青藤碱通过调控lncRNA XIST抑制角质形成细胞增殖和咪喹莫特诱导的银屑病样皮炎。

Sinomenine Suppressed Keratinocyte Proliferation and Imiquimod-Induced Psoriasis-Like Dermatitis by Regulating lncRNA XIST.

作者信息

Xiang Shoubao, Wu Xing, Xiang Yu

机构信息

Department of Dermatology, Xiangyang Central Hospital, Affiliated Hospital of Hubei University of Arts and Science, Xiangyang, China.

Department of Dermatology, The First Clinical Medical College of Guangzhou Medical University, Guangzhou, China,

出版信息

Skin Pharmacol Physiol. 2022;35(6):328-342. doi: 10.1159/000526420. Epub 2022 Aug 12.

DOI:10.1159/000526420

PMID:35960313

Abstract

BACKGROUND

Psoriasis is a chronic inflammatory skin disease. Sinomenine (SIN) has anti-inflammatory and antioxidant effects.

OBJECTIVE

The objective of this study was to confirm the anti-inflammatory effects and mechanism of SIN in imiquimod (IMQ)-induced psoriasis-like mouse model and IMQ-induced differentiated human keratinocytes (HaCaT) cells.

METHODS

BALB/c mice were treated with IMQ to construct a psoriasis-like mice model. PASI score and HE staining were used to observe pathology injury of skin tissue. The secretion of inflammatory factors and the oxidative stress level were detected by ELISA. HaCaT cells after induction of differentiation were treated with IMQ (100 μM) and SIN (10 μg/mL or 50 μg/mL), cell viability, the secretion of inflammatory factors, and the oxidative stress level were detected by MTT assay, ELISA, respectively. The expression of lncRNA XIST was detected by RT-qPCR. The relationship between XIST and EIF4G2 protein was detected by RNA immunoprecipitation (RIP) assay and ubiquitination experiment.

RESULTS

SIN significantly reduced PASI score, epidermal thickness, inflammatory response, and oxidative stress levels that increased by IMQ in vivo. SIN inhibited IMQ-induced HaCaT cell proliferation, inflammatory response, and oxidative stress levels and decreased the expression of XIST. Overexpression of XIST negated the protective effect of SIN on HaCaT cells. XIST interacted directly with EIF4G2 and regulated EIF4G2 expression via K48 ubiquitin. Knockdown of XIST reduced the half-life of EIF4G2 and decreased EIF4G2 protein stability. In addition, the E3 ubiquitin protein ligase MDM2 interacted with EIF4G2 and downregulated EIF4G2 expression. XIST reduced the interaction between MDM2 and EIF4G2, which mediated EIF4G2 K48 ubiquitination. Overexpression of XIST negated the protective effect of SIN on the inflammation of HaCaT cells through activating the NF-κB signaling pathway, while NF-κB pathway inhibitor PDTC reversed this result.

CONCLUSION

SIN had a protective effect on psoriasis and could inhibit HaCaT cell proliferation and inflammatory response via XIST/MDM2/EIF4G2/NF-κB axis.

摘要

背景

银屑病是一种慢性炎症性皮肤病。青藤碱（SIN）具有抗炎和抗氧化作用。

目的

本研究旨在证实青藤碱在咪喹莫特（IMQ）诱导的银屑病样小鼠模型及IMQ诱导的分化人角质形成细胞（HaCaT）中的抗炎作用及机制。

方法

用IMQ处理BALB/c小鼠构建银屑病样小鼠模型。采用银屑病面积和严重程度指数（PASI）评分及苏木精-伊红（HE）染色观察皮肤组织病理损伤。通过酶联免疫吸附测定（ELISA）检测炎症因子分泌及氧化应激水平。用IMQ（100 μM）和青藤碱（10 μg/mL或50 μg/mL）处理诱导分化后的HaCaT细胞，分别采用噻唑蓝（MTT）比色法、ELISA检测细胞活力、炎症因子分泌及氧化应激水平。通过逆转录定量聚合酶链反应（RT-qPCR）检测长链非编码核糖核酸XIST的表达。通过RNA免疫沉淀（RIP）实验和泛素化实验检测XIST与真核翻译起始因子4G2（EIF4G2）蛋白之间的关系。

结果

青藤碱显著降低了IMQ在体内诱导升高的PASI评分、表皮厚度、炎症反应及氧化应激水平。青藤碱抑制IMQ诱导的HaCaT细胞增殖、炎症反应及氧化应激水平，并降低XIST的表达。XIST过表达消除了青藤碱对HaCaT细胞的保护作用。XIST直接与EIF4G2相互作用，并通过K48泛素调节EIF4G2表达。敲低XIST可缩短EIF4G2的半衰期并降低EIF4G2蛋白稳定性。此外，E3泛素蛋白连接酶MDM2与EIF4G2相互作用并下调EIF4G2表达。XIST减少了MDM2与EIF4G2之间的相互作用，后者介导EIF4G2的K48泛素化。XIST过表达通过激活核因子κB（NF-κB）信号通路消除了青藤碱对HaCaT细胞炎症的保护作用，而NF-κB通路抑制剂吡咯烷二硫代氨基甲酸盐（PDTC）逆转了这一结果。