抑炎蛋白 CYLD 通过去泛素化修饰调控人牙龈成纤维细胞的炎症反应

The inhibitory effect of the deubiquitinase cylindromatosis (CYLD) on inflammatory responses in human gingival fibroblasts.

机构信息

Jiangsu Key Laboratory of Oral Diseases, Nanjing Medical University, Nanjing, China.

Department of Periodontics, The Affiliated Stomatological Hospital of Nanjing Medical University, Nanjing, China.

出版信息

Oral Dis. 2021 Sep;27(6):1487-1497. doi: 10.1111/odi.13672. Epub 2020 Oct 22.

DOI:10.1111/odi.13672

PMID:33031609

Abstract

OBJECTIVE

Experiments were performed to evaluate CYLD expression in human gingival tissue samples and to examine the effects of CYLD on inflammatory responses in lipopolysaccharide (LPS)- or TNF-α-stimulated human gingival fibroblasts (HGFs).

METHODS

Immunohistochemistry for CYLD and p65 expression was performed with healthy and inflamed gingival tissue samples. siRNA was used to knock down the expression of CYLD in HGFs. Upon LPS or TNF-α stimulation, NF-κB activation was detected in control and CYLD-knockdown HGFs. RT-PCR was applied to determine gene expression. Western blot analyses were employed to assess protein expression. Immunofluorescence staining was carried out to evaluate the nuclear translocation of p65.

RESULTS

Immunohistochemical staining showed the expression of CYLD in human gingival tissues. In addition, CYLD protein expression was reduced in inflamed gingival tissue samples compared with healthy tissue samples. CYLD knockdown greatly enhanced the mRNA expression of proinflammatory cytokines in LPS- or TNF-α-stimulated HGFs. Furthermore, knocking down CYLD expression increased LPS-stimulated NF-κB activation in HGFs. Unexpectedly, CYLD knockdown did not affect TNF-α-induced NF-κB activation.

CONCLUSIONS

Our results suggest that CYLD participates in periodontal inflammatory responses by negatively regulating LPS-induced NF-κB signalling.

摘要

目的

实验评估了 CYLD 在人牙龈组织样本中的表达，并研究了 CYLD 对脂多糖（LPS）或 TNF-α 刺激的人牙龈成纤维细胞（HGF）中炎症反应的影响。

方法

采用免疫组织化学方法检测 CYLD 和 p65 在健康和炎症牙龈组织样本中的表达。用 siRNA 敲低 HGF 中的 CYLD 表达。在 LPS 或 TNF-α 刺激后，检测对照和 CYLD 敲低的 HGF 中 NF-κB 的激活情况。采用 RT-PCR 确定基因表达。Western blot 分析评估蛋白表达。免疫荧光染色评估 p65 的核转位。

结果

免疫组织化学染色显示 CYLD 在人牙龈组织中的表达。此外，与健康组织样本相比，炎症性牙龈组织样本中的 CYLD 蛋白表达减少。CYLD 敲低显著增强了 LPS 或 TNF-α 刺激的 HGF 中促炎细胞因子的 mRNA 表达。此外，敲低 CYLD 表达增加了 LPS 刺激的 HGF 中 NF-κB 的激活。出乎意料的是，CYLD 敲低并不影响 TNF-α 诱导的 NF-κB 激活。