Effects Of Epigallocatechin-3-gallate On Macrophage Polarization In Murine Bone Marrow-derived Macrophages

Macrophages are a population of immune cells with a high degree of phenotypic and functional heterogeneity.They derive from circulating monocytes capable of changing their activation state in response to micro-environmental cues.Based on their functional phenotype,macrophages can be subdivided into classically activated macrophages(M1)and alternatively activated macrophages(M2).Like IFN-? as well as microbicidal stimuli such as lipopolysaccharides(LPS)polarize macrophages to an M1 phenotype,which secrete various of proinflammatory cytokines,reactive oxygen species,such as TNF-?,IL-1?,ROS etc to remove pathogens,necrotic tissue and activate other immune cells to help the body to resist infection.Activated macrophages are induced by interleukin-4(IL-4),and IL-13 and secrete a variety of anti-inflammatory factors,for example Ym-1 and IL-10 and so on,which mainly appear in the stage of inflammation and contribute to tissue repair and possess immune regulatory functions.(–)-Epigallocatechin-3-gallate(EGCG)is the most abundant and active compound in green tea.And the wide range of beneficial effects of green tea including anti-inflammatory,anticarcinogenic,antimicrobial,and anti-oxidative effects is mainly exhibited by EGCG.EGCG and its underlying mechanism of action have been extensively studied for anticancer and anti-inflammatory activities.However,it is unclear whether EGCG could affect macrophage polarization in murine bone marrow-derived macrophages.To determine the effect of epigallocatechin-3-gallate(EGCG)on macrophage polarization in murine bone marrow-derived macrophages,we analyzed mRNA and protein levels for marks in which M1(LPS and IFN-?-induced)and M2(IL-4-stimulated)secreted from bone marrow-derived macrophages,respectively.We collected bone marrow cells from 6~8 weeks C57BL/6 mice,which cultured RPMI1640 medium with 10% FBS with M-CSF(100 ng/mL)in vitro,and differentiated into macrophages,in which M1(LPS1 ?g/mLand50 ng/mL IFN-?-induced),or M2(20 ng/mL IL-4-stimulated).Meanwhile the cells were treated with various concentrations of EGCG(12.5-50 ?mol/L).7 days later,we examined the expression of pro-and anti-inflammatory factors,IL-1? TNF-?,iNOS,Arg-1,Ym-1and IL-10 in murine myeloid macrophages on gen and protein with RT-PCR and ELISA.We found that up-regulated the high levels of cytokines,IL-1??TNF-??iNOS in IFN-? and LPS-stimulated murine myeloid macrophages.However,EGCG effectively inhibited IFN-?/LPS-stimulated murine myeloid macrophages by reducing inflammatory cytokine levels in dose-dependent manner.Conversely,the cells were induced by IL-4,expressing high level of Arg-1,Ym-1and IL-10.In response to an M2 stimulus,EGCG-treated groups showed an increase expression of these cytokines on m RNA and protein in dose-dependent manner.These studies show that EGCG could attenuated marking factors expression of M1 macrophage,and promote the expression of M2 markers.