The Effects Of Bacille Calmette-Guerin's Compound Extracts On Airway Inflammation And Hyperresponsiveness In Allergic Murine Model

Part I Effect of different challenge methods on allergic murine modelObjective: To investigate the effect of different challenge methods on allergic murine model.Methods: BalB/c mice were sensitized with ovalbumin by intraperitoneal injection on days 0, 7 and 14, and firstly challenged with OVA on days 28. Animals were divided into 5 groups according to different challenge methods, including intranasal administration (2mg/mlOVA, 50μl)for 3 consecutive days (In.3d group), aerosol administrations (1%OVA) for 20 minutes once a day for 2 consecutive days (Ae.2d-20 group ), 20 minutes once a day for 3 consecutive days (Ae.3d-20 group), 30 minutes once a day for 3 consecutive days (Ae.3d-30 group), 20 minutes once a day for 4 consecutive days(Ae.4d-20 group). Mice in normal control group were untreated Airway responsiveness to increasing doses of methacholine was assessed 48 hours after the last ovalbumin challenge in conscious mice by using a single-chamber, whole-body plethysmograph, present as enhanced expiratory pause (Penh), followed by bronchoalveolar lavage with PBS. Cytology was analysed.Results: Airway hyperresponsiveness was showed in both of intranasal challenge and aerosol challenge. Eosinophils percentage (Eos%) in BALF of asthmatic mice was significantly increased compared with control(P<0.05). Airway hyperresponsiveness and Eos% in BALF of In.3d group were significantly increased compared with other groups of aerosol challenge(P<0.05). Eos% in BALF of In.3d group(46.30%±4.55%) were significantly increased compared with Ae.2d-20 group (31.19%±12.84%), Ae.3d-20 group (29.00%±12.33%) and Ae.4d-20 group (37.08%±8.44%)(P<0.05). Penh% of In.3d group was significantly increased than Ae.2d-20 group and Ae.4d-20 group when Mch was 3.125mg/ml. 2 mice died in the group of intranasal administration, there was no death in other groups.Conclusions: Allergic murine model can be established successfully by both of intranasal challenge and aerosol challenge, and more serious airway inflammation and hyperresponsiveness can be induced by the intranasal challenge than aerosol challenge.PartⅡThe effects of Bacille Calmette-Guerin's compound extracts on airway inflammation and hyperresponsiveness in allergic murine modelObjective To investigate the effect of BCG's compound extracts on airway hyperresponsiveness and airway inflammation in allergic murine model.Methods 1. BalB/c mice were sensitized and challenged by ovalbumin, as described previously. Mice were randomized and divided into untreated control group, dexamethasone group, DNA10 group, PSA10 group, BCG's compound extracts group. Dexamethasone (5mg/kg) was injected intraperitoneally 1 hour before every intranasal challenge. BCG's extracts (nucleic acid and polysaccharide) were administered by intraperitoneal injection 7 days before the first sensitizition.2. According to different BCG's compound proportions, BCG's compound extracts groups were divided into 16 groups.( table 1-1) 3. Airway hyperresponsiveness to increasing doses of methacholine was assessed 48 hours after the last ovalbumin challenge in conscious mice by using a single-chamber, whole-body plethysmograph, present as enhanced expiratory pause (Penh). Following this, bronchoalveolar lavage were performed with PBS. lungs were lavaged with PBS. Lungs of representative mice from each group were formalin-fixed, and sections were stained with H&E stain for examination by light microscopy.Results1. BCG's extracts with predominant proportion of PSA is the main part can significantly suppress antigen-induced airway eosinophilia and airway hyperresponsiveness(P<0.05). Eos% in BALF of P/D(10/1) group(41.48%±9.83%), P/D(40/10) group (43.05%±5.90%), P/D(100/40) group (42.58%±5.37%) were significantly lower than BCG-PSN group (50.55%±14.28%), DNA10 group (50.53%±8.13%) and PSA10 group (49.50%±4.35%)(P<0.05).2. BCG's extracts with predominant proportion of DNA is the main part can significantly suppress antigen-induced airway eosinophilia and airway hyperresponsiveness (P<0.05). Eos% in BALF of P/D(1/10) group (44.15%±7.76), P/D(10/40) group (43.35%±5.74%), P/D(40/100) group (45.55%±2.45%) were significantly lower than BCG-PSN group (P<0.05). Eos% in BALF of P/D(10/40) group was significantly lower than DNA10 group and PSA10 group(P<0.05)。Penh% of P/D(40/100) group was significantly lower than BCG-PSN group at 0.78, 3.125, 6.25mg/ml concentrations of Mch, and significantly lower than DNA10 group at 6.25-25mg/ml concentrations of Mch (P<0.05).3. Eos% in BALF were significantly suppressed by BCG's compound extracts with equal proportion, except P/D(1/1). Eos% in BALF of P/D(40/40) group (40.5%±7.97%) and P/D(100/100) group (44.35%±4.16%) were significantly lower than BCG-PSN group, DNA10 group, PSA10 group (P<0.05). Airway hyperresponsiveness of P/D=1:1 groups were significantly lower than the asthma group (P<0.05). Penh% of P/D(40/40) group, P/D(100/100) group were significantly lower than P/D(1/1) group, P/D(10/10) group, BCG-PSN group(P<0.05). Penh% of P/D(100/100) group was significantly lower than DNA10 group at 6.25-50mg/ml concentrations of Mch, and significantly lower than PSA10 group at 1.56 and 50mg/ml concentrations of Mch (P<0.05).4. The levels of IFN-γin P/D(40/40) group (24.42pg/ml±3.95pg/ml) and P/D(100/100) group (26.96pg/ml±7.98pg/ml) were significantly higher than asthma group (18.18pg/ml±6.06pg/ml)(P<0.05).Conclusion1. BCG's compound extracts can suppress the airway inflammation and hyperresponsiveness in allergic murine model. Early intervention with BCG's compound extracts based on 40μg or 100μg equal proportion are more effective than the BCG-PSN group and BCG single extracts groups.2. BCG's extracts can increase IFN-γlevels in BALF of allergic murine model.