The Role Of MK2 In Modulating Macrophage Activation And Pulmonary Inflammatory Injury And Its Drug Interference

Acute lung injury（ALI）and acute respiratory distress（ARDS）are serious pulmonary diseases,that are characterized by inflammatory cell infiltration,production of inflammatory cytokines and chemokines,respiratory disorder,and eventually lead patients to die.ALI is a complicated diseases in which activated macrophages release a large amount of inflammatory factors that promotes neutrophil accumulation and induces endothelial cell and epithelial cell damage.It is vital to determine how macrophage modulates the pathogenesis of ALI.Since ALI has high incidence,morbidity and mortality but there is no effective treatment,It is importance of illumination of the pathogenesis,identification of novel drug targets development and development of potential therapeutic agents for treatment of ALI.MK2 is an important downstream substrate of p38 MAPK,that participating in regulation of cytokine production,transcription factor activation,inflammatory receptor expression.Although MK2 play a critical role in inflammatory diseases,it has not been completely defined how MK2 regulates macrophage activation and ALI.Here,we found that MK2 deficient mice(MK2^-/-)protected against sepsis-induced ALI.In response to lipopolysaccharide（LPS）challenge,MK2^-/-mice and myeloid-specific MK2 deficient mice(MK2^Lyz2-KO)displayed attenuated inflammatory response,especially producing fewer amount of cytokines.LPS treatment in vitro resulted in reduction of cytokine expression in MK2^-/-bone marrow derived macrophages（BMDMs）.miRNA is a class of non-coding single stranded RNA that plays an critical role in regulation of inflammatory process.Despite both the importance of MK2 and miRNAs in macrophage activation and acute inflammation,it still remains unclear whether miRNAs contribute to MK2-mediated cytokine production in ALI.In this study,we demonstrated that LPS-induced microRNA lethal-7e（let-7e）expression was significantly increased in MK2^-/-and MK2^Lyz2-KO macrophages,indicating that let-7e is involved in the inflammatory response mediated by MK2.Transfection of let-7e mimics into MK2^+/+BMDM decreased cytokine expression,neutralization of let-7e in MK2^-/-BMDM with let-7e antagomirs rescued LPS-induced cytokine production.Meanwhile,LPS-induced phosphorylation of CREB,a substrate of MK2,was down-regulated in MK2^-/-BMDMs.Lin28,an specific inhibitory molecule of let-7,was significantly reduced in MK2^-/-macrophages.Furthermore,we revealed that CREB inhibitor downregulates Lin28 expression and upregulates let-7e expression.Our results suggested that MK2boosts LPS-induced macrophage activation and ALI via increasing activation of CREB and consequently the expression of Lin28 and down-regulation of let-7e,indicating that p38 MAPK/MK2/CREB/Lin28/let-7e may be a novel signaling pathway and interference of these potential drug target molecules may inhibit macrophage activation and alleviate ALI.Protostemonine（PSN）is one of the main components of Stemona sessilifolia,a traditional Chinese medicine which is used to treat respiratory diseases.In this study,we found that PSN alleviates LPS-induced acute lung injury in vivo,and attenuates LPS-induced macrophage activation and reduces the release of inflammatory mediators such as iNOS,NO,TNF-?,IL-6 and IL-1?in vitro.Furthermore,PSN inhibits the TLR4-mediated signaling pathway and phosphorylation of MK2 in macrophages,which suggested that PSN might be a potential drug candidate for treatment of ALI.In addition,we also found that PSN suppressed eosinophilic asthmatic inflammation,decreased IL-4-induced AAM polarization and relieved pathological damage of asthma.Collectively,in this study,we identified that（1）MK2 boosts LPS-induced macrophage activation and ALI via increasing activation of CREB and consequently the expression of Lin28 and down-regulation of let-7e;（2）PSN inhibits macrophage activation and ALI by suppressing TLR4-mediated signaling pathways and MK2phophorylation;（3）PSN attenuates eosinophilic inflammation by suppressing AAM polarization.This study clarified the molecular mechanism of MK2 in regulation of ALI,and provided a number of potential drug targets and a therapeutic candidate for treating ALI.