COX-2基因对TGF-β1诱导的人胚肺成纤维细胞生长及Notch信号通路的影响及其作用机制

The role of COX-2 gene expression in TGF-β1-induced growth and Notch signaling in human fetal lung fibroblasts

目的探讨抑制环氧合酶-2(COX-2)基因表达对TGF-β1诱导的人胚肺成纤维细胞系MRC-5活力、胶原合成和转化及Notch信号通路的影响。方法将MRC-5细胞分为对照组、TGF-β1组、NC组和COX-2-siRNA组,各组细胞处理48 h,Western blotting检测COX-2、胶原合成相关的COL-Ⅰ和COL-Ⅲ、 EMT标志物α-SMA及Notch信号通路受体Notch1及配体Jagged1的蛋白表达;CCK8法检测各组细胞活力。结果 TGF-β1组COX-2的表达显著高于对照组,COX-2-siRNA组COX-2的表达显著低于TGF-β1组(P<0.05),NC组COX-2的表达与TGF-β1组差异无统计学意义(P>0.05);与对照组比较,TGF-β1组细胞活力及COL-Ⅰ、COL-Ⅲ、α-SMA、Notch1和Jagged1的蛋白表达均显著升高(P<0.05),与TGF-β1组比较,COX-2-siRNA组细胞活力及COL-Ⅰ、COL-Ⅲ、α-SMA、Notch1和Jagged1的蛋白表达均显著降低(P<0.05)。结论抑制COX-2基因表达中可能通过降低MRC-5细胞活力、抑制细胞胶原合成和转化,并下调Notch信号通路,从而对肺纤维化起保护作用。

Objective To investigate the role of cyclooxygenase-2(COX-2) gene expression in TGF-β1-indcued collagen synthesis, transformation and Notch signaling in human fetal lung fibroblasts. Methods Human fetal lung fibroblasts MRC-5 cells transfected with non-targeting control or COX-2-specific siRNAs were treated with or without TGF-β1 for 48 h. Western blotting was used to detect COX-2 protein expression, alterations in the synthesis of COX-I and COX-III, and the EMT marker α-SMA, and changes in the expression of the Notch signal pathway receptor Notch1 and its cognate ligand Jagged1. Cell viability was assessed by CCK8 assay. Results COX-2 expression was significantly higher in the TGF-β1 group than the control group. Transfection of cells with COX-2-specific siRNAs significantly attenuated TGF-β1-dependent induction of COX-2 expression(P<0.05), while transfection of cells with control siRNAs had no effect. Cell viability and the expression of COX-I, COX-III, α-SMA, Notch1 and Jagged1 proteins were significantly increased in the TGF-β1 group(P<0.05), when compared with the TGF-β1 group, but were suppressed following the transfection of cells with COX-2 siRNA(P<0.05). Conclusions Inhibition of COX-2 gene expression may play a protective role in pulmonary fibrosis by reducing lung fibroblast viability and suppressing the ability of these cells to synthesize collagen and activate Notch pathway signaling.