| ObjectiveThe activation of interleukin-4(Interleukin-4,IL-4)in M2 type macrophages is helpful in the treatment of periodontitis and other related inflammatory diseases.Gallic acid(Gallic acid,GA)is a class of phenolic compounds that play various roles in the process of immune cell differentiation,activation and maintenance of the stability of the immune system,but its effect on M2 macrophage polarization and its mechanism have not been defined.Therefore,an in vitro polarization model of mouse peritoneal macrophages was established to explore the effects and mechanism of different concentrations of gallic acid on the polarization of M2 type macrophages.MethodsPeritoneal macrophages were extracted from eight-week-old C57BL/6female mice via intraperitoneal lavage,and treated by IL-4 to establish an M2 macrophage polarization model.The experiment was randomly divided into M group,IL-4 group,GA group(10,20,30,40 μg/m L)and p38 MAPK antagonist group.Cell morphology was observed under an inverted microscope.Cell viability was determined by MTT,Taipan blue staining and Annexin V-PE/7-AAD double staining.F4/80+CD206+ M2 macrophages were screened by flow cytometry.Real-time quantitative polymerase chain reaction(q RT-PCR)was used to detect m RNA expression of M2 macrophage-associated factor Arginase1(Arg1),Interleukin-10(IL-10)and Transforming growth factor-β1(TGF-β1).Protein expression of Arg1,IL-10,TGF-β1,β-actin,p38mitogen-activated protein kinase(p38MAPK),p38 mitogen-activated protein kinase(p-p38MAPK),signal transducer and activator of transcription 6(STAT6),phosphorylated signal transducer and activator of transcription(p-STAT6)were detected by western blot.ResultsCompared with the IL-4 group,disc-shaped cells in the GA group became larger and the number increased.There was no significant difference in macrophage activity in the GA group(P>0.05).The expression of F4/80+CD206+M2 macrophage expression showed significantly increase(P<0.01).The m RNA and protein expression of Arg1,IL-10,TGF-β1 were significantly increased.The protein expression levels of STAT6 and p38 MAPK were significantly increased.After adding SB202190,compared to IL-4 group,the percentage of M2 type macrophages and related factors expression were significantly decreased(P<0.05).ConclusionGA(10-40 μg/m L)had no significant effect on macrophage activity but can promot concentration-dependent polarization of macrophages to M2 via p38MAPK/STAT6 signaling pathway. |
