Please use this identifier to cite or link to this item: http://hdl.handle.net/11455/25058
標題: 應用反轉錄聚合酶鏈反應測定基質金屬蛋白酶及其抑制因子在人類乳癌石蠟組織中之表現
mRNA expressions of matrix metalloproteinases and tissue inhibitor metalloproteinases on formalin fixed paraffin embedded human breast tumor tissues using RT-PCR
作者: 謝昭漢
Hsieh, Chao-Han
關鍵字: 基質金屬蛋白西每;matrix metalloproteinases (MMP);基質金屬蛋白西每抑制因子;甲醛固定石蠟包埋組織;反轉錄聚合西每鏈反應;tissue inhibitors of metalloproteinases (TIMP);formalin fixation paraffin embedding (FFPE);reverse transcription polymerase chain reaction (RT-PCR)
出版社: 動物科學系所
引用: Abrahamsen, H. N., T. Steiniche, E. Nexo, S. J. Hamilton-Dutoit, and B. S. Sorensen. 2003. Towards quantitative mRNA analysis in paraffin-embedded tissues using real-time reverse transcriptase-polymerase chain reaction: a methodological study on lymph nodes from melanoma patients. J. Mol. Diagn. 5: 34-41. Ahokas, K, J. Lohi, H. Lohi, O. Elomaa, M. L. Karjalainen-Lindsberg, J. Kere, and U. Saarialho-Kere. 2002. Matrix metalloproteinase-21, the human orthologue for XMMP, is expressed during fetal development and in cancer. Gene. 301:31-41. Alvarez, O. A., D. F. Carmichael, and Y. A. DeClerck. 1990. Inhibition of collagenolytic activity and metastasis of tumor cells by a recombinant human tissue inhibitor of metalloproteinases. J. Natl. Cancer Inst. 82:589-595. Amălinei, C., I. D. Căruntu, S. E. Giuşcă, and R. A. Bălan. 2010. Matrix metalloproteinases involvement in pathologic conditions. Rom. J. Morphol. Embryol. 51: 215-228. Antonov, J., D. R. Goldstein, A. Oberli, A. Baltzer, M. Pirotta, A. Fleischmann, H. J. Altermatt, and R. Jaggi. 2005. Reliable gene expression measurements from degraded RNA by quantitative real-time PCR depend on short amplicons and a proper normalization. Lab. Invest. 85: 1040-1050. Aresu, L., M. Giantin, E. Morello, M. Vascellari, M. Castagnaro, R. Lopparelli, V. Zancanella, A. Granato, S. Garbisa, A. Arico, A. Bradaschia, F. Mutinelli, and M. Dacasto. 2011. Matrix metalloproteinases and their inhibitors in canine mammary tumors. BMC Vet. Res. 7: 33. Auerbach, C, M. Moutschen-Dahmen, and J. Moutschen. 1977. Genetic and cytogenetical effects of formaldehyde and related compounds. Mutat. Res. 39: 317-361. Atkinson, J.M., C. J. Pennington, S. W. Martin, V. A. Anikin, A. J. Mearns, P. M. Loadman, D. R. Edwards, and J. H. Gill. 2007. Membrane type matrix metalloproteinases (MMPs) show differential expression in non-small cell lung cancer (NSCLC) compared to normal lung: correlation of MMP-14 mRNA expression and proteolytic activity. Eur. J. Cancer. 43: 1764-1771. Baker, A.H., D. R. Edwards, and G. Murphy. 2002. Metalloproteinase inhibitors: biological actions and therapeutic opportunities. J. Cell. Sci. 115: 3719-3727. Bender, C., D. Zipeto, C. Bidoia, S. Costantini, A. Zamo, F. Menestrina, and U. Bertazzoni. 2009. Analysis of colorectal cancers for human cytomegalovirus presence. Infect. Agent. Cancer. 4: 6. Bonin, S., F. Hlubek, J. Benhattar, C. Denkert, M. Dietel, P. L. Fernandez, G. Hofler, H. Kothmaier, B. Kruslin, C. M. Mazzanti, A. Perren, H. Popper, A. Scarpa, P. Soares, G. Stanta, and P. J. Groenen. 2010. Multicentre validation study of nucleic acids extraction from FFPE tissues. Virchows Arch. 457: 309-317. Bresters, D., M. E. Schipper, H. W. Reesink, B. D. Boeser-Nunnink, and H. T. Cuypers. 1994. The duration of fixation influences the yield of HCV cDNA-PCR products from formalin-fixed, paraffin-embedded liver tissue. J. Virol. Methods. 48:267-272. Brummer, O., S. Athar, L. Riethdorf, T. Loning, and H. Herbst. 1999. Matrix-metalloproteinases 1, 2, and 3 and their tissue inhibitors 1 and 2 in benign and malignant breast lesions: an in situ hybridization study. Virchows Arch. 435: 566-573. Canete-Soler, R., L. Litzky, I. Lubensky, and R. J. Muschel. 1994. Localization of the 92 kd gelatinase mRNA in squamous cell and adenocarcinomas of the lung using in situ hybridization. Am. J. Pathol. 144: 518-527. Chambers, A. F., and L. M. Matrisian. 1997. Changing views of the role of matrix metalloproteinases in metastasis. J. Natl. Cancer Inst. 89: 1260-1270. Chang, T.W., and Y. L. Kuo. 2010. A model building exercise of mortality risk for Taiwanese women with breast cancer. BMC Med. Inform. Decis. Mak. 10: 43. Chaussain-Miller, C., F. Fioretti, M. Goldberg, and S. Menashi. 2006. The role of matrix metalloproteinases (MMPs) in human caries. J. Dent. Res. 85: 22-32. Chirco, R., X. W. Liu, K. K. Jung, and H. R. Kim. 2006. Novel functions of TIMPs in cell signaling. Cancer Metastasis Rev. 25: 99-113. Chu, W. S., Q. Liang, Y. Tang, R. King, K. Wong, M. Gong, M. Wei, J. Liu, S. H. Feng, S. C. Lo, J. A. Andriko, and M. Orr. 2006. Ultrasound-accelerated tissue fixation/processing achieves superior morphology and macromolecule integrity with storage stability. J. Histochem. Cytochem. 54: 503-513. Cronin, M., M. Pho, D. Dutta, J. C. Stephans, S. Shak, M. C. Kiefer, J. M. Esteban, and J. B. Baker. 2004. Measurement of gene expression in archival paraffin-embedded tissues: development and performance of a 92-gene reverse transcriptase-polymerase chain reaction assay. Am. J. Pathol. 164: 35-42. Davidson, B., S. Konstantinovsky, S. Nielsen, H. P. Dong, A. Berner, M. Vyberg, and R. Reich. 2004. Altered expression of metastasis-associated and regulatory molecules in effusions from breast cancer patients: a novel model for tumor progression. Clin. Cancer Res. 10: 7335-7346. Davies, B., D. W. Miles, L. C. Happerfield, M. S. Naylor, L. G. Bobrow, R. D. Rubens, Balkwill. F. R. 1993. Activity of type IV collagenases in benign and malignant breast disease. Br. J. Cancer. 67: 1126-1131. del Casar, J. M., G. Carreno, L. O. Gonzalez, S. Junquera, S. Gonzalez-Reyes, J. M. Gonzalez, M. Bongera, A.M. Merino, and F. J. Vizoso. 2010. Expression of metalloproteases and their inhibitors in primary tumors and in local recurrences after mastectomy for breast cancer. J. Cancer Res. Clin. Oncol.136: 1049-1058. Duffy, M. J. 1992. The role of proteolytic enzymes in cancer invasion and metastasis. Clin. Exp. Metastasis. 10: 145-155. Farragher, S. M., A. Tanney, R. D. Kennedy, and D. Paul Harkin. 2008. RNA expression analysis from formalin fixed paraffin embedded tissues. Histochem. Cell Biol. 130: 435-45. Feldman, M. Y. 1973. Reactions of nucleic acids and nucleoproteins with formaldehyde. Prog. Nucleic Acid Res. Mol. Biol. 13: 1-49. Feng, Y., B. Sun, X. Li, L. Zhang, Y. Niu, C. Xiao, L. Ning, Z. Fang, Y. Wang, L. Zhang, J. Cheng, W. Zhang, and X. Hao. 2007. Differentially expressed genes between primary cancer and paired lymph node metastases predict clinical outcome of node-positive breast cancer patients. Breast Cancer Res. Treat. 103: 319-329. Figueira, R. C., L. R. Gomes, J. S. Neto, F. C. Silva, I. D. Silva, and M. C. Sogayar. 2009. Correlation between MMPs and their inhibitors in breast cancer tumor tissue specimens and in cell lines with different metastatic potential. BMC Cancer. 14: 9-20. Finke, J., R. Fritzen, P. Ternes, W. Lange, and G. Dolken. 1993. An improved strategy and a useful housekeeping gene for RNA analysis from formalin-fixed, paraffin-embedded tissues by PCR. Biotechniques. 14: 448-453. Flenniken, A. M., and B. R. Williams. 1990. Developmental expression of the endogenous TIMP gene and a TIMP-lacZ fusion gene in transgenic mice. Genes Dev. 4: 1094-1106. Forsyth, P.A., H. Wong, T. D. Laing, N. B. Rewcastle, D. G. Morris, H. Muzik, K. J. Leco, R. N. Johnston, P. M. Brasher, G. Sutherland, and D. R. Edwards. 1999. Gelatinase-A (MMP-2), gelatinase-B (MMP-9) and membrane type matrix metalloproteinase-1 (MT1-MMP) are involved in different aspects of the pathophysiology of malignant gliomas. Br J Cancer. 79:1828-1835. Fujita, N., and P. A. Wade. 2004. Use of bifunctional cross-linking reagents in mapping genomic distribution of chromatin remodeling complexes. Methods. 33: 81-85. Galis, Z. S., G. K. Sukhova, M. W. Lark, and P. Libby. 1994. Increased expression of matrix metalloproteinases and matrix degrading activity in vulnerable regions of human atherosclerotic plaques. J. Clin. Invest. 94: 2493-2503. Gialeli, C., A. D. Theocharis, and N. K. Karamanos. 2011. Roles of matrix metalloproteinases in cancer progression and their pharmacological targeting. FEBS J.278: 16-27. Giannelli, G., C. Bergamini, F. Marinosci, E. Fransvea, M. Quaranta, L. Lupo, O. Schiraldi, and S. Antonaci. 2002. Clinical role of MMP-2/TIMP-2 imbalance in hepatocellular carcinoma. Int. J. Cancer. 97:425-431. Gnanapragasam, V. J. 2010. Unlocking the molecular archive: the emerging use of formalin-fixed paraffin-embedded tissue for biomarker research in urological cancer. BJU Int. 105:274-278. Godfrey, T. E., S. H. Kim, M. Chavira, D. W. Ruff, R.S. Warren, J. W. Gray, and R. H. Jensen. 2000. Quantitative mRNA expression analysis from formalin-fixed, paraffin-embedded tissues using 5'' nuclease quantitative reverse transcription polymerase chain reaction. J. Mol. Diagn. 2: 84-91. Gomez, D. E., D. F. Alonso, H. Yoshiji, and U. P. Thorgeirsson. 1997. Tissue inhibitors of metalloproteinases: structure, regulation and biological functions. Eur. J. Cell Biol. 74: 111-122. Greenlee, K. J., Z. Werb, and F. Kheradmand. 2007. Matrix metalloproteinases in lung: multiple, multifarious, and multifaceted. Physiol. Rev. 87: 69-98. Gross, J., and C. M. Lapierre. 1962. Collagenolytic activity in amphibian tissues: a tissue culture assay. Proc. Natl. Acad. Sci. 48: 1014-1022. Hawinkels, L. J., P. Kuiper, E. Wiercinska, H. W. Verspaget, Z. Liu, E. Pardali, C. F. Sier, and P. ten Dijke. 2010. Matrix metalloproteinase-14 (MT1-MMP)-mediated endoglin shedding inhibits tumor angiogenesis. Cancer Res. 15;70(10):4141-50. Huang, W. Y., T. M. Sheehy, L. E. Moore, A. W. Hsing, and M. P. Purdue. 2010. Simultaneous recovery of DNA and RNA from formalin-fixed paraffin-embedded tissue and application in epidemiologic studies. Cancer Epidemiol. Biomarkers Prev. 19: 973-977. Iwata, H., S. Kobayashi, H. Iwase, A. Masaoka, N. Fujimoto, and Okada Y. 1996. Production of matrix metalloproteinases and tissue inhibitors of metalloproteinases in human breast carcinomas. Jpn. J. Cancer Res. 87: 602–611. Jackson, D. P., P. Quirke, F. Lewis, A. W. Boylson, J. M. Sloan, D. Robertson, and G. R. Taylor. 1989. Detection of measules virus RNA in paraffin-embedded tissues. Lancet. 33: 1391. Jones, J. L., P. Glynn, and R. A. Walker. 1999. Expression of MMP-2 and MMP-9, their inhibitors and the activator MT1-MMP in primary breast carcinomas. J. Pathol. 189: 161–168. Katori, H., A. Nozawa, and M. Tsukuda. 2006. Increased expression of matrix metalloproteinase-2 and 9 and human papilloma virus infection are associated with malignant transformation of sinonasal inverted papilloma. J. Surg. Oncol. 93:80-85. Kessenbrock, K., V. Plaks, and Z. Werb. 2010. Matrix metalloproteinases: regulators of the tumor microenvironment. Cell. 141: 52-67. Kevorkian L, D. A. Young, C. Darrah, S. T. Donell, L. Shepstone, S. Porter, Brockbank S. M., D. R. Edwards, A. E. Parker, and I. M. Clark. 2004. Expression profiling of metalloproteinases and their inhibitors in cartilage. Arthritis. Rheum. 50: 131-141. Khokha, R. 1994. Suppression of the tumorigenic and metastatic abilities of murine B16-F10 melanoma cells in vivo by the overexpression of the tissue inhibitor of the metalloproteinases-1. J. Natl. Cancer Inst. 86: 299-304. Kiernan, J. A. 2000. Formaldehyde, formalin, paraformaldehyde and glutaraldehyde: What they are and what they do. Microsc. Today. 00: 8-12. Kim, H. J., C. I. Park, B. W. Park, H. D. Lee, and W. H. Jung. 2006. Expression of MT-1 MMP, MMP2, MMP9 and TIMP2 mRNAs in ductal carcinoma in situ and invasive ductal carcinoma of the breast. Yonsei Med. J. 47: 333-342. Kohrmann, A., U. Kammerer, M. Kapp, J. Dietl, and J. Anacker. 2009. Expression of matrix metalloproteinases (MMPs) in primary human breast cancer and breast cancer cell lines: New findings and review of the literature. BMC Cancer. 9: 188. Kossakowska, A. E., S. A. Huchcroft, S. J. Urbanski, D. R. Edwards. 1996. Comparative analysis of the expression patterns of metalloproteinases and their inhibitors in breast neoplasia, sporadic colorectal neoplasia, pulmonary carcinomas and malignant non-Hodgkin''s lymphomas in humans. Br. J. Cancer. 73: 1401-1408. Kuzuya, M., and A. Iguchi. 2003. Role of matrix metalloproteinases in vascular remodeling. J. Atheroscler. Thromb. 10: 275-282. Kwon, M. J., E. Oh, S. Lee, M. R. Roh, S. E. Kim, Y. Lee, Y. L. Choi, Y. H. In, T. Park, S. S. Koh, and Y. K. Shin. 2009. Identification of novel reference genes using multiplatform expression data and their validation for quantitative gene expression analysis. PLoS One. 4: e6162. Lambert, E., L. Bridoux, J. Devy, E. Dasse, M. L. Sowa, L. Duca, W. Hornebeck, L. Martiny, and E. Petitfrere-Charpentier. 2009. TIMP-1 binding to proMMP-9/CD44 complex localized at the cell surface promotes erythroid cell survival. Int. J. Biochem. Cell Biol. 41: 1102-1115. Lebeau, A., C. Muller-Aufdemkamp, C. Allmacher, U. Sauer, A. Nerlich, R. Lichtinghagen, and U. Lohrs. 2004. Cellular protein and mRNA expression patterns of matrix metalloproteinases-2, -3 and -9 in human breast cancer: correlation with tumour growth. J. Mol. Histol. 35: 443-455. Leonard, G. D., and S. M. Swain. 2004. Ductal carcinoma in situ, complexities and challenges. J. Natl. Cancer Inst. 96: 906-920. Li, Z., M. P. Clarke, M. D. Barker, and N. McKie. 2005. TIMP3 mutation in Sorsby''s fundus dystrophy: molecular insights. Expert Rev. Mol. Med. 7: 1-15. Liabakk, N. B., I. Talbot, R. A. Smith, K. Wilkinson, and F. Balkwill. 1996. Matrix metalloprotease 2 (MMP-2) and matrix metalloprotease 9 (MMP-9) type IV collagenases in colorectal cancer. Cancer Res. 56: 190-196. Liotta, L. A., W. G. Stetler-Stevenson. 1991. Tumor invasion and metastasis: an imbalance of positive and negative regulation. Cancer Res. 51: 5054-5059. Liu, Y. E., M. Wang, J. Greene, J. Su, S. Ullrich, H. Li, S. Sheng, P. Alexander, Q. A. Sang, and Y. E. Shi. 1997. Preparation and characterization of recombinant tissue inhibitor of metalloproteinase 4 (TIMP-4). J. Biol. Chem. 272: 20479-20483. Mannello, F., and G. Gazzanelli. 2001. Tissue inhibitors of metalloproteinases and programmed cell death: conundrums, controversies and potential implications. Apoptosis. 6: 479-482. Mannello, F., and M. Sebastiani. 2003. Zymographic analyses and measurement of matrix metalloproteinase-2 and -9 in nipple aspirate fluids. Clin. Chem. 49: 1546-1550. Massova, I., L. P. Kotra, R. Fridman, and S. Mobashery. 1998. Matrix metalloproteinases: Structures, evolution, and diversification. FASEB J. 12: 1075-1095. Masuda, N., T. Ohnishi, S. Kawamoto, M. Monden, and K. Okubo. 1999. Analysis of chemical modification of RNA from formalin-fixed samples and optimization of molecular biology applications for such samples. Nucleic Acids Res. 27: 4436-4443. McGhee, J. D., and P. H. von Hippel. 1977. Formaldehyde as a probe of DNA structure. 3. Equilibrium denaturation of DNA and synthetic polynucleotides. Biochem. 16: 3267-3276. Melendez-Zajgla, J., L. Del Pozo, G. Ceballos, and V. Maldonado. 2008. Tissue inhibitor of metalloproteinases-4. The road less traveled. Mol. Cancer. 7: 85. Mendes, O., H. T. Kim, and G. Stoica. 2005. Expression of MMP2, MMP9 and MMP3 in breast cancer brain metastasis in a rat model. Clin. Exp. Metastasis. 22: 237-246. Mimori, K., H. Ueo, C. Shirasaka, M. Mori. 2001. Clinical significance of MT1-MMP mRNA expression in breast cancer. Oncol. Rep. 8: 401-403. Mori, R., Q. Wang, K.D. Danenberg, J. K. Pinski, and P. V. Danenberg. 2008. Both beta-actin and GAPDH are useful reference genes for normalization of quantitative RT-PCR in human FFPE tissue samples of prostate cancer. Prostate. 68: 1555-1560. Murphy, G., V. Knauper, S. Cowell, R. Hembry, H. Stanton, G. Butler, J. Freije, A. M. Pendas, and C. Lopez-Otin. 1999. Evaluation of some newer matrix metalloproteinases. Ann. N. Y. Acad. Sci. 878: 25-39. Nakopoulou, L., I. Tsirmpa, P. Alexandrou, A. Louvrou, C. Ampela, S. Markaki, and P. S. Davaris. 2003. MMP-2 protein in invasive breast cancer and the impact of MMP-2/TIMP-2 phenotype on overall survival. Breast Cancer Res. Treat. 77: 145-55. Newby, A.C., M. Pauschinger, and F. G. Spinale. 2006. From tadpole tails to transgenic mice: metalloproteinases have brought about a metamorphosis in our understanding of cardiovascular disease. Cardiovasc. Res. 69: 559-561. Noseri, H., T. Erden, S. Toros, M. Habesoglu, E. Egeli, F. Aker, and S. Cetin. 2007. Intraductal papilloma of the parotid gland in a child. Eur. Arch. Otorhinolaryngol. 264: 1385-1386. Oberli, A., V. Popovici, M. Delorenzi, A. Baltzer, J. Antonov, S. Matthey, S. Aebi, H. J. Altermatt, and R. Jaggi. 2008. Expression profiling with RNA from formalin-fixed, paraffin-embedded material. BMC Med. Genomics. 1-9. Ohuchi, E., K. Imai, Y. Fujii, H. Sato, M. Seiki, Y. Okada. 1997. Membrane type1 matrix metalloproteinase digests interstitial collagens and other extracellular matrix macromolecules. J. Biol. Chem. 272: 2446-2451. Onisto, M., M. P. Riccio, P. Scannapieco, C. Caenazzo, L. Griggio, M. Spina, W. G. Stetler-Stevenson, and S. Garbisa. 1995. Gelatinase A/TIMP-2 imbalance in lymph-node-positive breast carcinomas, as measured by RT-PCR. Int. J. Cancer. 63: 621-626. O’Shaughnessy, J. 2005. Extending survival with chemotherapy in metastatic breast cancer. Oncologist. 10: 20-29. Papon, J. F., E. Lechapt-Zalcman, M. Abina, I. Abd-al-Sama, R. Peynegre, E. Escudier, and A. Coste. 2006. Matrix metalloproteinase-2 and -9 expression in sinonasal inverted papilloma. Rhinology. 44: 211-215. Park, Y. N., K. Abe, H. Li, T. Hsuih, S. N. Thung, and D. Y. Zhang. 1996. Detection of hepatitis C virus RNA using ligation-dependent polymerase chain reaction in formalin-fixed, paraffin-embedded liver tissues. Am. J. Pathol. 149: 1485-1491. Pilka, R, C. Whatling, H. Domanski, S. Hansson, P. Eriksson, and B. Casslen. 2003. Epithelial expression of matrix metalloproteinase-26 is elevated at mid-cycle in the human endometrium. Mol. Hum. Reprod. 9: 271-277. Ree ,A. H., V. A. Florenes, J. P. Berg, G. M. Maelandsmo, J. M. Nesland, and O. Fodstad. 1997. High levels of messenger RNAs for tissue inhibitors of metalloproteinases (TIMP-1 and TIMP-2) in primary breast carcinomas are associated with development of distant metastases. Clin. Cancer Res. 3: 1623-8. Ribeiro-Silva, A., H. Zhang, and S. S. Jeffrey. 2007. RNA extraction from ten year old formalin-fixed paraffin-embedded breast cancer samples: a comparison of column purification and magnetic bead-based technologies. BMC Mol. Biol. 8: 118. Ring, P., K. Johansson, M. Hoyhtya, K. Rubin, and G. Lindmark. 1997. Expression of tissue inhibitor of metalloproteinases TIMP-2 in human colorectal cancer: a predictor of tumour stage. Br. J. Cancer. 76: 805-811. Saarialho-Kere, U., E. Kerkela, T. Jahkola, S. Suomela, J. Keski-Oja, and J. Lohi. 2002. Epilysin (MMP-28) expression is associated with cell proliferation during epithelial repair. J. Invest. Dermatol. 119: 14-21. Sato, H., and T. Takino. 2010. Coordinate action of membrane-type matrix metalloproteinase-1 (MT1-MMP) and MMP-2 enhances pericellular proteolysis and invasion. Cancer Sci. 101: 843-847. Seo, D. W., H. Li, L. Guedez, P. T. Wingfield, T. Diaz, R. Salloum, B. Y. Wei, and W. G. Stetler-Stevenson. 2003. TIMP-2 mediated inhibition of angiogenesis: an MMP-independent mechanism. Cell. 114: 171-180. Shao, W., W. Wang, X. G. Xiong, C. Cao, T. D. Yan, G. Chen, H. Chen, W. Yin, J. Liu, Y. Gu, M. Mo, and J. He. 2011. Prognostic impact of MMP-2 and MMP-9 expression in pathologic stage IA non-small cell lung cancer. J. Surg. Oncol. Shibata, D. K., N. Arnheim, and W. J. Martin. 1988. Detection of human papilloma virus in paraffin-embedded tissue using the polymerase chain reaction. J. Exp. Med. 167: 225-230. Shipley, J. M., R. L. Wesselschmidt, D. K. Kobayashi, T. J. Ley, S. D. Shapiro. 1996. Etalloelastase is required for macrophage-mediated proteolysis and matrix invasion in mice. Proc. Natl. Acad. Sci. 93: 3942-3946. Sobral, L. M., A. Bufalino, M. A. Lopes, E. Graner, T. Salo, and R. D. Coletta. 2011. Myofibroblasts in the stroma of oral cancer promote tumorigenesis via secretion of activin A. Oral Oncol. Sopata, I. and J. Wize. 1979. A latent gelatin specific proteinase of human leucocytes and its activation. Biochim. Biophys. Acta. 571: 305-312. Stanta, G., and C. Schneider. 1991. RNA extracted from paraffin-embedded human tissues is amenable to analysis by PCR amplification. Biotechniques. 11: 304-308. Sun, Y., N. Lu, Y. Ling, Y. Gao, Y. Chen, L. Wang, R. Hu, Q. Qi, W. Liu, Y. Yang, Q. You, and Q. Guo. 2009. Oroxylin A suppresses invasion through down-regulating the expression of matrix metalloproteinase-2/9 in MDA-MB-435 human breast cancer cells. Eur. J. Pharmacol. 603: 22-28. Tapio S., and M. J. Atkinson. 2008. Molecular information obtained from radiobiological tissue archives: achievements of the past and visions of the future. Radiat. Environ. Biophys. 47:183-187. Tetu, B., J. Brisson, C. S. Wang, H. Lapointe, G. Beaudry, C. Blanchette, and D. Trudel. 2006. The influence of MMP-14, TIMP-2 and MMP-2 expression on breast cancer prognosis. Breast Cancer Res. 8: R28. Tsuchiya, Y., H. Sato, Y. Endo, Y. Okada, M. Mai, T. Sasaki, and M. Seiki. 1993. Tissue inhibitor of metalloproteinase 1 is a negative regulator of the metastatic ability of a human gastric cancer cell line, KKLS, in the chick embryo. Cancer Res. 53: 1397-1402. Ueng, S. H., T. Mezzetti, and F. A. Tavassoli. 2009. Papillary neoplasms of the breast: a review. Arch. Pathol. Lab. Med. 133: 893-907. Ueno, H., H. Nakamura, M. Inoue, K. Imai, M. Noguchi, H. Sato, M. Seiki, Y. Okada. 1997. Expression and tissue localization of membrane-types 1, 2, and 3 matrix metalloproteinases in human invasive breast carcinomas. Cancer Res. 57: 3159-3167. Vartio, T. and A. Vaheri. 1981. A gelatin-binding 70,000-dalton glycoprotein synthesized distinctly from fibronectin by normal and malignant adherent cells. J. Biol. Chem. 256: 13085-13090. Velasco, G., A. M. Pendas, A. Fueyo, V. Knauper, G. Murphy, and C. Lopez-Otin. 1999. Cloning and characterization of human MMP-23, a new matrix metalloproteinase predominantly expressed in reproductive tissues and lacking conserved domains in other family members. J. Biol. Chem. 274: 4570-4576. Vihinen, P., and V. M. Kahari. 2002. Matrix metalloproteinases in cancer: Prognostic markers and therapeutic targets. Int. J. Cancer 99: 157-166. Visse, R, and H. Nagase. 2003. Matrix metalloproteinases and tissue inhibitors of metalloproteinases: structure, function, and biochemistry. Circ. Res. 92: 827-839. von Weizsacker, F., S. Labeit, H. K. Koch, W. Oehlert, W. Gerok, and H. E. Blum. 1991. A simple and rapid method for the detection of RNA in formalin-fixed, paraffin-embedded tissues by PCR amplification. Biochem. Biophys. Res. Commun. 174: 176-180. Watanabe, M., Y. Takahashi, T. Ohta, M. Mai, T. Sasaki, and M. Seiki. 1996. Inhibition of metastasis in human gastric cancer cells transfected with tissue inhibitor of metalloproteinase 1 gene in nude mice. Cancer. 77: 1676-1680. Weidner, N., R. J. Cote, S. Suster, and L. M. Weiss. 2003. Modern surgical pathology. 1st ed. Philadelphia: SAUNDERS. Wiseman B. S., and Z. Werb. 2002. Stromal effects on mammary gland development and breast cancer. Science. 296:1046-1049. Woessner, J. F., and H. Nagase. 2000. Matrix metalloproteinases and TIMPs. OXFORD UNIVERSITY PRESS. Yu, J., R. Miller, W. Zhang, M. Sharma, V. Holtschlag, M. A. Watson, and H. L. McLeod. 2008. Copy-number analysis of topoisomerase and thymidylate synthase genes in frozen and FFPE DNAs of colorectal cancers. Pharmacogenomics. 9: 1459-1466. Zhang X., Y. Wang, G. Yamamoto, and T. Tachikawa. Expression of matrix metalloproteinases MMP-2, MMP-9 and their tissue inhibitors TIMP-1 and TIMP-2 in the epithelium and stroma of salivary gland pleomorphic adenomas. Histopathology. 55: 250-60. Zhao, Y. G., A. Z. Xiao, H. I. Park, R. G. Newcomer, M. Yan, Y. G. Man, S. C. Heffelfinger, and Q. X. Sang. 2004. Endometase/matrilysin-2 in human breast ductal carcinoma in situ and its inhibition by tissue inhibitors of metalloproteinases-2 and -4: a putative role in the initiation of breast cancer invasion. Cancer Res. 64: 590-598. 黃莉萍、曾嶔元。2010。分子醫學檢測之優良操作。生物醫學 3: 318-323。
摘要: 
甲醛固定石蠟包埋 (formalin fixed paraffin embedded, FFPE) 組織是醫院及研究單位用於保存樣品最常使用之方式,其提供了大量之樣品可做後續研究。然而,FFPE這個處理保存了組織完整性卻造成其內核酸嚴重地被破壞。因此,本實驗初步目的是建立以 RT-PCR 測定FFPE組織中RNA表現之條件。本實驗比較不同蛋白酶 K處理時間、引子濃度、反轉錄套組、amplicon size之引子在新鮮冷凍 (fresh frozen) 組織與經過甲醛浸泡1, 2及3天之石蠟包埋乳癌組織中GAPDH mRNA表現之差異。研究確認甲醛固定時間超過一天會顯著減少RNA回收量及品質,蛋白酶K作用時間延長至24小時能顯著增加RNA回收率。理想之amplicon size為100 bp以下。引子濃度5 μM經電泳與呈色後有可辨識之條帶,而200 nM則否。Biomi Biotechnology反轉錄套組較Invitrogen Superscript反轉錄套組在測定小amplicon size時更為敏感。35個PCR cycle number與31個相比,其RNA表現較不受到甲醛固定天數之影響。

基質金屬蛋白酶 (matrix metalloproteinases, MMPs) 為分解細胞外基質之酵素,其活性受到內源性之金屬蛋白酶抑制因子 (tissue inhibitors of metalloproteinases, TIMPs) 所調節,因此這兩者常被認為與腫瘤之轉移及侵犯有關。本試驗應用前述RT-PCR條件進一步測定乳癌石蠟組織中MMP-2, -9, -14與TIMP-1, -2之RNA表現。實驗中使用三十例2009年及兩例2007年之石蠟組織來進行實驗。分別為11例乳頭狀瘤 (intraductal papilloma, IP),11例腺管原位癌 (ductal carcinoma in situ, DCIS),及10例侵襲性腺管癌 (invasive ductal carcinoma, IDC)。其RT-PCR之電泳呈色結果經影像擷取系統定量,並統計後顯示,MMP-9與TIMP-2之mRNA表現在DCIS顯著較低。所有FFPE樣品均未能測出TIMP-1之RNA表現。各類型樣品結果經相關性分析顯示,MMP-2與MMP-9、MMP-2與TIMP-2及MMP-9與TIMP-2間在IP、DCIS及IDC中具有高度之相關性。而在DCIS中,MMP-2、MMP-9、MMP-14及TIMP-2彼此間均具有高度之相關性,綜合以上結果,推論MMP-2、MMP-14與TIMP-2之交互作用在原位癌中細胞外基質之降解扮演重要角色,而MMP-9與侵犯性乳癌具有較密切的相關。

Formalin fixed paraffin embedded (FFPE) is the most commonly used method worldwide for tissue storage; this resource represents a vast repository of tissue material with a long-term clinical follow-up. Although, FFPE preserves the tissue integrity it may cause extensive damage to nucleic acids stored within the tissue. Hence, the primary goal of this experiment is to set up the best condition for detecting mRNA expression in FFPE tissues by RT-PCR. To optimize the RT-PCR condition, we compare the GAPDH mRNA expression in fresh frozen tissues and tissues with formalin fixation for 1, 2 and 3 days under different proteinase K reaction time, primer concentration, commercial RT kits, amplicon size of primer treatment. We concluded that a shorten fixation time to less than one day, and extended proteinase K reaction time to 24 hours produced better RNA quality and recovery. The appropriate primer amplicon size is smaller than 100 bp and the concentration of primer is better at 5 μM in 1.5 μl. Biomi Biotech RT kit is more sensitive than Invitrogen Superscript RT kit in detecting small amplicon size primer.

Matrix metalloproteinases (MMPs) can degrade extracellular matrix, which is regulated by tissue inhibitors of metalloproteinases (TIMPs). Hence, those two factors are considered to play an important role in cancer metastasis and invasion. We applied the above optimal condition for RT-PCR to measure the RNA expressions of matrix metalloproteinases (MMP) -2, -9, -14 and tissue inhibitor of metalloproteinases (TIMP) -1, -2 on FFPE human breast cancer tissues. Thirty cases of FFPE human breast tumor from 2009 and two cases of FFPE human breast tumor from 2007 were adopted in this experiment. FFPE of eleven cases of intraductal papilloma (IP), eleven cases of ductal carcinoma in situ (DCIS), and ten cases of invasive ductal carcinoma (IDC) were included in current study. The RT-PCR results were quantified. The statistics of the results show that RNA expressions of MMP-9 and TIMP-2 were significantly lower in DCIS. TIMP-1 RNA expression was not detected in all samples studied. Correlation analysis show that the expression between MMP-2 and MMP-9, MMP-2 and TIMP-2, MMP-9 and TIMP-2 are highly related. The correlation analysis of MMP-2, MMP-9, MMP-14 and TIMP-2 are highly related in DCIS. Our results suggest that MMP-2, MMP-14 and TIMP-2 are important in extracellular matrix degradation in DCIS and MMP-9 is more relevant with invasive ductal carcinoma.
URI: http://hdl.handle.net/11455/25058
其他識別: U0005-2508201118462700
Appears in Collections:動物科學系

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