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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 林榮耀(Jung-Yaw Lin) | |
| dc.contributor.author | Tsung-Cheng Yang | en |
| dc.contributor.author | 楊琮誠 | zh_TW |
| dc.date.accessioned | 2021-06-15T00:59:39Z | - |
| dc.date.available | 2013-09-11 | |
| dc.date.copyright | 2008-09-11 | |
| dc.date.issued | 2008 | |
| dc.date.submitted | 2008-08-01 | |
| dc.identifier.citation | 1 Zbar B, Klausner R, Linehan WM: Studying cancer families to identify kidney cancer genes. Annu Rev Med 2003;54:217-233.
2 Jones J, Otu H, Spentzos D, Kolia S, Inan M, Beecken WD, Fellbaum C, Gu X, Joseph M, Pantuck AJ, Jonas D, Libermann TA: Gene signatures of progression and metastasis in renal cell cancer. Clin Cancer Res 2005;11:5730-5739. 3 Iliopoulos O, Kibel A, Gray S, Kaelin WG, Jr.: Tumour suppression by the human von hippel-lindau gene product. Nat Med 1995;1:822-826. 4 Hoshi S, Satoh M, Ohyama C, Hiramatu M, Watanabe R, Hagisawa S, Endo M, Arai Y: Active chemotherapy for bone metastasis in sarcomatoid renal cell carcinoma. Int J Clin Oncol 2003;8:113-117. 5 Sarkissian G, Fergelot P, Lamy PJ, Patard JJ, Culine S, Jouin P, Rioux-Leclercq N, Darbouret B: Identification of pro-mmp-7 as a serum marker for renal cell carcinoma by use of proteomic analysis. Clin Chem 2008;54:574-581. 6 Meyer HA, Tolle A, Jung M, Fritzsche FR, Haendler B, Kristiansen I, Gaspert A, Johannsen M, Jung K, Kristiansen G: Identification of stanniocalcin 2 as prognostic marker in renal cell carcinoma. Eur Urol 2008 7 Weiss RH, Borowsky AD, Seligson D, Lin PY, Dillard-Telm L, Belldegrun AS, Figlin RA, Pantuck AD: P21 is a prognostic marker for renal cell carcinoma: Implications for novel therapeutic approaches. J Urol 2007;177:63-68; discussion 68-69. 8 Tang SW, Chang WH, Chao YW, Lin CY, Chen HF, Lai YH, Zhan BW, Su YC, Jane SW, Chen YC, Hsu CI, Lin WC, Wang KC, Lai MK, Lin JY: Identification of differentially expressed genes in clear cell renal cell carcinoma by analysis of full-length enriched cdna library. J Biomed Sci 2006;13:233-240. 9 Aksoy S, Szumlanski CL, Weinshilboum RM: Human liver nicotinamide n-methyltransferase. Cdna cloning, expression, and biochemical characterization. J Biol Chem 1994;269:14835-14840. 10 Aksoy S, Brandriff BF, Ward A, Little PF, Weinshilboum RM: Human nicotinamide n-methyltransferase gene: Molecular cloning, structural characterization and chromosomal localization. Genomics 1995;29:555-561. 11 Hoshino J, Kuhne U, Kroger H: Methylation of nicotinamide in rat liver cytosol and its correlation with hepatocellular proliferation. Biochim Biophys Acta 1982;719:518-526. 12 Kang-Lee YA, McKee RW, Wright SM, Swendseid ME, Jenden DJ, Jope RS: Metabolic effects of nicotinamide administration in rats. J Nutr 1983;113:215-221. 13 Nakagawa K, Miyazaki M, Okui K, Kato N, Moriyama Y, Fujimura S: N1-methylnicotinamide level in the blood after nicotinamide loading as further evidence for malignant tumor burden. Jpn J Cancer Res 1991;82:1277-1283. 14 Naoi M, Dostert P, Yoshida M, Nagatsu T: N-methylated tetrahydroisoquinolines as dopaminergic neurotoxins. Adv Neurol 1993;60:212-217. 15 Yan L, Otterness DM, Weinshilboum RM: Human nicotinamide n-methyltransferase pharmacogenetics: Gene sequence analysis and promoter characterization. Pharmacogenetics 1999;9:307-316. 16 Roessler M, Rollinger W, Palme S, Hagmann ML, Berndt P, Engel AM, Schneidinger B, Pfeffer M, Andres H, Karl J, Bodenmuller H, Ruschoff J, Henkel T, Rohr G, Rossol S, Rosch W, Langen H, Zolg W, Tacke M: Identification of nicotinamide n-methyltransferase as a novel serum tumor marker for colorectal cancer. Clin Cancer Res 2005;11:6550-6557. 17 Xu J, Moatamed F, Caldwell JS, Walker JR, Kraiem Z, Taki K, Brent GA, Hershman JM: Enhanced expression of nicotinamide n-methyltransferase in human papillary thyroid carcinoma cells. J Clin Endocrinol Metab 2003;88:4990-4996. 18 Sartini D, Muzzonigro G, Milanese G, Pierella F, Rossi V, Emanuelli M: Identification of nicotinamide n-methyltransferase as a novel tumor marker for renal clear cell carcinoma. J Urol 2006;176:2248-2254. 19 Kitamura T, Asai N, Enomoto A, Maeda K, Kato T, Ishida M, Jiang P, Watanabe T, Usukura J, Kondo T, Costantini F, Murohara T, Takahashi M: Regulation of vegf-mediated angiogenesis by the akt/pkb substrate girdin. Nat Cell Biol 2008;10:329-337. 20 Yoeli-Lerner M, Toker A: Akt/pkb signaling in cancer: A function in cell motility and invasion. Cell Cycle 2006;5:603-605. 21 Kim D, Dan HC, Park S, Yang L, Liu Q, Kaneko S, Ning J, He L, Yang H, Sun M, Nicosia SV, Cheng JQ: Akt/pkb signaling mechanisms in cancer and chemoresistance. Front Biosci 2005;10:975-987. 22 Merseburger AS, Hennenlotter J, Kuehs U, Simon P, Kruck S, Koch E, Stenzl A, Kuczyk MA: Activation of pi3k is associated with reduced survival in renal cell carcinoma. Urol Int 2008;80:372-377. 23 Sternlicht MD, Werb Z: How matrix metalloproteinases regulate cell behavior. Annu Rev Cell Dev Biol 2001;17:463-516. 24 Stetler-Stevenson WG: The role of matrix metalloproteinases in tumor invasion, metastasis, and angiogenesis. Surg Oncol Clin N Am 2001;10:383-392, x. 25 Brinckerhoff CE, Matrisian LM: Matrix metalloproteinases: A tail of a frog that became a prince. Nat Rev Mol Cell Biol 2002;3:207-214. 26 Allgayer H, Babic R, Beyer BC, Grutzner KU, Tarabichi A, Schildberg FW, Heiss MM: Prognostic relevance of mmp-2 (72-kd collagenase iv) in gastric cancer. Oncology 1998;55:152-160. 27 Ellenrieder V, Alber B, Lacher U, Hendler SF, Menke A, Boeck W, Wagner M, Wilda M, Friess H, Buchler M, Adler G, Gress TM: Role of mt-mmps and mmp-2 in pancreatic cancer progression. Int J Cancer 2000;85:14-20. 28 Hanemaaijer R, Verheijen JH, Maguire TM, Visser H, Toet K, McDermott E, O'Higgins N, Duffy MJ: Increased gelatinase-a and gelatinase-b activities in malignant vs. Benign breast tumors. Int J Cancer 2000;86:204-207. 29 Kanayama H: Matrix metalloproteinases and bladder cancer. J Med Invest 2001;48:31-43. 30 Shimada T, Nakamura H, Yamashita K, Kawata R, Murakami Y, Fujimoto N, Sato H, Seiki M, Okada Y: Enhanced production and activation of progelatinase a mediated by membrane-type 1 matrix metalloproteinase in human oral squamous cell carcinomas: Implications for lymph node metastasis. Clin Exp Metastasis 2000;18:179-188. 31 Seifert R, Hoshino J, Kroger H: Nicotinamide methylation. Tissue distribution, developmental and neoplastic changes. Biochim Biophys Acta 1984;801:259-264. 32 Aoyama K, Matsubara K, Kondo M, Murakawa Y, Suno M, Yamashita K, Yamaguchi S, Kobayashi S: Nicotinamide-n-methyltransferase is higher in the lumbar cerebrospinal fluid of patients with parkinson's disease. Neurosci Lett 2001;298:78-80. 33 Iacobuzio-Donahue CA, Maitra A, Shen-Ong GL, van Heek T, Ashfaq R, Meyer R, Walter K, Berg K, Hollingsworth MA, Cameron JL, Yeo CJ, Kern SE, Goggins M, Hruban RH: Discovery of novel tumor markers of pancreatic cancer using global gene expression technology. Am J Pathol 2002;160:1239-1249. 34 Rogers CD, Fukushima N, Sato N, Shi C, Prasad N, Hustinx SR, Matsubayashi H, Canto M, Eshleman JR, Hruban RH, Goggins M: Differentiating pancreatic lesions by microarray and qpcr analysis of pancreatic juice rnas. Cancer Biol Ther 2006;5:1383-1389. 35 Lim BH, Cho BI, Kim YN, Kim JW, Park ST, Lee CW: Overexpression of nicotinamide n-methyltransferase in gastric cancer tissues and its potential post-translational modification. Exp Mol Med 2006;38:455-465. 36 Jang JS, Cho HY, Lee YJ, Ha WS, Kim HW: The differential proteome profile of stomach cancer: Identification of the biomarker candidates. Oncol Res 2004;14:491-499. 37 Xu J, Capezzone M, Xu X, Hershman JM: Activation of nicotinamide n-methyltransferase gene promoter by hepatocyte nuclear factor-1beta in human papillary thyroid cancer cells. Mol Endocrinol 2005;19:527-539. 38 Strongin AY, Collier I, Bannikov G, Marmer BL, Grant GA, Goldberg GI: Mechanism of cell surface activation of 72-kda type iv collagenase. Isolation of the activated form of the membrane metalloprotease. J Biol Chem 1995;270:5331-5338. 39 Deryugina EI, Ratnikov B, Monosov E, Postnova TI, DiScipio R, Smith JW, Strongin AY: Mt1-mmp initiates activation of pro-mmp-2 and integrin alphavbeta3 promotes maturation of mmp-2 in breast carcinoma cells. Exp Cell Res 2001;263:209-223. 40 Qin H, Sun Y, Benveniste EN: The transcription factors sp1, sp3, and ap-2 are required for constitutive matrix metalloproteinase-2 gene expression in astroglioma cells. J Biol Chem 1999;274:29130-29137. 41 Carnero A, Blanco-Aparicio C, Renner O, Link W, Leal JF: The pten/pi3k/akt signalling pathway in cancer, therapeutic implications. Curr Cancer Drug Targets 2008;8:187-198. 42 Vivanco I, Sawyers CL: The phosphatidylinositol 3-kinase akt pathway in human cancer. Nat Rev Cancer 2002;2:489-501. 43 Park MJ, Kwak HJ, Lee HC, Yoo DH, Park IC, Kim MS, Lee SH, Rhee CH, Hong SI: Nerve growth factor induces endothelial cell invasion and cord formation by promoting matrix metalloproteinase-2 expression through the phosphatidylinositol 3-kinase/akt signaling pathway and ap-2 transcription factor. J Biol Chem 2007;282:30485-30496. 44 Lee WJ, Chen WK, Wang CJ, Lin WL, Tseng TH: Apigenin inhibits hgf-promoted invasive growth and metastasis involving blocking pi3k/akt pathway and beta 4 integrin function in mda-mb-231 breast cancer cells. Toxicol Appl Pharmacol 2008;226:178-191. 45 Yasuda A, Sawai H, Takahashi H, Ochi N, Matsuo Y, Funahashi H, Sato M, Okada Y, Takeyama H, Manabe T: Stem cell factor/c-kit receptor signaling enhances the proliferation and invasion of colorectal cancer cells through the pi3k/akt pathway. Dig Dis Sci 2007;52:2292-2300. 46 Li J, Yen C, Liaw D, Podsypanina K, Bose S, Wang SI, Puc J, Miliaresis C, Rodgers L, McCombie R, Bigner SH, Giovanella BC, Ittmann M, Tycko B, Hibshoosh H, Wigler MH, Parsons R: Pten, a putative protein tyrosine phosphatase gene mutated in human brain, breast, and prostate cancer. Science 1997;275:1943-1947. | |
| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/42317 | - |
| dc.description.abstract | 腎細胞癌（Renal cell carcinoma）占成人所有癌症的百分之二，其中又以透明細胞（clear cell）腎細胞癌為最主要及最具侵略性之亞型。由於腎細胞癌對於傳統的化學治療或是輻射線治療等方式具有高度的抗性，所以手術切除仍是目前主要的治療方式；此外，當病人腎細胞癌有轉移現象出現後，五年內存活率將低於百分之二十，且臨床上至今也尚未發展出可作為預測診斷或治療癒後的分子指標。因此，我們希望藉由鑑定與腎細胞癌形成相關之基因來了解其致癌之分子機制並期望發展出新的診斷及治療方法。在先前的研究中，本實驗室已成功建構透明細胞腎細胞癌組織以及正常腎臟組織之全長互補去氧核醣核酸（cDNA）基因庫，藉由分析兩基因庫之基因表現，共比對出201個過度表現與182個表現受抑制之相關基因，而菸鹼胺-氮-甲基轉移 | zh_TW |
| dc.description.abstract | Renal cell carcinoma (RCC) accounts for 2 ~ 3% of all adult malignancies, and clear cell renal cell carcinoma (ccRCC) is the major and aggressive subtype of RCC. Because RCC responds poorly to traditional therapies such as chemotherapy and radiotherapy, surgery remains to be the main remedy. Therefore, it is important to identify RCC-associated genes involved in tumorigenesis and finally develop new approaches to diagnosis and therapy. In our previous studies, we have identified 201 up-regulated and 182 down-regulated genes in ccRCC by analyzing full-length enriched cDNA libraries of ccRCC and normal kidney. Nicotinamide N-methyltransferase (NNMT), one of the most up-regulated genes, was found to be overexpressed in ccRCC. Besides, recent studies have demonstrated that NNMT overexpression was also found in thyroid and colorectal tumors. Hence, the specific aim of our research is to investigate the roles of NNMT overexpression in ccRCC.
In this study, we used human embryonic kidney cells (HEK293) without endogenous NNMT expression for the following experiments. By using quantitative real-time PCR analysis, we observed that mRNA expression of matrix metalloprotease-2 (MMP-2) was up-regulated in the HEK293/NNMT cells. The results of western blot and gelatin zymography also showed that HEK293/NNMT cells had higher MMP-2 secretion. To further investigate the phenotype caused by MMP-2 up-regulation, we performed in vitro cell invasion assays to study the invasive activities with the HEK293/NNMT and HEK293/VEC cells, and the results revealed that NNMT overexpression promoted the HEK293 cell invasiveness. Besides, the invasive ability was suppressed by either MMP-2 inhibitor or MMP-2 siRNA. To delineate signaling pathway involved in MMP-2 up-regulation induced by NNMT, we used specific inhibitors to impede many signaling pathways and observed that either PI3K inhibitor (LY294002) or Akt inhibitor IV inhibited MMP-2 expression and MMP-2 promoter activity, and suppressed cell invasive ability. In addition, knockdown of Akt or NNMT by specific siRNAs inhibited MMP-2 expression and suppressed cell invasive ability. Taken together, our data suggest that NNMT overexpression might contribute to invasive phenotype in ccRCC by inducing MMP-2 expression via PI3K/Akt signaling pathway. | en |
| dc.description.provenance | Made available in DSpace on 2021-06-15T00:59:39Z (GMT). No. of bitstreams: 1 ntu-97-R95442012-1.pdf: 3985316 bytes, checksum: 4fbfdf59498fa53d51e64ee587417667 (MD5) Previous issue date: 2008 | en |
| dc.description.tableofcontents | 致謝 ii
Contents iii Abbreviations ix 摘要 xi Abstract xiii Introduction 1 Materials and Methods 6 1. Materials 6 2. Cell Culture 7 3. MTT Assay 7 4. RNA Extraction and Reverse Transcription 8 4.1 RNAs extraction 8 4.2 DNase treatment 9 4.3 Reverse transcription 9 5. Quantitative Real-time PCR（Q-PCR） 10 5.1 Real-time PCR assay 10 5.2 Quantification for expressive level of genes 11 6. Construction of Deleted MMP-2 Promoter 12 6.1 Preparation of Luria-Bertani (LB) medium and plates 12 6.2 Extraction of plasmid 12 6.3 Amplification of fragment of MMP-2 promoter by polymerase chain reaction (PCR) 12 6.4 Extraction of PCR and digestion DNA products by agarose gel electrophoresis 13 6.5 Cloning PCR products into yT&A cloning vector 14 6.6 Transformation 14 6.7 Colony PCR and restriction enzyme digestion 15 6.8 Ligation of digested products to pGL3-basic luciferase reporter vector. 16 7. Cell Invasion Assay 16 7.1 Preparation of cells and matrigel-coding chamber 16 7.2 Fixation and Staining 17 8. Transient transfection 17 9. Western blot 18 9.1 Preparation of cell lysates 18 9.2 Quantification of protein concentration by bicinchoninic (BCA) assay 19 9.3 Sodium-dodecyl-sulphate polyacrylamide gel electrophoresis (SDS-PAGE) 19 9.3.1 Preparation of SDS-polyacrylamide gels 20 9.3.2 Preparation of protein samples and performance of polyacrylamide gel electrophoresis 20 9.4 Transferring the protein by using Semi-Dry System 21 9.5 Immunoblotting 22 10. Gelatin Zymography 23 10.1 Sample preparation 23 10.2 Preparation of gelatin gel and performance of gel electrophoresis 24 10.3 Gel electrophoresis and development 24 10.4 Staining and de-staining of gel 25 11. Luciferase reporter assay 25 11.1 Preparation of cell lysates 25 11.2 Detection of luciferase activity 26 Results 27 1. Overexpression of NNMT has no effects on proliferation rate but up-regulates MMP-2 in HEK293 cells. 27 2. Augmentation of MMP-2 by NNMT over-expression promotes invasive ability of HEK293 cells. 29 3. NNMT-induced MMP-2 up-regulation and higher invasiveness are mediated by activation of the PI3K/Akt signaling pathway. 30 4. Inhibitors of PI3K and Akt significantly suppressed MMP-2 promoter activity. 31 5. Knockdown of Akt suppresses NNMT-mediated MMP-2 up-regulation and invasive ability in HEK293 cells. 32 6. Knockdown of NNMT suppresses MMP-2 up-regulation and invasive ability in HEK293/NNMT cells. 33 7. Possible transcription factors for enhancing MMP-2 promoter activity. 34 Discussion 35 Figures 41 Legends 54 References 63 | |
| dc.language.iso | en | |
| dc.subject | 菸鹼胺-氮-甲基轉移 | zh_TW |
| dc.subject | 腎細胞癌 | zh_TW |
| dc.subject | Nicotinamide N-methyltransferase | en |
| dc.subject | Akt | en |
| dc.subject | PI3K | en |
| dc.subject | Matrix metalloprotease-2 | en |
| dc.subject | Renal cell carcinoma | en |
| dc.title | 菸鹼胺-氮-甲基轉移酶藉由誘導第二型基質金屬蛋白酶表現促進透明細胞型腎細胞癌之侵犯能力 | zh_TW |
| dc.title | Nicotinamide-N-methyltransferase Enhances Invasive Ability of Clear Cell Renal Cell Carcinoma by Inducing Matrix Metalloprotease-2 Expression | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 96-2 | |
| dc.description.degree | 碩士 | |
| dc.contributor.oralexamcommittee | 呂紹俊(Shao-Chun Lu),周玉山(Yuh-Shan Jou) | |
| dc.subject.keyword | 腎細胞癌,菸鹼胺-氮-甲基轉移, | zh_TW |
| dc.subject.keyword | Renal cell carcinoma,Nicotinamide N-methyltransferase,Matrix metalloprotease-2,PI3K,Akt, | en |
| dc.relation.page | 66 | |
| dc.rights.note | 有償授權 | |
| dc.date.accepted | 2008-08-01 | |
| dc.contributor.author-college | 醫學院 | zh_TW |
| dc.contributor.author-dept | 生物化學暨分子生物學研究所 | zh_TW |
| 顯示於系所單位： | 生物化學暨分子生物學科研究所 | |
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