光学凝集测定法（出生类型）与流式细胞术在评估ADP诱导的血小板活化中的关系。

Relationships between optical aggregometry (type born) and flow cytometry in evaluating ADP-induced platelet activation.

作者信息

Sbrana Silverio, Della Pina Francesca, Rizza Antonio, Buffa Manuela, De Filippis Rossella, Gianetti Jacopo, Clerico Aldo

机构信息

Laboratory of Hematology and Flow Cytometry, CNR Institute of Clinical Physiology, Massa, Italy.

出版信息

Cytometry B Clin Cytom. 2008 Jan;74(1):30-9. doi: 10.1002/cyto.b.20360.

DOI:10.1002/cyto.b.20360

PMID:17630652

Abstract

BACKGROUND

Platelet response to activating agents is used to monitor the efficacy of anti-aggregation therapies. The aim of our study has been to demonstrate the existence of relationships between early events of ADP-induced platelet activation, measured by flow cytometry and platelet-rich plasma aggregation, quantified by optical aggregometry.

METHODS

We evaluated peripheral blood of 12 donors. The following parameters were quantified by cytometry after stimulation with adenosine diphosphate (ADP) (0.5, 1, 2, 5, 10, 20 muM): CD62P (P-selectin) and PAC-1 expression, and cytosolic Ca(2+) mobilization. Aggregation was measured by optical aggregometry. We also studied 13 patients, undergoing coronary stenting, treated with aspirin (before procedure) or with aspirin plus clopidogrel (after procedure). We evaluated CD62P and PAC-1 expression, aggregation, and vasodilator-stimulated phopshoprotein phosphorylation (platelet reactivity index, PRI).

RESULTS

Flow procedures were more sensitive than aggregometry, with a lowest interindividual variability. Linear relationships existed in donors between CD62P expression and Ca(2+) mobilization (P < 0.0001), and between aggregation and Ca(2+) mobilization (P < 0.0001). Linear relationships existed between aggregation and CD62P expression, as percentage (P < 0.0001), or relative fluorescence intensity (RFI) (P < 0.0001). Exponential equations related aggregation and PAC-1 expression, as percentage (P < 0.0001), or RFI (P < 0.0001). Linear relationships between aggregation and CD62P expression (as percentage) existed in the patients before (P = 0.0022) and after procedure (P = 0.0020). Exponential relationships between aggregation and PAC-1 expression (as percentage) existed before (P = 0.0012) and after procedure (P = 0.0024). Linear correlations related aggregation response predicted on CD62P expression, and measured aggregation inhibition after clopidogrel (P = 0.0013) as well as predicted aggregation and PRI inhibition (P = 0.0031).

CONCLUSIONS

Tight relationships between aggregation and cytometric quantification of platelet markers in whole blood, in particular CD62P, allow to predict aggregation response to ADP from flow data in patients treated with aspirin alone or with aspirin plus clopidogrel.

摘要

背景

血小板对激活剂的反应用于监测抗聚集治疗的疗效。我们研究的目的是证明通过流式细胞术测量的ADP诱导的血小板激活早期事件与通过光学聚集法量化的富血小板血浆聚集之间存在关联。

方法

我们评估了12名献血者的外周血。在用二磷酸腺苷（ADP）（0.5、1、2、5、10、20 μM）刺激后，通过细胞术对以下参数进行量化：CD62P（P-选择素）和PAC-1表达以及胞质Ca(2+)动员。通过光学聚集法测量聚集。我们还研究了13名接受冠状动脉支架置入术的患者，术前用阿司匹林治疗，术后用阿司匹林加氯吡格雷治疗。我们评估了CD62P和PAC-1表达、聚集以及血管舒张剂刺激的磷蛋白磷酸化（血小板反应性指数，PRI）。

结果

流式检测程序比聚集法更敏感，个体间变异性最低。在献血者中，CD62P表达与Ca(2+)动员之间存在线性关系（P < 0.0001），聚集与Ca(2+)动员之间也存在线性关系（P < 0.0001）。聚集与CD62P表达之间存在线性关系，以百分比表示（P < 0.0001）或相对荧光强度（RFI）表示（P < 0.0001）。指数方程将聚集与PAC-1表达相关联，以百分比表示（P < 0.0001）或RFI表示（P < 0.0001）。在患者术前（P = 0.0022）和术后（P = 0.0020），聚集与CD62P表达（以百分比表示）之间存在线性关系。术前（P = 0.0012）和术后（P = 0.0024），聚集与PAC-1表达（以百分比表示）之间存在指数关系。基于CD62P表达预测的聚集反应与氯吡格雷治疗后测量的聚集抑制之间存在线性相关性（P = 0.0013），以及预测的聚集与PRI抑制之间存在线性相关性（P = 0.0031）。