NiTi合金是一種新的生醫材料，具有形狀記憶效應，可藉著溫度改變造成的相變化來記憶不同的形狀，使其在外科手術上具有便利性，此外更具有良好的生物相容性，在植入人體後，不易引起生理的免疫反應，因此正逐漸被廣泛的應用在臨床醫療上。然而其本身有一顯著的缺點，即對血液的相容性不佳，容易導致血栓的發生。
在本實驗中，嘗試利用矽烷化反應來對NiTi合金的表面進行改質，使其具有一活性的-NH2官能基，接著利用偶合劑將具有抗凝血功能的血栓調節素，以化學鍵結的方式固定在合金表面上。並利用BCA、Lowry和Brafdord試劑來定量固定化的蛋白質，再藉由血小板吸附實驗及protein C活化測試，評估其生物活性和血液相容性。
實驗結果顯示，固定化後的血栓調節素仍然具有促使凝血酶活化protein C的能力，但與溶液相中的血栓調節素相比，固定化後的血栓調節素有活性降低的現象，經由血小板吸附實驗，可以發現表面固定血栓調節素後的NiTi合金，明顯減少了對血小板的吸附情形，因此可以證明經由血栓調節素的固定化後，確實能夠改善NiTi合金的血液相容性。

Nitinol is a newly developed biomaterial and is gaining popularity in many biomedical applications. Because nitiol has good biocompatibility, it will not induce the inflammatory response and the repulsion by the immunization after implanted in human body. Besides, nitinol is a kind of shape memory alloy that can memorize shapes under different temperature. This can improve the convenience in surgery. However, nitinol has poor blood compatibility that limits the applications of nitinol.

In this study, the surface of nitinol was modified by silanization with organosilane to introduce amino groups. The introduced functional groups were available for the subsequent covalent immobilization of thrombomodulin (TM) by using coupling reagent. The immobilized TM were determined by BCA method, Lowry method, and Braford method. The blood compatibility of various sample and the bioactivity of the immobilized TM were evaluated by protein C assay and platelet adhesion test.

Based on the result, it was found that the immobilized TM still had the ability to accelerate the activation of protein C, while its activity was lower than the free TM in solution. Furthermore, platelet adhesion test showed that the TM-immobilized surface had the least platelet adhesion. Therefore, it is possible to improve the blood compatibility of nitinol by the immobilization of thrombomodulin on surface.

參 考 文 獻

1. 何敏夫,血液學,合記出版社,1993.
2. 王文憲,人體生理學,合記出版社,1994.
3. B. D. Ratner, A. S. Hoffman, F. J. Schoen, and J. E. Lemons, Biomaterials Science, Academic Press, 1996.
4. Charles T. Esmon, Thrombomodulin as a model of molecular mechanisms that modulate protease specificity and function at the vessel surface, The FASEB journal Vol. 9 946-955, 1995.
5. Charles T. Esmon, Molecular events that control the protein C anticoagulant pathway, Thrombosis and Haemostasis, 70(1) 29-35.199
6. J. E. Sadler, S. R. Lentz, J. P. Sheehan, M. Tsiang, and Q. Wu, Structure-function relationships of the thrombin-thrombomodulin interation, Haemostasis 23, 183-193, 1993.
7. Charles T. Esmon, The Roles of protein C and thrombomodulin in the regulation of blood coagulation, The journal of biological chemistry Vol. 264 , 4743-4746, 1989.
8. A. Vindigni, C. E. White , E. A. komives, and E. D. Cera, Energetics of thrombin-thrombomodulin interaction, Biochemistry, 36, 6674-6681,1997.
9. Shaun R. Coughlin, Molecular mechanisms of Thrombin signaling, Seminars in Hematology , Vol. 31, 270-277, 1994.
10. W. J. Buehler, J. V. Gilfrich, and K. C. wiley, J. Appl. Phys.,34,1467,1963.
11. 形狀記憶金屬講習會,工研院材料所, 1988.
12. 廖南吉,形狀記憶合金與用途,南台圖書公司,1989.
13. 賴耿陽, 金屬鈦(理論與應用), 復漢出版社,1990.
14. Irena Gotman, Characteristics of metal used in implants, Journal of endourology, Vol 11, 383-389,December 1997.
15. H.-Y. Song, T. S. Shim, S.-G. Kang, G.-S. Jung, D.-Y. Lee, T.-H. Kim, S. Park, Y. M. Ahn, and W. S. Kim, Tracheobronchial strictures: Treatment with a polyurethane-covered Retrievable expandable nitiol stent-initial experience, Radiology, 213,905-912,1999.
16. E. Rechavia, F. Litvack, M. C. Fishbien, M. NaKamura, and N. Eigler, Biocompatibility of polyurethane-coated stents : Tissue and vascular aspects, Catheterization and cardiovascular diagnosis 45, 202-207, 1998.
17. T. B. McPherson, H. S. Shim, and K. Park, Grafting of PEO to glass, nitinol ,and pyrolytic carbon surfaces by γ irradiation, J. Biomed. Mater Res.( Appl. Biomater. )38,289-302, 1997.
18. P. Hildebrandt, M. Sayyad, A. Rzany, M. Schaldach, and H. Seiter, Prevention of surface encrustation of urological implants by coating with inhibitors, Biomaterials 22,503-507,2001.
19. J. Lahann, D. Klee, W. Pluester, and H. Hoecker, Bioactive immobilization of r-hirudin on CVD-coated metallic implant devices, Biomaterials 22, 817-826, 2001.
20. J. Lahann, D. Klee, and H. Höcker, Chemical vapour deposition polymerization of substituted [2,2] paracyclophanes, Macromol. Rapid Commun, 19, 441-444,1998.
21. X. Kong, R. G. Grabitz, W. van Oeveren, D. Klee, T. G. van Kooten, F. Freudenthal, Ma Qing, G. von Bernuth, M.-C. Seghaye, Effect of biologically active coating on biocompatibility of nitinol devices designed forr the closure of intra-atrial communications, Biomaterials,23, 1775-1783,2002.
22. M. Akashi, I. Maruyama, N. Fukudome, and E. Yashima, Immobilization of human thrombomodulin on glass beads and its anticoagulant activity, Bioconjugate Chem. Vol. 3, 363-365,1992.
23. K. Yagi, K. Hirota, S Yamasaki, A. Uwai, and Y. Miura, Anicoagulant activity of immobilized thrombomodulin, Chem. Pharm. Bull. 37(3) 732-734, 1989.
24. A. Kishida. Y. Akatshka, M. Yanag, T. Aikou, I. Maruyama, and M. Akashi, In vivo and ex vivo evaluation of the antithrombogenicity of human thrombomodulin immobilized biomaterials, ASAIO Journal 41, M369, 1995.
25. A. Kishida, Y. Ueno, N. Fukudome, E. Yashima, I. Maruyama and M. Akashi, Immobilization of human thrombomodulin onto poly(ether urethane urea) for developing antithrombogenic blood-contacting materials, Biomaterials Vol. 15 , 848-852, 1994.
26. A. Kishida, Y. Ueno, I. Maruyama, and M. Akashi, Immobilization of human thrombomodulin on biomaterials: evaluatiuon of the activity of immobilized human thrombomodulin, Biomaterials Vol. 15, 1170-1174, 1994.
27. V. N. Vasilets, G. Hermel, U. König, C. Werner, M. Müller, F. Simon, K. Grundke, Y. Ikada, and H-J. Jacobasch, Microwave CO2 plasma-initiated vapor phase graft polymerization of acrylic acid onto polytetrafluoroethylene for immobilization of human thrombomodulin, Biomaterials 18, 1139-1145, 1997.
28. S. Sano, K. Kato, and Y. Ikada, Introduction of functional groups onto the surface of polyethylene for protein immobilization, Biomaterials , 14, 817-822,1993.
29. R. F. Taylor, Protein immobilization : fundamentals and applications,1991.
30. H.-S. Han, S.-L. Yang, H.-Y. Yeh, J.-C. Lin, H.-L. Wu, and G.-Y. Shi, Studies of a novel human thrombomodulin immobilized substrate: surface characterization and anticoagulation activity evaluation, J. Biomater. Sci. Polymer Edn, 12,1075-1089,2001.
31. 國立成功大學化學工程研究所碩士論文, 楊曉玲撰。
32. E. Uchida, Y. Uyama, and Y. Ikada, Sorption of low-molecular-weight anions into thin polycation layers grafted onto a film, Langmuir 9, 1121-1124, 1993.
33. S. Sano, K. Kato, and Y. Ikada, Introduction of functional groups onto the surface of polyethylene for protein immobilization, Biomaterials, 14,817-822,1993.
34. P. K. Smirth, R. I. Krohn, G. T. Hermanson, A. K. Mallia, F. H. Garther, M.D. Provenzano, E. K. Fujimoto, N. M. Goeke, B. J. Olson, and D. C. Klenk, Measurement of protein using bicinchoninic acid, Analytical biochemistry 150,76-85, 1985.
35. O. H. Lowry , N. J. Rosebrough, A. L. Farr, and R. J. Randall, J. Biol. Chem. 193, 265-275, 1951.
36. M. M. Bradford, A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the priciple of protein-dye binding, Analytical biochemistry 72,248-254,1976.
37. S. P. J. Brooks, B. J. Lampi, G. Sarwar, and H. G. Botting, A Comparison of methods for determining total body protein, Analytical biochemistry 226, 26-30, 1995.
38. J. W. Jenzano, S. L. Hogan, C. M. Noyes, G. L. Featherstone, and R. L. Lundblad, Comparison of five techniques for the determination of protein content in mixed human saliva, Analytical Biochemistry 159, 370-376, 1986.
39. C. V. Sapan, R. L. Lundblad, and N. C. Price, Colorimetric protein assay techniques, Biotechnol. Appl. Biochem. 29, 99-108, 1999.
40. K. Mosbach, Methods in Enzymology, vol. 91.
41. B. O’Brien, W. M. Carroll, and M. J. Kelly, Passivation of nitinol wire for vascular implants―a demonstration of the benefits, Biomaterials 23, 1739-1748,2002.
42. http://www.nitinol.com