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目的探讨低盐(LS)培养对小鼠致密斑细胞(MMDD1)环氧化酶2(COX-2)表达、前列腺素E2(PGE2)释放的诱导作用及p38丝裂素激活蛋白激酶(MAPK)信号通路的调控作用。方法采用RT-PCR和免疫印迹方法检测正常盐(NS)与LS培养对MMDD1细胞COX-2表达的影响。用ELISA法检测上清液PGE2的含量。用免疫印迹检测细胞内p-p38 MAPK表达的变化。结果与NS培养相比,LS培养均能诱导MMDD1细胞COX-2 mRNA和蛋白表达增加(16 h时高峰mRNA 0．94±0．12比0．26±0．09,28 h时高峰蛋白0．59±0．02比0．25±0．07,P均< 0．01);PGE2分泌各时间点均显著升高,于24 h达高峰[(644．33±26．54)ng／L比(224．0±18．33) ng／L,P<0．01]。LS培养后,MMDD1细胞内p38MAPK的磷酸化程度显著上调,180 min时较高(从0．17±0．01升至0．28±0．01,P<0．01)。20μmol／L p38抑制剂SB-203580下调LS诱导的COX-2蛋白表达(从0．58±0．01降至0．19±0．02,P<0．01)。结论低盐培养促进MMDD1细胞COX-2的表达和PGE2的分泌。p38 MAPK激活介导了低盐诱导的MMDD1细胞COX-2表达。
Objective To investigate the effects of low salt (LS) culture on the expression of cyclooxygenase 2 (COX-2), the release of prostaglandin E2 (PGE2) and the expression of p38 mitogen-activated protein kinase (MAPK) Regulation of signaling pathways. Methods The effects of normal saline (NS) and LS culture on the expression of COX-2 in MMDD1 cells were detected by RT-PCR and Western blotting. The content of PGE2 in supernatant was detected by ELISA. The changes of intracellular p-p38 MAPK expression were detected by Western blotting. Results Compared with NS, LS cultured could induce the increase of COX-2 mRNA and protein expression in MMDD1 cells (peak mRNA 0.94 ± 0.12 vs. 0.26 ± 0.09 at 16 h, peak protein 0 at 28 h .59 ± 0.02 vs 0.25 ± 0.07, P <0.01). PGE2 secretion increased significantly at each time point and peaked at 24 h [(644.33 ± 26.54) ng / L ratio (224.0 ± 18.33) ng / L, P <0.01]. After LS cultured, the phosphorylation of p38MAPK in MMDD1 cells was significantly up-regulated (from 0.17 ± 0.01 to 0.28 ± 0.01, P <0.01) at 180 min. 20μmol / L p38 inhibitor SB-203580 down-regulated LS-induced COX-2 protein expression (from 0.58 ± 0.01 to 0.19 ± 0.02, P <0.01). Conclusion Low salt culture can promote the expression of COX-2 and the secretion of PGE2 in MMDD1 cells. p38 MAPK activation mediates low-salt-induced COX-2 expression in MMDD1 cells.