海水对肺腺癌细胞株A549细胞蛋白酶激活受体2表达的作用研究

目的 观察海水对肺腺癌细胞株A549细胞的炎性损伤及对蛋白酶激活受体2(PAR-2)表达的影响,探讨海水导致急性肺损伤的作用机制.方法 将培养的A549细胞接种于6孔板中,按随机数字表法把细胞分为对照组及海水处理2、4、8 h组,相应时间点用实时荧光定量逆转录-聚合酶链反应(RT-PCR)和蛋白质免疫印迹法(Western blotting)检测PAR-2的mRNA和蛋白表达;收集细胞培养上清液,用酶联免疫吸附法(ELISA)检测肿瘤坏死因子-α(TNF-α)和白细胞介素-8(IL-8)含量.结果 用海水处理后,A549细胞PAR-2 mRNA和蛋白表达均于2 h显著增加(P均<0.05),4 h达高峰,分别为对照组的1.8倍和2.2倍,随后下降,但仍显著高于对照组(P均<0.01);TNF-α和IL-8含量则于2 h即显著升高并达最高峰[分别为(214.35±20.85)ng/L,(55.86±5.65)ng/L],随后下降,但仍显著高于对照组[分别为(25.86±3.85)ng/L,(6.97±1.77)ng/L,P均<0.01].结论 海水可致A549细胞炎症反应明显,并可诱导A549细胞PAR-2表达增高.

The effect of seawater on expression of protease-activated receptor 2 in adenocarcinoma of lung cell line A549 cells

Objective To investigate the effect of seawater on expression of protease-activated receptor 2 (PAR-2) in adenocarcinoma of lung cell line A549 cells,and the inflammatory injury on A549 cells induced by seawater. Methods A549 cells were randomly divided into four groups: control group,2 hours group,4 hours group,8 hours group,in which cells were treated with seawater for 2,4 and 8 hours respectively. After seawater treatment,cells were collected for real-time quantitative reverse transcrition-polymerase chain reaction (RT-PCR) and Western blotting analysis to determine the expression of PAR-2 mRNA and its protein. Supernatant were collected for tumor necrosis factor-α (TNF-α) and interleukin-8 (IL-8) which were determined by enzyme-linked immunosobent assy (ELISA). Results The expression of PAR-2 mRNA and protein of A549 cells increased in a time-dependent manner after seawater treatment,significantly so after 2 hours in all groups (both P<0.05),and peaked at 4 hours after seawater treatment (1.8-fold and 2.2-fold respectively,both P<0.01),followed by a decrease though still higher than those of control group significantly (both P<0.01). TNF-α and IL-8 in supernatant increased significantly after seawater treatment,peaking at 2 hours after seawater treatment [(214.35±20.85) ng/L,(55.86±5.65) ng/L] and then followed by a slight decrease though still significantly higher than those of control group [(25.86±3.85) ng/L,(6.97±1.77) ng/L,all P<0.01]. Conclusion Seawater can induce significant inflammation of A549 cells and up-regulate the expression of PAR-2 on A549 cells.