内毒素血症犬部分去除白细胞后肺内凝血纤溶功能的变化

Changes in coagulation-fibrinolysis function in alveolar lavage fluid of endotoxemic dogs after partial removal of peripheral leukocytes

目的观察部分去除外周血白细胞后内毒素血症犬肺内凝血功能的变化,探讨活化白细胞对内毒素血症犬肺损伤的影响及其机制。方法雄性杂种犬30只,采用随机数字表法分为内毒素组（L组）、假白细胞去除组（S组）及白细胞去除处理组（T组）,每组10只。L组仅经静脉输注2mg/kg LPS建立内毒素血症犬模型,不行白细胞去除;T组在建模后12~14h使用自动连续血流血细胞分离机进行白细胞去除;S组建模后12~14h行假白细胞去除（将去除的终产物回输入体内）。建模后36h切取肺组织,采用ELISA法检测肺泡灌洗液中中性粒细胞弹性蛋白酶（NE）、可溶性血栓调节蛋白（sTM）、活化蛋白C（APC）、纤溶酶原激活物抑制物-1（PAI-1）水平,免疫组化染色观察肺组织血栓调节蛋白（TM）的表达,并行肺湿/干重检测及肺组织病理学观察。结果与L组及S组比较,T组犬肺泡灌洗液中APC水平显著增高,NEs、TM、PAI-1水平及肺湿/干重比值显著降低（P〈0.05）,肺组织内TM表达量也明显增加,而L组与S组各指标比较无显著差异（P〉0.05）。病理学观察显示,T组急性肺损伤发生率（2/10）显著低于L组（7/10）及S组（8/10,P〈0.05）。结论部分去除白细胞可降低肺泡灌洗液中的NE水平,阻止肺内TM表达进一步下降,从而改善肺泡灌洗液中凝血纤溶异常,降低内毒素犬急性肺损伤的发生率。

Objective To observe the effect of partial removal of peripheral leucocytes on the coagulation-fibrinolysis function of alveolar lavage fluid（ALF） in endotoxemic dogs,and explore the influence and mechanisms of activated leucocytes on lung injury in endotoxemic dogs.Methods Thirty male mongrel dogs were involved in present study and randomly divided into 3 groups（10 each）： LPS group（group L）,sham leukocytapheresis group（group S） and leukocytapheresis group（group T）.Endotoxemic model was reproduced in group L by administration of LPS（2mg/kg）,but the animals did not receive leukocytapheresis.Animals in group T received leukocytapheresis using a continuous-flow blood cell separator 12-14 hours after administration of LPS.Animals in group S received sham leukocytapheresis（the end products were transfused back into the dogs） at 12-14 hours after administration of LPS.At 36h after administration of LPS,the lung tissues were harvested to obtain ALF,and the levels of neutrophil elastase（NE）,soluble thrombomodulin（sTM）,activated protein C（APC） and plasminogen activator inhibitor-1（PAI-1） in ALF were determined,the expression of thrombomodulin in lung tissue was observed by immunohistochemical staining,while the routine pathological examination and wet/dry ratio of lung tissue were performed.Results The APC level in ALF was significantly higher,while the NE,sTM and PAI-1 levels in ALF and wet/dry ratio of lung tissue were significantly lower in group T than in group L and group S（P0.05）.Immunohistochemical examination revealed that the expression of thrombomodulin in lung tissue was higher in group T than in group L and group S.No significant difference was found between group L and group S in the indexes mentioned above.Pathological observation showed the incidence of acute lung injury was significantly lower in group T（2/10） than in group L（7/10） and group S（8/10,P0.05）.Conclusion Partial removal of peripheral leukocytes may lower the level of NE in ALF,prevent the lowering of sTM expression in lung tissue,thus improve the coagulation-fibrinolysis balance in ALF,and reduce the incidence of acute lung injury in dogs with endotoxemia.