臺灣博碩士論文加值系統

紫珠早在傳統中國醫療上用以止血化瘀，國外化學成分分析指出紫珠富含黃酮類、萜類和酚類化合物，而在前人研究中發現紫珠具有抗菌、抗腫瘤、抗氧化與抗發炎等活性，但其作用機制都未明。因此本篇利用不同的萃取條件評估紫珠萃取物之生物活性探討。
結果顯示，紫珠萃取物讓腫瘤細胞停滯在G2/M期進而抑制腫瘤細胞的增生，但紫珠萃取物對正常細胞不具有細胞毒性。在腫瘤細胞轉移試驗中發現，紫珠萃取物能有效降低腫瘤細胞經由PMA誘導之基質金屬蛋白酶(MMPs)的表現量。
在抗氧化試驗結果顯示，紫珠萃取物在H2O2誘導之中國倉鼠卵巢上皮細胞CHO-K1中具有降低活性氧ROS的效果。而在抗發炎試驗中，紫珠萃取物能抑制由LPS誘導之老鼠巨噬細胞Raw264.7細胞中NO之表現量。
綜合上述，紫珠萃取物具抑制腫瘤細胞生長、抑制腫瘤細胞轉移、抗氧化及抗發炎等生物活性。

Callicarpa sp. have been used in Chinese traditional medicine for a long time. Although some bioactivities from Callicarpa sp. have been studied, such as antioxidative, anti-bacterial, anti-metastasis and anti-inflammatory activities; unfortunately, there are very few scientific evidences to support their biological functions from Callicarpa sp. Many studies have shown that Callicarpa sp. was rich in terpenoids, flavonoids, and phenolic acid. In order to investigate the bioactivities of Callicarpa sp, the various types of extracts from Callicarpa sp. were evaluated during this study.
The results showed that the extract from Callicarpa sp. can significantly inhibit the growth of carcinoma cells but did not have effect on CHO-K1 cells via MTT assay. Moreover, the growth-inhibition effect on carcinoma cells was arrested in G2/M phase conducted by FACAM.
The effect of anti-metastasis was evaluated by gelatin zymography assay. The results showed that the-extract from Callicarpa sp. could effectively inhibit the matrix metalloproteinases (MMPs) secretions in carcinoma cells
The antioxidant activity assay showed that the extract from Callicarpa sp. decreased ROS level. And in the anti-inflammatory assay, the extract from Callicarpa sp. also decreased NO level.
The above results suggest that the extract from Callicarpa sp. can inhibit carcinoma cell growth, arrest the cell cycle and inhibit MMPs expression. And the extract from Callicarpa sp. also have the antioxidant activity and anti-inflammatory activity by decreasing ROS and NO levels.

中文摘要
英文摘要
第一章 文獻回顧
一、 紫珠
1.1紫珠的分類及特徵
1.2 紫珠的化學組成
1.3 紫珠的生物活性
二、 癌症
2.2 乳癌的治療
2.3 癌細胞的轉移
三、 細胞週期
四、 細胞凋亡
五、 免疫調節
六、 自由基
第二章 研究動機
第三章 實驗架構
第四章 材料與方法
一、 紫珠來源樣品製備
1.1 來源
1.2 樣品製備
二、 細胞株
三、 實驗方法
3.1 細胞存活率試驗( MTT assay )
3.3 細胞凋亡試驗，雙染( PI / Annexin V )
3.4 反轉綠聚合酶鏈鎖反應( PT-PCR )
3.5 西方墨點法(Western blot)
3.6 轉移試驗( Zymorgraphy )
3.7 抗氧化
3.7.1 化學法DPPH
3.7.2 化學法螯合亞鐵測定
3.7.3細胞法( DCFH-DA )
3.8 免疫調節
3.8.1 抗發炎
3.8.2 間接免疫
3.9 統計分析
第五章 實驗結果
一、抑癌試驗
1.1 抑制癌細胞增生
1.2 細胞凋亡變化
1.3 對細胞週期的影響
1.4 細胞週期素之RNA與蛋白質的表現
二、轉移試驗
三、抗氧化試驗
3.1 DPPH
3.2 螯合亞鐵能力
3.3 DCFH-DA
四、免疫調節試驗
4.1 抗發炎
4.2 間接免疫
第六章 討論
一、抑癌試驗
二、轉移試驗
三、抗氧化試驗
四、免疫調節試驗
第七章 結論
第八章 未來計畫
第九章 參考文獻

王本祥，現代中藥藥理學。紫珠：852-854，1997。
王祝年，韓壯，崔海濱，等，裸花紫珠的化學成分。熱帶亞熱帶植物學報，15(4)：359-362，2007。
世界衛生組織，2015。http://www.who.int/mediacentre/factsheets/fs297/en/
乳癌防治基金會，2015。http://www.breastcf.org.tw/index.php/knowledge-base/current-status
高飛鵬，汪豪，葉文才，等，裸花紫珠葉的化學成分。中國藥科大學學報，41(2)：120-123，2010。
馬燕春，張旻，徐文彤，等，裸花紫珠化學成分及細胞毒活性研究。中國中藥雜誌，39(16)，2014。
張宏達，中國紫珠屬植物之研究。植物分類學報，第一卷，2690-312，1951。
孫美，王立波，吳立軍，藥用紫珠屬植物的研究進展。安徽農業科學，Journal of Anhui Agri. Sci. 43(13)：101-104, 2015.
符健，鄺少軼，王世雄，裸花紫珠片的抗菌消炎和止血作用研究。海南大學學報，自然科學版，20(2)：154-157, 2002。
曾乙召，桑黃固態發酵產物之生物活性探討，南臺科技大學生物科技研究所碩士學位論文，2013。
董琳，王金輝，劉明生，裸花紫珠葉中的酚酸類化學成分。瀋陽藥科大學學報，27(4)：290-291，2010。
蔡金平，董琳，關薇薇，劉明生，裸花紫珠的研究進展。現代藥物與臨床，第27卷，第一期，2012。
Ado M.A., Abas F., Leong S.W., Shaari K. Ismail I.S., Ghazali H.M. and Lajis N.H. Chemical constituents and biological activities of Callicarpa maingayi leaves. South African Journal of Botany, 104:98-104, 2016.
Chavez K.J., Garimella S.V. and Lipkowtz S. Triple Negative Breast Cancer Cell Lines: One Tool in the Search for Better Treatment of Triple Negative Breast Cancer. Author Manuscript, 32(1-2):35-48, 2010.
Chen L., Qiu J., Yang C., Yang X., Chen X., Jiang J., and Luo X. Identification of a novel estrogen receptor beta1 binding partner, inhibitor of differentiation-1, and role of ERbeta1 in human breast cancer cells. Cancer Letters, 278(2):210-219, 2009.
Chhipa R.R. and Bhat M.K., Bystander killing of breast cancer MCF-7 cells by MDA-MB-231 cells exposed to 5-fluorouracil is mediated via Fas. Journal of Cellular Biochemistry, 101(1):68-79, 2007.
Chung P.Y., Chung L.Y. and Navaratnam P. Potential targets by pentacyclic triterpenoids from Callicarpa farinose against methicillin-resistant and sensitive Staphylococcus aureus. Fitoterapia, 94:48-54, 2014.
Cui J., Xing L., Li Z., Wu S., Wang J., Liu J., Wang J., Yan X. and Zhang X. Ochratoxin A induces G2 phase arrest in human gastric epithelium GES-1 cells in vitro. Toxicology Letters, 193:152-158, 2010.
Darby S.C., Ewertz M. and Hall P. Ischemic heart disease after breast cancer radiotherapy. New Engl J, 368:2523-2527, 2013.
Dong L., Zhang L., Zhang X., Liu M., Wang J. and Wang Y. Two new 3,4-seco-labdane diterpenoids from Callicarpa nudiflora and their inhibitory activities against nitric oxide production. Phytochemistry Letters, 10:127-131, 2014.
Foulkes W.D., Smith I.E. and Reis-Filho J.S. Triple-Negative Breast Cancer. The new england journal of medicine, 363:1938-48, 2010.
Fu J., Kuang S.T. and Wang S.X. Study on the effects of Callicarpa nudiflora tablets in the antibiosis and antiphlogosis and the hemostasis. Nat Sci J Hainan Univ, 2:154-7, 2002.
Gao F.P., Wang, H., Ye, W.C. and Zhao, S.X. Chemical constituents from the leaves of Callicarpa nudiflora. Zhongguo Yaoke Daxue Xuebao, 41:120-123, 2010.
Ghosh A., Bhowmik A., Bhandary S., Putatunda S., Laskar A., Biswas A., Dolui S., Banerjee B., Khan R., Das N., Chakraborty A., Ghosh M.K. and Sen P.C. Formulation and antitumorigenic activities of nanoencapsulated nifetepimine: A promising approach in treating triple negative breast carcinoma. Nanomedicine: Nanotechnology, Biology, and Medicine, 1973-1985, 2016.
Goodwin C.J., Holt S.J., Downes S. and Marshall N.J. Microculture tetrazolium assays: a comparison between two new tetrazolium salts, XTT and MTS. J Immunol Methods. 179(1):95-103, 1995.
Han K., Dai Y., Zou Z., Fu M., Wang Y. and Zhang Z. Molecular characterization and expression profiles of cdc2 and cyclin B during oogenesis and spermatogenesis in green mud crab (Scylla paramamosain). Comparative Biochemistry and Physiology Part B: Biochemistry and Molecular Biology, 163(3):292-302, 2012.
Hsieh W.T., Huang K.Y., Lin H.Y., and Chung J. G. Physalis angulata induced G2/M phase arrest in human breast cancer cells. Food and Chemical Toxicology 44(7): 974–983, 2006.
Huang X., Liu Y., Lu Y. and Ma C. Anti-inflammatory effects of eugenol on lipopolysaccharide-induced inflammatory reaction in acute lung injury via regulating inflammation and redox status. International Immunopharmacology, 26:265-271, 2015.
Hung W.I., Hsu B.Y., Tung Y.C., Ho C.T. and Hwang L.S. Inhibitory effects of antioxidant vitamins against thiyl radical-induced trans fatty acid formation in PC-12 cells. Journal of Functional Foods, 21:212-222, 2016.
Hwang Y.P., Kim H.G., Choi J.H., Park B.H., Jeong M.H., Jeong T.C. and Jeong H.G. Acteoside inhibits PMA‐induced matrix metalloproteinase‐9 expression via CaMK/ERK‐and JNK/NF‐κB‐dependent signaling. Molecular Nutrition & Food Research, 55(S1):S103-S116, 2011.
Jones W.P. and Kinghorn, A.D. Biologically active natural products of the genus Callicarpa. Current Bioactive Compounds, 4:15–32, 2008.
Kao Y.L., Kuo Y.M., Lee Y.I., Yang S.F., Chen W.R.. and Lee H.J. Apple polyphenol induces cell apoptosis, cell cycle arrest at G2/M phase, and mitotic catastrophe in human bladder transitional carcinoma cells. Journal of Functional Foods, 14:384-394, 2015.
Kim K.N., Heo S.J., Yoon W.J., Kang S.M., Ahn G., Yi T.H., and Jeon T.J. Fucoxanthin inhibits the inflammatory response by suppressing the activation of NF-kappaB and MAPKs in lipopolysaccharide-induced RAW 264.7 macrophages. Eur J Pharmacol, 649:369–75, 2010.
Klein G., Vellenga E, Fraaije M.W., Kamps W.A. and de Bont E.S. The possible role of matrix metalloproteinase MMP-2 and MMP-9 in cancer, e.g. acute leukemia. Crit Rev Oncol Hematol. 50(2):87-100, 2004.
Lim H.K., Choi Y.A., Park W., Lee T., Ryu S.H., Kim S.Y., Kim J.R., Kim J.H. and Baek S.H. Phosphatidic acid regulates systemic inflammatory responses by modulating the Akt-mammalian target of Rapamycin-p70 S6 kinase 1 pathway. The Journal of Biological Chemistry 278 (46): 45117–45127, 2003.
Lin, C. Z., Zhu, C. C., Zhao, Z. X., Li, X. H., Xiong, T. Q., Xia, Y. Y., & Ning, Y. Two new abietane diterpenoids from the caulis and leaves of Callicarpa kochiana. Fitoterapia, 83(1), 1-5, 2012.
Liu Y.W., Cheng Y.B., Liaw C.C., Chen C.H., Guh J.H., Hwang T.L., Tsai J.S., Wang W.B. and Shen Y.C. Bioactive Diterpenes from Callicarpa longissima. Journal of Natural Products, 75(4):689-693, 2012.
Luo Y.H., Zhou Z.Q., Ma S.C. and Fu H.Z. Three new antioxidant furofuran lignans from Callicarpa nudiflora. Phytochemistry Letters, 7:194-197, 2014.
Mei W.L., Han Z., Cui H.B., Zhao Y.X., Deng Y.Y. and Dai H.F. A new cytotoxic iridoid from Callicarpa nudiflora. Nat. Prod. Res, 24.10:899-904, 2010.
Milenic D.E. Monoclonal antibody-based therapy strategies: providing options for the cancer patient, Current pharmaceutical design 8.19:1749–1764, 2002.
Min Y., Sun T., Niu Z. and Liu F. Vitamin C and vitamin E supplementation alleviates oxidative stress induced by dexamethasone and improves fertility of breeder roosters. Animal Reproduction Science, 171:1-6, 2016.
Nicholson D.W. Thornberry N.A. Apoptosis. Life and death decisions. Science, 299(2504):214-215, 2003.
Ono M., Mishima K., Yamasaki T., Masuoka C., Okawa M., Kinjo J., Ikeda T. and Nohara T. A new lignin glucoside from the stems of Callicarpa japonica Thunb. var. luxurians Rehd. J Nat Med. 63:86–90, 2009.
Roskoski R. Jr. Cyclin-dependent protein kinase inhibitors including palbociclib as anticancer drugs. Pharmacological Research, 107:249-275, 2016.
Shin N.R., Shin I.S., Song H.H., Hong J.M., Kwon O.K., Jeon C.M., Kim J.H., Lee S.W., Lee J.K., Jin H., Li W.Y., Oh S.R., Hahn K.W. and Ahn K.S. Callicarpa japonica Thunb. reduces inflammatory responses: A mouse model of lipopolysaccharide-induced acute lung injury. International Immunopharmacology, 26:174-180, 2015.
Sinha S., Khan S., Shukla S., Lakra A.D., Kumar S., Das G., Maurya R. and Meeran S.M. Cucurbitacin B inhibits breast cancer metastasis and angiogenesis through VEGF-mediated suppression of FAK/MMP-9 signaling axis. The International Journal of Biochemistry & Cell Biology, 77:41-56, 2016.
Toyokuni S., Okamoto K., Yodoi J. and Hiai H. Persistent oxidative stress in cancer. FEBS, 358(1):1-3, 1995.
Wang G., Wang W., Zhou J. and Yang X. Correlation between telomerase activity and matrix metalloproteinases 2 expression in gastric cancer. Cancer Biomark, 13(1):21-8, 2013.
Wang J., Tan X., Yang Q., Zeng X., Zhou Y., Luo W., Lin X., Song L. Cai J., Wang T. and Wu X. Inhibition of autophagy promotes apoptosis and enhances anticancer efficacy of adriamycin via augmented ROS generation in prostate cancer cells. The International Journal of Biochemistry & Cell Biology, 77: 80-90, 2016.
Wang Z.N., Han, Z., Cui, H.B. and Dai, H.F. Chemical constituents from Callicarpa nudiflora. J. Trop. Subtrop. 15:359, 2007.
Wang Z.Y. and Li Y. Estrogen receptor alpha-36 (ER-α36): A new player in human breast cancer. Molecular and Cellular Endocrinology, 418:193-206, 2015
Werner F., Jain M.K., Feinberg M. W., Sibinga N.E.S., Pellacani A., Wieseli P., Chin, M.T., Topper J.N., Perrella M.A. and Lee M.E. Transforming growth factor-b1 inhibition of macrophage activation is mediated via Smad3. The Journal of Biological Chemistry 275 (47): 36653–36658, 2000.
Wong T.T., Sethi C, Daniels J.T., Limb G.A., Murphy G. and Khaw P.T. Matrix metalloproteinases in disease and repair processes in the anterior segment. Surv Ophthalmol. 47(3):239-56, 2002.
Wu, H. M. Studies on the Chemical Constituents and Antitubercular Activities from the Leaves and Twigs of Callicarpa pilosissima. 2009.
Xing X., Wang J., Xing L.X., Li Y.H., Yan X. and Zhang X.H. Involvement of MAPK and PI3K signaling pathway in sterigmatocystin-induced G2 phase arrest in human gastric epithelium cells. Mol Nutr Food Res.55(5):749-60, 2011.
Xu, Jing, et al. Diterpenoids from Callicarpa kwangtungensis and their NO inhibitory effects. Fitoterapia, 2016.
Yan K., Zhang C., Feng J., Hou L., Yan L., Zhou Z., Liu Z., Liu C., Fan Y., Zheng B. and Xu Z. Induction of G1 cell cycle arrest and apoptosis by berberine in bladder cancer cells. Eur J Pharmacol. 661(1-3):1-7, 2011.
Zhang L., Liu M.S., Huang J., Li G.Y., Zhang C., Dong L., Zhang K. and Wang J.H. A new 3,4-seco-labdane diterpenoid with potential anti-inflammatory activity from the leaves of Callicarpa nudiflora. J. Asian Nat. Prod, 216-221, 2014.
Zhiqiang Z., Wei X., Fu H. and Luo Y. Chemical constituents of Callicarpa nudiflora and their anti-platelet aggregation activity. Fitoterapia, 88:91-95, 2013.
Zhou, Zhiqiang, et al. Chemical constituents of Callicarpa nudiflora and their anti-platelet aggregation activity. Fitoterapia, 88: 91-95, 2013.