Department of Otolaryngology-Head and Neck Surgery, Eye and ENT Hospital, Fudan University, No. 83, Fenyang Road, Xuhui District, Shanghai, China.
CAS Key Laboratory of Tissue Microenvironment and Tumor, Shanghai Institute of Nutrition and Health, University of Chinese Academy of Sciences, Chinese Academy of Sciences, No. 320, Yueyang Road, Xuhui District, Shanghai, China.
Clin Immunol. 2022 Jan;234:108895. doi: 10.1016/j.clim.2021.108895. Epub 2021 Nov 24.
Serum/glucocorticoid-regulated kinase 1 (SGK1) has been identified as a crucial regulator in fibrotic disorders. Herein, we explored SGK1 role in tissue remodeling of chronic rhinosinusitis (CRS).
Lentivirus was employed to generate an SGK1-overexpressing human bronchial epithelial cell (16HBE) line. To screen SGK1 downstream genes, RNA sequencing was performed on SGK1-overexpressing and control cell lines. To determine protein and gene expression levels, immunohistochemistry, western blotting, and quantitative real-time polymerase chain reaction were employed. Correlation analysis was performed using mRNA expression levels of SGK1, transforming growth factor β1 (TGF-β1), and connective tissue growth factor (CTGF) derived from CRS mucosal tissue and GEO database. Gene set enrichment analysis was conducted using gene sets from Molecular Signatures Database. The severity of symptoms in CRS patients was assessed using the 22-Item Sinonasal Outcome Test.
SGK1 overexpression significantly increased the expression of connective tissue growth factor (CTGF) in 16HBE cells (P < 0.01). Consistently, CTGF protein level was considerably greater in mucosal tissue of CRS without nasal polyps (CRSsNP) than in CRS with nasal polyps (CRSwNP) (P < 0.05) or in control subjects (P < 0.01). TGF-β1 protein level was higher in mucosal tissue of CRSsNP patients than in CRSwNP patients (P < 0.001) or in the control group (P < 0.01). mRNA levels of SGK1 and CTGF (P < 0.05, r = 0.668; P = 0.001, r = 0.630), TGF-β1 and CTGF (P < 0.05, r = 0.560; P < 0.05, r = 0.420), as well as SGK1 and TGF-β1(P < 0.05, r = 0.612; P < 0.05, r = 0.524) were significantly correlated in CRS mucosal tissue and GSE36830 dataset, respectively. TGF-β1-induced upregulated genes were significantly enriched in SGK1 overexpression group. In vitro assays, TGF-β1 promoted SGK1 and CTGF expression in a concentration- and time-dependent manner. Administrating an SGK1 inhibitor, GSK650394, significantly inhibited TGF-β1-induced CTGF expression in 16HBE and dispersed primary nasal polyp cells.
TGF-β1 stimulation significantly increases SGK1 and CTGF expression. By regulating TGF-β1-CTGF pathway, SGK1 may participate in tissue remodeling in the pathological mechanism of CRS.
血清/糖皮质激素调节激酶 1（SGK1）已被确定为纤维化疾病的关键调节因子。在此，我们研究了 SGK1 在慢性鼻-鼻窦炎（CRS）组织重塑中的作用。
利用慢病毒构建 SGK1 过表达的人支气管上皮细胞（16HBE）系。为了筛选 SGK1 的下游基因，对 SGK1 过表达和对照细胞系进行 RNA 测序。利用免疫组织化学、Western blot 和实时定量聚合酶链反应（PCR）检测蛋白和基因表达水平。利用 CRS 黏膜组织和 GEO 数据库中 SGK1、转化生长因子-β1（TGF-β1）和结缔组织生长因子（CTGF）的 mRNA 表达水平进行相关性分析。利用分子特征数据库中的基因集进行基因集富集分析。采用 22 项鼻鼻窦结局测试（SNOT-22）评估 CRS 患者的症状严重程度。
SGK1 过表达显著增加了 16HBE 细胞中 CTGF 的表达（P<0.01）。同样，在无鼻息肉的 CRS（CRSsNP）患者的黏膜组织中 CTGF 蛋白水平显著高于有鼻息肉的 CRS（CRSwNP）患者（P<0.05）或对照组（P<0.01）。在 CRSsNP 患者的黏膜组织中 TGF-β1 蛋白水平高于 CRSwNP 患者（P<0.001）或对照组（P<0.01）。CRS 黏膜组织中 SGK1 和 CTGF（P<0.05，r=0.668；P=0.001，r=0.630）、TGF-β1 和 CTGF（P<0.05，r=0.560；P<0.05，r=0.420）以及 SGK1 和 TGF-β1（P<0.05，r=0.612；P<0.05，r=0.524）的 mRNA 水平均呈显著相关，在 GSE36830 数据集和 CRS 黏膜组织中，分别与 TGF-β1 和 CTGF 呈显著正相关。TGF-β1 诱导的上调基因在 SGK1 过表达组中显著富集。体外实验中，TGF-β1 以浓度和时间依赖的方式促进 SGK1 和 CTGF 的表达。给予 SGK1 抑制剂 GSK650394 可显著抑制 TGF-β1 诱导的 16HBE 和分散的原发性鼻息肉细胞中 CTGF 的表达。
TGF-β1 刺激显著增加 SGK1 和 CTGF 的表达。SGK1 通过调节 TGF-β1-CTGF 通路，可能参与 CRS 病理机制中的组织重塑。
