多巴酚丁胺对脓毒症休克兔急性肺损伤的影响

Effects of dobutamine on acute lung injury in rabbits of septic shock

目的探讨不同剂量多巴酚丁胺对脓毒症休克兔急性肺损伤（acute lung injury，ALI）的保护作用及其可能机制。方法采用盲肠结扎穿孔联合静脉注射内毒素制备脓毒症休克模型，将70只新西兰雄性大白兔随机（随机数字法）均分为假手术组（A组）、Au模型组（B组）、多巴酚丁胺低剂量组（C组）、多巴酚丁胺中剂量组（D组）和多巴酚丁胺高剂量组（E组），分别于造模后3h和6h点处死，留取肺组织标本。ELISA法检测环磷酸腺苷（cyclicadenosine monophosphate，cAMP）浓度；Western-blot法检测水通道蛋白5（Aquaporin5，AQP5）蛋白浓度；计算肺湿／干重（wettodryweight，W／D）比值；光镜和电镜下观察各组小鼠肺组织病理学改变，并对肺组织病理损伤进行评分。多组间差异比较采用单因素方差分析，组间两两比较采用LSD法。结果与A组相比，B组cAMP、AQP5蛋白浓度在3h及6h明显降低（3．53±0．43）pmol／mL vs．（21．18±0．62）pmol／mL；（0．44±0．04）pmol／mLvs．（0．99±0．06）pmol／mL；（2．71±0．56）pmol／mL vs．（21．78±0．62）pmol／mL；（0．29±0．05）pmol／mLvs．（0．91±0．06）pmol／mL；P〈0．001，W／D比值则明显升高P〈0．00。与B组相比，C组cAMP及AQP5蛋白水平在6h明显增加（8．48±0．61）pmol／mLvs．（2．71±0．56）pmol／mL，P〈0．001；（0．49±0．04）pmol／mLvs．（0．29±0．05）pmol／mL，（P=O．001），D、E组cAMP及AQP5蛋白水平在3h、6h明显增力口（10．86±0．66）pmol／mL vs．（3．53±0．43）pmol／mL；（0．60±0．05）pmol／mLvs．（0．44±0．04）pmol/mL；（13．80±0．49）pmol／mLvs．（2．71±0．56）pmol／mL；（0．64±0．03）pmol／mLvs．（0．29±O．05）pmol／mL；（15．57±O．60）pmol／mLvs．（3．53±0．43）pmol／mL；（0．91±0．05）pmol／mLvs．（0．44±O．04）pmol／mL；（19．30±0．42）pmol／mLvs．（2．71±0．56）pmol／mL；（0．89±0．08）pmol／mLvs．（0．29±0．05）pmol／mL；P〈0．01，E组W／D比值明显降低（P=0．002；P=0．001）。与C、D组相比，E组cAMP及AQP5水平在3h、6h均有所增加（P〈0．01）。光镜及电镜下，B组肺的病理形态及超微结构损伤均较重，肺病理学评分明显升高（P〈0．01）；多巴酚丁胺干预后，肺的病理形态及超微结构损伤有所减轻，肺组织病理学评分明显降低（P〈0．01）。结论多巴酚丁胺对内毒素诱导的急性肺损伤有一定程度保护作用，其机制可能与提高肺组织cAMP水平，上调AQP5蛋白表达量有关，且大剂量多巴酚丁胺效果更佳。

Objective To explore the effect of different doses of dobutamine on acute lung injury （ALI） in rabbits with septic shock and to clarify the possible mechanism. Methods The rabbits model of septic shock was made by cecal ligation and puncture combined with intravenous injection of endotoxin, 70 male New Zealand white rabbits were randomly divided into five groups （14 rabbits in each groups） ： sham operation group （group A）, ALI group （group B）, dobutamine low-dose group （group C）, dobutamine medium-dose group （group D） and dobutamine high-dose group （group E）, 7 rabbits from each group were sacrificed 3 h and 6 h after septic shock. The level of cyclic adenosine monophosphate （cAMP） in lung tissue was detected by ELISA . The expression of aquaporin 5 （AQPS） protein was determined by western blotting. The wet to dry weight （W/D） ratio was measured. The pathological and ultrastructural changes of lung tissue were evaluated by optical microscopy and electron microscope, and lung injury score was assessed. The differences among the different groups were analyzed by one-way ANOVA （LSD test ）. Results The level of cAMP and expression of AQP5 protein in lung tissue at 3 h and 6 h were dramatically lower in group B than those in group A （3.53 ±0.43） pmol/mLvs. （21.18 ±0.62） pmol/mL; （0.44 ± 0.04） pmol/mLvs. （0.99±0.06）pmol/mL; （2.71_±0.56）pmol/mLvs. （21.78±0.62） pmol/mL; （0. 29 ±0. 05） pmol/mLvs. （0. 91 ±0. 06） pmol/mL; all P 〈0. 001, while the W/D ratio was obviously higher in group B than those in group A （ all P 〈 O. 001 ） . Compared with group B, the level of cAMP and AQP5 protein expression in lung tissue were significantly increased at 6 h in group C （8. 48 ± 0. 61 ） pmol/ mLvs. （2.71 ±0.56） pmol/mL, P 〈0.01; （0.49±0.04） pmol/mLvs. （0.29 ±0.05） pmol/mL, P = 0. 001 and at 3 h and 6 h in group D and E （ 10. 86 ± 0. 66 ） pmol/mLvs. （ 3.53 ± 0. 43 ） pmol/mL ; （0.60±0.05） pmol/mLvs. （0.44±0.04） pmol/mL; （13.80±0.49） pmol/mLvs. （2.71 ±0.56） pmol/mL; （0.64 ±0.03） pmol/mLvs. （0.29 ±0.05） pmol/mL; （15.57 ±0.60） pmol/mL vs. （3.53±0.43） pmol/mL; （0.91±0.05） pmol/mLvs. （0.44 ±0.04） pmol/mL; （19.30±0.42） pmol/mL vs. （2. 71 ±0. 56） pmol/mL; （0. 89±0. 08） pmol/mL vs. （0. 29 ±0.05） pmol/mL; all P 〈 0.01, while the W/D ratio in group E was decreased obviously （ P = 0. 002 ; P = 0. 001 ）. Compared with group C and D, the level of cAMP and the expression of AQP5 protein at 3 h and 6 h in group E increased significantly （ all P 〈 0.01. The pathological and ultrastruetural changes of lung tissue were more intensive in group B than those in group A and the lung injury scores were obviously higher （ P 〈 O. 01 ）. The degree of lung pathological and ultrastrnctural lesion was mneliorated after administration of dobutanmine. Additionally, histological scores decreased significantly （P 〈 0.01 ）. Conclusions Our study demonstrated that dobutamine could improve ALl induced by endotoxin, the mechanism of protective effect may involve in increasing the level of cAMP and up-regulating the AQP5 protein expression, and high-dose dobutamine had better effects.