請用此 Handle URI 來引用此文件:
http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/54499
完整後設資料紀錄
DC 欄位 | 值 | 語言 |
---|---|---|
dc.contributor.advisor | 鄭永銘 | |
dc.contributor.author | Shu-Yuan Mai | en |
dc.contributor.author | 麥淑媛 | zh_TW |
dc.date.accessioned | 2021-06-16T03:00:33Z | - |
dc.date.available | 2017-09-25 | |
dc.date.copyright | 2015-09-25 | |
dc.date.issued | 2015 | |
dc.date.submitted | 2015-07-03 | |
dc.identifier.citation | 1. Venook AP, Papandreou C, Furuse J, et al., The incidence and epidemiology of hepatocellular carcinoma: a global and regional perspective. Oncologist, 2010. 15 Suppl 4: p. 5-13.
2. Ahmed F, Perz JF, Kwong S, et al., National trends and disparities in the incidence of hepatocellular carcinoma, 1998-2003. Prev Chronic Dis, 2008. 5(3): p. A74. 3. Llovet JM, Updated treatment approach to hepatocellular carcinoma. J Gastroenterol, 2005. 40(3): p. 225-35. 4. Gomaa AI, Khan SA, Toledano MB, et al., Hepatocellular carcinoma: epidemiology, risk factors and pathogenesis. World J Gastroenterol, 2008. 14(27): p. 4300-8. 5. Block TM, Mehta AS, Fimmel CJ, et al., Molecular viral oncology of hepatocellular carcinoma. Oncogene, 2003. 22(33): p. 5093-107. 6. El-Serag HB, Hepatocellular carcinoma. N Engl J Med, 2011. 365(12): p. 1118-27. 7. Alter MJ, Epidemiology of hepatitis C virus infection. World J Gastroenterol, 2007. 13(17): p. 2436-41. 8. Gao B and Bataller R, Alcoholic liver disease: pathogenesis and new therapeutic targets. Gastroenterology, 2011. 141(5): p. 1572-85. 9. Liu M, Jiang L, and Guan XY, The genetic and epigenetic alterations in human hepatocellular carcinoma: a recent update. Protein Cell, 2014. 5(9): p. 673-91. 10. Yang SF, Chang CW, Wei RJ, et al., Involvement of DNA damage response pathways in hepatocellular carcinoma. Biomed Res Int, 2014. 2014: p. 153867. 11. Farazi PA and DePinho RA, Hepatocellular carcinoma pathogenesis: from genes to environment. Nat Rev Cancer, 2006. 6(9): p. 674-87. 12. Sanyal AJ, Yoon SK, and Lencioni R, The etiology of hepatocellular carcinoma and consequences for treatment. Oncologist, 2010. 15 Suppl 4: p. 14-22. 13. Thorgeirsson SS and Grisham JW, Molecular pathogenesis of human hepatocellular carcinoma. Nat Genet, 2002. 31(4): p. 339-46. 14. Hanahan D and Weinberg RA, Hallmarks of cancer: the next generation. Cell, 2011. 144(5): p. 646-74. 15. Nault JC and Zucman-Rossi J, Genetics of hepatocellular carcinoma: the next generation. J Hepatol, 2014. 60(1): p. 224-6. 16. Nault JC, Mallet M, Pilati C, et al., High frequency of telomerase reverse-transcriptase promoter somatic mutations in hepatocellular carcinoma and preneoplastic lesions. Nat Commun, 2013. 4: p. 2218. 17. Cleary SP, Jeck WR, Zhao X, et al., Identification of driver genes in hepatocellular carcinoma by exome sequencing. Hepatology, 2013. 58(5): p. 1693-702. 18. Kan Z, Zheng H, Liu X, et al., Whole-genome sequencing identifies recurrent mutations in hepatocellular carcinoma. Genome Res, 2013. 23(9): p. 1422-33. 19. Fujimoto A, Totoki Y, Abe T, et al., Whole-genome sequencing of liver cancers identifies etiological influences on mutation patterns and recurrent mutations in chromatin regulators. Nat Genet, 2012. 44(7): p. 760-4. 20. Sporn MB and Liby KT, NRF2 and cancer: the good, the bad and the importance of context. Nat Rev Cancer, 2012. 12(8): p. 564-71. 21. Hayes JD and McMahon M, NRF2 and KEAP1 mutations: permanent activation of an adaptive response in cancer. Trends Biochem Sci, 2009. 34(4): p. 176-88. 22. Solis LM, Behrens C, Dong W, et al., Nrf2 and Keap1 abnormalities in non-small cell lung carcinoma and association with clinicopathologic features. Clin Cancer Res, 2010. 16(14): p. 3743-53. 23. Singh A, Misra V, Thimmulappa RK, et al., Dysfunctional KEAP1-NRF2 interaction in non-small-cell lung cancer. PLoS Med, 2006. 3(10): p. e420. 24. Shibata T, Kokubu A, Gotoh M, et al., Genetic alteration of Keap1 confers constitutive Nrf2 activation and resistance to chemotherapy in gallbladder cancer. Gastroenterology, 2008. 135(4): p. 1358-1368, 1368 e1-4. 25. Shibata T, Ohta T, Tong KI, et al., Cancer related mutations in NRF2 impair its recognition by Keap1-Cul3 E3 ligase and promote malignancy. Proc Natl Acad Sci U S A, 2008. 105(36): p. 13568-73. 26. Yoo NJ, Kim HR, Kim YR, et al., Somatic mutations of the KEAP1 gene in common solid cancers. Histopathology, 2012. 60(6): p. 943-52. 27. Konstantinopoulos PA, Spentzos D, Fountzilas E, et al., Keap1 mutations and Nrf2 pathway activation in epithelial ovarian cancer. Cancer Res, 2011. 71(15): p. 5081-9. 28. Sjoblom T, Jones S, Wood LD, et al., The consensus coding sequences of human breast and colorectal cancers. Science, 2006. 314(5797): p. 268-74. 29. Nioi P and Nguyen T, A mutation of Keap1 found in breast cancer impairs its ability to repress Nrf2 activity. Biochem Biophys Res Commun, 2007. 362(4): p. 816-21. 30. Kansanen E, Kuosmanen SM, Leinonen H, et al., The Keap1-Nrf2 pathway: Mechanisms of activation and dysregulation in cancer. Redox Biol, 2013. 1(1): p. 45-9. 31. Motohashi H, Katsuoka F, Engel JD, et al., Small Maf proteins serve as transcriptional cofactors for keratinocyte differentiation in the Keap1-Nrf2 regulatory pathway. Proc Natl Acad Sci U S A, 2004. 101(17): p. 6379-84. 32. Motohashi H and Yamamoto M, Nrf2-Keap1 defines a physiologically important stress response mechanism. Trends Mol Med, 2004. 10(11): p. 549-57. 33. Nioi P, Nguyen T, Sherratt PJ, et al., The carboxy-terminal Neh3 domain of Nrf2 is required for transcriptional activation. Mol Cell Biol, 2005. 25(24): p. 10895-906. 34. Apopa PL, He X, and Ma Q, Phosphorylation of Nrf2 in the transcription activation domain by casein kinase 2 (CK2) is critical for the nuclear translocation and transcription activation function of Nrf2 in IMR-32 neuroblastoma cells. J Biochem Mol Toxicol, 2008. 22(1): p. 63-76. 35. McMahon M, Thomas N, Itoh K, et al., Redox-regulated turnover of Nrf2 is determined by at least two separate protein domains, the redox-sensitive Neh2 degron and the redox-insensitive Neh6 degron. J Biol Chem, 2004. 279(30): p. 31556-67. 36. Kobayashi A, Kang MI, Okawa H, et al., Oxidative stress sensor Keap1 functions as an adaptor for Cul3-based E3 ligase to regulate proteasomal degradation of Nrf2. Mol Cell Biol, 2004. 24(16): p. 7130-9. 37. Tong KI, Katoh Y, Kusunoki H, et al., Keap1 recruits Neh2 through binding to ETGE and DLG motifs: characterization of the two-site molecular recognition model. Mol Cell Biol, 2006. 26(8): p. 2887-900. 38. Ma Q and He X, Molecular basis of electrophilic and oxidative defense: promises and perils of Nrf2. Pharmacol Rev, 2012. 64(4): p. 1055-81. 39. Magesh S, Chen Y, and Hu L, Small molecule modulators of Keap1-Nrf2-ARE pathway as potential preventive and therapeutic agents. Med Res Rev, 2012. 32(4): p. 687-726. 40. Villeneuve NF, Lau A, and Zhang DD, Regulation of the Nrf2-Keap1 antioxidant response by the ubiquitin proteasome system: an insight into cullin-ring ubiquitin ligases. Antioxid Redox Signal, 2010. 13(11): p. 1699-712. 41. Yamamoto T, Suzuki T, Kobayashi A, et al., Physiological significance of reactive cysteine residues of Keap1 in determining Nrf2 activity. Mol Cell Biol, 2008. 28(8): p. 2758-70. 42. Sekhar KR, Rachakonda G, and Freeman ML, Cysteine-based regulation of the CUL3 adaptor protein Keap1. Toxicol Appl Pharmacol, 2010. 244(1): p. 21-6. 43. Itoh K, Chiba T, Takahashi S, et al., An Nrf2/small Maf heterodimer mediates the induction of phase II detoxifying enzyme genes through antioxidant response elements. Biochem Biophys Res Commun, 1997. 236(2): p. 313-22. 44. Chan JY, Cheung MC, Moi P, et al., Chromosomal localization of the human NF-E2 family of bZIP transcription factors by fluorescence in situ hybridization. Hum Genet, 1995. 95(3): p. 265-9. 45. Moi P, Chan K, Asunis I, et al., Isolation of NF-E2-related factor 2 (Nrf2), a NF-E2-like basic leucine zipper transcriptional activator that binds to the tandem NF-E2/AP1 repeat of the beta-globin locus control region. Proc Natl Acad Sci U S A, 1994. 91(21): p. 9926-30. 46. Wakabayashi N, Shin S, Slocum SL, et al., Regulation of notch1 signaling by nrf2: implications for tissue regeneration. Sci Signal, 2010. 3(130): p. ra52. 47. Satoh H, Moriguchi T, Takai J, et al., Nrf2 prevents initiation but accelerates progression through the Kras signaling pathway during lung carcinogenesis. Cancer Res, 2013. 73(13): p. 4158-68. 48. Itoh K, Mimura J, and Yamamoto M, Discovery of the negative regulator of Nrf2, Keap1: a historical overview. Antioxid Redox Signal, 2010. 13(11): p. 1665-78. 49. Venugopal R and Jaiswal AK, Nrf1 and Nrf2 positively and c-Fos and Fra1 negatively regulate the human antioxidant response element-mediated expression of NAD(P)H:quinone oxidoreductase1 gene. Proc Natl Acad Sci U S A, 1996. 93(25): p. 14960-5. 50. Solis WA, Dalton TP, Dieter MZ, et al., Glutamate-cysteine ligase modifier subunit: mouse Gclm gene structure and regulation by agents that cause oxidative stress. Biochem Pharmacol, 2002. 63(9): p. 1739-54. 51. Hayes JD, Chanas SA, Henderson CJ, et al., The Nrf2 transcription factor contributes both to the basal expression of glutathione S-transferases in mouse liver and to their induction by the chemopreventive synthetic antioxidants, butylated hydroxyanisole and ethoxyquin. Biochem Soc Trans, 2000. 28(2): p. 33-41. 52. Rushmore TH, Morton MR, and Pickett CB, The antioxidant responsive element. Activation by oxidative stress and identification of the DNA consensus sequence required for functional activity. J Biol Chem, 1991. 266(18): p. 11632-9. 53. Motohashi H, O'Connor T, Katsuoka F, et al., Integration and diversity of the regulatory network composed of Maf and CNC families of transcription factors. Gene, 2002. 294(1-2): p. 1-12. 54. Kobayashi A, Ito E, Toki T, et al., Molecular cloning and functional characterization of a new Cap'n' collar family transcription factor Nrf3. J Biol Chem, 1999. 274(10): p. 6443-52. 55. Kwong M, Kan YW, and Chan JY, The CNC basic leucine zipper factor, Nrf1, is essential for cell survival in response to oxidative stress-inducing agents. Role for Nrf1 in gamma-gcs(l) and gss expression in mouse fibroblasts. J Biol Chem, 1999. 274(52): p. 37491-8. 56. Ohtsuji M, Katsuoka F, Kobayashi A, et al., Nrf1 and Nrf2 play distinct roles in activation of antioxidant response element-dependent genes. J Biol Chem, 2008. 283(48): p. 33554-62. 57. Chan JY, Kwong M, Lu R, et al., Targeted disruption of the ubiquitous CNC-bZIP transcription factor, Nrf-1, results in anemia and embryonic lethality in mice. EMBO J, 1998. 17(6): p. 1779-87. 58. Sankaranarayanan K and Jaiswal AK, Nrf3 negatively regulates antioxidant-response element-mediated expression and antioxidant induction of NAD(P)H:quinone oxidoreductase1 gene. J Biol Chem, 2004. 279(49): p. 50810-7. 59. DeNicola GM, Karreth FA, Humpton TJ, et al., Oncogene-induced Nrf2 transcription promotes ROS detoxification and tumorigenesis. Nature, 2011. 475(7354): p. 106-9. 60. Jaramillo MC and Zhang DD, The emerging role of the Nrf2-Keap1 signaling pathway in cancer. Genes Dev, 2013. 27(20): p. 2179-91. 61. Kim YR, Oh JE, Kim MS, et al., Oncogenic NRF2 mutations in squamous cell carcinomas of oesophagus and skin. J Pathol, 2010. 220(4): p. 446-51. 62. Shibata T, Kokubu A, Saito S, et al., NRF2 mutation confers malignant potential and resistance to chemoradiation therapy in advanced esophageal squamous cancer. Neoplasia, 2011. 13(9): p. 864-73. 63. Loignon M, Miao W, Hu L, et al., Cul3 overexpression depletes Nrf2 in breast cancer and is associated with sensitivity to carcinogens, to oxidative stress, and to chemotherapy. Mol Cancer Ther, 2009. 8(8): p. 2432-40. 64. Bauer AK, Hill T, 3rd, and Alexander CM, The involvement of NRF2 in lung cancer. Oxid Med Cell Longev, 2013. 2013: p. 746432. 65. Lister A, Nedjadi T, Kitteringham NR, et al., Nrf2 is overexpressed in pancreatic cancer: implications for cell proliferation and therapy. Mol Cancer, 2011. 10: p. 37. 66. Liao H, Zhou Q, Zhang Z, et al., NRF2 is overexpressed in ovarian epithelial carcinoma and is regulated by gonadotrophin and sex-steroid hormones. Oncol Rep, 2012. 27(6): p. 1918-24. 67. Wong TF, Yoshinaga K, Monma Y, et al., Association of keap1 and nrf2 genetic mutations and polymorphisms with endometrioid endometrial adenocarcinoma survival. Int J Gynecol Cancer, 2011. 21(8): p. 1428-35. 68. Sayin VI, Ibrahim MX, Larsson E, et al., Antioxidants accelerate lung cancer progression in mice. Sci Transl Med, 2014. 6(221): p. 221ra15. 69. Reddy NM, Kleeberger SR, Yamamoto M, et al., Genetic dissection of the Nrf2-dependent redox signaling-regulated transcriptional programs of cell proliferation and cytoprotection. Physiol Genomics, 2007. 32(1): p. 74-81. 70. Reddy NM, Kleeberger SR, Cho HY, et al., Deficiency in Nrf2-GSH signaling impairs type II cell growth and enhances sensitivity to oxidants. Am J Respir Cell Mol Biol, 2007. 37(1): p. 3-8. 71. Wang XJ, Sun Z, Villeneuve NF, et al., Nrf2 enhances resistance of cancer cells to chemotherapeutic drugs, the dark side of Nrf2. Carcinogenesis, 2008. 29(6): p. 1235-43. 72. Homma S, Ishii Y, Morishima Y, et al., Nrf2 enhances cell proliferation and resistance to anticancer drugs in human lung cancer. Clin Cancer Res, 2009. 15(10): p. 3423-32. 73. Mitsuishi Y, Motohashi H, and Yamamoto M, The Keap1-Nrf2 system in cancers: stress response and anabolic metabolism. Front Oncol, 2012. 2: p. 200. 74. Mitsuishi Y, Taguchi K, Kawatani Y, et al., Nrf2 redirects glucose and glutamine into anabolic pathways in metabolic reprogramming. Cancer Cell, 2012. 22(1): p. 66-79. 75. Harris IS, Treloar AE, Inoue S, et al., Glutathione and thioredoxin antioxidant pathways synergize to drive cancer initiation and progression. Cancer Cell, 2015. 27(2): p. 211-22. 76. Warburg O, On the origin of cancer cells. Science, 1956. 123(3191): p. 309-14. 77. Gatenby RA and Gillies RJ, Why do cancers have high aerobic glycolysis? Nat Rev Cancer, 2004. 4(11): p. 891-9. 78. Son J, Lyssiotis CA, Ying H, et al., Glutamine supports pancreatic cancer growth through a KRAS-regulated metabolic pathway. Nature, 2013. 496(7443): p. 101-5. 79. DeBerardinis RJ, Is cancer a disease of abnormal cellular metabolism? New angles on an old idea. Genet Med, 2008. 10(11): p. 767-77. 80. Maher JM, Dieter MZ, Aleksunes LM, et al., Oxidative and electrophilic stress induces multidrug resistance-associated protein transporters via the nuclear factor-E2-related factor-2 transcriptional pathway. Hepatology, 2007. 46(5): p. 1597-610. 81. Watson J, Oxidants, antioxidants and the current incurability of metastatic cancers. Open Biol, 2013. 3(1): p. 120144. 82. Klein EA, Thompson IM, Jr., Tangen CM, et al., Vitamin E and the risk of prostate cancer: the Selenium and Vitamin E Cancer Prevention Trial (SELECT). JAMA, 2011. 306(14): p. 1549-56. 83. Broos S, Hulpiau P, Galle J, et al., ConTra v2: a tool to identify transcription factor binding sites across species, update 2011. Nucleic Acids Res, 2011. 39(Web Server issue): p. W74-8. 84. Chorley BN, Campbell MR, Wang X, et al., Identification of novel NRF2-regulated genes by ChIP-Seq: influence on retinoid X receptor alpha. Nucleic Acids Res, 2012. 40(15): p. 7416-29. 85. Milde-Langosch K, The Fos family of transcription factors and their role in tumourigenesis. Eur J Cancer, 2005. 41(16): p. 2449-61. 86. Chiu R, Boyle WJ, Meek J, et al., The c-Fos protein interacts with c-Jun/AP-1 to stimulate transcription of AP-1 responsive genes. Cell, 1988. 54(4): p. 541-52. 87. Gruda MC, Kovary K, Metz R, et al., Regulation of Fra-1 and Fra-2 phosphorylation differs during the cell cycle of fibroblasts and phosphorylation in vitro by MAP kinase affects DNA binding activity. Oncogene, 1994. 9(9): p. 2537-47. 88. Tabor E, Tumor suppressor genes, growth factor genes, and oncogenes in hepatitis B virus-associated hepatocellular carcinoma. J Med Virol, 1994. 42(4): p. 357-65. 89. Bavner A, Matthews J, Sanyal S, et al., EID3 is a novel EID family member and an inhibitor of CBP-dependent co-activation. Nucleic Acids Res, 2005. 33(11): p. 3561-9. 90. Sahin U, Koslowski M, Tureci O, et al., Expression of cancer testis genes in human brain tumors. Clin Cancer Res, 2000. 6(10): p. 3916-22. 91. Condomines M, Hose D, Raynaud P, et al., Cancer/testis genes in multiple myeloma: expression patterns and prognosis value determined by microarray analysis. J Immunol, 2007. 178(5): p. 3307-15. 92. Gaugler B, Van den Eynde B, van der Bruggen P, et al., Human gene MAGE-3 codes for an antigen recognized on a melanoma by autologous cytolytic T lymphocytes. J Exp Med, 1994. 179(3): p. 921-30. 93. Patard JJ, Brasseur F, Gil-Diez S, et al., Expression of MAGE genes in transitional-cell carcinomas of the urinary bladder. Int J Cancer, 1995. 64(1): p. 60-4. 94. Caballero OL and Chen YT, Cancer/testis (CT) antigens: potential targets for immunotherapy. Cancer Sci, 2009. 100(11): p. 2014-21. 95. Axelsen JB, Lotem J, Sachs L, et al., Genes overexpressed in different human solid cancers exhibit different tissue-specific expression profiles. Proc Natl Acad Sci U S A, 2007. 104(32): p. 13122-7. 96. Goodman RH and Smolik S, CBP/p300 in cell growth, transformation, and development. Genes Dev, 2000. 14(13): p. 1553-77. 97. Iyer NG, Ozdag H, and Caldas C, p300/CBP and cancer. Oncogene, 2004. 23(24): p. 4225-31. 98. Sonis ST, Oral mucositis in cancer therapy. J Support Oncol, 2004. 2(6 Suppl 3): p. 3-8. 99. Kovac S, Angelova PR, Holmstrom KM, et al., Nrf2 regulates ROS production by mitochondria and NADPH oxidase. Biochim Biophys Acta, 2015. 1850(4): p. 794-801. 100. Han G, Bian L, Li F, et al., Preventive and therapeutic effects of Smad7 on radiation-induced oral mucositis. Nat Med, 2013. 19(4): p. 421-8 | |
dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/54499 | - |
dc.description.abstract | 肝細胞癌是肝癌的最常見的類型,現在是全世界癌症死亡的第二大原因。根據最近外顯子組定序研究,許多突變基因和訊息途徑與肝細胞癌的生成有關。最常突變的基因包括TERT(59%)、beta-catenin(10〜32%)、TP53(18〜35%)、MLL(17〜20%)、JAK1(9%)、KEAP1(8%)和NRF2(6%)。在肝細胞癌中NRF2的突變和異常堆積,在肝臟癌變過程中可能扮演促進惡性細胞生長的重要角色。NRF2執行細胞防禦的功能來抵抗氧化和親電子所造成的壓力並且保護細胞免受於化學性的癌變發生。然而,最近的許多研究發現,許多種癌症有NRF2蛋白質異常堆積和活化的現象。在此研究中,我們探討NRF2如何促進肝臟癌變。我們的研究發現,使用核糖核酸干擾(RNA interference)抑制NRF2的表現量在體外培養實驗和體內小鼠異種移植實驗中會抑制細胞增生、聚球(sphere formation)能力、移動和侵犯的能力。我們通過微陣列分析結果發現新穎的受NRF2調控的基因, c-Fos和EID3。使用西方墨漬法、染色質免疫沉澱法及發光脢法,我們發現NRF2會辨認且結合在c-Fos和EID3的啟動子上的抗氧化反應元件(ARE)以促進其表現。此外,我們建構了在角質細胞特定表達活化的NRF2的轉殖小鼠模型。我們也測試了是否NRF2過度活化能改善頭頸癌患者藉由輻射治療所出現的常見副作用口腔粘膜炎。 | zh_TW |
dc.description.abstract | Hepatocellular carcinoma (HCC) is the most common type of liver cancer and is now the second leading cause of cancer death worldwide. According to recently exome sequencing studies, many mutated genes and pathways implicated in HCC were identified. The most frequently mutated genes include TERT (59%), beta-catenin (10~32%), TP53 (18~35%), MLL (17~20%), JAK1 (9%), KEAP1 (8%) and NRF2 (6%). Mutation and aberrant accumulation of NRF2 in HCC may play important roles in promoting malignant progression during liver carcinogenesis. NRF2 executes cellular defenses against oxidative and electrophilic stresses and protects against chemical carcinogenesis. However, recent studies found that NRF2 is accumulated and activated in many types of cancer. In this study, we examine how NRF2 contributes to liver carcinogenesis. We found that knockdown of NRF2 by shRNA suppressed in vitro and in vivo cell proliferation, sphere formation, cell migration and cell invasion. Through microarray analysis, we identified novel NRF2-regulated genes, c-Fos and EID3. Using western blotting, chromatin immunoprecipitation, and promoter luciferase reporter assay, we found that NRF2 recognized an antioxidant response element (ARE) in the promoter of c-Fos and EID3 to upregulate their expression. In addition, we generated a mouse model of K14.NRF2 E82G, which express a constitutively active NRF2 transgene under the control of a keratin 14 promoter. We tested whether NRF2 overactivity improves oral mucositis, a common side effect of irradiation for patients with head and neck cancer. | en |
dc.description.provenance | Made available in DSpace on 2021-06-16T03:00:33Z (GMT). No. of bitstreams: 1 ntu-104-R02444001-1.pdf: 9187451 bytes, checksum: d38a4abd5197a2a77186cbf8053509b5 (MD5) Previous issue date: 2015 | en |
dc.description.tableofcontents | Page
口試委員審定書 I 謝辭 II 中文摘要 III Abstract IV Contents VI 1. Introduction 1 1.1 Hepatocellular carcinoma …….…….………………………….……………………1 1.2 The pathogenesis of hepatocellular carcinoma…………...……..…………………...2 1.3 Major pathways altered in hepatocellular carcinoma………….……………..……...2 1.4 Structures and interaction between Nrf2 and Keap1………………………………...3 1.5 The functions of NF-E2 family…………………………..………………………….5 1.6 Nrf2 acts as an oncogene…………………………………………………………….7 1.7 Aims of the study……………………………………………………………………10 2. Materials and Methods 11 2.1 Reagents....................................................................................................................11 2.2 Plasmids....................................................................................................................11 2.3 Cell culture……………………………………………………………………….11 2.4 RNA interference for knockdown experiments........................................................12 2.5 RNA isolation……………………………………………………………………....12 2.6 RT-PCR…………………………………………….................................................13 2.7 Real-time PCR...........................................................................................................13 2.8 Immunohistochemistry………………………………………………......................14 2.9 Western blotting........................................................................................................14 2.10 MTTassay................................................................................................................15 2.11 Soft agar colony-forming assay...............................................................................15 2.12 In vitro sphere formation..........................................................................................16 2.13 In vivo Xenograft tumor formation in mice..............................................................17 2.14 Wound-healing Assay.............................................................................................17 2.15 In vitro Boyden Chamber Invasion assay..................................................................17 2.16 Microarray analysis……………………………………..........................................18 2.17 Promoter luciferase reporter assay............................................................................19 2.18 Immunoprecipitation assay (IP)………………………………………………19 2.19 Chromatin immunoprecipitation assay (ChIP)........................................................20 2.20 Rescue experiments restores c-Fos expression in Nrf2 knockdown stable cells….22 2.21 Generation of the transgenes and transgenic mice...................................................22 2.22 Radiation-induced oral mucositis............................................................................23 3. Results 24 3.1 Overexpression of NRF2 in HCC………………………….......................................24 3.2 Knockdown of NRF2 in SK-Hep1 inhibited anchorage-independent growth, tumor sphere formation, and in vivo tumorgenicity…………………….................................24 3.3 Knockdown of NRF2 in SK-Hep1 suppressed cell migration and invasion..............25 3.4 Identification of differential gene expression between control and NRF2 knockdown cells..................................................................................................................................26 3.5 Induction of NRF2 expression in HepG2 by SFN......................................................26 3.6 c-Fos expression is activated by NRF2…………………..........................................27 3.7 NRF2 directly binds to the c-Fos and EID3 promoters..............................................27 3.8 Overexpression of c-Fos does not affect cell proliferation in NRF2-knockdown SK-Hep1 cells..................................................................................................................28 3.9 K14.NRF2 E82G mice are not resistant to radiation-induced oral mucositis.............28 4. Discussion 30 4.1 NRF2 is aberrantly activated in HCC specimens......................................................30 4.2 NRF2 plays a critical role in progression of HCC.....................................................30 4.3 The novel target genes of NRF2................................................................................31 4.4 The role of c-Fos in cancers………………………...................................................31 4.5 The functions of EID3………………………………………………………………33 4.6 Activation of NRF2-associated cancer signaling pathways promotes tumor growth………………………………………………………………………………...34 4.7 No significant effects of NRF2 overexpression in a mouse model of radiation-induced oral mucositis…………………………………………………………………….35 5. Figures and Tables 37 Figure 1. Immunohistochemical stain of NRF2 expression in representative hepatocellular carcinoma specimens……………………………………………………37 Figure 2. Expression analysis of NRF2 in hepatocellular carcinoma cell lines using real-time PCR and western blotting………………………………………...................................38 Figure 3. Knockdown efficiency of NRF2 shRNA in SK-Hep1 cells....................................39 Figure 4. Analysis of regulation of NRF2 downstream genes by shRNAs in SK-Hep1 cells…………………………………………………………………………………....40 Figure 5. In vitro proliferation of SK-Hep1 cells as examined by MTT assay………….41 Figure 6. NRF2 knockdown blocked the anchorage-independent growth ability of SK-Hep1 cells, as determined by in vitro soft agar colony formation assay.…...................43 Figure 7. NRF2 knockdown in SK-Hep1 cells inhibited tumor sphere formation……46 Figure 8. NRF2 knockdown inhibited tumor growth of subcutaneous xenograft of SK-Hep1 cells in NOD/SCID mice……………………………………………………47 Figure 9. NRF2 knockdown in SK-Hep1 cells inhibited cell migration by wound-healing assay……………………………………………………………………………50 Figure 10. NRF2 knockdown in SK-Hep1 cells inhibited cell invasion in Boyden chamber assay……………………………………………………………………………..51 Figure 11. Validation of the success of knockdown of Nrf2 and downregulation of Nrf2 targets in the samples sent for microarray analysis……………………………………53 Figure 12. Heat maps illustrated gene expression profiling by microarray analysis…….54 Figure 13. Validation of the genes regulated by NRF2 knockdown identified by microarray analysis..........................................................................................................55 Figure 14. Treatment of SFN and tBHQ stabilized and activated NRF2………………56 Figure 15. SFN-induced NRF2 activation and induction of NRF2 target genes...............57 Figure 16. Knockdown and induction of NRF2 results in up- and down-regulation of c-Fos………………………………………………………………………………….58 Figure 17. Schematics showing it’s the predicted AREs on c-Fos locus.......................60 Figure 18. Alteration of NRF2 expression changed luciferase activities of c-Fos promoter and AP-1 reporter…………………………………………………………………61 Figure19. Validation of the feasibility of the antibodies used in ChIP assay by immunoprecipitaion......................................................................................................62 Figure 20. ChIP-PCR assay confirms NRF2 binds to the putative ARE of c-Fos promoter........................................................................................................................63 Figure 21. ChIP-PCR assay confirms NRF2 binds to the putative ARE of EID3 promoter.........................................................................................................................65 Figure 22. Re-expression of c-Fos in Nrf2-knocked down SK-Hep1 cells...................66 Figure 23. c-Fos overexpression restored AP-1 promoter luciferase activities in NRF2-knockdown SK-Hep1 cells............................................................................................68 Figure 24. Overexpression of c-Fos did not restore anchorage-dependent or anchorage-independent growth ability in SK-Hep1 with knockdown of NRF2….........................69 Figure 25. Overexpression of c-Fos enhanced anchorage-dependent growth ability in PLC5 NRF2 knockdown cells.......................................................................................70 Figure 26. Evaluation of radiation-induced oral mucositis in K14. NRF2 E82G mice..72 Table 1. The primers used for PCR cloning....................................................................74 Table2. The clones of shRNA in lentiviral vectors used for knockdown of the endogenous Nrf2…………………………………………………………………..74 Table 3-1. The primers used for RT-PCR reaction............................................................75 Table 3-2. The primers used for Real time-PCR reaction.................................................75 Table 4.1. The known human target genes of Nrf2 detected microarrays........................76 Table 4-2. The genes down-regulated in Nrf2-depleted cells detected by microarrays analysis…………………………………………………………………………………77 Table 4-3. The primers used for Real time-PCR reaction to validate microarray results...............................................................................................................................78 Table 5. The primers used for ChIP...................................................................................79 Table 6.The primers used for transgenic mice genotyping and verified copy number of the transgene in the different transgenic lines................................................................80 6. Reference 81 | |
dc.language.iso | en | |
dc.title | 肝臟癌變過程NRF2經由活化癌症相關的訊息途徑促進細胞增生和轉移 | zh_TW |
dc.title | NRF2 accelerates cell proliferation and metastasis through activation of cancer-associated signaling cascades during liver carcinogenesis | en |
dc.type | Thesis | |
dc.date.schoolyear | 103-2 | |
dc.description.degree | 碩士 | |
dc.contributor.oralexamcommittee | 葉秀慧,張以承,劉兆蓮 | |
dc.subject.keyword | 肝細胞癌,NRF2,KEAP1,c-Fos,EID3,肝臟癌變, | zh_TW |
dc.subject.keyword | Hepatocellular carcinoma (HCC),NRF2,KEAP1,c-Fos,EID3,liver carcinogenesis, | en |
dc.relation.page | 87 | |
dc.rights.note | 有償授權 | |
dc.date.accepted | 2015-07-03 | |
dc.contributor.author-college | 醫學院 | zh_TW |
dc.contributor.author-dept | 病理學研究所 | zh_TW |
顯示於系所單位: | 病理學科所 |
文件中的檔案:
檔案 | 大小 | 格式 | |
---|---|---|---|
ntu-104-1.pdf 目前未授權公開取用 | 8.97 MB | Adobe PDF |
系統中的文件,除了特別指名其著作權條款之外,均受到著作權保護,並且保留所有的權利。