To Alleviate Airway Allergic Inflammation Diseases As Well As Mechanisms By IL-2 Combined With Dexamethasone

PurposeTo alleviate mouse allergic airway inflammatory diseases by administration of IL-2(IL-2) combined with dexamethasone.To explore the role of FoxO3a in glucocorticoid-induced apoptosis by silencing foxo3a siRNA gene and detection of lymphocyte apoptosis after dexamethasone treatment.To explore the mechanism of the expansion of regulatory T cells by IL-2 combined with dexamethasone pre-treatment.Methodsa) To establish a mouse model of allergic airway diseases and detect the number of BALF cells,cytokine levels,lung pathology.b) After different doses of IL-2 combined with dexamethasone,Treg changes and the duration of effect were measured by flow cytometry.c) After IL-2 combined with dexamethasone pretreatment,allergic airway disease model in mice was set up followed by a series of measurement.d) lymphocyte suspension was pretreated by different concentration of dexamethasone,collected and detected by western blot for the change of FoxO3a and phospho-FoxO3a.After foxo3a siRNA transfection and dexamethasone pre-treatment,lymphocyte apoptosis was detected by apoptosis detection kit. The dexamethasone-sensitive cell lines and dexamethasone-resistant cell lines were compared with each other for the change of FoxO3a and phospho-FoxO3a after dexamethasone treatment.e) CD4~+ CD25~+ cells and CD4~+ CD25~- cells were selected by magnetic selection kit.To observe these cells' response to different concentration of IL-2 and dexamethasone(detection of apoptosis).After dexamethasone pretreatment,FoxO3a,phospho-FoxO3a,Bim were detected by western blot analysis under different concentrations of IL-2 and dexamethasone. f) After Akt-Ⅳinhibitor pretreatment,phospho-FoxO3a changes were detected by western blot.FoxO3a,phospho-FoxO3a and Bim were also detected after Akt-Ⅳpretreatment.Results1.Allergic airway disease model was set up successfully because the total cell count,eosinophils,IL-4,IL-5 level in mice bronchoalveolar lavage fluid were increased while the IFN-γlevel was decreased.2.IL-2 combined with dexamethasone might expand regulatory T cells with appropriate dose of IL-2 400000IU each mouse,dexamethasone 0.1mg per mouse.3.After IL-2 combined with dexamethasone treatment,we found that mouse allergic airway disease had been alleviated.4.With increased dose of dexamethasone,the apoptosis rate and total FoxO3a expression were also increased,while phospho-FoxO3a was reduced. Dexamethasone-induced lymphocyte apoptosis was decreased after foxo3a gene silencing.5.After dexamethasone treatment,there were increased FoxO3a expression and decreased phospho-FoxO3a expression in dexamethasone-sensitive tumor cells,while the protein expression in dexamethasone-resistant tumor cell lines didn't change much.6.In the presence of appropriate dose of IL-2,after dexamethasone treatment,CD25~+ Tregs had higher expression of phospho-FoxO3a and lower apoptosis rate than CD25~- Teff.7.After Akt inhibitor treatment, phospho-FoxO3a expression in Treg and Teff was inhibited,then changing the dose of IL-2 couldn't affect phospho-FoxO3a expression and apoptosis rate of Treg and Teff.Conclusion1.IL-2 combined with dexamethasone could alleviate allergic airway inflammatory disease.2.IL-2 combined with dexamethasone could upregulate Tregs.3.FoxO3a activity might takes animportant part in dexamethasone-induced lymphocyte apoptosis. 4.The expression of CD25 might be the reason of that Treg could be protected from dexamethasone-induced lymphocyte apoptosis by IL-2.5.After IL-2/IL-2R activation,both Akt and SGK might phosphoate FoxO3a.