| Objective:asthmatic model and beta 2 adrenergic receptor desensitization asthmatic model were established in order to study the mechanism of beta 2 adrenergic receptor desensitization and effectiveness of corticosteroids.Methods:Thirty-two BALB/c(6-8 weeks old)mice were divided into four groups,which is,group A,phosphate buffered saline(PBS)-treated,group B, ovalbumin(OVA)-induced,group C,salbutamol-treated,and group D, dexamethasone-treated.Group B,C and D were sensitized on days 0,14 and 21 by intraperitoneal injection of 200μg chicken egg OVA(grade V,Sigma, St.Louis,MO,USA)together with 20mg aluminum hydroxide in a total volume of 200μl.From the twenty-eighth day on,mice were challenged with an aerosol of 1%OVA(W/V)in saline using an ultrasonic nebulizer 30min/d for 1 week.Group C and D underwent the same procedure as group B besides daily intraperitoneal injection of 60μg salbutamol and inhaling an aerosol of 0.01%salbutamol 30min/d for a week half an hour before they were challenged with OVA.based on this,group D mice were injected dexamethasone 5mg/kg/d for 1 week by intraperitoneal injection.Group A mice were sensitized and challenged with PBS.Results:airway inflammation of group B and group C was more severe than that of group A and group D(p＜0.05)by lung function and a series of pathophysiologic indexs and pathologic sections.The concentration of beta 2 receptor was lower than that of other 3 groups(p＜0.01).Conclusion:asthmatic model and beta 2 adrenergic receptor desensitization asthmatic model were successfully established.PartⅡTwo dimentional electrophoresis and mass spectrographic analysisObjective:To study the different expressive dots of mouse lung total protein between beta 2 adrenergic receptor desensitization asthmatic mice and asthmatic mice by comparative proteomics.At the same time,The distinct protein dots of beta 2 adrenergic receptor desensitization asthmatic mice and dexamethasone-treated mice were also studied in the same way.Methods: Total lung protein of each group was extracted,then they were separated by isoelectric focusing and SDS-PAGE electrophoresis,so every different protein dots were found by software and statistical analysis.Subsequently, variant protein dots were identified by mass spectrogram analysis.In the end, the vital protein dots were found between the two groups,respectively.Results:Twenty and 17 different protein dots were found between the two groups,respectively.Compared to asthmatic group,twelve dots were up-regulated and eight dots were down-regulated in beta 2 adrenergic receptor desensitization asthmatic group.Compared to beta 2 adrenergic receptor desensitization asthmatic group,thirteen dots were up-regulated and four dots were down-regulated in dexamethasone-treated group.Among these dots,Rho-GDI2,Peroxiredoxin 5 and Proteasome were interesting dots we concerned.Conclusion:Oxidative stress and small G protein regulators may play an important role in the procedure of beta 2 adrenergic receptor desensitization and proteasome may play a vital role in the mechanism of corticosteroids effectiveness on beta 2 adrenergic receptor desensitization.PartⅢQualification by immunoblottingObjective:Mass spectrogram results were deeply verified by Western blot analysis in order to increase validity and persuasion.Methods:After extracting the lung total protein of every group mice,the concentrations were tested and the target protein were probed by antigen and antibody conjugation reaction.The concentration changes of Rho-GDI2,Peroxiredoxin 5 and Proteasome were found by target strips.Results:Compared to asthmatic group,Rho-GDI2 and Peroxiredoxin 5 were distinctively increased in beta 2 adrenergic receptor desensitization asthmatic group.Compared to beta 2 adrenergic receptor desensitization asthmatic group,Proteasome was decreased in dexamethasone-treated group.Conclusion:Immunoblotting analysis progressively verified the results of mass spectrogram and it indicated that oxidative stress and small G protein regulators may be related to beta 2 adrenergic receptor desensitization and proteasome may be correlated with corticosteroid effectiveness on beta 2 adrenergic receptor desensitization.
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