1、預防性應用烏司他丁對健康大鼠呼吸機相關性肺損傷的保護徐州醫(yī)學院江蘇省麻醉學重點實驗室 221002 肖剛 趙文靜目的：機械通氣是ICU中治療危重病人的重要手段之一，有研究表明不恰當?shù)耐鈺T發(fā)機械通氣性相關肺損傷(ventilator-induced lung injury，VILI)。本實驗通過研究VILI的炎癥反應機制，觀察不同劑量烏司他丁（ulinastatin，Uti）對VILI的預防作用。方法：40只健康SD大鼠隨機分為3 組：自主呼吸組（C）、通氣損傷組（M）和應用烏司他丁組（U）；其中U組按給藥劑量又分為三個亞組：U1(1 萬單位/kg)、U5（5萬單位/kg）、U10(10 萬

2、單位/kg)。分組后大鼠稱重，腹腔注射10%水合氯醛（1ml/300g）麻醉，無菌行氣管切開和股動靜脈置管，術后靜置維持30min，測定血氣指標定為基礎值。除C組外，各組分別于機械通氣前10min給予等容積的生理鹽水和Uti,然后靜脈注射肌松藥并連接小動物呼吸機進行機械通氣(參數(shù)設定為：潮氣量=30ml/kg、頻率40次/分鐘、PEEP0),機械通氣時間設為4h。通氣中每隔1h從股動脈抽血監(jiān)測動脈血氣用于評估通氣功能。實驗結(jié)束后分別收集動脈血、支氣管肺泡灌洗液（BALF）和肺組織標本。取出BALF經(jīng)4，3000r/10min離心后，用考馬斯亮藍法測量上清液中總蛋白含量，沉渣用生理鹽水重懸后行瑞

3、氏染色計數(shù)炎性細胞總數(shù)和分類；用ELISA 法測量BALF和血清中TNF-a水平；留取的肺組織一部分用于測量肺組織中髓過氧化物酶(MPO)活性和濕/干重比（W/D），另一部分制成石蠟切片，HE染色后光鏡下觀察肺組織的病理學改變。結(jié)果：M組大鼠通氣過程中PaO2逐漸下降，通氣4h后同基礎值比較有明顯降低（73.24.1mmHg 對99.35.2mmHg，p0.05）。光鏡下發(fā)現(xiàn)M組肺組織中肺泡間隔斷裂，大量的炎性細胞浸潤，透明膜形成。同C組相比，M組肺組織W/D比值和MPO活性都明顯增加（分別為4.880.17;1.350.11,p均0.01），而在BALF中白細胞總數(shù)和蛋白含量明顯升高（分別是

4、7.42.2109/L對2.71.7109/L；79.318.4 對 50.715.6, p均0.01）；用ELISA測量BALF和血清中TNF-a升高程度也具有統(tǒng)計學意義（分別為431.786.6 對174.946.0；91.219.1 對50.917.4,p0.05），U5和U10則能明顯抑制肺的損傷程度，指標改善與MV組都有顯著性差異（P0.05）。結(jié)論：目前的研究認為VILI并不僅僅是肺泡結(jié)構的物理性損傷，還應該包括微血管和細胞參與的炎癥反應性損傷。本實驗通過大潮氣量通氣成功模擬了肺損傷，發(fā)現(xiàn)大潮氣量通氣可以明顯破壞肺組織結(jié)構，增強肺血管的通透性，誘導大量中性粒細胞在肺泡腔聚集和活化，

5、并且明顯上調(diào)BALF和血清中炎性介質(zhì)的水平，進一步證實機械通氣可以通過炎癥反應的激活和放大，造成肺組織的損傷加重，甚至可以誘發(fā)全身的炎癥反應。Uti作為一種蛋白酶抑制劑可以通過對炎性反應的抑制有效預防VILI的產(chǎn)生,且預防的效果呈劑量依賴性。【關鍵詞】機械通氣 呼吸機誘導肺損傷 烏司他丁 支氣管肺泡灌洗液 【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