摘要
目的探讨蛋白酶激活受体1(PAR1)激动肽和凝血酶对人肺上皮细胞白细胞介素-8(IL-8)分泌的影响。方法人肺上皮细胞系A549细胞分别接种于12孔培养板各孔内,并分别用不同浓度的PAR1激动肽SFLLR和反PAR1激动肽RLLFS以及不同浓度的凝血酶和/或凝血酶抑制剂水蛭素进行刺激,刺激时间为2和16h。用ELISA方法检测上清液中的IL-8水平。结果经过16h的培养,SFLLR可引起浓度相关性IL-8的释放增加,增加到300μmol/L时诱导IL-8的释放量比基础分泌量增加了近16倍,RLLFS不能引起IL-8的释放增加。凝血酶也可引起浓度相关性IL-8释放,凝血酶在浓度1kU/L时就可引起IL-8释放量增加,10kU/L时诱导IL-8释放量达高峰,为基础分泌量的7.5倍。水蛭素可以抑制凝血酶对IL-8的释放作用。时间相关曲线表明,PAR1介导的IL-8释放从2h起即可引起增加,16h达高峰。结论PAR1激动肽和凝血酶可促进人肺上皮细胞分泌IL-8,PAR1拮抗剂和凝血酶抑制剂可能具有抗炎作用。
Objective To investigate the actions of protease-activated receptor 1 (PAR1) agonists and thrombin on the secretion ofintedeukin-8 (IL-8) from human lung epithelial cells. Methods A549 cells were cultured in a 12-well culture plate. The challenge was performed by addition of various concentrations of PAR1 agonist peptides SFLLR and its reverse peptides RLLFS, thrombin or hirudin, a thrombin inhibitor, into each well, respectively. After 2 or 16 h, the reactions were terminated by removal of the supernatant from each well, A sandwich enzyme-linked immunosorbent assay (ELISA) was used to determine the levels oflL-8 in the supernatants. Results Following a 16-hour incubation, SFLLR was able to induce concentration-dependent secretion oflL-8. The maximum release oflL-8 was increased nearly 16 fold more than the baseline release. The reverse PAR1 agonists had little effects on IL-8 release. Thrombin was also able to induce concentrationdependent secretion of IL-8. As low as 1 kU/L thrombin was able to induce IL-8 release from the epithelial cells, and the maximum accumulated release of IL-8 was observed with 10 kU/L thrombin, which was 7.5 fold the baseline release. Thrombin inhibitor hirudin could inhibit thrombin-induced secretion of IL-8. The time course showed that the actions of PAR1 agonist peptides SFLLR and thrombin initiated at 2 h and reached the peak at 16 h. Conclusion PAR1 agonist peptides and thrombin are potent secretogogues of IL-8 release from cultured human lung epithelial cells, and PAR1 antagonists and thrombin inhibitor may possess the ability to inhibit airway inflammation.
出处
《第一军医大学学报》
CAS
CSCD
北大核心
2005年第8期963-966,共4页
Journal of First Military Medical University
基金
广东省科技计划重点项目(2003B31502)
李嘉诚基金资助项目(C0200001)~~