摘要
目的研究急性呼吸窘迫综合征（ARDS）气道上皮细胞屏障功能改变及其p38丝裂素活化蛋白激酶（p38MAPK）的调节作用。方法原代人类气道上皮细胞在气-液界面培养成熟后，分别与标准气道上皮细胞培养液、1μg／mL脂多糖（LPS）、1μg／mLLPS＋10nmol／Lp38MAPK阻断剂SB203580共同培养，于3、6、24和48h收获上述3组细胞。测量每个时间点的细胞存活率、经气道上皮电阻（TER）、上皮大分子通透性（荧光标记牛血清蛋白的荧光强度）和蛋白质组学变化。结果与对照组比较，LPS组气道上皮细胞存活率显著下降[24h：（62．5±12．0）％比（85．0±15．0）％，T=-5．681，P=0．001；48h：（67．5±17．0）％比（79．0±20．0）％，T=-4．260，p=0．001]，TER（Ω／cm^2）显著降低（3h：307±108比376±60，T=-3．606，P=0．049；48h：290．4-86比371±43，T=6．971，P=0．037）；上皮细胞大分子通透性明显增高（48h：（122．2±22．0）％比（100．0±18．0）％，T=3．182，p=0．036]，p38MAPK磷酸化明显增加（相对强度：0．34±0．16比0．28±0．10，T=4．303，p=0．029）。p38MAPK阻断剂SB203580可抑制由LPS导致的气道上皮细胞存活率下降[24h：（82．5±22．0）％比（62．5±25．0）％，T=-6．124，P=0．010；48h：（79．5±28．0）％比（67．5±16．0）％，T=-3．860，P=0．039]和LPS导致的TER（Ω／cm^2）降低（48h：411±118比290±97，T=3．560，P=0．022）；此外，SB203580还能明显抑制LPS诱导的p38MAPK磷酸化增加（相对强度：0．04±0．01比0134±0．16，T=3．840，P=0．011）。结论ARDS时气道上皮细胞数量减少和屏障功能障碍，p38MAPK可能介导LPS致ARDS的上皮细胞屏障功能改变。
Objective To observe airway epithelial barrier dysfunction in acute respiratory distress syndrome （ARDS）, and the possible regulatory role of p38 mitogen-activated protein kinase （p38MAPK）. Methods The primary human trachea-bronchial epifhelial cells were incubated either in 1 μg/mL lipopolysaccharide （LPS）, 1 μg/mL LPS ＋ 10 nmol/L p38MAPK inhibitor （SB203580）, or in control medium. Cells were harvested at 3, 6, 24 and 48 hours after incubation. The cell count, trans-epithelial electrical resistance （TER）, macromoleeular permeability （fluorescence staining）, and proteomics were measured at different time points. Results Compared with control group, the amount of airway epithelial cells in LPS group was significantly lowered at 24 hours and 48 hours [ 24 hours： （62.5 ± 12.0）% vs. （85.0 ± 15.0）%, T=-5.681, P=0.001 ; 48 hours： （67.5 ± 17.0）% vs. （79.0 ± 20.0）%, T=-4.260, P=0.001 ], TER was also significantly decreased at 3 hours and 48 hours （Ω/cm^2, 3 hours： 307 ± 108 vs. 376 ± 60, T=3.606, P= 0.049 ; 48 hours ： 290 ± 86 vs. 371 ± 43, T= 6.971, P=0.037 ）, and the macromoleeular permeability was shown to be increased in LPS group [ 48 hours ： （ 122.2 ± 22.0 ） % vs. （ 100.0 ± 18.0 ） %, T= 3.182, P= 0.036 ] ; and phosphorylation-p38MAPK （p-p38MAPK） was increased （relative intensity ： 0.34 ± 0.16 vs. 0.28 ± 0.10, T=4.303, P=0.029）. However, SB203580 attenuated the damages induced by LPS both in the amount of epithelial cells [24hours： （82.5±22.0）%vs. （62.5±25.0）%, T=-6.124, P=0.010; 48hours： （79.5±28.0）%vs. （67.5± 16.0）%, T=-3.860, P=0.0391, and TER of epithelial cells （Ω/cm^2, 48 hours： 411 ± 118vs. 290±97, T=3.560, P=0.022） . In addition, the down-regulation of expression of p-p38MAPK as induced by LPS was significantly alleviated by SB203580 （relative intensity： 0.04 ±0.01 vs. 0.34 ±0.16, T=3.840, P=0.011）. Conclusion LPS induced airway epithelial barrier dysfunction in ARDS, and p38MAPK may be involved in the oathwav of LPS induced airway eoithelial barrier dysfunction.
出处
《中华危重病急救医学》
CAS
CSCD
北大核心
2013年第10期589-593,共5页
Chinese Critical Care Medicine
基金
教育部留学回国启动（168006）
广东省自然科学基金（06021341）
广东省科技计划项目（20108031600314）
