布雷菲德菌素A在脂多糖诱导急性肺损伤中的作用

Effect of Brefeldin A in acute lung injury induced by lipopolysaccharide

目的探讨布雷菲德菌素A(Brefeldin A)在脂多糖(LPS)诱导急性肺损伤中的作用。方法小鼠肺泡巨噬细胞(MH-S)和上皮细胞(MLE-12)分别给予浓度为1、10、100μM的Brefeldin A后,立即用LPS 500 ng/ml处理,收集3、6、9、24 h的细胞上清并测定MH-S中的肿瘤坏死因子-α(TNF-α)含量和MLE-12中的趋化因子KC值;ICR小鼠随机分为生理盐水组(Normal组)、模型组(LPS组)、地塞米松组(Dex组,5 mg/kg)、Brefeldin A组(BFA组,10 mg/kg),每组12只,气道内2 mg/kg滴入LPS制备急性肺损伤模型,生理盐水组给予等体积生理盐水。6 h后观察肺组织病理改变,测定肺泡灌洗液(BALF)中白细胞、白蛋白含量和TNF-α、白介素-1β(IL-1β)、白介素-6(IL-6)等炎症因子含量,检测肺组织中髓过氧化物酶(MPO)、c AMP含量和MAPK信号通路中ERK、p38和JNK等蛋白激酶分子磷酸化水平的变化。结果 100μM的Brefeldin A能显著减少MH-S中TNF-α的释放和MLE-12细胞中KC的产生(P<0.001)。Brefeldin A显著改善肺组织病理变化,降低BALF中白细胞(P<0.001)和TNF-α(P<0.05)含量,对BALF中白蛋白、IL-1β和IL-6无显著影响,显著降低小鼠肺组织中MPO活性(P<0.05),升高c AMP的水平(P<0.001),同时能显著抑制ERK的磷酸化(P<0.05)。结论 Brefeldin A对急性肺损伤可产生保护作用,其机制与抑制相关炎症因子释放、升高细胞内c AMP的含量、抑制ERK磷酸化等途径有关。

Objective To explore the effect of Brefeldin A in acute lung injury induced by lipopolysaccharide. Methods Mice alveolar macrophages （MH-S） and epithelial cells （MLE-12） were treated with Brefeldin A of 1, 10, 100 μM respec- tively, and then treated with LPS 500 ng/ml. The cell supernatants of 3, 6, 9 and 24 h were collected and the content of tumor necrosis faetor-α （TNF-α） in MH-S and the chemokine KC in MLE-12 were determined. ICR mice were randomly divided into normal saline control group （group Normal）, model group （group LPS）, dexamethasone group （group Dex, 5 mg/kg） and brefeldin A group （group BFA, 10 mg/kg）. Each group had 12 mice. The ALI mouse model was induced by instilling intratraeheally LPS 2 mg/kg. The physiological saline group was given equal volume of normal saline. After 6 h, lung tissue and alveolar lavage fluid （BALF） were harvested, lung pathology changes were observed, white blood cell and albumin content and tumor necrosis factor α（ TNF-α）, interleukin-1 β （ IL-1 β）, interleukin 6 （IL-6） in BALF were deter- mined, myeloperoxidase （MPO） activity, cAMP content and the changes of phosphorylation levels of protein kinases of ERK, p38 and JNK in MAPK signaling pathway in lung homogenates were detected by ELISA. Results BFA significantly reduced the release of TNF-α in MH-S and the production of KC in MLE-12 cells （ P 〈 0. 001 ）. BFA could significantly improve the pathological changes of lung tissue and decrease the content of white blood cells（P 〈0.001 ） and TNF-α con- tent （ P 〈0. 05）. However, there were no significant effects on albuufin, IL-1 β and IL-6 in BALF, and the activity of MPO in lung tissue was significantly decreased （ P 〈 0.05 ）, the level of cAMP was significantly increased （ P 〈 0.001 ）. Brefel- din A could significantly inhibit the phosphorylation of ERK （ P 〈 0. 05）. Conclusion Brefeldin A may have a protective effect on acute lung injury, and its mechanism may be related to inhibiting the release of related inflammatory factors, in- creasing intracellular cAMP content and inhibiting the phosphorylation of ERK.