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系統識別號 U0026-2407201511300300
論文名稱(中文) 研究腫瘤內皮標誌一在血管新生作用中的角色
論文名稱(英文) Study on the Role of Tumor Endothelial Marker 1 in Angiogenesis
校院名稱 成功大學
系所名稱(中) 生物化學暨分子生物學研究所
系所名稱(英) Department of Biochemistry and Molecular Biology
學年度 103
學期 2
出版年 104
研究生(中文) 呂庭堯
研究生(英文) Ting-Yao Lu
學號 S16024034
學位類別 碩士
語文別 英文
論文頁數 74頁
口試委員 指導教授-施桂月
口試委員-吳華林
口試委員-林淑華
口試委員-江美治
中文關鍵字 腫瘤內皮標誌一  血管新生作用  纖維母細胞  傷口癒合 
英文關鍵字 tumor endothelial marker 1  angiogenesis  fibroblast  wound healing 
學科別分類
中文摘要 腫瘤內皮標誌一屬於第一型穿膜醣蛋白質,是凝血酶調節素家族的一員。腫瘤內皮標誌一具有六個功能區域,依序是類凝集素功能區、類壽司功能區、三重複類表皮生長因子功能區、黏液素功能區、穿膜功能區及細胞質功能區。先前的研究指出腫瘤周圍血管的腫瘤相關纖維母細胞和外被細胞會大量表現腫瘤內皮標誌一,且腫瘤內皮標誌一的表現量與腫瘤的進程和腫瘤血管新生相關。從已存在的血管衍生所產生新的血管此作用稱為血管新生,而在皮膚傷口癒合過程中血管新生作用扮演了重要的角色。然而,腫瘤內皮標誌一的類表皮生長因子功能區在血管新生和皮膚傷口癒合中的角色尚未清楚。在本次的實驗中,我們利用酵母菌純化系統純化出重組腫瘤內皮標誌一的類表皮生長因子功能區蛋白(rTEM1D3)。內皮細胞經過重組腫瘤內皮標誌一的類表皮生長因子功能區蛋白處理後,其生長、移動、和管狀結構的形成皆明顯增加。重組腫瘤內皮標誌一的類表皮生長因子功能區蛋白也參與活化與血管新生作用相關的訊息傳遞分子,例如ERK1/2、Akt、和p38。除此之外,在正常小鼠及由鏈佐黴素引發的糖尿病鼠的皮膚傷口給予重組腫瘤內皮標誌一的類表皮生長因子功能區蛋白後能有效加速皮膚傷口癒合。我們也發現全身性剔除腫瘤內皮標誌一類表皮生長因子功能區的小鼠(TEM1EGFd/EGFd)其皮膚傷口癒合的速度和傷口血管新生作用都相較於TEM1+/+小鼠緩慢。此外,降低纖維母細胞的腫瘤內皮標誌一表現量能顯著降低內皮細胞的趨化移動性和管狀結構的生成,由此可知纖維母細胞上所表現的腫瘤內皮標誌一可能是吸引內皮細胞移動和增強管狀結構形成的重要因子。綜合以上結果,我們證明腫瘤內皮標誌一的類表皮生長因子功能區會參與血管新生作用,且重組腫瘤內皮標誌一的類表皮生長因子功能區蛋白在治療糖尿病的傷口癒合中是一個具有潛力的治療藥物。
英文摘要 Tumor endothelial marker 1 (TEM1), a type I transmembrane glycoprotein, belongs to the thrombomodulin family and contains six domains, including an N-terminal C-type lectin-like domain, a sushi-like domain, a three-repeated epidermal growth factor (EGF)-like domain, a mucin-like domain, a transmembrane domain and a cytoplasmic domain. Previous studies indicate that TEM1 is highly expressed in tumor-associated fibroblasts and pericytes residing in blood vessels nearby tumors, and TEM1 expression is associated with tumor progression and neovascularization. Angiogenesis is characterized by a new blood vessel formation from pre-existing vessels and plays a critical role in cutaneous wound healing. However, the roles of TEM1 EGF-like domain in angiogenesis and skin wound healing are largely unknown. In this study, recombinant TEM1 EGF-like domain (rTEM1D3) was prepared by Pichia pastoris expression system. rTEM1D3 treatment increased endothelial cells proliferation, migration, and tube formation. Angiogenesis-related intracellular signaling pathways, including extracellular signal–regulated kinase 1/2, p38, and Akt, were activated after rTEM1D3 stimulation in endothelial cells. Furthermore, local administration of rTEM1D3 improved cutaneous wound healing in wild type C57BL/6 mice and STZ-induced diabetic mice. Mice with TEM1 EGF-like domain deletion (TEM1EGFd/EGFd) exhibited delayed wound closure and poor angiogenesis compared with TEM1+/+ mice. Knockdown of TEM1 in fibroblasts significantly reduced the chemotactic motility and tube formation ability of endothelial cells, indicating that TEM1 in fibroblasts may be an important factor to attract endothelial cell migration and to enhance tube formation. In conclusion, we demonstrate that TEM1 EGF-like domain is involved in angiogenesis and rTEM1D3 may have potential for the treatment of diabetic wound.
論文目次 Chapter Page
Abstract in Chinese 1
Abstract in English 2
Acknowledgement 3
Contents 4
List of Figures 5
Abbreviation 6
Instruments 8
Reagents and Chemicals 10
Introduction 14
Specific Aim 19
Materials and Methods 20
Results 48
Conclusion 53
Discussion 54
References 57
Figures 62
Appendixes 74

參考文獻 1. Pandya, N. M., Dhalla, N. S., and Santani, D. D. (2006) Angiogenesis--a new target for future therapy. Vascular pharmacology 44, 265-274
2. Otrock, Z. K., Mahfouz, R. A., Makarem, J. A., and Shamseddine, A. I. (2007) Understanding the biology of angiogenesis: review of the most important molecular mechanisms. Blood cells, molecules & diseases 39, 212-220
3. Singer, A. J., and Clark, R. A. F. (1999) Cutaneous Wound Healing. New England Journal of Medicine 341, 738-746
4. Gurtner, G. C., Werner, S., Barrandon, Y., and Longaker, M. T. (2008) Wound repair and regeneration. Nature 453, 314-321
5. Harper, D., Young, A., and McNaught, C.-E. (2014) The physiology of wound healing. Surgery (Oxford) 32, 445-450
6. Tonnesen, M. G., Feng, X., and Clark, R. A. F. (2000) Angiogenesis in Wound Healing. J Investig Dermatol Symp Proc 5, 40-46
7. Eming, S. A., Brachvogel, B., Odorisio, T., and Koch, M. (2007) Regulation of angiogenesis: wound healing as a model. Progress in histochemistry and cytochemistry 42, 115-170
8. Hardwicke, J., Schmaljohann, D., Boyce, D., and Thomas, D. (2008) Epidermal growth factor therapy and wound healing — past, present and future perspectives. The Surgeon 6, 172-177
9. Grotendorst, G. R., Soma, Y., Takehara, K., and Charette, M. (1989) EGF and TGF-alpha are potent chemoattractants for endothelial cells and EGF-like peptides are present at sites of tissue regeneration. Journal of Cellular Physiology 139, 617-623
10. Kwon, Y. B., Kim, H. W., Roh, D. H., Yoon, S. Y., Baek, R. M., Kim, J. Y., Kweon, H., Lee, K. G., Park, Y. H., and Lee, J. H. (2006) Topical application of epidermal growth factor accelerates wound healing by myofibroblast proliferation and collagen synthesis in rat. Journal of veterinary science 7, 105-109
11. Brazzell, R. K., Stern, M. E., Aquavella, J. V., Beuerman, R. W., and Baird, L. (1991) Human recombinant epidermal growth factor in experimental corneal wound healing. Investigative ophthalmology & visual science 32, 336-340
12. Galiano, R. D., Tepper, O. M., Pelo, C. R., Bhatt, K. A., Callaghan, M., Bastidas, N., Bunting, S., Steinmetz, H. G., and Gurtner, G. C. (2004) Topical vascular endothelial growth factor accelerates diabetic wound healing through increased angiogenesis and by Mobilizing and Recruiting Bone Marrow-Derived Cells. The American journal of pathology 164, 1935-1947
13. Mehta, V. B., and Besner, G. E. (2007) HB-EGF promotes angiogenesis in endothelial cells via PI3-kinase and MAPK signaling pathways. Growth factors 25, 253-263
14. Rettig, W. J., Garin-Chesa, P., Healey, J. H., Su, S. L., Jaffe, E. A., and Old, L. J. (1992) Identification of endosialin, a cell surface glycoprotein of vascular endothelial cells in human cancer. Proceedings of the National Academy of Sciences of the United States of America 89, 10832-10836
15. Bagley, R. G., Honma, N., Weber, W., Boutin, P., Rouleau, C., Shankara, S., Kataoka, S., Ishida, I., Roberts, B. L., and Teicher, B. A. (2008) Endosialin/TEM 1/CD248 is a pericyte marker of embryonic and tumor neovascularization. Microvascular research 76, 180-188
16. Christian, S., Winkler, R., Helfrich, I., Boos, A. M., Besemfelder, E., Schadendorf, D., and Augustin, H. G. (2008) Endosialin (Tem1) is a marker of tumor-associated myofibroblasts and tumor vessel-associated mural cells. The American journal of pathology 172, 486-494
17. MacFadyen, J., Savage, K., Wienke, D., and Isacke, C. M. (2007) Endosialin is expressed on stromal fibroblasts and CNS pericytes in mouse embryos and is downregulated during development. Gene expression patterns : GEP 7, 363-369
18. MacFadyen, J. R., Haworth, O., Roberston, D., Hardie, D., Webster, M. T., Morris, H. R., Panico, M., Sutton-Smith, M., Dell, A., van der Geer, P., Wienke, D., Buckley, C. D., and Isacke, C. M. (2005) Endosialin (TEM1, CD248) is a marker of stromal fibroblasts and is not selectively expressed on tumour endothelium. FEBS letters 579, 2569-2575
19. Rouleau, C., Curiel, M., Weber, W., Smale, R., Kurtzberg, L., Mascarello, J., Berger, C., Wallar, G., Bagley, R., Honma, N., Hasegawa, K., Ishida, I., Kataoka, S., Thurberg, B. L., Mehraein, K., Horten, B., Miller, G., and Teicher, B. A. (2008) Endosialin protein expression and therapeutic target potential in human solid tumors: sarcoma versus carcinoma. Clinical cancer research : an official journal of the American Association for Cancer Research 14, 7223-7236
20. Christian, S., Ahorn, H., Koehler, A., Eisenhaber, F., Rodi, H. P., Garin-Chesa, P., Park, J. E., Rettig, W. J., and Lenter, M. C. (2001) Molecular cloning and characterization of endosialin, a C-type lectin-like cell surface receptor of tumor endothelium. The Journal of biological chemistry 276, 7408-7414
21. Nanda, A., Karim, B., Peng, Z., Liu, G., Qiu, W., Gan, C., Vogelstein, B., St Croix, B., Kinzler, K. W., and Huso, D. L. (2006) Tumor endothelial marker 1 (Tem1) functions in the growth and progression of abdominal tumors. Proceedings of the National Academy of Sciences of the United States of America 103, 3351-3356
22. Maia, M., DeVriese, A., Janssens, T., Moons, M., Lories, R. J., Tavernier, J., and Conway, E. M. (2011) CD248 facilitates tumor growth via its cytoplasmic domain. BMC cancer 11, 162
23. Tomkowicz, B., Rybinski, K., Foley, B., Ebel, W., Kline, B., Routhier, E., Sass, P., Nicolaides, N. C., Grasso, L., and Zhou, Y. (2007) Interaction of endosialin/TEM1 with extracellular matrix proteins mediates cell adhesion and migration. Proceedings of the National Academy of Sciences of the United States of America 104, 17965-17970
24. Maia, M., de Vriese, A., Janssens, T., Moons, M., van Landuyt, K., Tavernier, J., Lories, R. J., and Conway, E. M. (2010) CD248 and its cytoplasmic domain: a therapeutic target for arthritis. Arthritis and rheumatism 62, 3595-3606
25. Rupp, C., Dolznig, H., Puri, C., Sommergruber, W., Kerjaschki, D., Rettig, W. J., and Garin-Chesa, P. (2006) Mouse endosialin, a C-type lectin-like cell surface receptor: expression during embryonic development and induction in experimental cancer neoangiogenesis. Cancer immunity 6, 10
26. Zelensky, A. N., and Gready, J. E. (2005) The C-type lectin-like domain superfamily. The FEBS journal 272, 6179-6217
27. Shi, C. S., Shi, G. Y., Chang, Y. S., Han, H. S., Kuo, C. H., Liu, C., Huang, H. C., Chang, Y. J., Chen, P. S., and Wu, H. L. (2005) Evidence of human thrombomodulin domain as a novel angiogenic factor. Circulation 111, 1627-1636
28. Kao, Y. C., Jiang, S. J., Pan, W. A., Wang, K. C., Chen, P. K., Wei, H. J., Chen, W. S., Chang, B. I., Shi, G. Y., and Wu, H. L. (2012) The epidermal growth factor-like domain of CD93 is a potent angiogenic factor. PloS one 7, e51647
29. Munoz-Chapuli, R., Quesada, A. R., and Angel Medina, M. (2004) Angiogenesis and signal transduction in endothelial cells. Cellular and molecular life sciences : CMLS 61, 2224-2243
30. Wietecha, M. S., Chen, L., Ranzer, M. J., Anderson, K., Ying, C., Patel, T. B., and DiPietro, L. A. (2011) Sprouty2 downregulates angiogenesis during mouse skin wound healing. American Journal of Physiology - Heart and Circulatory Physiology 300, H459-H467
31. Wu, K. K., and Huan, Y. (2001) Streptozotocin-Induced Diabetic Models in Mice and Rats. in Current Protocols in Pharmacology, John Wiley & Sons, Inc. pp
32. Cheng, T. L., Wu, Y. T., Lin, H. Y., Hsu, F. C., Liu, S. K., Chang, B. I., Chen, W. S., Lai, C. H., Shi, G. Y., and Wu, H. L. (2011) Functions of rhomboid family protease RHBDL2 and thrombomodulin in wound healing. The Journal of investigative dermatology 131, 2486-2494
33. Bagley, R. G., Rouleau, C., St Martin, T., Boutin, P., Weber, W., Ruzek, M., Honma, N., Nacht, M., Shankara, S., Kataoka, S., Ishida, I., Roberts, B. L., and Teicher, B. A. (2008) Human endothelial precursor cells express tumor endothelial marker 1/endosialin/CD248. Molecular cancer therapeutics 7, 2536-2546
34. Carson-Walter, E. B., Winans, B. N., Whiteman, M. C., Liu, Y., Jarvela, S., Haapasalo, H., Tyler, B. M., Huso, D. L., Johnson, M. D., and Walter, K. A. (2009) Characterization of TEM1/endosialin in human and murine brain tumors. BMC cancer 9, 417
35. Facciponte, J. G., Ugel, S., De Sanctis, F., Li, C., Wang, L., Nair, G., Sehgal, S., Raj, A., Matthaiou, E., Coukos, G., and Facciabene, A. (2014) Tumor endothelial marker 1-specific DNA vaccination targets tumor vasculature. The Journal of clinical investigation 124, 1497-1511
36. Carmeliet, P. (2003) Angiogenesis in health and disease. Nature medicine 9, 653-660
37. Folkman, J., and Shing, Y. (1992) Angiogenesis. The Journal of biological chemistry 267, 10931-10934
38. Yancopoulos, G. D., Davis, S., Gale, N. W., Rudge, J. S., Wiegand, S. J., and Holash, J. (2000) Vascular-specific growth factors and blood vessel formation. Nature 407, 242-248
39. Oliner, J., Min, H., Leal, J., Yu, D., Rao, S., You, E., Tang, X., Kim, H., Meyer, S., Han, S. J., Hawkins, N., Rosenfeld, R., Davy, E., Graham, K., Jacobsen, F., Stevenson, S., Ho, J., Chen, Q., Hartmann, T., Michaels, M., Kelley, M., Li, L., Sitney, K., Martin, F., Sun, J. R., Zhang, N., Lu, J., Estrada, J., Kumar, R., Coxon, A., Kaufman, S., Pretorius, J., Scully, S., Cattley, R., Payton, M., Coats, S., Nguyen, L., Desilva, B., Ndifor, A., Hayward, I., Radinsky, R., Boone, T., and Kendall, R. (2004) Suppression of angiogenesis and tumor growth by selective inhibition of angiopoietin-2. Cancer cell 6, 507-516
40. O'Reilly, M. S., Boehm, T., Shing, Y., Fukai, N., Vasios, G., Lane, W. S., Flynn, E., Birkhead, J. R., Olsen, B. R., and Folkman, J. (1997) Endostatin: an endogenous inhibitor of angiogenesis and tumor growth. Cell 88, 277-285
41. Jimenez, B., Volpert, O. V., Crawford, S. E., Febbraio, M., Silverstein, R. L., and Bouck, N. (2000) Signals leading to apoptosis-dependent inhibition of neovascularization by thrombospondin-1. Nature medicine 6, 41-48
42. Ishii, H., and Majerus, P. W. (1985) Thrombomodulin is present in human plasma and urine. The Journal of clinical investigation 76, 2178-2181
43. Takano, S., Kimura, S., Ohdama, S., and Aoki, N. (1990) Plasma thrombomodulin in health and diseases. Blood 76, 2024-2029
44. Bohlson, S. S., Silva, R., Fonseca, M. I., and Tenner, A. J. (2005) CD93 Is Rapidly Shed from the Surface of Human Myeloid Cells and the Soluble Form Is Detected in Human Plasma. The Journal of Immunology 175, 1239-1247
45. Larsen, A. K., Ouaret, D., El Ouadrani, K., and Petitprez, A. (2011) Targeting EGFR and VEGF(R) pathway cross-talk in tumor survival and angiogenesis. Pharmacology & therapeutics 131, 80-90
46. Garg, P., Yang, S., Liu, A., Pallero, M. A., Buchsbaum, D. J., Mosher, D. F., Murphy-Ullrich, J. E., and Goldblum, S. E. (2011) Thrombospondin-1 opens the paracellular pathway in pulmonary microvascular endothelia through EGFR/ErbB2 activation. American journal of physiology. Lung cellular and molecular physiology 301, L79-90
47. 李瑞庭. (2014) 探討腫瘤內皮標誌1在皮膚傷口癒合中扮演的角色. 碩士, 國立成功大學
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