Iguratimod alleviated Sjögren's syndrome through regulating macrophage polarization

Abstract Background The mechanism by which iguratimod (IGU) attenuates Sjögren's syndrome (SS) remains elusive. In this study, we aimed to investigate the relationship between the therapeutic effect of IGU and macrophage polarization in SS mice. Methods Female nonobese diabetic (NOD) mice were selected as the model of SS. Forty‐five female NOD mice (8‐week‐old) and five female Institute of Cancer Research (ICR) mice (8‐week‐old) were applied in this study. Five ICR mice served as control group. Five NOD mice served as model group. The remaining 40 NOD mice were randomly divided into four groups and were administrated with IGU (10, 20, and 40 mg/kg) or vehicle (model control group). After 4 weeks, the salivary flow rate of mice in each group was measured. The histopathology of submandibular glands (SG) was determined by hematoxylin and eosin staining. The markers of polarization of M1 and M2 macrophages in the SG of NOD mice were detected by quantitative real time‐polymerase chain reaction (qRT‐PCR) and Western blot analysis. The polarization of M1 and M2 macrophages in the spleen of NOD mice was detected by flow cytometric analysis. Lipopolysaccharide and interleukin‐4 were used to stimulate RAW264.7 cells, qRT‐PCR and flow cytometry analysis were used to detect the mRNA expression of M1 and M2 macrophage markers. The activation of key proteins in mitogen‐activated protein kinases (MAPKs) signaling pathway was determined by Western blot analysis. Results Compared to ICR mice, NOD mice exhibit SS‐like symptoms, lymphocyte infiltration in SG, macrophage polarization in SG. After IGU treatment, SS‐like symptoms and lymphocyte infiltration were reduced, along with a decreased proportion of M1 macrophages and an increased proportion of M2 macrophages in the SGs. Compared with the vehicle‐treated model control group, IGU reduced percentages of M1 macrophages (20 mg/kg, 40 mg/kg vs. vehicle, 21.42 ± 1.79%, 21.03 ± 1.79% vs. 28.68 ± 2.23%), whereas increased M2 macrophages (20 mg/kg vs. vehicle, 15.84 ± 0.77% vs. 11.99 ± 1.03%) in NOD mice. IGU (1 µg/mL) also suppressed M1 macrophage polarization (IGU vs. vehicle, 29.01 ± 3.70% vs. 59.67 ± 1.30%), whereas promoted M2 macrophage polarization (IGU vs. vehicle, 67.33 ± 4.78% vs. 52.30 ± 3.68%) in RAW264.7 cells. IGU also significantly regulated macrophage polarization in vitro and IGU significantly reduced the phosphorylation levels of p38, extracellular regulated protein kinase 1/2, and c‐Jun N‐terminal kinase proteins in macrophages. Conclusions IGU might alleviate SS‐like symptoms and regulate macrophage polarization through inhibiting the MAPKs pathway in SS mice. These findings clarified the mechanism underlying the therapeutic effects of IGU on SS, and provided new evidence for the further application of IGU in the treatment of SS patients.