Foxp3 基因沉默对炎性环境中牙周膜成纤维细胞炎症因子表达及增殖、迁移的影响。
Effects of Foxp3 gene silencing on the expression of inflammatory cytokines and the proliferation and migration of human periodontal ligament fibroblasts in an inflammatory environment.
机构信息
School of Stomatology, Hangzhou Normal University, Hangzhou 310000, China.
Dept. of Periodontal Clinic, The Affiliated Hospital of Hangzhou Normal University, Hangzhou 310000, China.
出版信息
Hua Xi Kou Qiang Yi Xue Za Zhi. 2023 Jun 1;41(3):269-275. doi: 10.7518/hxkq.2023.2022409.
OBJECTIVES
This study aimed to clarify the effects of Foxp3 silencing on the expression of inflammatory cytokines in human periodontal ligament cells (hPDLFs) in an inflammatory environment and on cell proliferation and invasiveness, as well as to explore the role of Foxp3 gene in the development of periodontitis.
METHODS
An small interfering RNA (siRNA) construct specific for Foxp3 was transfected into hPDLFs. Foxp3 silencing efficiency was verified by reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting, and the siRNA with the optimum silencing effect of Foxp3 gene was screened. Using lipopolysaccharide to simulate an inflammatory environment , CCK-8 detected the effect of silencing Foxp3 on hPDLFs proliferation under inflammatory conditions. Wound-healing experiments and transwell assays were conducted to detect the effect of silencing Foxp3 on hPDLF migration under inflammatory conditions. The expression of the inflammatory cytokines interleukin (IL)-6 and IL-8 was detected by RT-PCR and Western blotting under inflammatory conditions.
RESULTS
After siRNA transfection, RT-PCR and Western blotting analyses showed that the expression of Foxp3 mRNA in the Foxp3-si3 group decreased significantly (=21.03, <0.000 1), and the protein expression of Foxp3 also decreased significantly (=12.8, <0.001). In the inflammatory environment, Foxp3 gene silencing had no significant effect on hPDLFs proliferation (>0.05), and Foxp3 gene silencing promoted hPDLFs migration (<0.05). Moreover, the expression of IL-6 and IL-8 increased (<0.05).
CONCLUSIONS
In an inflammatory environment, Foxp3 gene silencing promoted hPDLFs migration but had no significant effect on hPDLFs proliferation. The expression of inflammatory factors expressed in hPDLFs increased after Foxp3 gene silencing, indicating that Foxp3 gene inhibited inflammation in periodontitis.
目的
本研究旨在阐明 Foxp3 沉默对炎症环境下人牙周韧带细胞（hPDLFs）中炎症细胞因子表达、细胞增殖和侵袭的影响，并探讨 Foxp3 基因在牙周炎发病机制中的作用。
方法
将 Foxp3 的小干扰 RNA（siRNA）构建体转染到 hPDLFs 中。通过逆转录-聚合酶链反应（RT-PCR）和 Western blot 验证 Foxp3 沉默效率，并筛选出 Foxp3 基因具有最佳沉默效果的 siRNA。用脂多糖模拟炎症环境，CCK-8 检测炎症条件下沉默 Foxp3 对 hPDLFs 增殖的影响。划痕实验和 Transwell 实验检测炎症条件下沉默 Foxp3 对 hPDLFs 迁移的影响。RT-PCR 和 Western blot 检测炎症条件下炎症细胞因子白细胞介素（IL）-6 和 IL-8 的表达。
结果
siRNA 转染后，RT-PCR 和 Western blot 分析显示 Foxp3-si3 组 Foxp3 mRNA 的表达显著降低（=21.03，<0.0001），Foxp3 蛋白表达也显著降低（=12.8，<0.001）。在炎症环境中，Foxp3 基因沉默对 hPDLFs 增殖没有显著影响（>0.05），但促进了 hPDLFs 的迁移（<0.05）。此外，IL-6 和 IL-8 的表达增加（<0.05）。
结论
在炎症环境中，Foxp3 基因沉默促进 hPDLFs 迁移，但对 hPDLFs 增殖没有显著影响。沉默 Foxp3 基因后，hPDLFs 中炎症因子的表达增加，表明 Foxp3 基因抑制了牙周炎的炎症反应。
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