Project description:Pulmonary innate immunity is required for host defense; however, excessive neutrophil inflammation can cause life-threatening acute lung injury. B lymphocytes can be regulatory, yet little is known about peripheral transitional IgM+ B cells in terms of regulatory properties. Using single cell RNA sequencing, we discovered eight IgM+ B cell subsets with unique gene regulatory networks in the lung circulation dominated by transitional type 1 (T1B) and 2 (T2B) B cells. Lung intravital confocal microscopy revealed that T2B marginate in the pulmonary capillaries via CD49e and required CXCL13 and CXCR5. During lung inflammation, marginated T2B dampened excessive neutrophil vascular inflammation via the specialized proresolving molecule lipoxin A4 (LXA4). Exogenous CXCL13 dampened excessive neutrophilic inflammation by increasing marginated B cells and LXA4 recapitulated neutrophil regulation in B-cell deficient mice during inflammation and fungal pneumonia. Thus, the lung microvasculature is enriched in multiple IgM+ B cell subsets with marginating capillary T2B that dampen neutrophil responses.
Project description:Identifying host factors that contribute to pneumonia incidence and severity are of utmost importance to guiding the development of more effective therapies. Lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) is a scavenger receptor known to promote vascular injury and inflammation, but it is unknown whether and how LOX-1 functions in the lung. Here, we provide evidence of substantial accumulation of LOX-1 in the lungs of ARDS patients and in mice with pneumonia. Unlike previously described injurious contributions of LOX-1, we found that LOX-1 is uniquely protective in the pulmonary airspaces, limiting proteinaceous edema and inflammation. We also identified alveolar macrophages and recruited neutrophils as two prominent sites of LOX-1 expression in the lungs, whereby macrophages are capable of further induction during pneumonia and neutrophils exhibit a rapid, but heterogenous elevation of LOX-1 in the infected lung. Blockade of LOX-1 led to dysregulated immune signaling in alveolar macrophages, marked by alterations in activation markers and a concomitant elevation of inflammatory gene networks. However, bone marrow chimeras also suggested a prominent role for neutrophils in LOX-1-mediated lung protection, further supported by LOX-1+ neutrophils exhibiting transcriptional changes consistent with reparative processes. Taken together, this work establishes LOX-1 as a tissue-protective factor in the lungs during pneumonia, possibly mediated by its influence on immune signaling in alveolar macrophages (AMs) and LOX-1+ airspace neutrophils.
Project description:These studies profiled the expression of mRNAs and microRNAs (miRs) in lung neutrophils in WT mice during S. pneumoniae pneumonia and performed in depth in silico analyses. Lung neutrophils were isolated 24 hours after intratracheal instillation of PBS or S. pneumoniae, and mRNAs and miRs differentially expressed (DE) between S. pneumoniae- and PBS-treated samples were identified using microarrays.
Project description:The transcription factor Nrf2 (gene symbol Nfe2l2) regulates the transcriptional response to oxidative stress and plays a critical protective role in the lungs. These studies tested the role of Nrf2 during pneumonia induced by Streptococcus pneumoniae (SP) at 24 hours in mice and identified Nrf2-dependent genes and pathways in lung tissue and in recruited neutrophils.
Project description:These studies profiled the expression of mRNAs and microRNAs (miRs) in lung neutrophils in WT mice during S. pneumoniae pneumonia and performed in depth in silico analyses. Lung neutrophils were isolated 24 hours after intratracheal instillation of PBS or S. pneumoniae, and mRNAs and miRs differentially expressed (DE) between S. pneumoniae- and PBS-treated samples were identified using microarrays.
Project description:Host-directed therapy, using nasal administration of the Toll-like receptor 5 agonist flagellin in combination with antibiotics, has demonstrated effectiveness against pneumococcal pneumonia. In this study, we investigated the immune mechanisms underlying the protective effects. Transcriptomic analysis of lung tissue during infection revealed that flagellin not only enhanced pathways associated with myeloid cell infiltration into the airways and antimicrobial functions, but also promoted the early and transient mobilization of neutrophils and inflammatory monocytes. Neutrophils were identified as crucial for the protective effects of flagellin. The adjunct activity of flagellin correlated with the increased recruitment of neutrophils into airways, their localization at the periphery of bronchi, alveoli, and lung vessels, along with alterations in phagocytic activity. Lastly, single-cell analysis highlighted that the treatment stimulated the recruitment of specific neutrophil subsets capable of enhancing antibacterial effectiveness. In conclusion, this study highlights neutrophils as an appealing target for host-directed therapy in infection.
Project description:Pneumonia caused by Streptococcus pneumoniae is a leading cause of death worldwide, and bacterial resistance to antimicrobial drugs has become a major issue. A growing body of evidence indicates that the successful treatment of bacterial infections results from synergy between antibiotic-mediated direct antibacterial activity and the host’s immune defenses. However, the mechanisms underlying the protective immune responses induced by amoxicillin (a β-lactam antibiotic used as the first-line treatment of S. pneumoniae infections) have not been characterized. A better understanding of amoxicillin’s effects on host-pathogen interactions might facilitate the development of other treatment options. Given the crucial role of neutrophils in the control of S. pneumoniae infections, we decided to investigate amoxicillin’s impact on neutrophil development in a mouse model of pneumococcal superinfection. Although a single therapeutic dose of amoxicillin prevented local and systemic inflammatory responses, it did not impair the emergency granulopoiesis triggered in the bone marrow by S. pneumoniae. Importantly, treatment of pneumonia with amoxicillin was associated with a greater mature neutrophil count in the bone marrow; these neutrophils had specific transcriptomic and proteomic profiles. Furthermore, amoxicillin-conditioned, mature neutrophils in the bone marrow had a less activated phenotype and might be rapidly mobilized in peripheral tissues in response to systemic inflammation. Thus, by revealing a novel effect of amoxicillin on the development and functions of bone marrow neutrophils during Streptococcus pneumoniae pneumonia, our findings provide new insights into the impact of amoxicillin treatment on host immune responses.
Project description:Vaccination reduces morbidity and mortality from pneumonia but its effect on the tissue-level response to infection is still poorly understood. We evaluated pneumonia disease progression, acute phase response and lung gene expression profiles in mice inoculated intranasally with virulent gram-positive Streptococcus pneumoniae serotype (ST) 3, with and without prior immunization with pneumococcal polysaccharide ST 3 (PPS3), or co-immunization with PPS3 and with a low dose of lipopolysaccharide (LPS). Pneumonia severity was assessed in the acute phase, 5, 12, 24 and 48 h post-inoculation (p.i.) and the resolution phase of 7 days p.i. Primary PPS3 specific antibody production was upregulated and IgM binding to pneumococci increased in PPS3-immunized mice. Immunizations with PPS3 or PPS3 + LPS decreased bacterial recovery the lung and blood at 24 and 48 h and increased survival. Microarray analysis of whole lung RNA revealed significant changes in the acute phase protein serum amyloid A (SAA) between noninfected and infected mice, which were attenuated by immunization. SAA transcripts were higher in the liver and lungs of infected controls, and SAA protein was elevated in serum, but decreased in PPS3-immunized mice. Thus, during a virulent pneumonia infection, prior immunization with PPS3 in an IgM-dependent manner as well as co-immunization with PPS3 + LPS attenuated pneumonia severity and promoted resolution of infection, concomitant with significant regulation of cytokine gene expression in the lungs, and acute phase proteins in the lungs, liver and serum. Each lung RNA sample represented an individual mouse, creating biological repeats for each treatment. In-vivo treatments were as follows: non-infected lung (vehicle-immunized) control (n=5), infected lung (vehicle-immunized) control at 48 hr post-inoculation (n=5), PPS3 and LPS co-immunized lung at 48 hr post-inoculation (n=4) and PPS3 and LPS co-immunized lung at 7 days post-inoculation (n=4).
Project description:Vaccination reduces morbidity and mortality from pneumonia but its effect on the tissue-level response to infection is still poorly understood. We evaluated pneumonia disease progression, acute phase response and lung gene expression profiles in mice inoculated intranasally with virulent gram-positive Streptococcus pneumoniae serotype (ST) 3, with and without prior immunization with pneumococcal polysaccharide ST 3 (PPS3), or co-immunization with PPS3 and with a low dose of lipopolysaccharide (LPS). Pneumonia severity was assessed in the acute phase, 5, 12, 24 and 48 h post-inoculation (p.i.) and the resolution phase of 7 days p.i. Primary PPS3 specific antibody production was upregulated and IgM binding to pneumococci increased in PPS3-immunized mice. Immunizations with PPS3 or PPS3 + LPS decreased bacterial recovery the lung and blood at 24 and 48 h and increased survival. Microarray analysis of whole lung RNA revealed significant changes in the acute phase protein serum amyloid A (SAA) between noninfected and infected mice, which were attenuated by immunization. SAA transcripts were higher in the liver and lungs of infected controls, and SAA protein was elevated in serum, but decreased in PPS3-immunized mice. Thus, during a virulent pneumonia infection, prior immunization with PPS3 in an IgM-dependent manner as well as co-immunization with PPS3 + LPS attenuated pneumonia severity and promoted resolution of infection, concomitant with significant regulation of cytokine gene expression in the lungs, and acute phase proteins in the lungs, liver and serum.