Proteomic and functional characterisation of platelet microparticle size classes.
Proteomic and functional characterisation of platelet microparticle size classes.
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DOI：
10.1160/th09-04-243
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发表时间：
2009-10
影响因子：
6.7
通讯作者：
Powell DW
中科院分区：
文献类型：
--
作者：
Dean WL;Lee MJ;Cummins TD;Schultz DJ;Powell DW
活化的血小板释放称为微粒的大的脂质-蛋白质复合物。这些血小板微粒（PMP）由质膜的囊泡碎片和α颗粒组成。PMP促进凝血，促进血小板和白细胞粘附于内皮下基质，支持血管生成和刺激血管平滑肌增殖。PMP分为4个大小类别，以便于鉴定活性蛋白和脂质组分。从活化的人血小板中获得PMP，并通过凝胶过滤色谱法将其分成4个大小级。使用二维液相色谱串联质谱法鉴定蛋白质。使用PFA-100®测定对血小板的功能作用，并通过测量跨内皮细胞电阻测定对内皮细胞的功能作用。PMP大小级别在其质膜受体和粘附分子、趋化因子、生长因子和蛋白酶抑制剂的含量上存在显著差异。两个最小尺寸的类（3和4）抑制胶原/腺苷二磷酸介导的血小板血栓形成，而组分2和4刺激内皮细胞形成屏障。热变性阻断组分4对内皮细胞功能的影响，但不阻断组分2，这意味着组分4中的活性组分是蛋白质，组分2中的活性组分是热稳定蛋白质或脂质，但不是鞘氨醇-1-磷酸。PMP大小级分的蛋白质组学和功能分析表明，PMP可以分成不同的大小类别，不同的蛋白质组分，蛋白质/脂质比，以及对血小板和内皮细胞的功能影响。这项分析将有助于识别PMP中的活性成分，并阐明它们与动脉粥样硬化和癌症等疾病的关系。
Activated platelets release large lipid-protein complexes termed microparticles. These platelet microparticles (PMP) are composed of vesicular fragments of the plasma membrane and α-granules. PMP facilitate coagulation, promote platelet and leukocyte adhesion to the subendothelial matrix, support angiogenesis and stimulate vascular smooth muscle proliferation. PMP were separated into 4 size classes to facilitate identification of active protein and lipid components. PMP were obtained from activated human platelets and separated into 4 size classes by gel filtration chromatography. Proteins were identified using 2-dimensional, liquid chromatography tandem mass spectrometry. Functional effects on platelets were determined using the PFA-100® and on endothelial cells by measuring transendothelial cell electrical resistance. PMP size classes differed significantly in their contents of plasma membrane receptors and adhesion molecules, chemokines, growth factors and protease inhibitors. The two smallest size classes (3 and 4) inhibited collagen/adenosine-diphosphate-mediated platelet thrombus formation, while fractions 2 and 4 stimulated barrier formation by endothelial cells. Heat denaturation blocked the effect of fraction 4 on endothelial cell function, but not fraction 2 implying that the active component in fraction 4 is a protein and in fraction 2 is a heat-stable protein or lipid but not sphingosine-1-phosphate. Proteomic and functional analysis of PMP size fractions has shown that PMP can be separated into different size classes that differ in protein components, protein/lipid ratio, and functional effects on platelets and endothelial cells. This analysis will facilitate identification of active components in the PMP and clarify their involvement in diseases such as atherosclerosis and cancer.