The Regulation Of Allergic Asthma By Commensal Bacteria

The human mucosa is colonized by trillions of microorganisms,most of which have co-evolved with the host in a symbiotic relationship.These microorganisms can both promote the development and functions of the mucosal immune system as well as contribute to tissue damage and diseases.Allergic asthma is a common respiratory immune disease that has become increasingly popular in recent years,especially in developed countries.Epidemiological data indicate that antibiotics exposure during infants and young children increases the risk of allergic asthma.The antibiotics exposure is one of the major causes of commensal bacteria disorder.These views are consistent with the Hygiene Hypothesis,which means that growing up in an environment with fewer microbiota can lead to an increase in the prevalence of allergic diseases.The data from mouse asthma model provided evidence for the Hygiene Hypothesis:Germ-free mice and antibiotic-treated neonatal mice were more susceptible to OVA(ovalbumin)or House Dust Mite-induced asthma,which are consistent with conclusions from clinical studies.However,the composition and function of commensal bacteria change over time,especially in the early time of life such as from birth to weaning(pre-weaning),which indicate that the commensal bacteria in different periods of life may have different effects on asthma development.Previous mouse studies have focused on the protective role of commensal bacteria in pre-weaning mice on the development of asthma,but it is unclear whether the bacteria colonized after weaning also play a similar role.Until now,it is still incompletely understood why some people develop asthma but others do not even though both populations are exposed to allergens to the same degree.In fact,allergic sensitization determines whether a person will develop asthma or not.Allergic sensitization is a process of complex interactions between the host immune and environmental factors,and polarizes an allergen-specific Th2 immune response.However,whether the commensal bacteria can regulate this key process remains to be explored.Here are main results of our study:1.Commensal bacteria aggravate OVA-induced asthma by affecting allergic sensitization in post-weaning mice.To investigate the effect of dysbiosis of commensal bacteria on OVA-induced asthma in post-weaning mice,we administered a combination of antibiotics to 3-week-old post-weaning mice in drinking water(referred to as antibiotic(ATB)mice)to disrupt commensal bacteria.Then,we established asthma by peritoneal sensitization using OVA/alum and intranasal challenge with OVA.Compared with non-antibiotic-treated control mice(referred to as ATB-free mice),ATB mice exhibited reduced lung injury and infiltration of fewer eosinophils as well as lower IgE and Th2-type cytokines in pulmonary alveoli after asthma establishment.These results suggested that,different from the protective function of commensal bacteria in pre-weaning mice,commensal bacteria in post-weaning mice could aggravate OVA-induced asthma.However,if we advanced OVA/alum sensitization to the time point of initiation of antibiotic administration(referred to as ATB’ mice),these mice exhibited aggravated lung injury and Th2 response after asthma establishment when compared with non-antibiotic-treated control mice(referred to as ATB-free’ mice),which.is different to the results from ATB mice.However,there were no significant differences between ATB-free mice and ATB-free’ mice after asthma establishment.These results suggested that commensal bacteria in post-weaning mice affected the sensitization phase of OVA-induced asthma and might have two opposite functions.2.Commensal bacteria promote sensitization through maintenance of NLRP3/IL-1β expression in pMφ.Commensal bacteria influenced the sensitization phase of OVA-induced asthma in post-weaning mice,so we next explore which types of cells in the peritoneal cavity were involved in this process and how commensal bacteria affected target cells.We found that depletion of peritoneal macrophages(pMφ)or deficiency of NLRP3/IL-1β signaling attenuates OVA-induced asthma.Further studies showed that pMcp rapidly activate and secrete large amounts of IL-1β within a few hours after sensitization.Inflammatory monocytes were gradually recruited into peritoneal cavity to induce Th2 responses under IL-1β signal from pMφ.And antibiotic treatment can cause downregulation of NLRP3/IL-1β expression in pMcp,which in turn reduce recruitment of inflammatory monocytes in the sensitization phase and further alleviate asthma.3.Commensal bacteria activate TLR signaling to control recruited inflammatory monocyte-mediated excessive inflammation.Treating post-weaning mice with antibiotic at the time point of OVA/alum sensitization could aggravate the severity of asthma in the ATB’ mice with normal pMφ activation,and there was no difference in recruitment of inflammatory cells between ATB-free’ and ATB’ mice,implying that,in addition to maintain NLRP3/IL-1β signaling to promote sensitization,commensal bacteria might play other roles to control excessive inflammatory response in OVA-induced asthma.Further studies found that after sensitization,the expression of TLRs on inflammatory monocytes recruited to the peritoneal cavity gradually increased.On the contrary,the expression of TLR on pMφ rapidly decreased.The protective effect of TLR signaling on asthma has been widely reported.We found that TLRs signaling-mediated the protective effect acted on the early stage of sensitization phase in OVA-induced asthma.Inflammatory monocyte transfer could induce asthma in the recipient mice after OVA challenge even without OVA/alum sensitization indicating that recruited inflammatory monocytes after sensitization played a critical role in OVA-induced asthma.Furthermore,we treated the purified inflammatory monocytes with TLR stimuli in vitro before transferring into the recipient mice and found that treated inflammatory monocytes induced milder asthma disease in recipient mice after OVA challenge compared with untreated inflammatory monocytes.Previous studies have demonstrated TLRs signaling activation prevent OVA-induced asthma through promoting Thl response.Therefore,these data suggested that activation of TLRs signaling on recruited inflammatory monocytes could attenuate OVA-induced asthma disease.Further studies showed that there were lower levels of Thl-associated cytokines IL-6 and TNF-a in recruited inflammatory monocytes in the peritoneal cavity of ATB’ mice compared with their counterparts from ATB-free’ mice.Altogether,these results indicated that,in addition to maintain NLRP3/IL-1β signaling in pMφ to recruit inflammatory cells after allergic sensitization,commensal bacteria also activated TLR signaling on inflammatory monocytes to inhibit excessive Th2 response in a chronological order.4.NLRP3/IL-1β signaling and TLRs signaling are regulated by different bacterial species.To explore which bacterial species regulated NLRP3/IL-1βsignaling and/or TLR signaling in the mouse model of OVA-induced asthma,we treated WT mice with an individual antibiotic or paired antibiotics in drinking water.The results showed that treatment with ampicillin alone or metronidazole alone attenuated OVA-induced asthma,whereas treatment with vancomycin alone or neomycin alone aggravated asthma,suggesting that,ampicillin-or metronidazole-sensitive bacteria might function to maintain NLRP3/IL-1β signaling,whereas vancomycin-or neomycin-sensitive bacteria might function to activate TLR signaling.Consistent with our hypotheses,we found that treatment with metronidazole alone down-regulated expression of NLRP3 and 1L-1β in pMφ significantly.However,unexpectedly,ampicillin treatment alone had no influence,indicated that commensal bacteria might influence other signaling to influence OVA-induced asthma.Treatment with vancomycin alone displayed the same effect as metronidazole treatment,which contradicted our speculation.Further studies showed that treatment with vancomycin alone also down-regulated TLR expression in pMcp,which could be why treatment with vancomycin alone continued to aggravate asthma even though NLRP3/IL-1β signaling had been down-regulated.In conclusion,our study found that NLRP3/IL-1β signaling and TLR signaling are regulated by different bacterial components.