背景:组蛋白去乙酰化酶(HDAC)2的表达和活性降低可能导致重度哮喘患者的炎症增加。结缔组织生长因子(CTGF)是严重哮喘气道纤维化的关键介质。然而,HDAC2/Sin3A/甲基-CpG结合蛋白(MeCP)2共阻遏复合物在调节肺成纤维细胞CTGF表达中的作用尚不清楚.
方法:研究了HDAC2/Sin3A/MeCP2辅抑制物复合物在人肺成纤维细胞(WI-38)中内皮素(ET)-1刺激的CTGF产生中的作用。我们还评估了HDAC2,Sin3A和MeCP2在卵清蛋白诱导的气道纤维化模型肺中的表达。
结果:HDAC2抑制WI-38细胞中ET-1诱导的CTGF表达。ET-1处理以时间依赖性方式降低HDAC2活性并增加H3乙酰化。此外,HDAC2的过表达抑制了ET-1诱导的H3乙酰化。抑制c-Jun氨基末端激酶,细胞外信号调节激酶,或p38通过抑制HDAC2磷酸化和降低HDAC2活性来减弱ET-1诱导的H3乙酰化。Sin3A和MeCP2的过表达均减弱了ET-1诱导的CTGF表达和H3乙酰化。ET-1诱导HDAC2/Sin3A/MeCP2共阻遏复合物的破坏,然后促使HDAC2,Sin3A,和来自CTGF启动子区的MeCP2。HDAC2,Sin3A,或MeCP2减弱ET-1刺激的AP-1-荧光素酶活性。此外,Sin3A-或MeCP2抑制的ET-1诱导的H3乙酰化和AP-1-荧光素酶活性通过HDAC2siRNA的转染而逆转。在卵清蛋白诱导的气道纤维化模型中,HDAC2和Sin3A的蛋白质水平低于对照组;然而,MeCP2表达无显著差异。该模型肺组织中磷酸-HDAC2/HDAC2和H3乙酰化的比率高于对照组。总的来说,没有刺激,HDAC2/Sin3A/MeCP2共阻遏子复合物通过调节人肺成纤维细胞中CTGF启动子区域的H3去乙酰化来抑制CTGF的表达。用ET-1刺激,HDAC2/Sin3A/MeCP2共阻遏物复合物被破坏并从CTGF启动子区解离;随后是AP-1激活和CTGF产生的最终开始。
结论:HDAC2/Sin3A/MeCP2共阻遏复合物是肺成纤维细胞中CTGF的内源性抑制剂。此外,HDAC2和Sin3A在气道纤维化的发病机制中可能比MeCP2更重要。
BACKGROUND: Reduction of histone deacetylase (HDAC) 2 expression and activity may contribute to amplified inflammation in patients with severe asthma. Connective tissue growth factor (CTGF) is a key mediator of airway fibrosis in severe asthma. However, the role of the HDAC2/Sin3A/methyl-CpG-binding protein (MeCP) 2 corepressor complex in the regulation of CTGF expression in lung fibroblasts remains unclear.
METHODS: The role of the HDAC2/Sin3A/MeCP2 corepressor complex in endothelin (ET)-1-stimulated CTGF production in human lung fibroblasts (WI-38) was investigated. We also evaluated the expression of HDAC2, Sin3A and MeCP2 in the lung of ovalbumin-induced airway fibrosis model.
RESULTS: HDAC2 suppressed ET-1-induced CTGF expression in WI-38 cells. ET-1 treatment reduced HDAC2 activity and increased H3 acetylation in a time-dependent manner. Furthermore, overexpression of HDAC2 inhibited ET-1-induced H3 acetylation. Inhibition of c-Jun N-terminal kinase, extracellular signal-regulated kinase, or p38 attenuated ET-1-induced H3 acetylation by suppressing HDAC2 phosphorylation and reducing HDAC2 activity. Overexpression of both Sin3A and MeCP2 attenuated ET-1-induced CTGF expression and H3 acetylation. ET-1 induced the disruption of the HDAC2/Sin3A/MeCP2 corepressor complex and then prompted the dissociation of HDAC2, Sin3A, and MeCP2 from the CTGF promoter region. Overexpression of HDAC2, Sin3A, or MeCP2 attenuated ET-1-stimulated AP-1-luciferase activity. Moreover, Sin3A- or MeCP2-suppressed ET-1-induced H3 acetylation and AP-1-luciferase activity were reversed by transfection of HDAC2 siRNA. In an ovalbumin-induced airway fibrosis model, the protein levels of HDAC2 and Sin3A were lower than in the control group; however, no significant difference in MeCP2 expression was observed. The ratio of phospho-HDAC2/HDAC2 and H3 acetylation in the lung tissue were higher in this model than in the control group. Overall, without stimulation, the HDAC2/Sin3A/MeCP2 corepressor complex inhibits CTGF expression by regulating H3 deacetylation in the CTGF promoter region in human lung fibroblasts. With ET-1 stimulation, the HDAC2/Sin3A/MeCP2 corepressor complex is disrupted and dissociated from the CTGF promoter region; this is followed by AP-1 activation and the eventual initiation of CTGF production.
CONCLUSIONS: The HDAC2/Sin3A/MeCP2 corepressor complex is an endogenous inhibitor of CTGF in lung fibroblasts. Additionally, HDAC2 and Sin3A may be of greater importance than MeCP2 in the pathogenesis of airway fibrosis.