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研究生:楊怡萱
研究生(外文):Yi-Hsuan Yang
論文名稱:Sulforaphane促進前列腺素之抗血小板活性探討
論文名稱(外文):Sulforaphane enhances the antiplatelet activity of prostaglandins
指導教授:吳志中吳志中引用關係
指導教授(外文):Chih-Chung Wu
學位類別:碩士
校院名稱:高雄醫學大學
系所名稱:天然藥物研究所
學門:醫藥衛生學門
學類:藥學學類
論文種類:學術論文
論文出版年:2014
畢業學年度:102
語文別:中文
論文頁數:87
中文關鍵詞:血小板sulforaphaneprostaglandinscAMPPI3K
外文關鍵詞:plateletssulforaphaneprostaglandinscAMPPI3K
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血小板在動脈粥狀硬化的病程或是血栓形成等心血管疾病皆扮演著重要的角色。因此,在治療和預防心血管疾病上,抑制血小板活化已是確立的目標。Sulforaphane是一種異硫氰酸酯,常見於十字花科的蔬菜中,已知sulforaphane在體外及體內模式具有抑制血小板活化及血栓形成的作用。人體血管中的內皮細胞會釋放prostaglandin I2 (PGI2),增加血小板cAMP含量而抑制血小板活化。在研究中,我們想探討sulforaphane是否會增強prostaglandins抗血小板的效果,並進一步研究其作用機轉。

實驗結果顯示,sulforaphane (10-20 μM) 可協同PGI2類似物 (iloprost)、PGE1及forskolin抑制protease activated receptor-1 (PAR1) 致效劑和thromboxane A2 受體致效劑 U46619所引起的血小板凝集。Sulforaphane不會促進PGE1所引起的cAMP的增加,且protein kinase A (PKA) 抑制劑並不會防止sulforaphane促進PGE1抑制血小板凝集;此結果顯示cAMP/PKA路徑並非sulforaphane的主要作用標的。Sulforaphane可促進PGE1抑制PAR1致效劑所引起的phosphoinositide-3 kinase (PI3K)/Akt路徑活化,但不影響protein kinase C (PKC)/Ca2+路徑。Sulforaphane併用PGE1不會抑制PAR1致效劑所造成ADP的釋放,卻可抑制2MesADP (ADP類似物) 引起之PI3K/Akt路徑活化。此外,PI3K抑制劑wortmannin也可促進PGE1對血小板凝集的抑制作用。綜合上述結果,可推論sulforaphane藉由抑制PI3K/Akt路徑促進前列腺素抑制血小板活化的作用。

Platelets play an important role in cardiovascular diseases (CVDs) both in the pathogenesis of atherosclerosis and in the development of acute thrombotic events. Therefore, platelet inhibition is an established target in treatment and prevention of CVDs. Sulforaphane is an isothiocyanate found naturally in cruciferous vegetables, it has been shown to possess antiplatelet activity and prevent thromboembolism in vitro and in vivo.
Prostaglandin I2 (PGI2) is an endogenous antiplatelet substance released from endothelial cells in human blood vessel, it causes an increase in cAMP levels in platelets leading to inhibition of platelet activation. In the present study, we want to explore whether sulforaphane enhances the antiplatelet effect of prostaglandins, and to investigate the mechanism of the actions.
The results show that sulforaphane (10-20 μM) enhanced PGI2 analog (iloprost), PGE1, and forskolin to inhibit platelet aggregation caused by protease activated receptor-1 (PAR1) agonist and thromboxane A2 receptor agonist (U46619). Sulforaphane did not enhance the increase in cAMP by PGE1. Futhermore, the protein kinase A (PKA) inhibitor H89 did not reverse the inhibition of platelet aggregation caused by sulforaphane plus PGE1. Therefore, the enhancement of antiplatelet activity of PGE1 by sulforaphane is not mainly related to the cAMP/PKA pathway. Sulforaphane enhanced PGE1 to inhibit the activation of phosphoinositide-3 kinase (PI3K)/Akt pathway, but not PKC/Ca2+ pathway, caused by PAR1 agonist. The combination of sulforaphane and PGE1 did not inhibit ADP release caused by PAR1 agonist but inhibited 2MesADP (an ADP analog)-induced PI3K/Akt activation. Similar to sulforaphane, the PI3K inhibitor wortmannin also enhanced PGE1 to inhibit platelet aggregation. Taken together, our results suggest that sulforaphane enhances the antiplatelet effect of prostaglandins by inhibiting PI3K/Akt pathway.

縮寫表 6
摘要 9
Abstract 11
第一章 緒論 13
血小板的活化 15
血小板活化的作用機制 19
抑制血小板活化的作用機制 26
抗血小板藥物 28
Sulforaphane藥物簡介 32
研究動機與背景 39
第二章 實驗材料與方法 40
實驗材料 41
實驗方法 42
第三章 實驗結果 47
Sulforaphane合併PGE1對血小板凝集反應的影響 48
Sulforaphane合併forskolin對血小板凝集反應的影響 48
Sulforaphane合併PGI2對血小板凝集反應的影響 48
Sulforaphane合併PGE1對血小板內cyclic nucleotides含量的影響 49
Sulforaphane 合併 PGE1對VASP磷酸化的影響 50
Sulforaphane與PKA抑制劑作用探討 50
Sulforaphane 合併 PGE1對血小板內PI3K/Akt活化的影響 51
Sulforaphane 合併 PGE1對protein kinase C (PKC)的影響 51
Sulforaphane 合併 PGE1對血小板內鈣離子的影響 52
Sulforaphane與PI3K抑制劑作用比較 53
Sulforaphane 合併 PGE1對ATP釋放的影響 53
第四章 圖表 54
第五章 綜合討論與未來展望 73
第六章 參考文獻 78


Albers GW. Role of ticlopidine for prevention of stroke. Stroke. 1992; 23: 912-916.

Andrews RK, Arthur JF, Gardiner EE. Targeting GPVI as a novel antithrombotic strategy. J Blood Med. 2014; 5: 59-68.

Barry FA, Graham GJ, Fry MJ, Gibbins JM. Regulation of glycogen synthase kinase 3 in human platelets: a possible role in platelet function? FEBS Lett. 2003; 553: 173-178.

Bennett JS, Berger BW, Billings PC. The structure and function of platelet integrins. J Thromb Haemost. 2009; 7 Suppl 1: 200-205.

Blair P, Flaumenhaft R. Platelet alpha-granules: basic biology and clinical correlates. Blood Rev. 2009; 23: 177-89.

Boddupalli S, Mein JR, Lakkanna S, James DR. Induction of phase 2 antioxidant enzymes by broccoli sulforaphane: perspectives in maintaining the antioxidant activity of vitamins a, C, and e. Front Genet. 2012; 3: 7.

Bozulic L, Hemmings BA. PIKKing on PKB: regulation of PKB activity by phosphorylation. Curr. Opin. Cell Biol. 2009; 21: 256-261.

Bynagari-Settipalli YS, Lakhani P, Jin J, Bhavaraju K, Rico MC, Kim S, WoulfeD, Protein kinase C isoform ε negatively regulates ADP-induced calcium mobilization and thromboxane generationin platelets. Arterioscler Thromb Vasc Biol. 2012; 32: 1211-9.

Cantley LC. The phosphoinositide 3-kinase path way. Science. 2002; 296: 1655-1657.

Chen D, Lemons PP, Schraw T, Whiteheart SW. Molecular mechanisms of platelet exocytosis: role of SNAP-23 and syntaxin 2 and 4 in lysosome release. Blood. 2000; 96: 1782-8.

Cheung KL, Kong AN. Molecular targets of dietary phenethyl isothiocyanate and sulforaphane for cancer chaemoprevention. AAPS J. 2010; 12: 87-97.

Chuang WY , Kung PH, Kuo CY, Wu CC. Sulforaphane prevents human platelet aggregation through inhibiting the phosphatidylinositol 3-kinase/Akt pathway. Thromb Haemost. 2013; 109: 1120-30.

Cruz MA, Chen J, Whitelock JL, Morales LD, Lopez JA. The platelet glycoprotein Ib-von Willebrand factor interaction activates the collagen receptor alpha 2 beta 1 to bind collagen: activation-dependent conformational change of the alpha2-I domain. Blood. 2005; 105: 1986-1991.

De Candia E. Mechanisms of platelet activation by thrombin: a short history. Thromb Res. 2012; 129: 250-6.

Dittrich M, Birschmann I, Mietner S, Sickmann A, Walter U, Dandekar T. Platelet protein interactions: map, signaling components, and phosphorylation groundstate. Arterioscler Thromb Vasc Biol. 2008; 28: 1326-31.

Elzagallaai A, Rose SD, Brandan NC, Trifaro JM. Myristoylated alanine-rich C kinase substrate phosphorylation is involved in thrombin-
induced serotonin releasefrom platelets. Br J Haematol. 2001; 112: 593-602

Fahey JW, Zhang Y, Talalay P. Broccoli sprouts: an exceptionally rich source of inducers of enzymes that protect against chemical carcinogens.
Proc Natl Acad Sci U S A. 1997; 94: 10367-72.

Fahey JW, Zalcmann AT, Talalay P. The chemical diversity and distribution of glucosinolates and isothiocyanates among plants.
Phytochemistry. 2001; 56: 5-51.

Ferreiro JL, Angiolillo DJ. Clopidogrel response variability: current status and future directions. Thromb Haemost. 2009; 102: 7-14.

Ferreiro JL, Angiolillo DJ. New directions in antiplatelet therapy. Circ Cardiovasc Interv. 2012; 5: 433-45.

Fontana P, Zufferey A, Daali Y, Reny JL. Antiplatelet therapy: targeting the TxA2 pathway. J Cardiovasc Transl Res. 2014; 7: 29-38.

Furchgott RF and Zawadzki JV. The obligatory role of endothelial cells in the relaxation of arterial smooth muscle by acetylcholine. Nature. 1980; 288: 373-376.

Gertler FB, Niebuhr K, Reinhard M, Wehland J, Soriano P. Mena, a relative of VASP and Drosophila Enabled, is implicated in the control of microfilament dynamics. Cell. 1996; 87: 227-239.

Gresele P, Momi S, Falcinelli E, et al. Anti-platelet therapy: phosphodiesterase inhibitors. Br J Clin Pharmacol. 2011; 72: 634-646.

Hardy AR, Jones ML, Mundell SJ, Poole AW. Reciprocal cross-talk between P2Y1 and P2Y12 receptors at the level of calcium signaling in human platelets. Blood. 2004; 104: 1745-52.

Hanada M, Feng J, Hemmings BA. Structure, regulation and function of PKB/AKT- a major therapeutic target. Biochim. Biophys. Acta. 2004; 1697: 3–16.

Healy AM, Pickard MD, Pradhan AD, Wang Y, Chen Z, Croce K, Sakuma M, Shi C, Zago AC, Garasic J, Damokosh AI, Dowie TL, Poisson L, Lillie J, Libby P,Ridker PM, Simon DI. Platelet expression profiling and clinical validation of myeloid-related protein-14 as a novel determinant of cardiovascular events. Circulation. 2006; 113: 2278-84.

Hechler B, Gachet C. P2 receptors and platelet function. Purinergic Signal. 2011; 7: 293-303.

Jackson SP, Schoenwaelder SM, Goncalves I, Nesbitt WS, Yap CL, Wright CE, Kenche V, Anderson KE, Dopheide SM,Yuan Y, Sturgeon SA, Prabaharan H, Thompson PE, Smith GD, Shepherd PR, Daniele N, Kulkarni S, Abbott B, Saylik D,Jones C, Lu L, Giuliano S, Hughan SC, Angus JA, Robertson AD, Salem HH. PI3-kinase p110beta: a new target for antithrombotic therapy. Nat Med. 2005; 11: 507-14.

Jang EK, Azzam JE, Dickinson NT, Davidson MM, Haslam RJ. Roles for both cyclic GMP and cyclic AMP in the inhibition of collagen-induced platelet aggregation by nitroprusside. Br J Haematol. 2002; 117: 664-675.

Jayakumar T , Chen WF, Lu WJ, Chou DS, Hsiao G, Hsu CY, Sheu JR, Hsieh CY. A novel antithrombotic effect of sulforaphane via activation of platelet adenylate cyclase: ex vivo and in vivo studies. J Nutr Biochem. 2013; 24: 1086-95.

Jennifer Yeung and Michael Holinstat. Newer agents in antiplatelet therapy: a review J Blood Med. 2012; 3: 33-42.

Jin J, Kunapuli SP. Coactivation of two different G protein-coupled
Receptors is essential for ADP-induced Platelet aggregation. Proc Natl Acad Sci U S A. 1998; 95: 8070-4.

Joel S. Bennett Structure and function of the platelet integrin αIIbβ3. J Clin Invest. 2005; 115: 3363–69.

Kehrel, B. Platelet-collagen interactions. Semin Thromb Hemost. 1995; 21: 123-9.

Keum YS. Regulation of the Keap1/Nrf2 system by chaemopreventive sulforaphane: implications of posttranslational modifications. Ann NY Acad Sci. 2011; 1229: 184-189.

Kim S, Kunapuli SP. P2Y12 receptor in platelet activation. Platelets. 2011; 22: 54-58.

Kleiman NS,Freedman JE,Tracy PB,Furie BC,Bray PF,Rao SV,Phillips DR,Storey RF,Rusconi CP,French PA,Steinhubl SR,Becker RC. Platelets: developmental biology, physiology, and translatable platforms for preclinical investigation and drug development. Platelets. 2008; 19: 239-51.

Lahav J, Jurk K, Hess O, Barnes MJ, Farndale RW, Luboshitz J, Kehrel BE. Sustained integrin ligation involves extracellular free sulfhydryls and enzymatically catalyzed disulfide exchange. Blood. 2002; 100: 2472-8.

Lechtenberg BC, Freund SM, Huntington JA. GpIbα interacts exclusively with exosite II of thrombin. J Mol Biol. 2014; 426: 881-93.

Li D, August S, Woulfe DS. GSK3beta is a negative regulator of platelet function and thrombosis. Blood. 2008; 111: 3522-30.

Li W, Tang X, Yi W, Li Q, Ren L, Liu X, Chu C, Ozaki Y, Zhang J, Zhu L. Glaucocalyxin A inhibits platelet activation and thrombus formation preferentially via GPVI signaling pathway. PLoS One. 2013; 8: e85120.

Mackman N. Triggers, targets and treatments for thrombosis. Nature. 2008; 451: 914-8

McManus EJ, Sakamoto K, Armit LJ, et al. Role that phosphorylation of GSK3 plays in insulin and Wnt signalling defined by knockin analysis. Embo J. 2005; 24: 1571-83.

Miller OV, Gorman RR. Evidence for distinct prostaglandin I2 and D2 receptors in human platelets. J Pharmacol Exp Ther. 1979; 210: 134-40.

Moncada S and Higgs EA. The discovery of nitric oxide and its role in vascular biology. Br J Pharmacol. 2006; 147 Suppl 1: S193-201.

Morrell CN, Matsushita K, Chiles K, et al. Regulation of platelet granule exocytosis by S-nitrosylation. Proc Natl Acad Sci U S A. 2005; 102: 3782-87.

Naik MU, Stalker TJ, Brass LF, Naik UP. JAM-A protects from thrombosis by suppressing integrin αIIbβ3-dependent outside-in signaling in platelets. Blood. 2012; 119: 3352-60.

O’Brien KA, Stojanovic-Terpo A, Hay N, Du X. An important role for Akt3 in platelet activation and thrombosis. Blood. 2011; 118: 4215-4223.

Offermanns S. Activation of platelet function through G protein-coupled receptors. Circ Res. 2006; 99: 1293-304.

Oh CH, Shin JI, Mo SJ, Yun SJ, Kim SH, Rhee YH. Antiplatelet activity of L-sulforaphane by regulation of platelet activation factors, glycoprotein IIb/IIIa and thromboxane A2. Blood Coagul Fibrinolysis. 2013; 24: 498-504.

Ostrowska M, Adamski P, Koziński M, Navarese EP, Fabiszak T, Grześk G, Paciorek P, Kubica J. Off-target effects of glycoprotein IIb/IIIa receptor inhibitors. Cardiol J. 2014

Paul BZ1,Daniel JL, Kunapuli SP. Platelet shape change is mediated by both calcium-dependent and -independent signaling pathways. Role of p160 Rho-associated coiled-coil-containing protein kinase in platelet shape change. J Biol Chem. 1999; 274: 28293-300.

Patrono C, Garcia Rodriguez LA, Landolfi R, Baigent C. Low-dose aspirin for the prevention of atherothrombosis. N Engl J Med. 2005; 353:
2373-383.

Ramaschi G, Torti M. Lectin-induced calcium mobilization in human platelets : use of fluorescent probes. Methods Mol Med. 1998; 9: 433-9.

Reinhard M, Jarchau T, Walter U. Actin-based motility: stop and go with Ena/VASP proteins. Trends Biochem Sci. 2001; 26: 243-249.

Riboli E, Norat T. Epidemiologic evidence of the protective effects of fruit and vegetables on cancer risk. Am J Clin Nutr. 2003; 78: 559-695.

Roberts W, Magwenzi S, Aburima A, Naseem KM. Thrombospondin-1 induces platelet activation through CD36-dependent inhibition of the cAMP/protein kinase A signaling cascade. Blood. 2010 18; 116: 4297-306.

Rondina MT, Weyrich AS. Targeting phosphodiesterases in anti-platelet therapy. Handb Exp Pharmacol. 2012;225-38.

Schlessinger J. Cell signaling by receptor tyrosine kinases. Cell. 2000; 103: 211-25.

Shror K. The pharmacology of cilostazol. Diabetes Obes Metab. 2002; 4: 14-9.

Siess W, Lapetina EG. Functional relationship between cyclic AMP-
dependent protein phosphorylation and platelet inhibition. Biochem J. 1990; 271: 815-819.

Smolenski A, Bachmann C, Reinhard K, et al. Analysis and regulation of vasodilator-stimulated phosphoprotein serine 239 phosphorylation in vitro and in intact cells using a phosphospecific monoclonal antibody. J Biol Chem. 1998; 273: 20029-35.

Smolenski A. Novel roles of cAMP/cGMP-dependent signaling in platelets. J Thromb Haemost. 2012; 10: 167-176.

Smyth SS, Woulfe DS, Weitz JI, Gachet C, Conley PB, Goodman SG, Roe MT, Kuliopulos A, Moliterno DJ, French PA,Steinhubl SR, Becker RC; 2008 Platelet Colloquium Participants. Arterioscler Thromb Vasc Biol. 2009 ; 29: 449-57.

Stephens L, Williams R, Hawkins P. Phosphoinositide 3-kinases as drug targets in cancer. Curr Opin Pharmacol. 2005; 5: 357-65.

Storey RF, Bliden KP, Patil SB, Karunakaran A, Ecob R, Butler K, Teng
R, Wei C, Tantry US, Gurbel PA: ONSET/OFFSET investigators. Incidence of dyspnea and assessment of cardiac and pulmonary function
in patients with stable coronary artery disease receiving ticagrelor, clopidogrel,or placebo in the ONSET/OFFSET study. J Am Coll Cardiol.
2010; 56: 185-193.

Vanhaesebroeck B, Waterfield MD.Signaling by distinct classes of phosphoinositide 3-kinases. Exp Cell Res. 1999; 253: 239-54.

Versteeg HH, Heemskerk JW, Levi M, Reitsma PH. New Fundamentals in Hemostasis. Physiol Rev. 2013; 93: 327-58.
Wangorsch G, Butt E, Mark R, Hubertus K, Geiger J, Dandekar T, Dittrich M. Time-resolved in silico modeling of fine-tuned cAMP signaling in platelets: feedback loops, titrated phosphorylations and pharmacological modulation. BMC Syst Biol. 2011; 5: 178.

Wang TY, Su NY, Shih PC, Tsai PS, Huang CJ. Anti-inflammation effects of naloxone involve phosphoinositide 3-kinase delta and gamma. J Surg Res. 2014: pii: S0022-4804(14)00572-1.

Wentworth JK, Pula G, Poole AW. Vasodilator-stimulated phosphor protein (VASP) is phosphorylated on Ser157 by protein kinase C- dependent and-independent mechanisms in thrombin-stimulated humun platelets. Biochem J. 2006; 393 : 555-64.

Weyrich A, Cipollone F, Mezzetti A, Zimmerman G. Platelets in atherothrombosis: new and evolving roles. Curr Pharm Des. 2007; 13: 1685-91.

Williams RL. Mammalian phosphoinositide-specific phospholipase C. Biochim Biophys Acta.1999; 1441: 255-67.

Wiviott SD, Trenk D, Frelinger AL, ODonoghue M, Neumann FJ, Mich-
elson AD, Angiolillo DJ, Hod H, Montalescot G, Miller DL,Jakubowski JA, Cairns R, Murphy SA, McCabe CH, Antman EM, Braunwald E; PRINCIPLE-TIMI 44 Investigators. Prasugrel compared with high loading- and maintenance-dose clopidogrel in patients with planned percutaneous coronary intervention: the prasugrel in comparison to clopidogrel for inhibition of platelet activation and aggregationthrombolysis in myocardial infarction 44 trial. Circulation. 2007; 116: 2923-2932.

Zapata JC, Cox D, Salvato MS. The Role of Platelets in the Pathogenesis of Viral Hemorrhagic Fevers. PLoS Negl Trop Dis. 2014; 8: e2858.

Zhao J, Kobori N, Aronowski J, et al. Sulforaphane reduces infarct volume following focal cerebral ischaemia in rodents. Neurosci Let.t 2006; 393: 108-112.

Zhang W, Colman RW. Thrombin regulates intracellular cAMP concentration in human platelets through phosphorylation/activation of PDE3A. Blood. 2007; 110: 1475-1482.

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