预防性应用乌司他丁对健康大鼠呼吸机相关性肺损伤的保

1、预防性应用乌司他丁对健康大鼠呼吸机相关性肺损伤的保护徐州医学院江苏省麻醉学重点实验室 221002 肖刚 赵文静目的：机械通气是ICU中治疗危重病人的重要手段之一，有研究表明不恰当的通气会诱发机械通气性相关肺损伤(ventilator-induced lung injury，VILI)。本实验通过研究VILI的炎症反应机制，观察不同剂量乌司他丁（ulinastatin，Uti）对VILI的预防作用。方法：40只健康SD大鼠随机分为3 组：自主呼吸组（C）、通气损伤组（M）和应用乌司他丁组（U）；其中U组按给药剂量又分为三个亚组：U1(1 万单位/kg)、U5（5万单位/kg）、U10(10 万

2、单位/kg)。分组后大鼠称重，腹腔注射10%水合氯醛（1ml/300g）麻醉，无菌行气管切开和股动静脉置管，术后静置维持30min，测定血气指标定为基础值。除C组外，各组分别于机械通气前10min给予等容积的生理盐水和Uti,然后静脉注射肌松药并连接小动物呼吸机进行机械通气(参数设定为：潮气量=30ml/kg、频率40次/分钟、PEEP0),机械通气时间设为4h。通气中每隔1h从股动脉抽血监测动脉血气用于评估通气功能。实验结束后分别收集动脉血、支气管肺泡灌洗液（BALF）和肺组织标本。取出BALF经4，3000r/10min离心后，用考马斯亮蓝法测量上清液中总蛋白含量，沉渣用生理盐水重悬后行瑞

3、氏染色计数炎性细胞总数和分类；用ELISA 法测量BALF和血清中TNF-a水平；留取的肺组织一部分用于测量肺组织中髓过氧化物酶(MPO)活性和湿/干重比（W/D），另一部分制成石蜡切片，HE染色后光镜下观察肺组织的病理学改变。结果：M组大鼠通气过程中PaO2逐渐下降，通气4h后同基础值比较有明显降低（73.24.1mmHg 对99.35.2mmHg，p0.05）。光镜下发现M组肺组织中肺泡间隔断裂，大量的炎性细胞浸润，透明膜形成。同C组相比，M组肺组织W/D比值和MPO活性都明显增加（分别为4.880.17;1.350.11,p均0.01），而在BALF中白细胞总数和蛋白含量明显升高（分别是

4、7.42.2109/L对2.71.7109/L；79.318.4 对 50.715.6, p均0.01）；用ELISA测量BALF和血清中TNF-a升高程度也具有统计学意义（分别为431.786.6 对174.946.0；91.219.1 对50.917.4,p0.05），U5和U10则能明显抑制肺的损伤程度，指标改善与MV组都有显著性差异（P0.05）。结论：目前的研究认为VILI并不仅仅是肺泡结构的物理性损伤，还应该包括微血管和细胞参与的炎症反应性损伤。本实验通过大潮气量通气成功模拟了肺损伤，发现大潮气量通气可以明显破坏肺组织结构，增强肺血管的通透性，诱导大量中性粒细胞在肺泡腔聚集和活化，

5、并且明显上调BALF和血清中炎性介质的水平，进一步证实机械通气可以通过炎症反应的激活和放大，造成肺组织的损伤加重，甚至可以诱发全身的炎症反应。Uti作为一种蛋白酶抑制剂可以通过对炎性反应的抑制有效预防VILI的产生,且预防的效果呈剂量依赖性。【关键词】机械通气 呼吸机诱导肺损伤 乌司他丁 支气管肺泡灌洗液 【Abstact】 Objection: Mechanical ventilation(MV) with inappropriate ventilator setting been shown to initiate ventilator-induced lung injury(VILI).

6、 We examined the mechanism that this procedure induces lung injury with inflammatory features and explore the protect role against VILI in healthy rat by pretreated with different doses of ulinastatin(Uti). Method: Forty healthy SD rats were randomly assigned to three groups：control group(C),injurou

7、s ventilation group(M),pretreated with Uti group(U)；U group also was divided into three sub-groups according to doses:U1(10,000u/kg),U5(50,000u/kg),U10(100,000u/kg).The last four groups were ventilated for 4h with same strategy: VT=30 ml/kg,RR=40time/kg,I/E=3:1,Fio2=21%,PEEP=0.In addition, the blood

8、 gas analysis was monitored to value the state of the rat in the every hour of the ventilation.After the experiment ending,we observed the damage degree of the lung tissue under light microscope,and the chang of pulmonary permeability.Moreover,the myloperoxidase(MPO) activities in the lung tissue an

9、d the level of TNF-a in the serum and BALF were measured respectively.Result:our study demonstrates that injurious ventilation might cause maked pulmonary tissue damage,enhanced pulmonary peameability and induced neutrophil recruitment into the alveolar space and formed pulmonary edema.Po2 in the M

10、group were significantly reduced compared with baseline value (p0.05).Furthermore,compared with C group,the MPO activity in BALF ,the concentrations of TNF-a in the serum and BALF in MV group were all significantly increase (p0.01).In contrast, pretreatment with Uti could lessened lung injury from h

11、igh tidal volume ventilation in different degrees.Of all above indexes ,there were no statistical differences between the U1 group and the M group,but there were significantly differences in the U5 and U10 group compared with M group(p0.05). Conclusion:Mechanical ventilation with high tidal volume could cause injures to the normal lung tissue which was closed related to the recruitment and activation of neutrophil ,and releasing of a number of inflammatory mediators.Uti could attenuate the lung inj