永生化人支气管上皮细胞中TGF-β_1对MAPK家族ERK通路的活化效应

Activation effect of TGF-β_1 in human bronchial epithelial BEP2D cells on ERK MAPK pathway in signal transduction

目的研究转化生长因子-β1(TGF-β1)在永生化人支气管上皮细胞BEP2D信号转导中MAPK家族ERK通路的激活情况及其引起的细胞增殖、凋亡和形态学的改变。方法用Western blot方法检测TGF-β1对ERK的活化特点;用荧光染料染色分析和流式细胞术检测TGF-β1引起ERK活化时细胞的凋亡变化;用细胞克隆形成率分析TGF-β1引起ERK活化时细胞的增殖情况;观察TGF-β1引起ERK活化时细胞的形态改变。结果TGF-β1可在短时间内激活ERK1/2,1h达峰值,然后逐渐减弱;TGF-β1诱导BEP2D细胞凋亡,抑制其增殖,使其体积和间距增大;用U0126抑制ERK通路,能促进TGF-β1对细胞在凋亡、增殖方面的作用,但抑制其对细胞形态的影响。结论TGF-β1诱导的ERK1/2的磷酸化在调节BEP2D细胞的增殖和凋亡间的平衡方面起着重要作用。

Objective To study the effect of TGF β1 on the activation of ERK MAPK in human bronchial epithelial BEP2D cells. Methods Westem blot was employed to examine the time-dependent activation of ERK MAPK by TGF-β1 BEP2D cells were harvested after treatment of human bronchial epithelial cells with 2 ng/ml TGF-β1 for 0, 10, 30, 60, 120, 240 and 480 min, respectively. Fluores- cent dye staining and flow cytometry were employed to assess the apoptosis of BEP2D cells treated with vehicle, or with 2ng/ml TGF-β1, or co-treated with 2ng/ml TGF-β1 and 5μM U0126. Proliferation of BEP2D cells treated with vehicle, or with 2ng/ml TGF-β1 or 5μM U0126, or co-treated with 2ng/ml TGF-β1 and 5μM U0126 was assayed with colony-forming test, respectively. Morphological observation was performed to observe the morphological changes in BEP2D cells treated with vehicle, or with 5ng/ml TGF-β1 or 5μM U0126, or co- treated with 5ng/ml TGF-β1 and 5μM U0126, respectively. Results TGF-β1 activated ERK MAPK in BEP2D cell. The maximal activation of ERK MAPK took place at 60min after stimulation with 2ng/ml TGF-β1. TGF β1 treatment effectively inhibited cell proliferation, and induced their apoptosis and epithelial-mesenchymal transition. Pretreatment with U0126, an inhibitor of ERK MAPK, significantly enhanced the TGF-β1-mediated anti-proliferation and apoptosis effects, and inhibited the effect of epithelial-mesenchymal transition of TGF-β1 in BEP2D cells. Conclusion TGF-β1-induced phosphorylation of ERK MAPK may participate in BEP2D cell proliferation and apoptosis regulation.