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完整後設資料紀錄
DC 欄位 | 值 | 語言 |
---|---|---|
dc.contributor.advisor | 黃德富 | |
dc.contributor.author | Chiang-Hui Chen | en |
dc.contributor.author | 陳薔卉 | zh_TW |
dc.date.accessioned | 2021-06-15T12:42:18Z | - |
dc.date.available | 2019-08-26 | |
dc.date.copyright | 2016-08-26 | |
dc.date.issued | 2016 | |
dc.date.submitted | 2016-07-27 | |
dc.identifier.citation | 1. Jackson, S.P. Arterial thrombosis[mdash]insidious, unpredictable and deadly. Nat
Med 17, 1423-1436 (2011). 2. Ruggeri, Z.M. Platelets in atherothrombosis. Nat Med 8, 1227-1234 (2002). 3. Angiolillo, D.J., et al. Basic principles of platelet biology and clinical implications. Circulation journal : official journal of the Japanese Circulation Society 74, 597-607 (2010). 4. Marcus, A.J., et al. The endothelial cell ecto-ADPase responsible for inhibition of platelet function is CD39. Journal of Clinical Investigation 99, 1351-1360 (1997). 5. Varga-Szabo, D., et al. Cell adhesion mechanisms in platelets. Arteriosclerosis, thrombosis, and vascular biology 28, 403-412 (2008). 6. Jennings, L.K. Mechanisms of platelet activation: need for new strategies to protect against platelet-mediated atherothrombosis. Thrombosis and haemostasis 102, 248-257 (2009). 7. Jackson, S.P. & Schoenwaelder, S.M. Antiplatelet therapy: in search of the 'magic bullet'. Nat Rev Drug Discov 2, 775-789 (2003). 8. Brass, L.F. Thrombin and Platelet Activation. Chest 124, 18S-25S (2003). 9. Sakariassen, K.S., et al. The role of platelet membrane glycoproteins Ib and IIb-IIIa in platelet adherence to human artery subendothelium. British journal of haematology 63, 681-691 (1986). 107 10. Hagedorn, I., et al. Arterial thrombus formation. Novel mechanisms and targets. Hamostaseologie 30, 127-135 (2010). 11. Santoro, S.A. Identification of a 160,000 dalton platelet membrane protein that mediates the initial divalent cation-dependent adhesion of platelets to collagen. Cell 46, 913-920 (1986). 12. Falati, S., et al. Real-time in vivo imaging of platelets, tissue factor and fibrin during arterial thrombus formation in the mouse. Nat Med 8, 1175-1181 (2002). 13. Kulkarni, S., et al. A revised model of platelet aggregation. Journal of Clinical Investigation 105, 783-791 (2000). 14. Phillips, D.R., et al. Beta3 tyrosine phosphorylation in alphaIIbbeta3 (platelet membrane GP IIb-IIIa) outside-in integrin signaling. Thrombosis and haemostasis 86, 246-258 (2001). 15. Estevez, B., et al. Targeting integrin and integrin signaling in treating thrombosis. Arteriosclerosis, thrombosis, and vascular biology 35, 24-29 (2015). 16. Shattil, S.J., et al. The final steps of integrin activation: the end game. Nat Rev Mol Cell Biol 11, 288-300 (2010). 17. Plow, E.F., et al. Ligand binding to integrins. The Journal of biological chemistry 275, 21785-21788 (2000). 18. Hynes, R.O. Integrins: Bidirectional, Allosteric Signaling Machines. Cell 110, 673-687 (2002). 19. Tadokoro, S., et al. Talin Binding to Integrin ß Tails: A Final Common Step in Integrin Activation. Science 302, 103-106 (2003). 108 20. Calderwood, D.A., et al. Talins and kindlins: partners in integrin-mediated adhesion. Nat Rev Mol Cell Biol 14, 503-517 (2013). 21. Ginsberg, M.H., et al. Platelet integrins. Thrombosis and haemostasis 70, 87-93 (1993). 22. Thornton, M.A., et al. The Human Platelet αIIb Gene Is Not Closely Linked to Its Integrin Partner β3. Blood 94, 2039-2047 (1999). 23. Xiong, J.P., et al. Crystal structure of the extracellular segment of integrin alpha Vbeta3 in complex with an Arg-Gly-Asp ligand. Science 296, 151-155 (2002). 24. Xiao, T., et al. Structural basis for allostery in integrins and binding to fibrinogen-mimetic therapeutics. Nature 432, 59-67 (2004). 25. Calvete, J.J. Clues for understanding the structure and function of a prototypic human integrin: the platelet glycoprotein IIb/IIIa complex. Thrombosis and haemostasis 72, 1-15 (1994). 26. Hynes, R.O. Integrins: versatility, modulation, and signaling in cell adhesion. Cell 69, 11-25 (1992). 27. Bennett, J.S. Structure and function of the platelet integrin alphaIIbbeta3. Journal of Clinical Investigation 115, 3363-3369 (2005). 28. Haas, T.A. & Plow, E.F. Integrin-ligand interactions: a year in review. Current opinion in cell biology 6, 656-662 (1994). 29. Vinogradova, O., et al. A Structural Mechanism of Integrin αIIbβ3 “Inside-Out” Activation as Regulated by Its Cytoplasmic Face. Cell 110, 587-597 (2002). 109 30. Du, X., et al. Long range propagation of conformational changes in integrin alpha IIb beta 3. The Journal of biological chemistry 268, 23087-23092 (1993). 31. Kim, M., et al. Bidirectional transmembrane signaling by cytoplasmic domain separation in integrins. Science 301, 1720-1725 (2003). 32. Lau, T.L., et al. The structure of the integrin alphaIIbbeta3 transmembrane complex explains integrin transmembrane signalling. The EMBO journal 28, 1351-1361 (2009). 33. Li, Z., et al. Signaling during platelet adhesion and activation. Arteriosclerosis, thrombosis, and vascular biology 30, 2341-2349 (2010). 34. Kahner, B.N., et al. Kindlins, integrin activation and the regulation of talin recruitment to alphaIIbbeta3. PloS one 7, e34056 (2012). 35. Moers, A., et al. G13 is an essential mediator of platelet activation in hemostasis and thrombosis. Nat Med 9, 1418-1422 (2003). 36. Tucker, K.L., et al. Clot retraction. Methods in molecular biology (Clifton, N.J.) 788, 101-107 (2012). 37. Shen, B., et al. A directional switch of integrin signalling and a new anti-thrombotic strategy. Nature 503, 131-135 (2013). 38. Gould, R.J., et al. Disintegrins: a family of integrin inhibitory proteins from viper venoms. Proceedings of the Society for Experimental Biology and Medicine. Society for Experimental Biology and Medicine (New York, N.Y.) 195, 168-171 (1990). 110 39. Huang, T., et al. Trigramin. A low molecular weight peptide inhibiting fibrinogen interaction with platelet receptors expressed on glycoprotein IIb-IIIa complex. Journal of Biological Chemistry 262, 16157-16163 (1987). 40. Huang, T.-F., et al. Characterization of a potent platelet aggregation inhibitor from Agkistrodon rhodostoma snake venom. Biochimica et Biophysica Acta (BBA)-General Subjects 925, 248-257 (1987). 41. Ouyang, C., et al. Characterization of the platelet aggregation induced and inhibitor from Echis carinarus snake venom. Biochimica et Biophysica Acta (BBA)-General Subjects 841, 1-7 (1985). 42. Calvete, J.J., et al. Snake venom disintegrins: evolution of structure and function. Toxicon 45, 1063-1074 (2005). 43. Koh, C.Y. & Kini, R.M. From snake venom toxins to therapeutics – Cardiovascular examples. Toxicon 59, 497-506 (2012). 44. Calvete, J.J., et al. Snake venom disintegrins: novel dimeric disintegrins and structural diversification by disulphide bond engineering. The Biochemical journal 372, 725-734 (2003). 45. Calvete, J.J., et al. The disulfide bond pattern of catrocollastatin C, a disintegrin-like/cysteine-rich protein isolated from Crotalus atrox venom. Protein Science 9, 1365-1373 (2000). 46. Knight, D.M., et al. The immunogenicity of the 7E3 murine monoclonal Fab antibody fragment variable region is dramatically reduced in humans by substitution of human for murine constant regions. Molecular immunology 32, 111 1271-1281 (1995). 47. Phillips, D.R. & Scarborough, R.M. Clinical pharmacology of eptifibatide. The American journal of cardiology 80, 11b-20b (1997). 48. Egbertson, M.S., et al. Non-peptide fibrinogen receptor antagonists. 2. Optimization of a tyrosine template as a mimic for Arg-Gly-Asp. Journal of medicinal chemistry 37, 2537-2551 (1994). 49. Stangl, P.A. & Lewis, S. Review of Currently Available GP IIb/IIIa Inhibitors and Their Role in Peripheral Vascular Interventions. Seminars in Interventional Radiology 27, 412-421 (2010). 50. Aster, R.H., et al. DRUG-INDUCED IMMUNE THROMBOCYTOPENIA: PATHOGENESIS, DIAGNOSIS AND MANAGEMENT. Journal of thrombosis and haemostasis : JTH 7, 911-918 (2009). 51. Peter, K., et al. Platelet activation as a potential mechanism of GP IIb/IIIa inhibitor-induced thrombocytopenia. American Journal of Cardiology 84, 519-524. 52. Chong, B.H. Drug-induced thrombocytopenia: MIBS trumps LIBS. Blood 119, 6177-6178 (2012). 53. Frelinger, A.L., 3rd, et al. Monoclonal antibodies to ligand-occupied conformers of integrin alpha IIb beta 3 (glycoprotein IIb-IIIa) alter receptor affinity, specificity, and function. The Journal of biological chemistry 266, 17106-17111 (1991). 112 54. Gulino, D., et al. Identification of a monoclonal antibody against platelet GPIIb that interacts with a calcium-binding site and induces aggregation. The Journal of biological chemistry 265, 9575-9581 (1990). 55. Gao, C., et al. Eptifibatide-induced thrombocytopenia and thrombosis in humans require FcγRIIa and the integrin β3 cytoplasmic domain. The Journal of clinical investigation 119, 504-511 (2009). 56. Slupsky, J.R., et al. Role of Fc gamma RII in platelet activation by monoclonal antibodies. Journal of immunology (Baltimore, Md. : 1950) 148, 3189-3194 (1992). 57. Huang, T.-F., et al. FcγRII mediates platelet aggregation caused by disintegrins and GPIIb/IIIa monoclonal antibody, AP2. Experimental hematology 36, 1704-1713 (2008). 58. Chen, Y.-C., et al. Expression in Pichia pastoris and characterization of echistatin, an RGD-containing short disintegrin. Toxicon 60, 1342-1348 (2012). 59. Laemmli, U.K. Cleavage of Structural Proteins during the Assembly of the Head of Bacteriophage T4. Nature 227, 680-685 (1970). 60. MONLEÓ N, D., et al. Conformation and concerted dynamics of the integrin-binding site and the C-terminal region of echistatin revealed by homonuclear NMR. Biochemical Journal 387, 57-66 (2005). 61. Yamada, T. & Kidera, A. Tailoring echistatin to possess higher affinity for integrin α IIb β 3. FEBS letters 387, 11-15 (1996). 113 62. Pidard, D., et al. Interaction of AP-2, a monoclonal antibody specific for the human platelet glycoprotein IIb-IIIa complex, with intact platelets. Journal of Biological Chemistry 258, 12582-12586 (1983). 63. Frenette, P.S., et al. Platelets roll on stimulated endothelium in vivo: an interaction mediated by endothelial P-selectin. Proceedings of the National Academy of Sciences of the United States of America 92, 7450-7454 (1995). 64. Merten, M. & Thiagarajan, P. P-selectin expression on platelets determines size and stability of platelet aggregates. Circulation 102, 1931-1936 (2000). 65. Leytin, V., et al. Quantification of platelet activation status by analyzing P-selectin expression. Biochemical and biophysical research communications 273, 565-570 (2000). 66. Mitchell, W.B., et al. Mapping early conformational changes in alphaIIb and beta3 during biogenesis reveals a potential mechanism for alphaIIbbeta3 adopting its bent conformation. Blood 109, 3725-3732 (2007). 67. Law, D.A., et al. Integrin cytoplasmic tyrosine motif is required for outside-in [alpha]IIb[beta]3 signalling and platelet function. Nature 401, 808-811 (1999). 68. Chen, Y.-L., et al. Determination of the structure of two novel echistatin variants and comparison of the ability of echistatin variants to inhibit aggregation of platelets from different species. Biochemical Journal 305, 513-520 (1995). 69. Scarborough, R.M., et al. Platelet glycoprotein IIb/IIIa antagonists. What are the relevant issues concerning their pharmacology and clinical use? Circulation 100, 437-444 (1999). 114 70. Use of a monoclonal antibody directed against the platelet glycoprotein IIb/IIIa receptor in high-risk coronary angioplasty. The EPIC Investigation. The New England journal of medicine 330, 956-961 (1994). 71. Gan, Z., et al. Echistatin. A potent platelet aggregation inhibitor from the venom of the viper, Echis carinatus. Journal of Biological Chemistry 263, 19827-19832 (1988). 72. Marcinkiewicz, C., et al. Significance of RGD loop and C-terminal domain of echistatin for recognition of αIIbβ3 and αvβ3 integrins and expression of ligand-induced binding site. Blood 90, 1565-1575 (1997). 73. Schneider, D.J., et al. Paradoxical inhibition of fibrinogen binding and potentiation of alpha-granule release by specific types of inhibitors of glycoprotein IIb-IIIa. Cardiovascular research 45, 437-446 (2000). 74. Peter, K., et al. Induction of fibrinogen binding and platelet aggregation as a potential intrinsic property of various glycoprotein IIb/IIIa (alphaIIbbeta3) inhibitors. Blood 92, 3240-3249 (1998). 75. Honda, S., et al. Topography of ligand-induced binding sites, including a novel cation-sensitive epitope (AP5) at the amino terminus, of the human integrin beta 3 subunit. The Journal of biological chemistry 270, 11947-11954 (1995). 76. Huang, T.F., et al. Halysin, an antiplatelet Arg-Gly-Asp-containing snake venom peptide, as fibrinogen receptor antagonist. Biochemical pharmacology 42, 1209-1219 (1991). 115 77. Huang, T.F., et al. A potent antiplatelet peptide, triflavin, from Trimeresurus flavoviridis snake venom. The Biochemical journal 277 ( Pt 2), 351-357 (1991). 78. Worth, R.G., et al. Platelet FcgammaRIIA binds and internalizes IgG-containing complexes. Exp Hematol 34, 1490-1495 (2006). 79. McKenzie, S.E., et al. The role of the human Fc receptor Fc gamma RIIA in the immune clearance of platelets: a transgenic mouse model. Journal of immunology (Baltimore, Md. : 1950) 162, 4311-4318 (1999). 80. Masarachia, P., et al. Histomorphometric Evidence for Echistatin Inhibition of Bone Resorption in Mice with Secondary Hyperparathyroidism. Endocrinology 139, 1401-1410 (1998). 81. Blue, R., et al. Structural and therapeutic insights from the species specificity and in vivo antithrombotic activity of a novel alphaIIb-specific alphaIIbbeta3 antagonist. Blood 114, 195-201 (2009). 82. Negri, A., et al. Structure-based virtual screening of small-molecule antagonists of platelet integrin αIIbβ3 that do not prime the receptor to bind ligand. Journal of computer-aided molecular design 26, 1005-1015 (2012). 83. Zhu, J., et al. Closed headpiece of integrin alphaIIbbeta3 and its complex with an alphaIIbbeta3-specific antagonist that does not induce opening. Blood 116, 5050-5059 (2010). | |
dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/50475 | - |
dc.description.abstract | 血小板在正常生理下的凝血功能扮演重要的角色,然而在病理情況下,不正常的血栓形成可能會造成嚴重的心血管疾病。Disintegrins 是一種從蛇毒蛋白中發現的一群具有RGD 或是KGD 氨基酸序列之胜肽血小板抑制劑,作用機轉為抑制血小板上的integrin αIIbβ3,它可專一地接在受器上,目前有以此結構為基礎發展成在臨床上使用的血小板抑制劑,然而現行臨床上使用的αIIbβ3 拮抗劑普遍存在著增加出血風險及血小板低下等危及生命的副作用。Echistatin 是一種短鏈的disintegrin,具有49 個氨基酸及4 對雙硫鍵。本篇從蘇氏鋸鱗蝰(Echis carinatus sochureki)中分離出ECS-f1 及ECS-f3,以及藉由酵母菌表現系統合成出Echistatin wild type 與數種C 端氨基酸進行變異的echistatin突變衍生物(由國立成功大學莊偉哲教授提供),探討不同echistatins 之間抗血栓作用及作用機轉。在活性方面,ECS-f1、ECS-f3、ECH WT 及echistatin 突變衍生物都具有濃度相關性地有效抑制人類富含血小板的血漿和人類血小板懸浮液中所引發的凝集反應。藉由流式細胞儀的分析,也可以觀察到echistatin 會增強在collagen 和thrombin 活化下血小板的p-selectin 表現;另外,這些echistatin 都會競爭單株抗體7E3 在integrin αIIbβ3 上的結合位置,但增加單株抗體AP5 的結合程度。而當echistatins 加入integrin αIIbβ3 的抑制性單株抗體AP2 的情況下,會造成FcγRII 的靠近,引發血小板凝集反應,造成下游訊息傳遞分子的磷酸化,例如PLCγ2、FAK、Syk 及Src。從clot retraction 試驗中可以觀察到,ECS-f3 可以顯著地抑制血塊的凝縮。動物體外實驗也可以看出,ECS-f3、ECH WT、ECHP47A、ECH K45A 也都呈現藥物濃度相關性地有效抑制collagen 在小鼠富含血小板的血漿中所引發的血小板凝集反應。另外,在尾靜脈給予抗血栓劑量下,四組也都會延長小鼠尾部出血時間,然而卻不會影響血小板數量。活體實驗中,以氯化鐵造成小鼠頸動脈傷害引發血栓產生,ECS-f3、ECH WT、ECHP47A、ECH K45A 皆可延長血栓生成及血管栓塞的時間。在本篇的實驗當中,雖自行分離出的disintegrin 和echistatin 突變衍生物的藥理作用和抗血栓作用,大體上都具有一定的有效性,但是由於C 端序列的不同,造成在抑制血栓活性、造成αIIbβ3 構型改變和其他其他實驗中具有特性上的差異。因此我們希望能夠找出C 端序列對於echistatin 的影響性,可以在未來設計較不會造成αIIbβ3 構型改變及較少出血風險的新一代integrin αIIbβ3 拮抗劑中提供一些有價值的資訊。 | zh_TW |
dc.description.abstract | Physiological haemostasis is mediated by platelets and coagulation system. However, in pathological situation, inappropriate thrombus formation leads to vessels occlusion in various cardiovascular diseases. Disintegrins are small-mass platelet aggregation inhibitors found in the snake venom. Disintegrins are potent antithrombotic agents, acting through the blockage of integrin αIIbβ3, an essential fibrinogen receptor in mediating platelet aggregation. Echistatin is a small-sized disintegrin which is a 49-amino acid RGD-containing peptide linked by four disulfide bonds. In this article, we purified disintegrin ECS-f1 and ECS-f3 from snake venom of Echis Carinatus sochureki. Furthermore, previous studies support a functional role for the C-terminal residues of echistatin in modulating its binding affinity towards integrin αIIbβ3, thus, different mutants of echistatin were expressed for comparative study (by Prof. Woei-Jer Chuang, National Cheng Kung University College of Medicine). ECS-f1, ECS-f3, ECH WT and echistatin mutants concentration-dependently inhibited platelet aggregation both in human platelet-rich plasma and platelet suspension. By using flow cytometry, we found that ECS-f1, ECS-f3, ECH WT and echistatin mutants enhanced the expression of P-selectin in collagen or thrombin-stimulated human platelet. In direct binding assay, echistatin significantly inhibited 7E3, however, enhanced AP5 mAb binding to platelet. In the presence of AP2, an inhibitory mAb of integrin αIIbβ3, both ECS-f1, ECS-f3 and echistatin mutants induced platelet aggregation around two-fold IC50 value, due to FcγRII recruitment and further caused the phosphorylation of downstream signaling molecules, including PLCγ2, FAK, Syk and Src. Furthermore, ECS-f3 could significantly affect clot retraction of platelet-rich plasma. Moreover, ECS-f3, ECH WT, ECH P47A and ECH K45A concentration-dependently inhibited platelet aggregation in mice PRP in vitro and dose-dependently inhibited platelet aggregation of PRP in response to collagen and ex vivo. Furthermore, all members of echistatins significantly prolonged the mice tail bleeding time as they were intravenously administered at antithrombotic doses (300 ng/g). However, all of them did not alter the platelet counts. In the in vivo ferric chloride (FeCl3)-induced arterial thrombosis model, administration of ECS-f3, ECH WT, ECH P47A and ECH K45A could prolong the occlusion time. In summary, the purified disintegrin echistatins and the echistatin mutants exhibited antithrombotic effects as tested in platelets and mice models. However, they showed various potency in antithrombotic activity and induction of conformational change as recognized by mAb AP2, suggesting that C-terminal residues are essential for modulating its binding activity. Further investigation of the structure-activity relationship of echistatins is needed for providing more information for the design of novel integrin αIIbβ3 inhibitors. | en |
dc.description.provenance | Made available in DSpace on 2021-06-15T12:42:18Z (GMT). No. of bitstreams: 1 ntu-105-R03443008-1.pdf: 3981004 bytes, checksum: c5cae03695f10b1040ef5df8f07cb7ec (MD5) Previous issue date: 2016 | en |
dc.description.tableofcontents | Content .............................................................................................................................. i
Abbreviations .................................................................................................................. vi Tables ............................................................................................................................. viii Figures ............................................................................................................................. ix 中文摘要 ....................................................................................................................... xiii Abstract ........................................................................................................................... xv Chapter 1 Introduction ...................................................................................................... 1 1.1 Platelets in hemostasis and thrombosis .................................................................. 1 1.2 Stages in the formation of platelet plugs................................................................ 2 1.3 Multiple receptors and ligands on platelet ............................................................. 4 1.4 Intergrins ................................................................................................................ 5 1.5 Structure and signaling of platelet integrins αIIbβ3 .............................................. 6 1.6 Disintegrins ............................................................................................................ 8 1.7 Integrin αIIbβ3 antagonists .................................................................................. 10 1.8 Thrombocytopenia induced by integrin αIIbβ3 antagonists ................................ 10 1.9 Motivation ............................................................................................................ 12 Chapter 2 Materials and Methods ................................................................................... 26 2.1 Materials .............................................................................................................. 26 2.1.1 Reagents ....................................................................................................... 26 2.1.2 Animals ........................................................................................................ 27 2.2 Methods ............................................................................................................... 28 2.2.1 Purification of platelet aggregation inhibitor from Echis carinatus sochureki ..................................................................................................................... 28 2.2.2 BCA assay for protein quantification ........................................................... 29 2.2.3 SDS-polyacrylamide gel electrophoresis (SDS-PAGE) ............................... 29 2.2.4 Coomassie blue staining ............................................................................... 30 2.2.5 Matrix assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) ...................................................................................... 30 2.2.6 Expression of echistatin wild type and its mutants in P. pastoris and purification ................................................................................................... 30 2.2.7 Preparation of human platelet-rich plasma (PRP) ........................................ 33 2.2.8 Preparation of human platelet suspension (PS) ............................................ 33 2.2.9 Platelet aggregation ...................................................................................... 34 2.2.10 Flow cytometric analysis of ECS-f3 and echistatin mutants binding to platelets ........................................................................................................ 34 2.2.11 Analysis of P-selectin expression ............................................................... 35 2.2.12 Western blot analysis .................................................................................. 36 2.2.13 Clot retraction ............................................................................................. 36 2.2.14 In vitro and ex vivo mouse platelet aggregation ......................................... 37 2.2.15 Tail bleeding assay in mice ........................................................................ 38 2.2.16 Platelet counts in mice ............................................................................... 38 2.2.17 Ferric chloride (FeCl3) induced arterial thrombosis model ........................ 38 2.2.18 Statistical analysis ...................................................................................... 39 Chapter 3 Results ............................................................................................................ 40 3.1 Purification of ECS-f1 and ECS-f3 from Echis carinatus sochureki snake venom ............................................................................................................................... 40 3.2 Determination of molecular mass of ECS-f1 and ECS-f3 ................................... 41 3.3 Protein identification of ECS-f1 and ECS-f3 ...................................................... 41 3.4 Effect of ECS-f1 and ECS-f3 on platelet aggregation of human platelet suspension ............................................................................................................................... 41 3.5 Effect of ECS-f1 and ECS-f3 on platelet aggregation of human platelet-rich plasma ............................................................................................................................... 42 3.6 Concentration-dependent inhibition of ECS-f1, ECS-f3 and echistatin mutants in human platelet-rich plasma and platelet suspension ............................................. 43 3.7 Combination of ECS-f1, ECS-f3 or other echistatin mutants and AP2 antibody induced platelet aggregation ................................................................................. 44 3.8 The signal transduction of ECS-f3 and AP2 antibody induced platelet aggregation ............................................................................................................................... 44 3.9 Effect of ECS-f1, ECS-f3 and echistatin mutants on P-selectin expression ........ 45 3.10 Measuring the binding epitopes on integrin αIIbβ3 of ECS-f1, ECS-f3 and other echistatin mutants .................................................................................................. 46 3.11 The effect of ECS-f3 and other echistatin mutants on clot retraction. ............... 47 3.12 In vitro and ex vivo effect of ECS-f3 and echistatin mutants on mice platelet-rich plasma ................................................................................................................... 48 3.13 Effect of ECS-f3 and echistatin mutants on tail bleeding time and platelet counts ............................................................................................................................... 49 3.14 Anti-thrombotic activity of ECS-f3, ECH WT, ECH P47A and ECH K45A in FeCl3-induced arterial thrombosis ......................................................................... 49 Chapter 4 Discussion ...................................................................................................... 91 Chapter 5 Conclusions and Perspectives ...................................................................... 103 References .................................................................................................................... 106 | |
dc.language.iso | en | |
dc.title | 蛇毒蛋白Echistatin和其突變衍生物之抗血栓作用及
作用機轉之探討 | zh_TW |
dc.title | The antithrombotic effects and action mechanisms of
disintegrin, echistatin and its mutants | en |
dc.type | Thesis | |
dc.date.schoolyear | 104-2 | |
dc.description.degree | 碩士 | |
dc.contributor.oralexamcommittee | 鄧哲明,楊春茂,吳文彬 | |
dc.subject.keyword | 蛇毒蛋白,抗黏著蛋白,抗血栓, | zh_TW |
dc.subject.keyword | snake venom,disintegrin,antithrombotic, | en |
dc.relation.page | 115 | |
dc.identifier.doi | 10.6342/NTU201601493 | |
dc.rights.note | 有償授權 | |
dc.date.accepted | 2016-07-27 | |
dc.contributor.author-college | 醫學院 | zh_TW |
dc.contributor.author-dept | 藥理學研究所 | zh_TW |
顯示於系所單位: | 藥理學科所 |
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