Effects Of Iloprost On Acute Allergic Airway Inflammation Mediated By ILC2s In Mice

BackgroundAllergic inflammation is the main pathological feature of type 2 immunological disorders,asthma is an airway inflammatory disease,which is characterized by mucus hyperproduction,airway hyperresponsiveness(AHR)and airway remodeling.The traditional view of asthma was induced by IgE-mediated mast cell proliferation,Th2cell activation and release of a large number of Th2 cytokines.Allergic airway inflammation has also been caused in response to allergens such as protease without the involvement of Th2 cells,suggesting that innate immune cells can produce large quantities of type 2 signature cytokines in acute stages of allergic responses.However,it has turned out that group 2 innate lymphoid cells(ILC2s),the mirror image cells of Th2 cells,play a crucial role in the initiation of acute allergic airway inflammation.ILC2s were first identified in intestinal-associated mucosal tissue,and depended on the transcription factors GATA3 and RORα,which was essential for its development.Moreover,ILC2s were found expressing IL-33 receptor(called ST2)and IL-25 receptor(called IL17RB).Published reviews have described that ILC2s responding to allergens or epithelium-derived cytokines IL-33 and IL-25 could produce Th2 cytokines,such as IL-5 and IL-13.Therefore,IL-33-ILC2-IL-5/IL-13axis is considered extremely important pathway in the acute allergic airway inflammation induced by protease.PGI2 is a metabolism of the arachidonicacid via cyclooxygenase(COX)pathway and produces biological effect through the G protein-coupled receptor IP.Iloprost,as the PGI2 analogue,has been used in the context of pulmonary hypertension.Animal studies have shown that inhibiting COX can enhance the inflammatory response and the expression of type 2 cytokines in lung CD4~+Th2 cells,suggesting that COX products can suppress inflammatory response induced by allergen.In the present study,iloprost also can inhibit the secretion of IL-4,IL-10 and IL-13 from Th2 cells in a dose dependent fashion,which can regulate type 2 immune response.Considering all of this,it is notable to determine whether iloprost has negative regulation on the function of ILC2s.ObjectiveIn this study,we established an acute allergic airway inflammation mice model by intranasally administration of IL-33,and interfered the mice with iloprost to explore the inhibitory effects of iloprost on ILC2s and the production of type 2cytokines,such as IL-5 and IL-13,which could provide us with potential novel means to acute allergic airway inflammation.Materials and methodsTwenty-four female C57BL/6 mice aged 6-8 weeks were randomly divided into four groups:DMSO control group,IL-33 stimulation group,IL-33+iloprost treament group and single iloprost group.Inflammatory cell infiltration in lung tissue was observed by HE staining,and mucus secretion was observed by PAS staining.The number of eosinophils(EOS)and group 2 innate lymphoid cells(ILC2s)in broncho alveolar lavage fluid(BALF)and lung tissues was analyzed by flow cytometry.The cytokine levels of IL-5 and IL-13 were detected by ELISA.The expressions of IL-5,IL-13,GATA3 and ST2 at mRNA levels were tested by real-time quantitative PCR.Results1.Pathological changesHE and PAS staining of lung sections revealed that the infiltration of inflammatory cells,hypersevere plasia of goblet cells and mucus secretion in IL-33stimulation group was more severe than DMSO control group and single iloprost group.The IL-33 and iloprost treatment group was treated with iloprost intranasally30 minutes prior to each challenge with IL-33 for 3 consecutive days.Compared with the IL-33 stimulation group,the IL-33 and iloprost treatment group had less mucus secretion,inflammatory cells infiltration and airway goblet cells hyperplasia.2.Flow cytometric analysis resultsCompared with the DMSO control group,the percentages and the number of eosnophils and ILC2s,in BALF and lung suspension,had increased in IL-33stimulation group(P<0.05);Compared with the IL-33 stimulation group,the percentages and the number of eosnophils and ILC2s had decreased in IL-33 and iloprost treatment group(P<0.05).3.ELISA resultsIn BALF and lung tissue homogenate,the levels of IL-5 and IL-13 in IL-33stimulation group were significantly higher than DMSO control group(P<0.05);when intervented with iloprost,the levels of IL-5 and IL-13 were decreased in IL-33and iloprost treatment group(P<0.05).4.Real-time PCR resultsThe expression of IL-5、IL-13、GATA3 and ST2 at mRNA level were increased in IL-33 stimulation group,compared with the DMSO control group(P<0.05);when intervented with iloprost,the expression of IL-5,IL-13,GATA3 and ST2 at mRNA level were decreased in IL-33 and iloprost treatment group(P<0.05).ConclusionIntranasal administration of IL-33 contributes to create the acute allergic airway inflammation mice model.Besides,iloprost can significantly restrain the proliferation of ILC2s and eosinophils,as well as the levels of type 2 cytokine such as IL-5 and IL-13,which suggests that iloprost has an inhibitory effect on ILC2s in a mouse model of acute allergic airway inflammation.