Department of Periodontology, Nihon University School of Dentistry at Matsudo, Matsudo, Chiba, 271-8587, Japan.
Stomatological Hospital of Anhui Medical University, Hefei, Anhui, China.
Inflamm Res. 2018 Dec;67(11-12):965-973. doi: 10.1007/s00011-018-1192-1. Epub 2018 Oct 10.
MicroRNAs (miRNAs) play important roles in biological processes such as cell differentiation, development, infection, immune response, inflammation and tumorigenesis. We previously reported that the expression of miR-200b was significantly increased in inflamed gingiva compared with non-inflamed gingiva. To elucidate the roles of miR-200b in the inflamed gingiva, we have analyzed the effects of miR-200b on the expression of IL-6 in human gingival fibroblasts (HGF).
Total RNA and protein were extracted from HGF after stimulation by interleukin-1β (IL-1β; 1 ng/ml) or tumor necrosis factor-α (TNF-α; 10 ng/ml) and transfected with miR-200b expression plasmid or miR-200b inhibitor. IL-6, IL-1β, inhibitor of nuclear factor kappa-B kinaseβ (IKKβ), Zinc-finger E-box-binding homeobox 1 (ZEB1) and E-cadherin mRNA and protein levels were analyzed by real-time PCR and Western blot.
IL-1β and TNF-α increased IL-6 mRNA and protein levels, and they were significantly suppressed by miR-200b overexpression, whereas they were further increased by miR-200b inhibitor in HGF. IKKβ and ZEB1 which are target genes of miR-200b negatively regulate E-cadherin. MiR-200b suppressed the expression of IKKβ and ZEB1 and increased E-cadherin mRNA and protein levels in HGF.
These results suggest that miR-200b attenuates inflammatory response via IKKβ and ZEB1 in periodontal tissue.
微小 RNA（miRNA）在细胞分化、发育、感染、免疫反应、炎症和肿瘤发生等生物学过程中发挥重要作用。我们之前报道过，与非炎症性牙龈相比，miR-200b 在炎症性牙龈中的表达显著增加。为了阐明 miR-200b 在炎症性牙龈中的作用，我们分析了 miR-200b 对人牙龈成纤维细胞（HGF）中 IL-6 表达的影响。
用白细胞介素-1β（IL-1β；1ng/ml）或肿瘤坏死因子-α（TNF-α；10ng/ml）刺激 HGF 后提取总 RNA 和蛋白质，并转染 miR-200b 表达质粒或 miR-200b 抑制剂。通过实时 PCR 和 Western blot 分析 IL-6、IL-1β、核因子 κB 激酶β（IKKβ）抑制剂、锌指 E 盒结合同源盒 1（ZEB1）和 E-钙粘蛋白 mRNA 和蛋白水平。
IL-1β 和 TNF-α 增加了 IL-6 mRNA 和蛋白水平，miR-200b 的过表达显著抑制了它们，而 miR-200b 抑制剂则进一步增加了 HGF 中的它们。miR-200b 的靶基因 IKKβ 和 ZEB1 负调控 E-钙粘蛋白。miR-200b 抑制了 IKKβ 和 ZEB1 的表达，并增加了 HGF 中 E-钙粘蛋白 mRNA 和蛋白水平。
这些结果表明，miR-200b 通过 IKKβ 和 ZEB1 抑制牙周组织中的炎症反应。
