Segura Dídac, Monreal Luis, Pérez-Pujol Sílvia, Pino Marcos, Ordinas Antonio, Brugués Rosa, White James G, Escolar Ginés
Departament de Medicina i Cirurgia Animals, Facultat de Veterinària, Universitat Autònoma de Barcelona, Bellaterra, Spain.
J Vet Intern Med. 2006 May-Jun;20(3):581-8. doi: 10.1892/0891-6640(2006)20[581:aopfih]2.0.co;2.
We studied equine platelet function and activation using ultrastructural examination, flow cytometry, and perfusion. The main aim of the study was to evaluate hemostatic mechanisms in horses using these techniques. Ultrastructural observations were done on resting and activated platelets. Flow cytometry was used to evaluate binding of antibodies to major platelet glycoproteins (GPIIb-IIIa, GPIV, and GPIb) and activation-dependent antigens (P-selectin and lysosomal integral membrane protein [LIMP]). Perfusion techniques were used to evaluate the interaction between platelets and damaged subendothelium. Aggregation experiments were done to identify the best agonists for flow cytometry. Ultrastructural observations confirmed that equine platelets lack a developed open canalicular system and that release of granule contents occurs by fusion of adjacent granule membranes that ultimately connect with external membranes. Flow cytometry identified a 2-fold increase in binding of antibodies against GPIIb-IIIa and GPIV after activation. Binding of antibodies against P-selectin and LIMP increased from 2.12 and 1.74% to 15.5 and 11.6%, respectively, in response to thrombin and to 21.86 and 10.50%, respectively, in response to collagen. Annexin V binding increased moderately after activation. Perfusion experiments with citrated blood indicated that equine platelets react more strongly to subendothelium than do human platelets. When blood was anticoagulated with low molecular weight heparin, a marked impairment of platelet interactions was observed. In conclusion, although some differences were observed between human and equine platelet function, some techniques currently used to assess human platelet function may be useful to assess equine platelets.
我们使用超微结构检查、流式细胞术和灌注技术研究了马血小板的功能和活化情况。本研究的主要目的是利用这些技术评估马的止血机制。对静息和活化的血小板进行了超微结构观察。流式细胞术用于评估抗体与主要血小板糖蛋白（GPIIb-IIIa、GPIV和GPIb）以及活化依赖性抗原（P-选择素和溶酶体整合膜蛋白[LIMP]）的结合情况。灌注技术用于评估血小板与受损内皮下层之间的相互作用。进行聚集实验以确定用于流式细胞术的最佳激动剂。超微结构观察证实，马血小板缺乏发达的开放小管系统，颗粒内容物的释放是通过相邻颗粒膜的融合发生的，这些颗粒膜最终与外部膜相连。流式细胞术显示，活化后针对GPIIb-IIIa和GPIV的抗体结合增加了2倍。针对P-选择素和LIMP的抗体结合在凝血酶作用下分别从2.12%和1.74%增加到15.5%和11.6%，在胶原作用下分别增加到21.86%和10.50%。活化后膜联蛋白V结合适度增加。用枸橼酸盐血进行的灌注实验表明，马血小板对内皮下层的反应比人血小板更强。当用低分子量肝素抗凝血液时，观察到血小板相互作用明显受损。总之，虽然在人和马血小板功能之间观察到了一些差异，但目前一些用于评估人血小板功能的技术可能有助于评估马血小板。
