The Effects Of Bacillus Calmette-Guerin Vaccinated At Ovalbumin Sensitization On Airway Inflammation In A Mouse Model Of Asthma

Background Asthma is a chronic airways inflammatory disorder in which many cells and cytokines participate associated with variable airflow limitation, airway hyperresponsiveness(AHR),airway inflammation and remodel, airway mucus hypersecretion. Asthma is characterized by a T-helper 2(Th2)type immune response. In recent decade,many research have proved that Bacillus Calmette-Guerin(BCG) is a strong inducer of a T-helper type 1(Th1) immune response. It can promote cytokines from Th1 cells such as IFN-γ synthesis and inhibits cytokines from Th2 cells such as IL-4/IL-5, which regulate the Th1/Th2 balance to Th1 response and inhabit airway eosinophilia and AHR to antigens, besides inhibits the Goblet cells(GC) hyperplasia and metaplasia(GCH/M) and the mucus secretion. So BCG can provide a preventive and therapy in asthma. In most animal researches,BCG is vaccinated before sensitized or challenged and few inculated after sensitized or challenged.But it's rare to study the effects of BCG inculated at allergen sensitization.An external study found that AHR in asthman model mice were inhibited by the treatment of BCG and Dermatophagoides pteronyssinus,but no inhibition action by only BCG. Moreover,several studies shew cooperative and enhanced inhibition effect on inflammation of asthma models by vaccinated with BCG and allergen synchronously,but the causation was not shown. Can BCG inhibit airway inflammation in asthma model more effectively at allergen sensitization than before and why?Objective Based on a mouse model of asthma we created previously, vaccinate BCG two times at or before allergen sensitization with different doses. We observe that effect of intraperitoneal BCG on the airway inflammation and mucus production, and investigate the difference of inhibitive effect on airway inflammation between at sensitization and before and research the potential reason.Methods Male C57BL/6 mice,6-8 weeks old,were sensitized two times and challenged three times by ovalbumin(OVA) as asthma model group, and mice were treated by normal saline as control group.Intraperitonally vaccinated BCG twice at or before allergen sensitization with 10~5, 10~6 colony-forming units(CFUs).(Al group:10~5 CFUs before sensitization, A2 group:10~6 CFUs before sensitization, B1 group:10~5 CFUs at sensitization, B2 group:10~6 CFUs at sensitization). BALF and PB were collected at 48 hours after the final allergen challenge respectively. Eosinophil and leukocyte was determined in BALF, IL-5,IFN-γ was measured by ELISA in PB and BALF. To show eosinophils infiltration,the histology of lung tissues was also observed.Results1.The changes of total number of leukocytes and the percent and number of eosinophils of BALF after BCG vaccination1.1 We found total number of leukocytes and the percent and number of eosinophils in model group increased obviously compared with control group(P<0.01);1.2 After BCG vaccination, total number of leukocytes and the percent and number of eosinophils in the vaccinated groups decreased obviously compared with model group,no significant differences in vaccinated groups(P>0.05).2.The effects of BCG vaccination on the changes of pulmonary pathology2.1 In control group, there are no Eos accumulation around airways and vessels, however there are large amount of inflammatory cells and Eos around airways and vessels of model group, while in vaccinated groups there are still a lot of inflammatory cells, but Eos infiltration in the alveolar wall decreased significantly compared with model group.2.2 The changes of BCG vaccination on airway mucus overproductionIn control group, few GC was found in the epithelial cells, few mucus storing in GC, no GC hyperplasia /hypertrophy(GCH/M) occurred. While large GCH/M was found in the airway epithelium, large mucus production in GC in model group. In vaccinated groups, the GCH/M and mucus were decreased to a large extent in airway epithelium.2.3 The semiquantity of mucus in the airway epithelium2.3.1 MOR and HR increased obviously in model group compared with control group(P<0.01,P<0.05);2.3.2 MOR decreased obviously in vaccinated groups compared with model group(P<0.01); MOR of B2 group decreased more obviously than MOR of A1 and A2 groups (P<0.05);2.3.3HR were restrained in B1,B2 groups compared with model group (P<0.05)and no inhibtion effect be found in A1,A2 groups(P>0.05).3.The effect of IL-5 in BALF and PB after BCG vaccination3.1 The concentration of IL-5 in BALF and PB of model group increased obviously compared with control group(P<0.01);3.2The concentration of IL-5 in BALF in the vaccinated groups but A group decreased obviously compared with model group(P<0.01), and The concentration of IL-5 in B2 group decreased more obviously than the one of A1 and A2 groups(P<0.01, P<0.05);3.3 To B1,B2 group,the concentration of IL-5 in PB were lower than model group(P<0.05); but no significant decrease have been found in A1,A2 groups(P>0.05).4.The effect of IFN-γ in BALF and PB after BCG vaccination4.1 No significant differences of the concentration of IFN-γ in BALF and PB have been found between model group and control group(P>0.05);4.2 In vaccinated groups, the concentration of IFN-γ in BALF increased obviously only be found in B2 group compared with model group(P<0.01);the one in B2 group more higer than the one in A1 group(P<0.01);4.3 The concentration of IFN-γ in PB increased obviously in B1,B2 groups comparedwith model group(P<0.05,P<0.01); the one in B2 group more higer than the one in A1 group(P<0.05).5.The change of IL-5/IFN-γ in BALF and PB after BCG vaccination5.1 The change of IL-5/IFN-γ in BALF after BCG vaccinationIn vaccinated groups, IL-5/IFN-γ in BALF decreased obviously compared with model group with except of A1 group, more significant change has be found in B2 group(P<0.01); significant differences have been found between B2 and A1,A2 groups(P<0.01), between B1 and A1 group(P<0.05), between A2 and A1 group(P<0.01).5.2 The change of IL-5/IFN-γ in PB after BCG vaccinationIn model group,IL-5/IFN-γ in PB increased obviously compared with control group;in vaccinated groups, IL-5/IFN-γ in PB decreased obviously compared with model group(A1 group P<0.05,the others P<0.01),and no statistics differences have been found in vaccinated groups (P>0.05).Conclusion1. Intraperitoneal BCG significantly inhibited the accumulation of eosinophilic in BALF and reduced eosinophilic influx into the airway lumen either at or before allergen sensitization. Intraperitonally vaccinated BCG at sensitization inhibited the mucus production in the airway epithelium more significantly than before sensitization.2. Intraperitoneal BCG at allergen sensitization significantly inhibited the airway inflammation in asthma model compared with vaccinated before allergen sensitization,and there was a dose-dependent manner in the preventive effect.3.Enhanced inhibition effect on the airway inflammation with intraperitoneal BCG at allergen sensitization may relate with Th1/Th2 balance in asthma model adjusted by BCG vaccination.