Department of Ophthalmology, The Affiliated Hospital of Qingdao University, NO. 16 Jiangsu Road, Qingdao, Shandong Province 266000, China.
Department of Biochemistry, Microbiology and Immunolog, Wayne State University School of Medicine, Detroit, MI 48201, USA.
Int Immunopharmacol. 2021 Jul;96:107785. doi: 10.1016/j.intimp.2021.107785. Epub 2021 May 24.
To explore the anti-inflammatory effect of lipoxin A4 (LXA4) in Aspergillus fumigatus (A. fumigatus) keratitis and the underlying mechanisms.
In A. fumigatus keratitis mouse models, enzyme-linked immunosorbent assay (ELISA) was used to detect the level of LXA4. Clinical scores were utilized to evaluate fungal keratitis (FK) severity. Fungal load was assessed by plate count. Immunofluorescence staining, HE staining and myeloperoxidase (MPO) assays were carried out to evaluate the neutrophil infiltration and activity. In A. fumigatus infected mouse corneas and inactivated A. fumigatus-stimulated RAW264.7 cells, quantitative real time polymerase chain reaction (qRT-PCR) and ELISA were applied to assess the expression of pro-inflammatory mediators and anti-inflammatory factors.Reactive oxygen species (ROS) was determined by 2',7'-dichlorodihydrofluorescein diacetate (DCFH-DA) staining in RAW264.7 cells.
LXA4 level was significantly increased in mice with A. fumigatus keratitis. In an A. fumigatus keratitis mouse model, LXA4 treatment alleviated FK severity, reduced fungal load and repressed neutrophil infiltration and activity. Additionally, LXA4 inhibited the expression of pro-inflammatory mediators including IL-1β, TNF-α, IL-6, cyclooxygenase-2 (COX-2), TLR-2, TLR-4, Dectin-1 and iNOS, and promoted the expression of anti-inflammatory factors IL-10 and Arg-1. In RAW264.7 cells, LXA4 receptor/formyl peptide receptor 2 (ALX/FPR2) blockade reversed the anti-inflammatory effect of LXA4. LXA4 suppressed inactivated A. fumigatus induced elevated ROS production in RAW264.7 cells, which was abrogated by ALX/FPR2 antagonist Boc-2.
LXA4 ameliorated inflammatory response by suppressing neutrophil infiltration, downregulating the expression of pro-inflammatory mediators and ROS production through ALX/FPR2 receptor in A. fumigatus keratitis.
探讨脂氧素 A4（LXA4）在烟曲霉菌角膜炎中的抗炎作用及其机制。
在烟曲霉菌角膜炎小鼠模型中，采用酶联免疫吸附试验（ELISA）检测 LXA4 水平。临床评分用于评估真菌性角膜炎（FK）的严重程度。平板计数法评估真菌负荷。免疫荧光染色、HE 染色和髓过氧化物酶（MPO）测定评估中性粒细胞浸润和活性。在烟曲霉菌感染的小鼠角膜和失活的烟曲霉菌刺激的 RAW264.7 细胞中，采用实时定量聚合酶链反应（qRT-PCR）和 ELISA 评估促炎介质和抗炎因子的表达。在 RAW264.7 细胞中通过 2'，7'-二氯二氢荧光素二乙酸酯（DCFH-DA）染色测定活性氧（ROS）。
烟曲霉菌角膜炎小鼠中 LXA4 水平显著升高。在烟曲霉菌角膜炎小鼠模型中，LXA4 治疗减轻 FK 严重程度，降低真菌负荷，抑制中性粒细胞浸润和活性。此外，LXA4 抑制促炎介质的表达，包括白细胞介素-1β（IL-1β）、肿瘤坏死因子-α（TNF-α）、白细胞介素-6（IL-6）、环氧化酶-2（COX-2）、Toll 样受体-2（TLR-2）、Toll 样受体-4（TLR-4）、Dectin-1 和诱导型一氧化氮合酶（iNOS），并促进抗炎因子白细胞介素-10（IL-10）和精氨酸酶-1（Arg-1）的表达。在 RAW264.7 细胞中，LXA4 受体/甲酰肽受体 2（ALX/FPR2）阻断剂逆转了 LXA4 的抗炎作用。LXA4 抑制失活的烟曲霉菌诱导的 RAW264.7 细胞中升高的 ROS 产生，该作用被 ALX/FPR2 拮抗剂 Boc-2 所阻断。
LXA4 通过 ALX/FPR2 受体抑制烟曲霉菌角膜炎中中性粒细胞浸润、下调促炎介质表达和 ROS 产生，从而改善炎症反应。
