Therapeutic effects of low molecular weight heparin and aspirin on the murine acute lung injury LUAN Zheng-gang, NARA Pulu, ZHANG Zhi-dan, MA Xiao-chun. *Department of ICU, The First Affiliated Hospital of China Medical University，Shenyang 110001，Liaoning, China.
Abstract

Objective To investigate the possible effects of nuclear factor-κB (NF-κB) activation on the expression of intercellular adhesion molecule-1 (ICAM-1) and P-selectin in murine acute lung injury (ALI) and assess the potential beneficial effects and mechanism of low molecular weight heparin (LMWH) and aspirin (ASA).

Methods Rat ALI model was established by injection of lipopolysaccharide into tail veinous. Rats were divided randomly into four groups: Control group, ALI group, LMWH group and ASA group. The change of NF-κB activity in the lung to show its relation with (ICAM-1) and P-selectin were observed by ABC immunohistochemistry.

Results Compared with control group, the activity of NF-κB and the expression of ICAM-1 and P-selectin increased significantly in the lung tissue of ALI group. Compared with ALI group , in LMWH and ASA administration groups the level of NF-κB activity and the expression of ICAM-1 and P-selectin were obviously down regulated and The pathological lesion and inflammatory response of lung were improved in LMWH and ASA groups. However the therapeutic effects of ASA were bigger than that of LMWH. all P<0.05.

Conclusion NF-κB activation in ALI plays a important role. NF-κB takes part in the activation of many kinds of inflammatory cells such as neutrophil, endothelial cells and so on and adjusts the genetic expression of ICAM-1 and P-selectin. LMWH and ASA showed their beneficial effects on lung injury in both functional and morphologic alterations. But the mechanism is different. LMWH can indirectly inhibit the activation of NF-κB. LMWH can improve lung microcirculation and decrease the adhesion of neutrophil and platelet. Regulatory roles of ASA could be considered as the inhibitor of NF-κB activation.
[Key Words] lung injury; nuclear factor-kappaB; low molecular weight heparin; adhesion molecule

       目前研究表明，机体遭受各种严重损伤可引起全身炎症反应综合征而导致急性肺损伤（ALI），其中由感染导致的全身炎症反应综合征是引起急性肺损伤的最常见的病因。我们通过急性肺损伤模型观察NF-κB活化在ALI中的作用及其对粘附分子ICAM-1、P-selectin表达的影响；观察低分子肝素（LMWH）和阿司匹林（ASA）对肺损伤的保护作用并探讨其机制。
1.材料和方法
1.1实验动物和主要试剂：健康雄性Wistar大鼠60只，体重200－250g（由中国医科大学动物中心提供）。LPS、ASA购自美国Sigma公司，LMWH购自法国赛诺菲－温莎公司。NF-κB（P65）和ICAM-1、P-selectin一抗、免疫组化试剂盒购自武汉博士德生物工程有限公司。髓过氧化物酶（MPO）测定试剂盒、血清丙二醛（MDA）测定试剂盒购自南京建成生物工程研究所。
1.2实验方法：（1）动物分组与ALI模型制备：将大鼠60只随机分成四组，对照组（n=15），ALI组（n=15），LMWH组（n＝15），ASA组（n＝15）。先用75％酒精洗鼠尾，再涂适量二甲苯，待尾静脉充盈可辨，即从尾静脉注射LPS 5mg/kg制备ALI模型，对照组尾静脉注入等量生理盐水。LMWH组于建模后，立即皮下注射低分子肝素20U/20g，对照组、ALI组、ASA组皮下注射等量生理盐水；ASA组于建模后，立即腹腔注射ASA 100mg/kg，对照组、ALI组、LMWH组腹腔注入等量生理盐水。（2）观察指标：模型成功后6小时，取材待测。PaO2测定：血气分析仪进行测定。血清MDA测定：分光光度法。肺组织学观察；肺组织MPO活力测定：比色法。肺组织NF-κB与ICAM-1、P-selectin活性检测：石蜡切片，常规处理。按试剂盒说明行ABC染色，一抗用兔抗大鼠NF-κB P65抗体，DAB显色。NF-κB活化后在单核巨噬细胞、中性粒细胞、内皮细胞核染有黄褐色颗粒者定为阳性；ICAM-1、P-selectin主要在血管内皮细胞和支气管上皮细胞膜染有黄褐色颗粒者定为阳性。图象分析以积分光密度值表示NF-κB和ICAM-1、P-selectin表达强度。
1.3统计学处理：数据以均数±标准差（ ±s ）表示，Spss10.0统计分析软件对数据进行处理，p<0.05认为差异有显著统计学意义。