C-Phycocyanin抑制血小板凝集作用之機轉探討

Mechanisms Involved in the Antiplatelet Activity of C-Phycocyanin

周伯修(Po-Hsiu Chou)

指導教授 ： 許準榕

臺北醫學大學/醫學院/醫學科學研究所/碩士(2004年)

摘要

藻藍素(C-phycocyanin, C-PC)是一種藻膽色素蛋白質(phyco- biliprotein)，為藍綠藻類中螺旋藻(Spirulina platensis)的主要成分。此ㄧ蛋白質色素已被證實具有抗癌、自由基清除、抗氧化及抗發炎的特性。然而C-PC在血小板上的藥理學功效尚未明確，因此我們有意探討C-PC在血小板活化過程中，對於訊息傳遞方面的抑制作用。研究結果顯示，C-PC隨著濃度的增加(0.5 - 10 nM)，能有效地抑制collagen (1 ?g/ml)與U46619 (1 ?M)所引起的人類血小板凝集反應以及ATP釋放反應；C-PC (4和8 nM)亦可顯著地抑制collagen (1 ?g/ml)所引起的細胞內鈣離子移動以及thromboxane A2 (TxA2)的生合成。另外，C-PC (4和8 nM)可有意義的增加人類血小板細胞內nitrite與cyclic GMP的含量，但對於cyclic AMP的含量並沒有影響；另一方面，C-PC (4和8 nM)可有效地清除由collagen (1 ?g/ml)刺激所產生的hydroxyl radicals以及誘發vasodilator-stimulated phosphopretein (VASP)之磷酸化。PDBu (150 nM)可誘發protein kinase C的活化，並且將47 kDa proteins磷酸化，C-PC (4和8 nM)可有效地抑制已標記[?-32P] ATP的人類血小板發生此磷酸化反應。 由結果證實，C-PC抑制血小板活性的作用可能牽涉下列路徑：(一) C-PC會刺激NO的生成，接著增加血小板細胞內cyclic GMP的含量，並且誘發VASP磷酸化、抑制protein kinase C的活性以及47 kDa proteins磷酸化反應。(二) C-PC利用其清除hydroxyl radicals的作用，影響phospholipase A2-cyclooxygenase路徑的反應，進一步抑制TxA2的生合成。綜合以上結果，導致C-PC抑制血小板細胞內鈣離子的移動以及濃度的增加，最後因而抑制血小板的凝集反應。此項作用意味著C-PC可有效地應用在治療與血小板過度活化相關之疾病。

關鍵字

藻藍素 ； 訊息傳遞

並列摘要

C-phycocyanin (C-PC), a phycobiliprotein, is one of the major constituents of blue-green algae, Spirulina platensis. This protein pigment has been reported to have anti-cancer, free radical scavenging, anti-oxidant and anti-inflammatory properties. However, the pharmacological functions of C-PC on platelets were not yet understood, we are interesting in investigating the inhibitory effects of C-PC on cellular signal transduction during the process of platelet activation. In this study, C-PC concentration-dependently (0.5-10 nM) inhibited collagen (1 ?g/ml)- and U46619 (1 ?M)-induced human platelets aggregation and ATP-release reaction. In addition, C-PC (4 and 8 nM) markedly inhibited intracellular Ca2+ mobilization in Fura-2 AM-loaded platelets and thromboxane A2 formation stimulated by collagen (1 ?g/ml). Furthermore, C-PC (4 and 8 nM) significantly increased the formations of nitrate and cyclic GMP but not cyclic AMP in human platelets. Moreover, C-PC (4 and 8 nM) obviously scavenged collagen (1 ?g/ml)-induced hydroxyl radicals and induced the phosphorylation of vasodilator- stimulated phosphoprotein (VASP). Rapid phosphorylation of a protein of Mr 47,000 (P47), a marker of protein kinase C activation, was triggered by PDBu (150 nM). This phosphorylation was inhibited by C-PC (4 and 8 nM) in [?-32P] ATP-labeled human platelets. In conclusion, our study suggested that the possible pathways of anti-platelet activity of C-PC may involve the following pathway: (1) C-PC stimulated nitrate formation, followed by increasing the amount of cyclic GMP and then induced VASP phosphorylation, inhibited protein kinase C activation and 47 kDa protein phosphorylation. (2) C-PC significantly inhibited thromboxane A2 formation through scavenging the hydroxyl radicals to inhibit phospholipase A2-cyclooxygenase pathway, and intracellular Ca2+ mobilization. Taken together, C-PC may be used as an effective tool in treating pathological disorder associated with platelet hyperaggregability.

並列關鍵字

C-phycocyanin ； signal transduction

參考文獻

Adelstein RS and Conti MA. (1975) Phosphorylation of platelet myosin increases actin-activated myosin ATPase activity. Nature. 256: 597-598.

Amatruda JF, Gattenneia DJ, Karpoua TS and Cooper JA. (1991) Effects of null mutations and overexpression of capping proteins on morphogenesis, actin distribution and polarized secretion in yeast. J Cell Biol. 119: 1151-1162.

Andersson PB, Perry VH and Gordon S. (1991) The kinetics and morphological characteristics of the macrophagemicroglial response to kainic acid-induced neuronal degeneration. Neuroscience. 42: 201-214.

Antiplatelet Trialists Collaboration. (1988) Secondary prevention of vascular events by prolonged antiplatelet therapy. Br Med J. 296: 320-331.

Armstrong RA, Jones RL and Wilson NH. (1985) Mechanism of the inhibition of platelet aggregation produced by prostaglandin F 2 alfa. Prostaglandins. 29: 601-610.