来氟米特调控巨噬细胞极化及其机制研究

doi:10.3969/j.issn.1004-616x.2025.02.005

Abstract

Abstract: OBJECTIVE：To investigate the effect and molecular mechanism of leflunomide on the anti-inflammatory differentiation of macrophages. METHODS：Mouse mononuclear macrophages (RAW264.7) were treated with 200 mmol/L leflunomide for 0，1，3，6，12 and 24 h. From these cells，mRNA expression levels of anti-inflammatory factors (IL-10，Arg-1，CD206)，pro-inflammatory factors (TNF-α，IL-6) and mitochondrial fusion genes (MFN1 and MFN2) in cell pellet were detected by real-time quantitative PCR (qPCR). ELISA was used to detect the content of anti-inflammatory factors in cell supernatants after flumid treatment for different years. DCFH-DA and Mito-LX staining were used to detect levels of total reactive oxygen species (ROS) and mitochondrial ROS. Mitotracker-Green staining laser confocal was used to determine cell mitochondrial morphology. Kits were used to determine content of malondialdehyde (MDA)，an intracellular oxidation product. The mRNA and protein expressions of peroxisome proliferator-activated receptor-γ (PPAR-γ) genes were determined by qPCR and Western blot，respectively. PPAR-γ inhibitor T0070907 (40 mmol/L) were pretreated for 4 h，and the changes of anti-inflammatory factors were detected by qPCR and Western blot，respectively. RESULTS：Results from the qPCR and ELISA analyses showed that the relative mRNA expression levels of anti-inflammatory factors IL-10，Arg-1 and CD206 treated with leflunomide were significantly higher than those in the control group (P<0.05). Expression of mitochondrial fusion genes MFN1 and MFN2 increased，mitochondrial length increased，and contents of ROS and MDA in cells and mitochondria decreased (P<0.05). Compared with the blank control group，mRNA and protein expression of PPAR-γ in the leflunomide group increased，and its inhibitors T0070907 reversed the increase of anti-inflammatory factors and the decrease of pro-inflammatory factors induced by leflunomide (P<0.05). CONCLUSION：The mitochondrial fusion agonist leflunomide may be useful for anti-inflammatory therapy by reducing ROS production and activating PPAR-γ to regulate the anti-inflammatory differentiation of macrophages.

Key words: leflunomide, macrophages, anti-inflammatory, peroxisome proliferator-activated receptor-γ, mitochondrial fusion

CLC Number:

R114

ZHANG Yujiao, JIANG Xiaoxu, BAI Ruiping, HUANG Xinyi, WANG Zhao, WU Hao, YU Weihua, LIU Qiling. Regulation and mechanism of leflunomide on anti-inflammatory differentiation of macrophages[J]. Carcinogenesis, Teratogenesis & Mutagenesis, 2025, 37(2): 121-127,133.

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Regulation and mechanism of leflunomide on anti-inflammatory differentiation of macrophages

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