Anti-inflammatory Effects Of BuShen YiQi Formula In RSV-induced Asthma Exacerbation Mice

Part 1 Dual effects of RSV-infections in OVA-challenged miceBackgroud and ObjectiveThe prevalence of asthma has dramatically increased and became a worldwide public health problem. About 20% allergic asthma patients have experienced acute asthma exacerbations. RSV infection has already been regarded as the leading cause of asthma exacerbation in children. However, the mechanisms responsible for asthma exacerbation induced by RSV infection fhave not been fully elucidated. In the present study, we aimed to investigate the effects of RSV infection in OVA-challenged mice via regulation of Th17/Treg cell responses.MethodsBALB/c mice were challenged with Ovalbumin (OVA), followed by Respiratory Syncytial Virus (RSV) infections for twice, and received with the same volume of PBS instead as control. Airway hyperresponsiveness (AHR) was examined by direct airway resistance analysis. Bronchoalveolar lavage fluid (BALF) was assessed for inflammatory cell counts and secreted cytokine levels. Lung tissues were detected for cell infiltration and mucus hypersecretion. Subsequently, CD4+T cells were sorted and purified from mice lungs in different groups. And CD4+T cell subpopulations including the expression levels of important transcription factors in T lymphocyte polarization were examined.ResultsThe data indicated that initial RSV-infection significantly reduced airway hyperresponsiveness and inhibited leukocytes infiltration, especially for eosinophils, as compared with OVA-challenged mice without infection (p<0.05). Histopathological analysis showed that initial RSV-infection could suppress airway inflammation induced by allergen (p<0.05). The significant decrease of IL-4, IL-5, IL-17a and increase of IL-10 were observed in BALF of OVA-challenged mice after initial RSV-infection (p<0.05). Consistently, the findings in vivo, confirmed initial RSV-infection could remarkably down-regulate Th17 cell proportion with lower expression of RORy T, and up-regulate Treg cell proportion along with higher expression of Foxp3 (p<0.05). However, RSV re-infection could strengthen airway hyperresponsiveness and inflammation, especially for lymphocyte infiltration, as compared with OVA-challenged mice without infection. The obvious increase of IL-5, IL-17a and decrease of IL-12 were observed in BALF of OVA-challenged mice after RSV re-infection (p<0.05). Moreover, RSV re-infection could up-regulate Thl7 cell proportion with higher expression of RORy T, and down-regulate Treg cell proportion along with lower expression of Foxp3 in vivo experiments.ConclusionsUpon all results revealed that RSV-induced respiratory infections may produce dual effects pertaining to asthma. Initial RSV-infection could attenuate OVA-induced airway inflammation with reversing the imbalance of Th17/Treg responses. In contrast, RSV re-infection was able to enhance the allergic inflammation with exacerbating the pathological Th17/Treg-response imbalance.Part 2 Anti-inflammatory effects of BuShenYiQi Formula in RSV-induced asthma exacerbation miceBackgroud and ObjectiveThe prevalence of allergic asthma has been increased rapidly in recent years. About 20% in all these sufferers have experienced asthma exacerbation. Although corticosteroids and β-agonists therapy can improve serious asthma symptoms, they can’t completely cure the disease with non-neglected side effects. BuShenYiQi Formula (BSYQF) has been widely used to treat bronchial asthma and its exacerbation for decades in Huashan Hospital of Fudan University, China. Nevertheless, the mechanisms through which BSYQF as a whole exerts antiasthmatic effects have not been fully elucidated. In this study, we attempted to evaluate the involvement of Thl, Th2 and Thl7 cells in the antiasthmatic effects of BSYQF in RSV-induced asthma exacerbation mice.MethodsBALB/c mice were challenged with OVA, followed by RSV infections for establishing asthmatic exacerbation model, and treated with BSYQF, receiving with the same volume of PBS instead as control. AHR was examined by direct airway resistance analysis. BALF was assessed for inflammatory cell counts and secreted cytokine levels. OVA sepcif ic IgE in BALF and serun was also tested. Lung tissues were detected for cell infiltration and mucus hypersecretion. Subsequently, CD4+T cells and alveolar macrophages were sorted and purified from mouse lungs in different groups. And CD4+T cell subpopulations including the expression levels of important transcription factors in T lymphocyte polarization were examined. In asthma exacerbation group, after co-culture of purified CD4+T cells with macrophages, we further analyzed changes of co-cultured cells with BSYQF treatment in vitro.ResultsOur results indicated that BSYQF significantly reduced airway hyperresponsiveness and inhibited leukocytes infiltration, especially for eosinophils and neutrophils in BALF (p<0.05). Histopathological analysis showed that BSYQF could suppress airway inflammation, as well as RSV replication (p<0.05). The decrease of antigen-specific IgE, IL-4, IL-5, IL-6, IL-17a and increase of IFN-γ, IL-12 were observed in BALF and serum after treatment with BSYQF (p<0.05). Consistently, the findings in vivo and in vitro, confirmed BSYQF could down-regulate Th2-Th17 cell proportions with lower expressions of GATA3, STAT6 and RORγ T (p<0.05), and up-regulate Th1 cell proportion along with higher expression of T-bet (p<0.05). And as a result of Th1-response strengthening, activated macrophages could be detected by marked enhancement of signature gene expressions and phagocytosis (p<0.05).ConclusionsUpon all results revealed that BSYQF had a regulatory role in the imbalance between Th1 and Th2-Th17 responses to achieve an alternative therapeutic effect on serious asthmatic symptoms. In addition, BSYQF also had the anti-virus effect. Part 3 Anti-inflammatory effects of BuShenYiQi Formula VS QingReHuoXue Formula in RSV-induced asthma exacerbation miceBackgroud and ObjectiveTCMs have been demonstrated with preventive and therapeutic effects on allergic asthma and its exacerbation. Summarizing the reported studies, we could find that investigated TCM treatments for asthma have been widely used of diaphoreticrecipes, heat-clearing and detoxicating drug during early time, which are suggested the thoughts of eliminating evil(祛邪). Subsequently, other studies have been attracted in prevention and cure for asthma and its exacerbation by strengthening the body resistance (扶正) in China. The results of these studies indicate that strengthening the body resistance is also an important treatment as well as eliminating evil. Especially for individuals who are prone to recurrent asthma, treatment with strengthening the body resistance, such as tonifying kidney and Qi, could obtain more satisfied curative effects. BSYQF as a representative prescription of strengthening the body resistance has been proven that its anti-asthmatic effects should be associated with reversing the imbalanced responses between Thl and Th2-Th17 in the study of Part 2.QingReHuoXue Formula (QRHXF) has also been used to treat bronchial asthma and its exacerbation for our team. Nevertheless, the mechanisms through which QRHXF as a whole exerts antiasthmatic effects have not also been really established. In this study, we attempted to evaluate the involvement of Thl, Th2 and Th17 cells in the antiasthmatic effects of QRHXF in RSV-induced asthma exacerbation mice. By contrast between BSYQF and QRHXF, we aimed to investigate the possible differences of anti-inflammatory effects in RSV-induced asthma exacerbation mice after treatment with BSYQF and QRHXF.MethodsBALB/c mice were challenged with OVA, followed by RSV-infections for establishing asthmatic exacerbation model, and treated with QRHXF, receiving with the same volume of PBS instead as control. AHR was examined by direct airway resistance analysis. BALF was assessed for inflammatory cell counts and secreted cytokine levels. OVA specific IgE in BALF and serum was also tested. Lung tissues were detected for cell infiltration and mucus hypersecretion. Subsequently, CD4+T cells and alveolar macrophages were sorted and purified from mouse lungs in different groups. And CD4+T cell subpopulations including the expression levels of important transcription factors in T lymphocyte polarization were examined. In asthma exacerbation group, after co-culture of purified CD4+T cells with macrophages in vitro, we further analyzed changes of co-cultured cells with QRHXF treatment.ResultsOur results indicated that QRHXF significantly reduced airway hyperresponsiveness and inhibited leukocytes infiltration, especially for eosinophils, neutrophils and lymphocytes in BALF (p<0.05). Histopathological analysis showed that BSYQF could suppress airway inflammation, as well as RSV replication (p<0.05). The marked decrease of antigen-specific IgE, IL-4, IL-5, IL-6, IL-17a and IFN-γ were observed in BALF or serum after treatment with QRHXF (p<0.05). Consistently, the findings in vivo and in vitro, confirmed QRHXF could remarkably down-regulate Th2-Thl7 cell proportions with lower expressions of GATA3, STAT6 and RORy T (p<0.05). Furthermore, the slight down-regulation of Thl cell proportion and T-bet expression were also detected after QRHXF adminstration. And as a result of Thl-response suppressed, macrophages could also be mildly inhibited, including down-regulation of signature gene expressions and phagocytosis.ConclusionsUpon all results revealed that QRHXF had a suppressive role in Th2-Th17 responses to achieve an alternative therapeutic effect on serious asthmatic symptoms. And the anti-virus effect of QRHXF was much stronger than that of BSYQF, as well as the anti-inflammation in airway. However, therapeutic effects of QRHXF in general symptoms of asthma exacerbation mice were weaker than that of BSYQF.