasis in non-small cell lung cancer H1299

周佳德(Jia-De Zhou)

指導教授 ： 謝易修

中山醫學大學/醫學院/生化微生物免疫研究所/碩士(2017年)

摘要

根據衛生福利部統計2016年國人十大死因以慢性疾病為主，死亡率依序為(1)惡性腫瘤(2)心臟疾病(3)腦血管疾病(4)肺炎(5)糖尿病(6)事故傷害(7)慢性下呼吸道疾病(8)高血壓性疾病(9)腎炎、腎病症候群及腎病變(10)慢性肝病及肝硬化。2016年癌症（惡性腫瘤）死亡人數為47,760人，占所有死亡人數的27.7%。肺癌在國人癌症死亡率一直高居不下。肺癌患者的死亡往往伴隨著癌症的轉移，所以癌症的轉移為癌症致死的主因。近年來很多學者研究中草藥，在許多的中草藥中發現，中草藥抗癌的來源幾乎都是來自植物鹼(plant alkaloids)，他們可以抑制有絲分裂作用，而抑制細胞再生所必蛋白合成，植物抗癌藥主要有長春鹼(vincaalkaloid)、紫杉醇(taxol)、喜樹鹼(camptothecin)等，也發現中草藥具有抗轉移、抗血管新生等功能。本實驗利用50%的酒精萃取蛇莓，透過MTT assay發現DIE對H1299肺癌細胞不具有毒殺作用，在wound healing assay、Boyden chamber assay、cell adhesion assay發現H1299處理Duchesnea indica extract (DIE)(0、25、50、75、100 µg/Ml)24小時後，H1299癌細胞移動(migration)能力隨著DIE濃度的上升有明顯被抑制的現象。透過zymography assay DIE 24小時處理之下肺癌H1299細胞MMP-2、u-PA活性有顯著被抑制的現象，所以DIE能抑制金屬基質蛋白酶和尿激酶行血纖維蛋白溶解酶原活化因子，在Western blotting assay發現DIE 24小時處理之下，隨著濃度上升肺癌H1299細胞p-Paxillin Tyr118、p-FAK Tyr397、p-ERK 1、p-ERK 2有隨著濃度上升而有被抑制的現象，Fibronectin、Vimentin、N-cadherin有隨著濃度上升而有被抑制的現象，E-cadherin有隨著濃度上升而有所增加，細胞核內c-Fos、c-Jun有隨著濃度上升而有被抑制的現象。所以利用50%的乙醇萃取之DIE能有效抑制H1299移動及侵襲能力。

關鍵字

蛇莓草 ； 非小細胞肺癌 ； 遠端轉移

並列摘要

According to the Ministry of Health and Welfare statistics in 2016 the top ten causes of chronic diseases, mortality rate (1) malignant neoplasms (2) heart disease (3) cerebrovascular disease (4) pneumonia (5) diabetes (6) accidental injury (7) vhronic lower respiratory tract disease (8) hypertensive disease (9 ) nephritis, nephrotic syndrome and renal disease (10) chronic liver disease and cirrhosis. the number of deaths for cancer in 2016 was 47,760, accounting for 27.7% of all deaths, (1) trachea, bronchus and lung cancer (2) liver and intrahepatic cholangiocarcinoma (3) colon, rectum and anal cancer (4) female breast cancer (5) oral cancer (6) prostate cancer (7) gastric cancer (8) pancreatic cancer (9) esophageal cancer (10) ovary cancer, So lung cancer in rural cancer mortality has been high. Lung cancer deaths are often accompanied by the transfer of cancer, so the transfer of cancer is the main cause of cancer death. in recent years, many scholars have studied Chinese herbal medicine. Among many chinese herbal medicines, chinese herbal medicine has almost all of which are derived from plant alkaloids. They can inhibit mitosis and inhibit the protein synthesis of cell regeneration. Vincaalkaloid, taxol, camptothecin, also found that Chinese herbal medicine with anti-metastasis, anti-angiogenesis and other functions.in this study, 50% alcohol was used to Duchesnea indica extract (DIE). DIE was found to have no toxic effect on H1299 lung cancer cells by MTT assay. H1299 was treated with wound healing assay, Boyden chamber assay and Cell adhesion assay. DIE (0, 25, 50, 75,100 μg / mL) 24 hours later, H1299 cancer cell migration with the increase in DIE concentration was significantly inhibited the phenomenon. The activity of MMP-2 and u-PA in lung cancer H1299 cells was significantly inhibited by 24 hours of zymography assay. Therefore, DIE could inhibit the expression of fibrinolytic enzyme activator of metalloproteinase and urokinase in Western blotting The expression of p-Paxillin Tyr118, p-FAK Tyr397, p-ERK1 and p-ERK2 in lung cancer H1299 cells were inhibited with the increase of the concentration of Fibronectin, Vimentin , N-cadherin has increased with the concentration of inhibition of the phenomenon, E-cadherin with the increase in concentration has increased, the nucleus c-Fos, c-Jun has increased with the concentration of the phenomenon has been suppressed. So the use of 50% alcohol extract DIE can effectively inhibit the H1299 mobile and invasive ability.

並列關鍵字

Duchesnea indica extracts ； metastasis ； non-small cell lung cancer

參考文獻

[7] 陳育民(民100).認識肺癌(第一版)。財團法人台灣癌症臨床研究發展基金會。

[62]黃士漢。(2013)。蛇莓抑制轉型生長因子ß1誘導人類肺腺癌細胞A549間質上皮轉型之機制研究。中山醫學大學生化暨生物科技研究所碩士論文，未出版，臺灣臺中市。

[3] J. Didkowska, U. Wojciechowska, M. Manczuk, J. Lobaszewski, Lung cancer epidemiology: contemporary and future challenges worldwide, Ann Transl Med, 4 (2016) 150.

[4] J. Stojsic, I. Milovanovic, J. Radojicic, B. Milenkovic, Lung cancer in women: histological type and patient age from 1985 to 2005, Med Oncol, 26 (2009) 265-268.

[8] T. Hanagiri, S. Shinohara, M. Takenaka, H. Shimokawa, Y. Nagata, M. Nakagawa, H. Uramoto, T. So, F. Tanaka, Clinical Characteristics of Resected T3 Non-small Cell Lung Cancer Characterized by Parietal Pleural Invasion or Chest Wall Invasion, Indian J Surg, 76 (2014) 354-358.