小窝蛋白－1通过 p38 MAPK 信号链通路对大鼠机械通气所致肺损伤的保护机制研究

The protective effect of caveolin-1 on lung injury in rats ventilated through p38 MAPK signaling pathway

目的：研究大鼠肺组织在不同潮气量机械通气的情况下小窝蛋白－1（ Cav－1）及p38 MAPK表达的变化。方法56只雄性健康成年SD大鼠，体质量220-330 g，随机均分为7组，自主呼吸对照组（ A组）：仅做气管切开；不同通气时间保护性机械通气组（ B1组和B2组）：潮气量（ VT ）为6 mL／kg，呼吸频率（ RR）为60次／min，B1组通气1 h，B2组通气2 h；不同通气时间大潮气量组（ C1组和C2组）；不同时间大潮气量＋SB203580组（ D1组和D2组）。 C和D组VT为30 mL／kg，RR为40次／min，C1和D1组通气1 h，C2和D2组通气2 h。机械通气前30 min D组大鼠给予注射SB203580溶液，实验中7组大鼠均给予动脉血气和血流动力学监测。 A组行气管切开即刻处死，剩余各组大鼠于预定时间机械通气结束时处死，光镜下观察肺病理改变，免疫组织化学染色法检测肺组织中Cav －1、p38 MAPK及AP－1蛋白的表达，测定Cav －1、MKK3、p38 MAPK mRNA表达，测定髓过氧化物酶（MPO）活性及肺湿干质量比值（W／D），ELISA法测定支气管肺泡灌洗液（ BALF）中总蛋白及IL－6含量。结果与A组比较，C组Cav－1、p38 MAPK蛋白及Cav－1、p38 MAPK、MKK3 mRNA表达上调，W／D比值、MPO、总蛋白浓度、IL－6含量升高，蛋白激酶（AP）－1蛋白表达下降（P＜0．05）。与B组比较，C组Cav－1、p38 MAPK蛋白及p38 MAPK、Cav－1、MKK3 mRNA表达上调，W／D比值、MPO、总蛋白浓度、IL－6含量升高，AP－1蛋白表达水平下降（ P＜0畅05）。与C组比较，D组p38 MAPK蛋白及p38 MAPK、MKK3 mRNA表达下降，Cav－1、AP－1蛋白及Cav－1 mRNA表达上升，W／D比值、MPO、总蛋白浓度、IL－6含量升高（P＜0．05）。结论 Cav－1及其p38 MAPK信号链在机械通气中表达的上调，参与了机械通气相关性肺损伤的保护作用。

Objective To investigate the expression of caveolin -1（Cav -1） under different ventilator conditions and to explain clearly the relationship between the expressions of Cav -1 and the activation of p38 MAPK signal in lung tissue in stress injury model .Methods Fifty-six healthy male SD rats （weighting 220-330 g） were randomly assigned into 7 groups according to the tidal volume （VT）, respiratory rate （RR） and duration of mechanical ventilation.They were spontaneous breathing control group （group A） which was only done tracheotomy;protective mechanical ventilation group （group B1 and B2） with VT of 6 mL/kg and RR of 60 bpm, and B1 and B2 group ventilation duration was 1 h and 2 h, respectively;Large-tidal volume ventilation group （ group C1, C2, D1, D2, VT 30 mL/kg, RR 40 bpm, ventilation duration 1 h, 2 h）, but only rat in group D1 and D2 were intraperitoneally injected with SB203580, a specific inhibitor of p38 MAPK.Hemodynamic parameters and arterial blood gases of all the rats in the 7 groups were measured throughout the study period .After maintaining ventilation for 1 h or 2 h, all rats were sacrificed and specimens of lung tissues and bronchoalveolar lavage fluid （BALF） were harvested. Lung pathological changes were observed by microscopy .The protein levels of Cav -1, p38 MAPK, and AP-1 in lung tissues were assayed with immunohistochemistry staining .The expression levels of Cav-1, MKK3, p38 MAPK mRNA in lung tissue were measured by reverse transcription -polymerase chain reaction （RT-PCR）.Lung myeloperoxidase （MPO） activity and wet-dry weight ratio （W/D） were measured.The levels of total protein and interleukin-6 （IL-6） in BALF were measured by enzyme-link immunosorbent assay （ELISA）.Results Compared with normal control group A, the expression of p38 MAPK, MKK3, Cav-1 mRNA and Cav -1, p38 MAPK protein was significantly up -regulated; the W/D ratio, MPO, concentration of IL -6 in BALF and total protein increased in group C; however, AP -1 protein was significantly down-regulated.The expression of p38 MAPK, MKK3, Cav-1 mRNA and Cav -1, p38 MAPK protein were significantly higher in group C than those in group of B; the W/D ratio, MPO, concentration of IL-6 in BALF and total protein were increased, but AP-1 protein was significantly lower in group C compared with group B.Compared with group C, expression of p38 MAPK, MKK3 mRNA and p38 MAPK protein was significantly lower in group D while Cav-1 mRNA, Cav-1 and AP-1 protein.W/D ratio, MPO, concentration of IL-6 in BALF and total protein were increased.Conclusion Cav-1 and the activation of p38 MAPK signal in lung tissue participate in the protective effects of the ventilator -induced lung injury.