Shin Jae-Min, Um Ji-Young, Lee Seoung-Ae, Park Il-Ho, Lee Soo-Hyung, Lee Heung-Man
1 Department of Otorhinolaryngology-Head and Neck Surgery, Korea University College of Medicine, Seoul, South Korea.
2 Division of Brain Korea 21 Program for Biomedical Science, Korea University College of Medicine, Seoul, South Korea.
Am J Rhinol Allergy. 2018 Jul;32(4):228-235. doi: 10.1177/1945892418770291. Epub 2018 Apr 20.
Purpose Methyl-CpG-binding protein 2 (MeCP2), known as a transcriptional regulator, has been suggested to play an important role in myofibroblast differentiation in the lung. The purpose of this study was to investigate the role of MeCP2 in transforming growth factor (TGF)- β1-induced myofibroblast differentiation and extracellular matrix (ECM) production in nasal polyp-derived fibroblasts (NPDFs). Methods To identify the expression of MeCP2 in nasal polyp tissues, immunohistochemistry staining and Western blot were performed. TGF- β1-induced NPDFs were treated with 5-azacytidine, a DNA methylation inhibitor, and the expression levels of α-SMA and fibronectin were determined by semiquantitative reverse transcription polymerase chain reaction, immunofluorescent staining, and Western blotting. The total soluble collagen was analyzed by the Sircol collagen assay. MeCP2 silenced by MeCP2-specific small interference ( si) RNA was verified by Western blot. Results The expression levels of MeCP2 increased in nasal polyp tissues compared to normal inferior turbinate tissues. 5-Azacytidine significantly inhibited the expression of α-SMA and fibronectin mRNA in a dose-dependent manner. In addition, 5-azacytidine suppressed collagen production and the expression of MeCP2 in the same manner. The expression levels of a-SMA and collagen production were significantly blocked by MeCP2 silencing in TGF- β1-induced NPDFs. Conclusions Our data suggest that MeCP2 plays an essential role in TGF- β1-induced myofibroblast differentiation and ECM production in NPDFs.
目的 甲基化CpG结合蛋白2(MeCP2)作为一种转录调节因子,已被认为在肺成肌纤维细胞分化中起重要作用。本研究旨在探讨MeCP2在转化生长因子(TGF)-β1诱导鼻息肉来源的成纤维细胞(NPDFs)向肌成纤维细胞分化及细胞外基质(ECM)产生过程中的作用。方法 通过免疫组织化学染色和蛋白质印迹法检测鼻息肉组织中MeCP2的表达。用DNA甲基化抑制剂5-氮杂胞苷处理TGF-β1诱导的NPDFs,通过半定量逆转录聚合酶链反应、免疫荧光染色和蛋白质印迹法检测α-平滑肌肌动蛋白(α-SMA)和纤连蛋白的表达水平。采用Sircol胶原分析法分析总可溶性胶原蛋白。通过蛋白质印迹法验证MeCP2特异性小干扰(si)RNA沉默MeCP2的效果。结果 与正常下鼻甲组织相比,鼻息肉组织中MeCP2的表达水平升高。5-氮杂胞苷以剂量依赖性方式显著抑制α-SMA和纤连蛋白mRNA的表达。此外,5-氮杂胞苷以同样的方式抑制胶原蛋白的产生和MeCP2的表达。在TGF-β1诱导的NPDFs中,MeCP2沉默显著阻断了α-SMA的表达和胶原蛋白的产生。结论 我们的数据表明,MeCP2在TGF-β1诱导的NPDFs向肌成纤维细胞分化及ECM产生过程中起重要作用。