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Phospholipase A2 Activates Hemostasis

机译：磷脂酶A2激活止血

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Background: Phospholipases A2 (PLA2) are aggressive enzymes that can destroy phospholipids of cell membranes. The resulting cell fragments trigger the kallikrein—mediated contact phase of coagulation. The aim of the present study was to expose citrated whole blood to PLA2 and to quantify thrombin generation in recalcified plasma.Methods: Normal citrated blood was exposed to bovine pancreatic or snake PLA2, lipopolysaccharide (LPS), or zymosan A for 30–45 min (RT). After centrifugation the plasma samples were recalcified (10 + 1) with 250 mM CaCl2 in the recalcified coagulation activity assay (RECA). After 0–45 min coagulation reaction time (CRT at 37°C) 1.6 M arginine (final test concentration) was added to stop hemostasis activation and to depolymerize non-crosslinked fibrin. The generated thrombin activity was chromogenically determined.Results: 100 ng/ml bovine pancreatic or snake PLA2 generates about 0.2–0.8 IU/ml thrombin after 15 min CRT. This thrombin generation is similar as that induced by 200 ng/ml LPS or 20 μg/ml zymosan A. Up to 60 ng/ml bovine pancreatic PLA2 the generated thrombin activity is proportional to the PLA2 activity used; 1 μg/ml PLA2 induces much less thrombin, but PLA2 at 10 μg/ml again results into thrombin generation of 0.1–3 IU/ml at 10–15 min CRT. As control, in pooled normal citrated plasma there is no significant change in thrombin generation when exposed to up to 10 μg/ml bovine pancreatic PLA2.Discussion: Elevated plasmatic PLA2 activities (occurring e.g. in trauma, pancreatitis, or sepsis) activate the blood hemostasis system resulting in pathologic disseminated intravascular coagulation (PDIC). It is suggested to diagnose these life threatening states as early as possible, screening all patients for plasmatic thrombin activity.Abbreviations: ΔA/t, absorbance increase per time; CRT, coagulation reaction time (37°C); CS-IIa, chromogenic substrate HD-cyclohexylglycyl-alanyl-arginyl-paranitroanilide for thrombin; F-well, fl at bottom well Polysorp?; IU, international units; LPS, lipopolysaccharide; mA, milli absorbance units; PDIC, pathologic disseminated intravascular coagulation; PLA2, phospholipase A2; RECA, recalcified coagulation activity assay; RT, room temperature; sPLA2, secretory PLA2; U, units; ZyA, zymosan A.
机译：背景：磷脂酶A2（PLA2）是攻击性酶，可以破坏细胞膜的磷脂。产生的细胞片段触发激肽释放酶介导的凝结接触阶段。本研究的目的是将柠檬酸全血暴露于PLA2并定量重新钙化血浆中的凝血酶生成。方法：正常柠檬酸血暴露于牛胰或蛇PLA2，脂多糖（LPS）或酵母聚糖A 30-45分钟。 （RT）。离心后，在重新钙化的凝血活性测定（RECA）中用250 mM CaCl2将钙样品重新钙化（10 +1）。在0–45分钟的凝固反应时间（37°C的CRT）后，添加1.6 M的精氨酸（最终测试浓度）以止血止血并解聚未交联的血纤蛋白。结果：100ng / ml牛胰腺或蛇PLA2在CRT 15分钟后产生约0.2–0.8 IU / ml凝血酶。凝血酶的产生与200 ng / ml LPS或20μg/ ml酵母聚糖A诱导的相似。高达60 ng / ml的牛胰腺PLA2产生的凝血酶活性与所用的PLA2活性成正比。 1μg/ ml的PLA2诱导的凝血酶要少得多，但是10μg/ ml的PLA2在CRT 10-15分钟时又会产生0.1–3 IU / ml的凝血酶。作为对照，在合并的正常柠檬酸血浆中，当暴露于高达10μg/ ml的牛胰腺PLA2中时，凝血酶的生成没有显着变化。讨论：血浆PLA2活性升高（例如在创伤，胰腺炎或败血症中发生）可激活血液止血系统导致病理性弥散性血管内凝血（PDIC）。建议尽早诊断这些威胁生命的状态，筛查所有患者的血浆凝血酶活性。缩写：ΔA/ t，每次吸光度增加; CRT，凝固反应时间（37°C）; CS-IIa，用于凝血酶的生色底物HD-环己基甘氨酰基-丙氨酰基-精氨酰基-对硝基苯胺; F井，fl井底是Polysorp？ IU，国际单位; LPS，脂多糖; mA，毫吸光度单位; PDIC，病理性弥散性血管内凝血; PLA2，磷脂酶A2; RECA，重钙化凝血活性测定;室温，室温; sPLA2，分泌型PLA2; U，单位; ZyA，酵母聚糖A。

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