The Regulatory Functions And Mechanisms Of GPR84 In Acute Respiratory Distress Syndrome

Acute respiratory distress syndrome?ARDS?is a kind of acute pulmonary disease,clinical characterizations of which include hyperinflammation,non-cardiac pulmonary edema and respiratory failure.Based on epidemiological data,several direct or indirect causes can induce ARDS,among which pneumonia,sepsis and trauma are the major clinical causes.Subject to the current limited knowledge on ARDS and lack of specific therapeutic strategy,ARDS is still an acute disease with high mortality.ICU mortality of severe ARDS patients is approximate to 46%,and mechanical ventilation is the common clinical therapy to relieve the respiratory dysfunction of ARDS patients.The pathogenesis of ARDS is closely related to the imbalance of pulmonary microenvironment's immune homeostasis.Different with other local tissue microenvironments,pulmonary microenvironment faces direct exposure to the external environment.The pulmonary immune homeostasis is maintained and regulated by a complex system,in which several cell populations and commensal bacteria are involved.At steady state,pulmonary microenvironment is kept at the status of immunological tolerance,which are regulated by type?alveolar epithelial cells,alveolar macrophages and regulatory T cells.Under pathological conditions such as ARDS,the immune homeostasis is disrupted.Infiltrated neutrophils produce amount of reactive oxygen species and accelerate the respiratory burst;macrophages,monocytes and dendritic cells are all capable to release several pro-inflammatory cytokines and cause cytokine storm;the existence of respiratory burst and cytokine storm may exacerbate the imbalance of immune homeostasis,and finally result in the impaired epithelium and dysfunction of alveolar units.Within the regulatory network of pulmonary immune homeostasis,alveolar macrophages are indispensably involved.Alveolar macrophages are self-renewal lung-resident macrophages and have long life spans.At quiescent state,alveolar macrophages are regulated by several negative signals in the alveolar niches and keep tolerant to the commensal bacteria and particles with low immunogenicity in the pulmonary microenvironment.During the process of pulmonary diseases,once alveolar macrophages recognize and acquire enough stimulatory signals,alveolar macrophages overcome the tolerance status,participate in the immune responses and tissue recovery,regulate functions of different cell populations and rescue the pulmonary immune homeostasis in the end.During the pathogenesis of different lung diseases,alveolar macrophages may play various roles with different immune regulatory functions.Traditional theory of macrophage polarization divides the activated status of macrophages under extreme conditions into M1?classical activation?and M2?alternative activation?.M1macrophages are mainly related with pro-inflammatory immune responses,while M2macrophages are involved in the immune regulation and tissue recovery.The process of macrophage polarization is regulated by different signals,including metabolites such as free fatty acids.However,previous studies have confirmed that the traditional theory of macrophage polarization?M1/M2?failed to perfectly elucidate the in vivo phenotypes of alveolar macrophages during the pathogenesis of different lung diseases.Analysis of the microarray data from GEO database shows that the expression of Gpr84,a member of free fatty acid receptors,is upregulated on lung tissues under different pathological conditions.GPR84 is a metabolic-sensing G protein-coupled receptor,and its potential endogenous ligands are medium-chain free fatty acids.Free fatty acids are essential for physiological activities.Free fatty acids mainly come from the metabolic products of intestinal commensal bacteria and are transported to different tissues through blood vessels in different forms.Recent studies have shown that free fatty acids can not only be absorbed by macrophages and involved in the metabolic reprogramming,but also act as signals to regulate the functions of macrophages via free fatty acid receptors.Several researches indicated that the activation of GPR84 and its downstream signal pathways may take part in the regulation of immune cell populations,such as macrophages and neutrophils.In the study,we intended to further explore the lung cell populations during ARDS process,and tried to reveal whether and how GPR84 was involved in the ARDS.With the establishment of LPS-induced ARDS mouse model,we found Gpr84 was upregulated on the lung tissues from ARDS mice.Gpr84^-/-mice shew alleviated ARDS symptoms with intact pulmonary histology,well-balanced air-blood barrier,limited respiratory burst and cytokine storm.Analysis on the ARDS process demonstrated that myeloid monocyte-derived cells were not the main source of macrophages in the alveolar niches during ARDS process,while alveolar macrophages transformed from CD11b^lo to CD11b^hi status,the trend of which was basically consistent with the trends of pro-inflammatory cytokines.Temporary depletion of alveolar macrophages coincided with the phenomena that infiltration of neutrophils reduced and sustained release of pro-inflammatory cytokines almost vanished.We firstly found that alveolar macrophages dominated the interaction with LPS in the pulmonary microenvironment,and CD11b^hi alveolar macrophages exhibited pro-inflammatory phenotype.Alveolar macrophages and their switch of subtypes played pivotal roles in the regulation of ARDS process.Gpr84^-/-mice shew downregulated switch of alveolar macrophage subtypes and blunted chemotaxis of neutrophils,and both were considered as possible key regulatory approaches of Gpr84 knockout to ARDS.Deep analysis on the regulatory mechanisms of neutrophil infiltration in ARDS shew that the reduced infiltration of neutrophils in Gpr84^-/-mice was mainly attributed to the downregulated alveolar macrophage-mediated recruitment via chemokines.Recent studies have already shown the preliminary regulatory effects of GPR84on the myeloid macrophages,however,the mechanisms remain further exploration.Alveolar macrophages are lung-resident macrophages under the regulation of pulmonary microenvironment and have different origins,phenotypes and transcriptional profiles.In the study,we intended to ascertain the regulatory effects and mechanisms of GPR84 on alveolar macrophages.Treatment with LPS in vitro shew that Gpr84^-/-alveolar macrophages displayed significantly downregulated expressions of ARDS-related cytokines,phagocytic capacity and switch of subtypes.We hence hypothesized that GPR84 may be involved in the LPS recognition process of alveolar macrophages via TLR4 and the activation of TLR4 downstream signal pathways.In spite of no obvious differences between wild-type and Gpr84^-/-alveolar macrophages at quiescent state,Gpr84^-/-alveolar macrophages shew downregulated expression or release of CD14 and LBP in response to LPS challenge.The downregulated LPS interaction of Gpr84^-/-alveolar macrophages with lower extent of CD14 expression in vivo also verified the results in vitro.Further analysis on the signal pathways shew that the downstream signal pathways of GPR84 on alveolar macrophages were referred to the phosphorylated level of Akt,Erk and Stat3.Overall,it came to a conclusion that GPR84 was involved in the LPS-induced switch of alveolar macrophage subtypes during ARDS due to their regulatory effects on the LPS recognition process of alveolar macrophages via Akt/Erk-Stat3-CD14/LBP.In 2018,Prometic Life Science Company declared that a potential GPR84antagonist had been approved by FDA to enter Phase 3 clinical trial of idiopathic pulmonary fibrosis.Combined with our results,GPR84 may be a potential therapeutic target to relieve the hyperinflammation in ARDS,rescue the pulmonary immune homeostasis and protect the physiological functions of alveolar units,all of which may jointly lead to the lower mortality and improved life quality of ARDS patients.