大蒜素和双硫仑可增强血小板整合素αIIbβ3与纤维蛋白原的结合。

Allicin and disulfiram enhance platelet integrin alphaIIbbeta3-fibrinogen binding.

作者信息

Manaster Yoav, Shenkman Boris, Rosenberg Nurit, Savion Naphtali

机构信息

Goldschleger Eye Research Institute and Department of Human Molecular Genetics and Biochemistry, Sackler Faculty of Medicine, Tel Aviv University, Israel.

出版信息

Thromb Res. 2009 Sep;124(4):477-82. doi: 10.1016/j.thromres.2009.06.019. Epub 2009 Jul 26.

DOI:10.1016/j.thromres.2009.06.019

PMID:19632706

Abstract

INTRODUCTION

Activation of the platelet receptor alphaIIbbeta3 (glycoprotein IIbIIIa) involves a change in the disulfide bonds pattern in the extra-cellular domain of the receptor. The disulfide-bond reducing agent, dithiothreitol (DTT), can increase integrin activity, and point mutations of specific cysteine residues of the integrin can cause its lockage at the high affinity state. The present study is aimed to support the hypothesis that prevention of specific alphaIIbbeta3 intra-molecular disulfide bond formation increases receptor-ligand binding activity.

METHODS

Platelet aggregation was induced by collagen or ADP and epinephrine. Integrin alphaIIbbeta3-fibrinogen binding was evaluated on prostaglandins E(1) (PGE(1))-treated washed platelets or baby hamster kidney (BHK) cells expressing human alphaIIbbeta3. Integrin was directly activated by an anti-ligand induced binding site (LIBS) PT25-2 antibody. The effect of sulfhydryl-reactive agents, such as allicin, glutathione, dithiobis nitrobenzoic acid (DTNB) and disulfiram, was tested on alphaIIbbeta3 activity.

RESULTS

Allicin (40 microM) completely inhibited washed platelets agonist-induced aggregation. Both allicin and disulfiram (40 microM) inhibited alphaIIbbeta3-fibrinogen binding and P-selectin expression in washed platelets. However, there was an increase in alphaIIbbeta3-fibrinogen binding but not P-selectin expression in PGE(1)-treated washed platelets activated by PT25-2 antibody. At a high concentration (400 microM) both inhibited alphaIIbbeta3-fibrinogen binding. Similarly, in BHK cells expressing alphaIIbbeta3 activated by PT25-2 antibody, allicin at a low concentration increased alphaIIbbeta3 activity.

CONCLUSIONS

Allicin and disulfiram inhibit agonist-induced washed platelet activation probably via inhibition of platelet signaling, but enhance PT25-2 antibody-induced alphaIIbbeta3 integrin activity most likely by preventing reformation of disulfide bridges thereby stabilizing the active conformation of the integrin.

摘要

引言

血小板受体αIIbβ3（糖蛋白IIbIIIa）的激活涉及受体胞外结构域中二硫键模式的改变。二硫键还原剂二硫苏糖醇（DTT）可增加整合素活性，整合素特定半胱氨酸残基的点突变可使其锁定在高亲和力状态。本研究旨在支持以下假说：防止特定的αIIbβ3分子内二硫键形成可增加受体-配体结合活性。

方法

用胶原蛋白或ADP和肾上腺素诱导血小板聚集。在前列腺素E1（PGE1）处理的洗涤血小板或表达人αIIbβ3的幼仓鼠肾（BHK）细胞上评估整合素αIIbβ3-纤维蛋白原结合。整合素通过抗配体诱导结合位点（LIBS）PT25-2抗体直接激活。测试了巯基反应性试剂如大蒜素、谷胱甘肽、二硫代双硝基苯甲酸（DTNB）和双硫仑对αIIbβ3活性的影响。

结果

大蒜素（40微摩尔）完全抑制洗涤血小板激动剂诱导的聚集。大蒜素和双硫仑（40微摩尔）均抑制洗涤血小板中αIIbβ3-纤维蛋白原结合和P-选择素表达。然而，在PT25-2抗体激活的PGE1处理的洗涤血小板中，αIIbβ3-纤维蛋白原结合增加，但P-选择素表达未增加。在高浓度（400微摩尔）时，二者均抑制αIIbβ3-纤维蛋白原结合。同样，在PT25-2抗体激活的表达αIIbβ3的BHK细胞中，低浓度的大蒜素增加αIIbβ3活性。