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目的观察支原体巨噬细胞活化脂肽-2(MALP-2)诱导人气道上皮细胞分泌粘蛋白MUC5AC的分子机制。方法体外培养人气道上皮细胞NCI-H292,分别采用0、0.1、1.0和5.0μg/m L MALP-2刺激NCI-H292细胞24 h,采用酶联免疫吸附测定(ELISA)检测培养上清中MUC5AC和基质金属蛋白酶9(MMP-9)的含量;Western blot检测表皮生长因子受体(EGFR)磷酸化水平。同时采用EGFR抑制剂AG-1478或MMP-9抑制剂(MMP-9 Inhibitor I)处理细胞,观察其对MUC5AC分泌的影响。结果 NCI-H 292细胞未刺激时,MUC 5 AC以及MMP-9分泌水平极低。当给予0.1~5μg/m L MALP-2作用18 h后,MUC 5 AC的分泌水平显著增高。此外,5μg/m L MALP-2作用NCI-H 292细胞1 h后可诱导EGFR磷酸化。采用AG-1478预处理细胞1 h后,MMP-9及MUC 5 AC分泌水平明显减少,同时,采用10 nmol/L MMP-9抑制剂处理后也能下调MUC 5 AC水平。结论支原体MALP-2经EGFR/MMP-9诱导气道上皮细胞分泌MUC5AC。
Objective To investigate the molecular mechanism of MALP-2 induced by Mycoplasma macrophage activating human lipopeptide-2 (MIP-2) in human airway epithelial cells. Methods Human airway epithelial cells NCI-H292 were cultured in vitro. NCI-H292 cells were stimulated with MALP-2 at concentrations of 0, 0.1, 1.0 and 5.0 μg / mL for 24 h respectively. MUC5AC was detected by enzyme-linked immunosorbent assay (ELISA) And matrix metalloproteinase 9 (MMP-9) were detected by Western blot. The phosphorylation of epidermal growth factor receptor (EGFR) was detected by Western blot. The cells were treated with EGFR inhibitor AG-1478 or MMP-9 Inhibitor I simultaneously to observe the effect on MUC5AC secretion. Results When NCI-H 292 cells were not stimulated, MUC 5 AC and MMP-9 secretion was extremely low. MUC 5 AC secretion increased significantly when MALP-2 was treated with 0.1 ~ 5 μg / ml L for 18 h. In addition, EGFR phosphorylation was induced by NCG-H292 cells treated with 5μg / mL MALP-2 for 1 h. After pretreatment of cells with AG-1478 for 1 h, the secretion of MMP-9 and MUC 5 AC significantly decreased. At the same time, MUC 5 AC level was down-regulated by 10 nmol / L MMP-9 inhibitor. Conclusion Mycoplasma MALP-2 secreted MUC5AC through EGFR / MMP-9-induced airway epithelial cells.