Effect Of Atorvastatin On Airway Inflammation And Immune Regulation In Asthmatic Mice Modle

Objective: BALB/c mice were sensitized and challenged by ovalbumin to induce asthma, and to observe the effect of atorvastatin on Th1, Th2, Th17 cytokines IFN-γ, IL-4, IL-17, and to further investigate effect of atorvastatin on airway inflammation and it’s immune regulation in asthmatic mice modle for the experimental basis of atorvastatin in treatment of bronchial asthma. Methods: Twenty-four Specific pathogen free female BALB/c mice were randomly divided into three groups(8 mice/group): control group(NS group), asthma model group(OVA group) and atorvastatin treatment group(AT group). Mice were sensitized with OVA/alum(containing 50μg of OVA and 2mg of aluminum hydroxide) in 0.2 m L normal saline by intraperitoneal plus subcutaneous injection on days 0, 7, 14 in all groups mice except negative control sensitized with 0.2 m L normal saline. One week after the final injection, mice in OVA group and AT group were challenged with intranasal inhalation of OVA 50μL(containing 20μg of OVA), once daily, for consecutive 7 days from day 21 to 27. Control mice were challenged with 50μL normal saline at the same time. AT group mice were treated with intraperitoneal injection of atorvastatin(10mg/kg) at 30 min before ovalbumin provocation at every time, for consecutive 7 day, while NS group and OVA group mice were recevied normal saline. All mice were sacrificed 24 h after the last provocation. Then BALF and lung tissue were collected. The total white cells, neutrophils, eosinophils, lymphocytes in BALF were counted. The pathomorphological change of lung tissue was observed by hematoxylin-eosin(HE)stain. Concentration of IFN-γ, IL-4 and IL-17 in BALFwas measured by ELISA. The expression of RORγt and IL-17 in lung tissue was observed by immunohistochemistry. Results:(1) General observation: mice in asthmatic group showed various types of typical asthma-like symptoms, such as oral lip cyanosis, scratching nose, dyspnea, nodded breathing, abdominal breathing and even gatism. Symptoms of acute asthma was eased in AT group, while mice in NS group showed no these symptoms.(2) Pathomorphological change: in OVA group, numerous inflammatory cell infiltrated around the airway and blood vessels, bronchial stenosis, mucus plug accmulated in the lumen bronchial as well as epithelium shedding were identified, and the changes described above were eased in AT group, while in NS group showed normal.(3) Count and categorization of cells in BALF: the number of total cells, eosinophils, neutrophils, lymphocytes in OVA group(24.3±2.1, 7.9±0.6, 2.8±0.2, 0.9±0.04) were much higher than those in NS group(7.1±0.4, 0.0±0.0, 1.4±0.08, 0.3±0.02) and OVA group(13.6±1.5, 2.5±0.12, 1.9±0.09, 0.7±0.05), and the differences were statistically significant(P<0.05); but the number of cells in AT group was still higher than those in NS group, and the difference bewteen them was statistically significant(P<0.05).(4) Concentrations of IFN-γ, IL-4 and IL-17 in BALF: the concentration of IL-4 and IL-17 in OVA group(157.45±9.01, 195.68 ±20.68) was significantly increased than those in NS group(37.58±1.62, 29.76 ±3.54) and AT group(72.82±6.98, 111.378±8.75), and the differences were statistically significant(all P<0.05), but level of IL-4, IL-17 in AT group was still higher than those in NS group, and the difference were statistically significant(P<0.05). The concentration of IFN-γ in OVA group(248.47±21.62) was decreased than that in NS group(256.00±17.71), with no statistic significance between them(P>0.05); IFN-γ in AT group(213.23±10.37) was decreased than that in OVA group and NS group(P<0.05), and there were statistic significance between them(all P<0.05). IFN-γ/IL-4 in OVA group(1.57±0.10) was significantly decreased than that in NS group(6.81±0.56) andAT group(2.93±0.19), and there were statistic significance among three groups(P<0.05), while IFN-γ/IL-4 in AT group was still lower than that in NS group(P<0.05).(5)The expression of RORγt and IL-17 in lungs: the OD value of RORγt and IL-17 in OVA group(0.523±0.060, 0.258±0.035)were higher than those in NS group(0.096±0.010, 0.032±0.004) and AT group(0.300±0.033, 0.115±0.019), and there was statistical significance among three groups(all P <0.05), and OD value of RORγt and IL-17 in AT group were still higher than those in NS group, and the differences were statistically significant(all P<0.05).(6) Correlation analysis demonstrated that concentration of IL-17 in BALF was positively correlated with the number of neutrophils(r=0.832, P<0.05) and eosinophils(r=0.960, P<0.05) in BALF. The expression of RORγt and IL-17 in lung tissue was positive correlation(r=0.865, P<0.05). Conclusions:(1) Asthmatic mice model was successfully established by sensitization of intraperitoneal plus subcutaneous injection and provocation of intranasal inhalation of ovalbumin.(2) Asthmatic mice were in Th1/Th2 imbalance, with Th2, Th17 hyperfunction.(3) Atorvastatin could relieve airway inflammation in asthmatic mice.(4) Atorvastatin exert immune regulation through correcting the imbalance of Th1/Th2, decreasing the level of IL-17 by suppressing the expression of RORγt, which might be one of atorvastatin’s anti-inflammatory mechanism in asthmatic mice.