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研究生(外文):Yi-Yeh Lai
論文名稱(外文):Thymoquinone induces autophagy and apoptosis in oral cancer cells
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百里醌是從果黑種草(Nigella sative)的種籽中萃取出來的化合物,在回教醫學中是一種常用藥材。在最近的研究中指出,百里醌在癌細胞增生、轉移和腫瘤血管新生有抑制的作用。然而,百里醌在頭頸部鱗狀細胞癌(HNSCC)的影響,至今仍不清楚。在本篇研究中,我們證實了百里醌對SASVO3(高度惡化的頭頸部鱗狀癌細胞株,從原位腫瘤連續移植三次)引發了強烈的毒殺作用,隨著處理藥物濃度增加,細胞的毒殺性也隨之增加。SASVO3細胞處理百里醌後,Bax和cleaved-caspase 7及9的蛋白表現量增加,證明百里醌會誘導細胞凋亡。另一方面,利用AVO stain assay,細胞處理百里醌後,觀察到自噬囊泡(autolysosome)的增加,此外,這些細胞自噬的特定標記物, LC3-I、LC3-II蛋白表現量也明顯增加,而Bcl-2,Bid,PARP,Akt和mTOR這些細胞自噬與細胞凋亡相關蛋白的表現量,也受到百里醌處理後下降。綜觀上述研究結果,百里醌透過誘導細胞凋亡和細胞自噬這兩種不同抗腫瘤途徑的機轉引發口腔癌細胞死亡。綜合以上,百里醌誘導口腔癌細胞死亡的機轉,在未來或許可應用在植物化學預防及癌症治療。

Thymoquinone (TQ) is a phytochemical compound extracted from the plant Nigella sative, one in Islamic medicine commonly used herbs. Recent studies reported that TQ exhibited inhibitory effects on cell proliferation, migration, and tumor angiogenesis in various cancer cell lines. However, the effects of TQ on anti-cancer properties in head and neck squamous cell carcinoma (HNSCC) remain unclear. In this study, we demonstrated that the TQ induced a strong cytotoxic effect toward a highly malignant HNSCC cell line, SASVO3, from primary tumors using three sequential rounds of xenotransplantation. The mechanisms of the cytotoxic effect were concentration-dependent. TQ induced apoptotic cell death in SASVO3 cells, as evidenced by increases in the expression of Bax and activation of caspase-7 and -9. On the other hand, these cells showed increased levels of autophagic vacuoles and LC3-I and LC3-II protein expression, specific markers of autophagy. The levels of Bcl-2, Bid, PARP, Akt, and mTOR, which have been implicated in the down-regulation of autophagy and induction of apopotosis, were decreased upon TQ treatment. Taken together, these findings indicate that TQ induced cell death in oral cancer cells via two distinct antineoplastic activities, the ability to induce apoptosis and autophagocytosis, and TQ represent promising candidates for future phytochemical-based mechanistic pathway-targeted cancer prevention strategies.

壹、 中英文摘要…………………………………………...2
貳、 縮寫表………………………………………………...4
參、 緒論…………………………………………………...6
肆、 研究動機…………………………………………….23
伍、 實驗材料與方法…………………………………….24
陸、 實驗結果…………………………………………….39
柒、 討論………………………………………………….48
捌、 參考文獻…………………………………………….52
玖、 圖表與圖表說明…………………………………….59
拾、 實驗流程圖………………………………………….80
拾壹、 附圖表…………………………………………….81

Abdelfadil, E., Y. H. Cheng, D. T. Bau, W. J. Ting, L. M. Chen, H. H. Hsu, Y. M. Lin, R. J. Chen, F. J. Tsai, C. H. Tsai and C. Y. Huang (2013). "Thymoquinone induces apoptosis in oral cancer cells through p38beta inhibition." Am J Chin Med 41(3): 683-696.

Ahmad, I., K. M. Muneer, I. A. Tamimi, M. E. Chang, M. O. Ata and N. Yusuf (2013). "Thymoquinone suppresses metastasis of melanoma cells by inhibition of NLRP3 inflammasome." Toxicol Appl Pharmacol 270(1): 70-76.

Al-Majed, A. A., F. A. Al-Omar and M. N. Nagi (2006). "Neuroprotective effects of thymoquinone against transient forebrain ischemia in the rat hippocampus." Eur J Pharmacol 543(1-3): 40-47.

Aoki, H., E. Iwado, M. S. Eller, Y. Kondo, K. Fujiwara, G. Z. Li, K. R. Hess, D. R. Siwak, R. Sawaya, G. B. Mills, B. A. Gilchrest and S. Kondo (2007). "Telomere 3'' overhang-specific DNA oligonucleotides induce autophagy in malignant glioma cells." FASEB J 21(11): 2918-2930.

Aoki, H., Y. Takada, S. Kondo, R. Sawaya, B. B. Aggarwal and Y. Kondo (2007). "Evidence that curcumin suppresses the growth of malignant gliomas in vitro and in vivo through induction of autophagy: role of Akt and extracellular signal-regulated kinase signaling pathways." Mol Pharmacol 72(1): 29-39.

Ashkenazi, A. and V. M. Dixit (1998). "Death receptors: signaling and modulation." Science 281(5381): 1305-1308.

Attoub, S., O. Sperandio, H. Raza, K. Arafat, S. Al-Salam, M. A. Al Sultan, M. Al Safi, T. Takahashi and A. Adem (2012). "Thymoquinone as an anticancer agent: evidence from inhibition of cancer cells viability and invasion in vitro and tumor growth in vivo." Fundam Clin Pharmacol.
Baehrecke, E. H. (2005). "Autophagy: dual roles in life and death?" Nat Rev Mol Cell Biol 6(6): 505-510.

Bednarek, I., D. Sypniewski, N. Gawlik, A. Galilejczyk and K. Goraus (2012). "The efficiency of silencing expression of the gene coding STAT3 transcriptional factor and susceptibility of bladder cancer cells to apoptosis." Contemp Oncol (Pozn) 16(4): 316-321.

Bhogal, R. H., C. J. Weston, S. M. Curbishley, D. H. Adams and S. C. Afford (2012). "Autophagy: a cyto-protective mechanism which prevents primary human hepatocyte apoptosis during oxidative stress." Autophagy 8(4): 545-558.

Bredesen, D. E., R. V. Rao and P. Mehlen (2006). "Cell death in the nervous system." Nature 443(7113): 796-802.

Chen, Y., E. McMillan-Ward, J. Kong, S. J. Israels and S. B. Gibson (2008). "Oxidative stress induces autophagic cell death independent of apoptosis in transformed and cancer cells." Cell Death Differ 15(1): 171-182.

Chen, Y. J., J. T. Chang, C. T. Liao, H. M. Wang, T. C. Yen, C. C. Chiu, Y. C. Lu, H. F. Li and A. J. Cheng (2008). "Head and neck cancer in the betel quid chewing area: recent advances in molecular carcinogenesis." Cancer Sci 99(8): 1507-1514.

Chen, Y. J., S. C. Lin, T. Kao, C. S. Chang, P. S. Hong, T. M. Shieh and K. W. Chang (2004). "Genome-wide profiling of oral squamous cell carcinoma." J Pathol 204(3): 326-332.

Cirera-Salinas, D., M. Pauta, R. M. Allen, A. G. Salerno, C. M. Ramirez, A. Chamorro-Jorganes, A. C. Wanschel, M. A. Lasuncion, M. Morales-Ruiz, Y. Suarez, A. Baldan, E. Esplugues and C. Fernandez-Hernando (2012). "Mir-33 regulates cell proliferation and cell cycle progression." Cell Cycle 11(5): 922-933.

Coates, J. M., J. M. Galante and R. J. Bold (2010). "Cancer therapy beyond apoptosis: autophagy and anoikis as mechanisms of cell death." J Surg Res 164(2): 301-308.

Costea, D. E., O. Tsinkalovsky, O. K. Vintermyr, A. C. Johannessen and I. C. Mackenzie (2006). "Cancer stem cells - new and potentially important targets for the therapy of oral squamous cell carcinoma." Oral Dis 12(5): 443-454.

Cregan, S. P., V. L. Dawson and R. S. Slack (2004). "Role of AIF in caspase-dependent and caspase-independent cell death." Oncogene 23(16): 2785-2796.

Danial, N. N. and S. J. Korsmeyer (2004). "Cell death: critical control points." Cell 116(2): 205-219.
Degenhardt, K., R. Mathew, B. Beaudoin, K. Bray, D. Anderson, G. Chen, C. Mukherjee, Y. Shi, C. Gelinas, Y. Fan, D. A. Nelson, S. Jin and E. White (2006). "Autophagy promotes tumor cell survival and restricts necrosis, inflammation, and tumorigenesis." Cancer Cell 10(1): 51-64.

Fadeel, B. and S. Orrenius (2005). "Apoptosis: a basic biological phenomenon with wide-ranging implications in human disease." J Intern Med 258(6): 479-517.

Forastiere, A. A., H. Goepfert, M. Maor, T. F. Pajak, R. Weber, W. Morrison, B. Glisson, A. Trotti, J. A. Ridge, C. Chao, G. Peters, D. J. Lee, A. Leaf, J. Ensley and J. Cooper (2003). "Concurrent chemotherapy and radiotherapy for organ preservation in advanced laryngeal cancer." N Engl J Med 349(22): 2091-2098.

Fu, L., Y. A. Kim, X. Wang, X. Wu, P. Yue, S. Lonial, F. R. Khuri and S. Y. Sun (2009). "Perifosine inhibits mammalian target of rapamycin signaling through facilitating degradation of major components in the mTOR axis and induces autophagy." Cancer Res 69(23): 8967-8976.

Gajate, C. and F. Mollinedo (2005). "Cytoskeleton-mediated death receptor and ligand concentration in lipid rafts forms apoptosis-promoting clusters in cancer chemotherapy." J Biol Chem 280(12): 11641-11647.

Gali-Muhtasib, H., M. Ocker, D. Kuester, S. Krueger, Z. El-Hajj, A. Diestel, M. Evert, N. El-Najjar, B. Peters, A. Jurjus, A. Roessner and R. Schneider-Stock (2008). "Thymoquinone reduces mouse colon tumor cell invasion and inhibits tumor growth in murine colon cancer models." J Cell Mol Med 12(1): 330-342.

Gao, M., P. Y. Yeh, Y. S. Lu, C. H. Hsu, K. F. Chen, W. C. Lee, W. C. Feng, C. S. Chen, M. L. Kuo and A. L. Cheng (2008). "OSU-03012, a novel celecoxib derivative, induces reactive oxygen species-related autophagy in hepatocellular carcinoma." Cancer Res 68(22): 9348-9357.

Gong, K., C. Chen, Y. Zhan, Y. Chen, Z. Huang and W. Li (2012). "Autophagy-related gene 7 (ATG7) and reactive oxygen species/extracellular signal-regulated kinase regulate tetrandrine-induced autophagy in human hepatocellular carcinoma." J Biol Chem 287(42): 35576-35588.

Green, D. R., R. A. Knight, G. Melino, A. Finazzi-Agro and S. Orrenius (2004). "Ten years of publication in cell death." Cell Death Differ 11(1): 2-3.

Green, D. R. and J. C. Reed (1998). "Mitochondria and apoptosis." Science 281(5381): 1309-1312.

Guo, Y., S. M. Srinivasula, A. Druilhe, T. Fernandes-Alnemri and E. S. Alnemri (2002). "Caspase-2 induces apoptosis by releasing proapoptotic proteins from mitochondria." J Biol Chem 277(16): 13430-13437.

Hayashi-Nishino, M., N. Fujita, T. Noda, A. Yamaguchi, T. Yoshimori and A. Yamamoto (2009). "A subdomain of the endoplasmic reticulum forms a cradle for autophagosome formation." Nat Cell Biol 11(12): 1433-1437.

Hoyer-Hansen, M., L. Bastholm, P. Szyniarowski, M. Campanella, G. Szabadkai, T. Farkas, K. Bianchi, N. Fehrenbacher, F. Elling, R. Rizzuto, I. S. Mathiasen and M. Jaattela (2007). "Control of macroautophagy by calcium, calmodulin-dependent kinase kinase-beta, and Bcl-2." Mol Cell 25(2): 193-205.

Hung, J. Y., Y. L. Hsu, C. T. Li, Y. C. Ko, W. C. Ni, M. S. Huang and P. L. Kuo (2009). "6-Shogaol, an active constituent of dietary ginger, induces autophagy by inhibiting the AKT/mTOR pathway in human non-small cell lung cancer A549 cells." J Agric Food Chem 57(20): 9809-9816.

Kerr, J. F., A. H. Wyllie and A. R. Currie (1972). "Apoptosis: a basic biological phenomenon with wide-ranging implications in tissue kinetics." Br J Cancer 26(4): 239-257.

Kim, R., M. Emi, K. Tanabe, S. Murakami, Y. Uchida and K. Arihiro (2006). "Regulation and interplay of apoptotic and non-apoptotic cell death." J Pathol 208(3): 319-326.

Lang, M., M. Borgmann, G. Oberhuber, R. Evstatiev, K. Jimenez, K. W. Dammann, M. Jambrich, V. Khare, C. Campregher, R. Ristl and C. Gasche (2013). "Thymoquinone attenuates tumor growth in ApcMin mice by interference with Wnt-signaling." Mol Cancer 12(1): 41.

Lee, Y. J., H. O. Song, Y. H. Lee, J. S. Ryu and M. H. Ahn (2013). "Proliferation of Toxoplasma gondii Suppresses Host Cell Autophagy." Korean J Parasitol 51(3): 279-287.

Lei, X., X. Lv, M. Liu, Z. Yang, M. Ji, X. Guo and W. Dong (2012). "Thymoquinone inhibits growth and augments 5-fluorouracil-induced apoptosis in gastric cancer cells both in vitro and in vivo." Biochem Biophys Res Commun 417(2): 864-868.

Letai, A. G. (2008). "Diagnosing and exploiting cancer''s addiction to blocks in apoptosis." Nat Rev Cancer 8(2): 121-132.

Levine, B. (2007). "Cell biology: autophagy and cancer." Nature 446(7137): 745-747.

Levine, B. and G. Kroemer (2008). "Autophagy in the pathogenesis of disease." Cell 132(1): 27-42.

Levine, B. and J. Yuan (2005). "Autophagy in cell death: an innocent convict?" J Clin Invest 115(10): 2679-2688.

Liao, C. T., J. T. Chang, H. M. Wang, S. H. Ng, C. Hsueh, L. Y. Lee, C. H. Lin, I. H. Chen, S. F. Huang, A. J. Cheng and T. C. Yen (2008). "Analysis of risk factors of predictive local tumor control in oral cavity cancer." Ann Surg Oncol 15(3): 915-922.

Lin, W. H., C. H. Hung, C. I. Hsu and J. Y. Lin (1997). "Dimerization of the N-terminal amphipathic alpha-helix domain of the fungal immunomodulatory protein from Ganoderma tsugae (Fip-gts) defined by a yeast two-hybrid system and site-directed mutagenesis." J Biol Chem 272(32): 20044-20048.

Locke, M., M. Heywood, S. Fawell and I. C. Mackenzie (2005). "Retention of intrinsic stem cell hierarchies in carcinoma-derived cell lines." Cancer Res 65(19): 8944-8950.

Lockshin, R. A. and Z. Zakeri (2001). "Programmed cell death and apoptosis: origins of the theory." Nat Rev Mol Cell Biol 2(7): 545-550.

Ogata, M., S. Hino, A. Saito, K. Morikawa, S. Kondo, S. Kanemoto, T. Murakami, M. Taniguchi, I. Tanii, K. Yoshinaga, S. Shiosaka, J. A. Hammarback, F. Urano and K. Imaizumi (2006). "Autophagy is activated for cell survival after endoplasmic reticulum stress." Mol Cell Biol 26(24): 9220-9231.

Paroni, G., C. Henderson, C. Schneider and C. Brancolini (2001). "Caspase-2-induced apoptosis is dependent on caspase-9, but its processing during UV- or tumor necrosis factor-dependent cell death requires caspase-3." J Biol Chem 276(24): 21907-21915.
Pentenero, M., S. Gandolfo and M. Carrozzo (2005). "Importance of tumor thickness and depth of invasion in nodal involvement and prognosis of oral squamous cell carcinoma: a review of the literature." Head Neck 27(12): 1080-1091.

Ravikumar, B., K. Moreau, L. Jahreiss, C. Puri and D. C. Rubinsztein (2010). "Plasma membrane contributes to the formation of pre-autophagosomal structures." Nat Cell Biol 12(8): 747-757.

Real, P. J., Y. Cao, R. Wang, Z. Nikolovska-Coleska, J. Sanz-Ortiz, S. Wang and J. L. Fernandez-Luna (2004). "Breast cancer cells can evade apoptosis-mediated selective killing by a novel small molecule inhibitor of Bcl-2." Cancer Res 64(21): 7947-7953.

Sakaki, K., J. Wu and R. J. Kaufman (2008). "Protein kinase Ctheta is required for autophagy in response to stress in the endoplasmic reticulum." J Biol Chem 283(22): 15370-15380.

Shi, Y. (2002). "Mechanisms of caspase activation and inhibition during apoptosis." Mol Cell 9(3): 459-470.

Sutton, K. M., C. D. Doucette and D. W. Hoskin (2012). "NADPH quinone oxidoreductase 1 mediates breast cancer cell resistance to thymoquinone-induced apoptosis." Biochem Biophys Res Commun 426(3): 421-426.

Wang, S. H., Y. L. Shih, W. C. Ko, Y. H. Wei and C. M. Shih (2008). "Cadmium-induced autophagy and apoptosis are mediated by a calcium signaling pathway." Cell Mol Life Sci 65(22): 3640-3652.

Weng, C., Y. Li, D. Xu, Y. Shi and H. Tang (2005). "Specific cleavage of Mcl-1 by caspase-3 in tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-induced apoptosis in Jurkat leukemia T cells." J Biol Chem 280(11): 10491-10500.

Widlak, P. and W. T. Garrard (2005). "Discovery, regulation, and action of the major apoptotic nucleases DFF40/CAD and endonuclease G." J Cell Biochem 94(6): 1078-1087.

Woo, C. C., A. P. Kumar, G. Sethi and K. H. Tan (2012). "Thymoquinone: potential cure for inflammatory disorders and cancer." Biochem Pharmacol 83(4): 443-451.

Woo, C. C., S. Y. Loo, V. Gee, C. W. Yap, G. Sethi, A. P. Kumar and K. H. Tan (2011). "Anticancer activity of thymoquinone in breast cancer cells: possible involvement of PPAR-gamma pathway." Biochem Pharmacol 82(5): 464-475.

Xu, M. Y., D. H. Lee, E. J. Joo, K. H. Son and Y. S. Kim (2013). "Akebia saponin PA induces autophagic and apoptotic cell death in AGS human gastric cancer cells." Food Chem Toxicol.

Yamanaka, Y., K. Shiraki, T. Inoue, K. Miyashita, H. Fuke, Y. Yamaguchi, N. Yamamoto, K. Ito, K. Sugimoto and T. Nakano (2006). "COX-2 inhibitors sensitize human hepatocellular carcinoma cells to TRAIL-induced apoptosis." Int J Mol Med 18(1): 41-47.

Yoshimori, T. (2004). "Autophagy: a regulated bulk degradation process inside cells." Biochem Biophys Res Commun 313(2): 453-458.

Zhou, W. J., R. Deng, X. Y. Zhang, G. K. Feng, L. Q. Gu and X. F. Zhu (2009). "G-quadruplex ligand SYUIQ-5 induces autophagy by telomere damage and TRF2 delocalization in cancer cells." Mol Cancer Ther 8(12): 3203-3213.

Zubair, H., H. Y. Khan, A. Sohail, S. Azim, M. F. Ullah, A. Ahmad, F. H. Sarkar and S. M. Hadi (2013). "Redox cycling of endogenous copper by thymoquinone leads to ROS-mediated DNA breakage and consequent cell death: putative anticancer mechanism of antioxidants." Cell Death Dis 4: e660.

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