Ye Huifang, Yang Xi, Chen Xiong, Shen Lijun, Le Rongrong
Eye Hospital, Wenzhou Medical University, Wenzhou, Zhejiang, China.
Eye Hospital, Wenzhou Medical University, Wenzhou, Zhejiang, China; The First People's Hospital of Yichang, Yichang, Hubei, China.
Exp Eye Res. 2020 Oct;199:108146. doi: 10.1016/j.exer.2020.108146. Epub 2020 Jul 26.
To examine the protective effects of Isoliquiritigenin (ISL) in angiotensin II (ANG II)-induced inflammation and fibrosis on Human Tenon's capsule Fibroblasts (HTFs) and Mouse Peritoneal Macrophages (MPMs). This study also investigated the potential mechanism of action of ISL.
Methyl-thiazolyl tetrazolium (MTT) assay was used to test ISL toxicity. An ELISA and an RT-qPCR assay detected the inflammatory cytokines (TNF-α, IL-6, COX-2, and ICAM-1). A Western blot investigated the expression levels of inflammation-related signals [nuclear factor-κB (NF-κB), peroxisome proliferator-activated receptor γ (PPARγ)], and fibrogenesis, including fibronectin and alpha-smooth muscle actin (α-SMA)]. Protein expressions of α-SMA were measured by immunofluorescence.
Pre-treatment with ISL (10 or 20 μM) dose-dependently decreased the mRNA levels of TNF-α, IL-6, ICAM-1, and COX-2 induced by ANG II (1 μg/ml) in both MPMs and HTFs. ANG II remarkably increased the amount of P65 in the nuclei and decreased the amount of P65 in the cytoplasm. Additionally, ANG II reduced PPARγ expression levels in a time-dependent manner. Furthermore, these effects which were induced by ISL were remarkably neutralized by ISL pre-treatment. Finally, ANG II markedly elevated the expression of fibronectin and α-SMA.
ISL could alleviate ANG II-induced fibrogenesis by inhibiting the NF-κB/PPARγ inflammatory pathway. In addition, ISL may be a potential agent for the treatment of conjunctival fibrosis. Most importantly, the NF-κB/PPARγ signaling pathway could be an effective therapeutic target for the prevention and treatment of conjunctival fibrosis after glaucoma surgery.
研究异甘草素（ISL）对血管紧张素II（ANG II）诱导的人Tenon囊成纤维细胞（HTFs）和小鼠腹腔巨噬细胞（MPMs）炎症和纤维化的保护作用。本研究还探讨了ISL的潜在作用机制。
采用甲基噻唑基四氮唑（MTT）法检测ISL毒性。酶联免疫吸附测定（ELISA）和逆转录定量聚合酶链反应（RT-qPCR）检测炎症细胞因子（肿瘤坏死因子-α、白细胞介素-6、环氧合酶-2和细胞间黏附分子-1）。蛋白质印迹法检测炎症相关信号[核因子-κB（NF-κB）、过氧化物酶体增殖物激活受体γ（PPARγ）]以及包括纤连蛋白和α平滑肌肌动蛋白（α-SMA）在内的纤维化形成相关蛋白的表达水平。通过免疫荧光法检测α-SMA的蛋白表达。
在MPMs和HTFs中，用ISL（10或20 μM）预处理呈剂量依赖性降低ANG II（1 μg/ml）诱导的肿瘤坏死因子-α、白细胞介素-6、细胞间黏附分子-1和环氧合酶-2的mRNA水平。ANG II显著增加细胞核中P65的量，降低细胞质中P65的量。此外，ANG II以时间依赖性方式降低PPARγ表达水平。此外，ISL预处理可显著中和这些由ANG II诱导的效应。最后，ANG II显著提高纤连蛋白和α-SMA的表达。
ISL可通过抑制NF-κB/PPARγ炎症通路减轻ANG II诱导的纤维化。此外，ISL可能是治疗结膜纤维化的潜在药物。最重要的是，NF-κB/PPARγ信号通路可能是预防和治疗青光眼手术后结膜纤维化的有效治疗靶点。
