Alveolar Macrophage Function Research Articles

Vitamin D3-vitamin D receptor axis suppresses pulmonary emphysema by maintaining alveolar macrophage homeostasis and function.

BackgroundChronic obstructive pulmonary disease (COPD) is characterized by emphysema and/or obstructive bronchiolitis. Deficiency in vitamin D3 (VD3), which regulates gene expression through binding to vitamin D receptor (VDR), is associated with high risks of COPD susceptibility. Alveolar macrophages (AM), which are generated during early ontogeny and maintained in alveoli by self-renewal in response to cytokine GM-CSF, are positively correlated with severity of emphysema. However, whether and how VD3, VDR and AM interact to contribute to COPD pathogenesis at the molecular and cellular levels are largely unknown.MethodsWe used systems biology approaches to analyze gene expression in mouse macrophages from different tissues to identify key transcription factors (TF) for AM and infer COPD disease genes. We used RNA-seq and ChIP-seq to identify genes that are regulated by VD3 in AM. We used VDR-deficient (Vdr−/−) mice to investigate the role of VD3-VDR axis in the pathogenesis of COPD and characterized the transcriptional and functional alterations of Vdr−/− AM.FindingsWe find that VDR is a key TF for AM and a COPD disease gene. VDR is highly expressed in AM and in response to VD3 inhibits GM-CSF-induced AM proliferation. In Vdr−/− AM, genes involved in proliferation and immune response are upregulated. Consistently, Vdr−/− mice progressively accumulate AM and concomitantly develop emphysema without apparent infiltration of immune cells into the lung tissue. Intratracheal transfer of Vdr−/− AM into wildtype mice readily induces emphysema. The production of reactive oxygen species at basal level and in response to heme or lipopolysaccharide is elevated in Vdr−/− AM and suppressed by VD3 in wildtype AM.InterpretationThese results show that the VD3-VDR axis is critical to counteract GM-CSF-induced AM proliferation and defect in this regulation leads to altered AM homeostasis and function. Our findings identify that VD3 deficiency contributes to emphysema by altering AM function without contributing to bronchiolitis. Our findings also suggest possibilities of modulating the VD3-VDR axis for inhibiting emphysema in COPD patients.

The effects of aging and exercise on lung mechanics, surfactant and alveolar macrophages

Purpose: Advancing age leads to changes to the respiratory system associated with increased susceptibility to lung diseases, and exercise may counteract this effect. To explore the underlying processes, we investigated the effects of aging and exercise on lung mechanics, alveolar macrophage function, and surfactant pools and activity, in mice. It was hypothesized that aging would impact lung mechanics, macrophage polarization, and the status of the surfactant system, and that these changes would be mitigated by exercise. Methods: Male C57BL/6 mice were housed from 2-3 to 22 months, for the aged group, or until 4 months of age for young mice. Mice in both groups were randomized to voluntarily running exercise or to non-exercise, for a 2-month period. Mice were euthanized and lung mechanics were analyzed using a flexiVent ventilator. Subsequently, the lungs were lavaged to obtain pulmonary surfactant and alveolar macrophages. Pulmonary surfactant was analyzed for pool sizes and activity whereas alveolar macrophages were examined for response to pro and anti-inflammatory stimuli. Results: Changes in lung mechanics, such as increased compliance and decreased airway resistance, were associated with aging but were not affected by exercise. The quantity as well as the biophysical activity of the pulmonary surfactant system was unaffected by either aging or exercise. More alveolar macrophages were recovered from exercising aged mice compared to both the young and non-exercising groups. Macrophages in this aged exercise group were more responsive to an anti-inflammatory stimulus. Conclusions: Our data supports previous literature that suggest the development of emphysema-like alterations to lung mechanics with aging. This effect was independent of exercise. Our data also indicates that surfactant is unaffected by aging and exercise. Alveolar macrophage properties and numbers were affected by exercise in the aging lung and may represent the main, potentially beneficial, effect of exercise on the pulmonary system.

Hydrogen gas inhalation enhances alveolar macrophage phagocytosis in an ovalbumin-induced asthma model

BackgroundMaintaining an airway clear of bacteria, foreign particles and apoptotic cells by alveolar macrophages is very essential for lung homeostasis. In asthma, the phagocytic capacity of alveolar macrophages is significantly reduced, which is thought to be associated with increased oxidative stress. Hydrogen (H2) has been shown to exert potent antioxidant and anti-inflammatory effects, yet its effects on phagocytosis of alveolar macrophages are unknown. This study is aimed to evaluate the beneficial effects of hydrogen gas inhalation on alveolar macrophage phagocytosis in an ovalbumin (OVA)-induced murine asthma model. MethodsFemale C57BL/6 mice were intraperitoneally sensitized with OVA before they were subject to airway challenge with aerosolized OVA. Hydrogen gas was delivered to the mice through inhalation twice a day (2 h once) for 7 consecutive days. Phagocytic function of alveolar macrophages isolated from bronchoalveolar lavage fluid was assessed by fluorescence-labeled Escherichia coli as well as flow cytometry. ResultsAlveolar macrophages isolated from OVA-induced asthmatic mice showed decreased phagocytic capacity to Escherichia coli when compared with those of control mice. Defective phagocytosis in asthmatic mice was reversed by hydrogen gas inhalation. Hydrogen gas inhalation significantly alleviated OVA-induced airway hyperresponsiveness, inflammation and goblet cell hyperplasia, diminished TH2 response and decreased IL-4 as well as IgE levels, reduced malondialdehyde (MDA) production and increased superoxide dismutase (SOD) activity. Concomitantly, hydrogen gas inhalation inhibited NF-κB activation and markedly activated Nrf2 pathway in OVA-induced asthmatic mice. ConclusionsOur findings demonstrated that hydrogen gas inhalation enhanced alveolar macrophage phagocytosis in OVA-induced asthmatic mice, which may be associated with the antioxidant effects of hydrogen gas and the activation of the Nrf2 pathway.

Histone Deacetylase 3 regulates the ontogeny and maintenance of tissue-resident macrophages

Abstract Tissue-resident macrophages (TRMs) are essential for tissue homeostasis, immune-surveillance, and resolution of inflammation, and contribute to the pathogenesis of cancer, autoimmune, and inflammatory diseases. Most adult TRMs are derived from embryonic yolk sac erythro-myeloid progenitors and self-maintain independently from adult hematopoiesis. A few genes have been recently identified to be required for TRM tissue-specific development. However, the master regulator that centrally controls TRM ontogeny remains a mystery. Using a fate-mapping strategy, here we report that conditional deletion of histone deacetylase 3 (HDAC3) in mice essentially abrogates epidermal Langerhans cells and TRMs in dermis, lung, kidney, and heart at embryonic stage and after birth. In contrast, HDAC3 deletion promotes liver TRM ontogeny, while without significant effects on brain TRM microglia. Using lung TRM alveolar macrophage (AM) as a model, we further indicate that HDAC3 controls metabolism and cell death, at least by directly targeting PPAR-γ during AM embryonic development. Moreover, single-cell RNA sequencing identifies four distinct AM sub-clusters and lack of HDAC3 dysregulates AM functions in a cluster-specific manner. Overall, this is the first study to demonstrate that HDAC3 serves as a key epigenetic regulator of embryonic TRM ontogeny and maintenance in a spatial- and tissue-specific manner. Not only do these findings provide a new conceptual framework for our understanding of epigenetic regulation in TRM ontogeny and homeostasis, they also shed light on HDAC3 as a potential therapeutic target for TRM-based intervention in cancer and autoimmune diseases.

Distinctive Effects of GM-CSF and M-CSF on Proliferation and Polarization of Two Major Pulmonary Macrophage Populations.

GM-CSF is required for alveolar macrophage (AM) development shortly after birth and for maintenance of AM functions throughout life, whereas M-CSF is broadly important for macrophage differentiation and self-renewal. However, the comparative actions of GM-CSF and M-CSF on AMs are incompletely understood. Interstitial macrophages (IMs) constitute a second major pulmonary macrophage population. However, unlike AMs, IM responses to CSFs are largely unknown. Proliferation, phenotypic identity, and M1/M2 polarization are important attributes of all macrophage populations, and in this study, we compared their modulation by GM-CSF and M-CSF in murine primary AMs and IMs. CSFs increased the proliferation capacity and upregulated antiapoptotic gene expression in AMs but not IMs. GM-CSF, but not M-CSF, reinforced the cellular identity, as identified by surface markers, of both cell types. GM-CSF, but not M-CSF, increased the expression of both M1 and M2 markers exclusively in AMs. Finally, CSFs enhanced the IFN-γ- and IL-4-induced polarization ability of AMs but not IMs. These first (to our knowledge) data comparing effects on the two pulmonary macrophage populations demonstrate that the activating actions of GM-CSF and M-CSF on primary AMs are not conserved in primary IMs.

Alveolar macrophage maturation is required for efficient killing of GBS in the lung

Abstract Group B Streptococcus (GBS) is the most common pathogen in neonates but rarely causes disease in adults. GBS causes pneumonia in newborns, suggesting a defect in lung innate immunity at birth. Using a novel neonatal mouse model of GBS pneumonia, we showed that newborn mice have reduced killing of GBS in the lung and more severe and persistent lung injury. Here we test the hypothesis that defects in alveolar macrophage (AM) maturation in newborns leads to GBS susceptibility. Clodronate depletion of AM from adult mice prevented GBS killing, supporting the requirement of AM for GBS protection. AM maturation and function requires GM-CSF, which is abundantly expressed in the lung. While AM maturation occurred on day 4 in WT mice, GM-CSF knockout mice (Csf2−/−) failed to develop AM. Consistent with the role of AM in GBS immunity, neonatal Csf2−/− mice had reduced GBS killing and lung macrophages failed to phagocytose GBS. Csf2−/− macrophages also had reduced expression of multiple Siglecs, cell surface receptors implicated in protection against encapsulated bacteria. Newborn lung macrophages expressed primarily Siglec-E (SigE), whereas expression of other Siglecs were only detected in more mature AM. We used SigE−/− mice to test its role in neonatal GBS pneumonia. GBS killing in SigE−/− neonates was similar to WT. However SigE−/− macrophages appeared to phagocytose GBS more efficiently in vivo, suggesting GBS may employ SigE to avoid detection by neonatal macrophages. Our data show that mature alveolar macrophages are required for protection against pulmonary GBS infection and potentially implicate the developmental regulation of macrophage Siglec expression in neonatal lung immunity.

PPAR-γ in Macrophages Limits Pulmonary Inflammation and Promotes Host Recovery following Respiratory Viral Infection.

Alveolar macrophages (AM) play pivotal roles in modulating host defense, pulmonary inflammation, and tissue injury following respiratory viral infections. However, the transcriptional regulation of AM function during respiratory viral infections is still largely undefined. Here we have screened the expression of 84 transcription factors in AM in response to influenza A virus (IAV) infection. We found that the transcription factor PPAR-γ was downregulated following IAV infection in AM through type I interferon (IFN)-dependent signaling. PPAR-γ expression in AM was critical for the suppression of exaggerated antiviral and inflammatory responses of AM following IAV and respiratory syncytial virus (RSV) infections. Myeloid PPAR-γ deficiency resulted in enhanced host morbidity and increased pulmonary inflammation following both IAV and RSV infections, suggesting that macrophage PPAR-γ is vital for restricting severe host disease development. Using approaches to selectively deplete recruiting monocytes, we demonstrate that PPAR-γ expression in resident AM is likely important in regulating host disease development. Furthermore, we show that PPAR-γ was critical for the expression of wound healing genes in AM. As such, myeloid PPAR-γ deficiency resulted in impaired inflammation resolution and defective tissue repair following IAV infection. Our data suggest a critical role of PPAR-γ expression in lung macrophages in the modulation of pulmonary inflammation, the development of acute host diseases, and the proper restoration of tissue homeostasis following respiratory viral infections.IMPORTANCE Respiratory viral infections, like IAV and respiratory syncytial virus (RSV) infections, impose great challenges to public health. Alveolar macrophages (AM) are lung-resident immune cells that play important roles in protecting the host against IAV and RSV infections. However, the underlying molecular mechanisms by which AM modulate host inflammation, disease development, and tissue recovery are not very well understood. Here we identify that PPAR-γ expression in AM is crucial to suppress pulmonary inflammation and diseases and to promote fast host recovery from IAV and RSV infections. Our data suggest that targeting macrophage PPAR-γ may be a promising therapeutic option in the future to suppress acute inflammation and simultaneously promote recovery from severe diseases associated with respiratory viral infections.

The effects of aging and exercise on lung mechanics, surfactant and alveolar macrophages

IntroductionIt has been well established that advancing age leads to changes to the respiratory system and an associated susceptibility to lung diseases. Furthermore, exercise may counteract this pulmonary disease susceptibility. To start to investigate the underlying processes of these effects we explored the baseline characteristics due to aging and exercise on lung mechanics and on two important components of a healthy pulmonary environment, alveolar macrophage function and the pulmonary surfactant system. It was hypothesized that aging would impact lung mechanics, macrophage polarization and the status of the surfactant system, and that these changes would be mitigated by exercise.Experimental approach: Male C57BL/6 mice were housed from 2–3 to 22 months, for the aged group, or until 4 months of age for young control mice. Mice in both the young and aged groups were randomized to either voluntarily running exercise or non‐exercise for a 2 month period. Mice were euthanized and lung mechanics were analyzed using a Flexivent ventilator. Subsequently, the lungs were lavaged to obtain pulmonary surfactant and alveolar macrophages. Pulmonary surfactant was analyzed for surfactant pool sizes and activity whereas alveolar macrophages were examined for a response to pro‐and anti‐inflammatory stimuli.ResultsChanges in lung mechanics, such as increased compliance and decreased airway resistance, were observed in the aged cohorts. These changes were not affected by exercise.The quantity as well as the biophysical activity of the pulmonary surfactant system was unaffected by either aging or exercise. More alveolar macrophages were recovered from exercising aged mice as compared to both the young and non‐exercising groups.Support or Funding InformationWestern Strategic Support, Lawson Internal Research FundsThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Reduced transient receptor potential vanilloid 2 expression in alveolar macrophages causes COPD in mice through impaired phagocytic activity

BackgroundDefective phagocytosis in alveolar macrophages is associated with chronic obstructive pulmonary disease (COPD). Transient receptor potential cation channel subfamily V member 2 (TRPV2), a type of nonselective cation channel pertinent to diverse physiological functions, regulates macrophage phagocytosis. However, the role of TRPV2 in COPD remains poorly understood. Here, we explored the role of TRPV2 in the development of COPD.MethodsMacrophage TRPV2 expression and phagocytosis function were measured in MH-S cells (a murine alveolar macrophage cell line) and a cigarette smoke exposure mouse model.ResultsTRPV2 expression and phagocytosis function were reduced when MH-S cells were exposed to cigarette smoke extract (CSE). TRPV2 knockdown by siRNA decreased phagocytosis in MH-S cells. Consistently, TRPV2 expression was reduced in alveolar macrophages prepared from bronchoalveolar lavage samples of mice which were exposed to cigarette smoke for 2 months. In addition, the alveolar space was progressively enlarged during development in TRPV2 knockout (TRPV2KO) mice. Moreover, exposure to cigarette smoke for 2 months significantly induced alveolar space enlargement in TRPV2KO mice, but not in wild-type (WT) mice. The phagocytic function of alveolar macrophages from TRPV2KO mice was reduced, compared with macrophages from WT mice.ConclusionsTRPV2 expression is profoundly downregulated in alveolar macrophages at early time points of cigarette smoke exposure. Reduced TRPV2-mediated phagocytic function renders the lung susceptible to cigarette smoke-induced alveolar space enlargement. TRPV2 may provide a therapeutic target for COPD induced by cigarette smoke.

Persisting Donor Alveolar Macrophages Have Increased Expression of Scavenger Receptor CD206 Compared to Graft-Infiltrating Recipient-Derived Macrophages Following Lung Transplantation

Purpose Alveolar macrophages play vital roles in attenuating lung injury through immune modulatory effects and improving gas exchange by scavenging cellular debris within the alveoli. A recent analysis of lung biopsy specimens suggests that donor-derived macrophages persist within the allograft after lung transplantation. The phenotypic differences between donor and graft-infiltrating recipient macrophages remains unreported. Methods Single cell suspensions obtained from bronchoalveolar lavage (BAL) of 15 lung transplant recipients at various times following transplantation were analyzed by flow cytometry. Macrophages were defined as CD45+CD3-CD19-CD64+CD14+CD11chisinglets. Cell origin (donor versus recipient) was identified by staining for human leukocyte antigen discrepancies. Paired t test was used to compare cell surface marker expression between donor and recipient macrophages. Results The proportion of donor macrophages within the BAL was variable across patients but did not vary over the time following transplantation (p = 0.5). The median percentage of donor macrophages was 10.5% (range 1-64). Donor macrophages had increased expression of CD206 (Figure: Rep. flow cytometry plot from BAL one month following transplantation and cumulative data from 10 participants, p = 0.004), with trends in increased expression of HLADR (p = 0.07), and no difference in CD163 expression (p = 0.17). Conclusion The scavenger function of alveolar macrophages is vital in maintaining gas exchange in the setting of severe influenza infection. Following lung transplantation, persisting donor-derived alveolar macrophages have significantly increased expression of the scavenger receptor CD206 when compared to graft-infiltrating recipient-derived macrophages. Further investigation is warranted to identify whether this has implications for graft function in the perioperative setting.

Alveolar Macrophages Treated With Bacillus subtilis Spore Protect Mice Infected With Respiratory Syncytial Virus A2.

Respiratory syncytial virus (RSV) is a major pathogen that infects lower respiratory tract and causes a common respiratory disease. Despite serious pathological consequences with this virus, effective treatments for controlling RSV infection remain unsolved, along with poor innate immune responses induced at the initial stage of RSV infection. Such a poor innate defense mechanism against RSV leads us to study the role of alveolar macrophage (AM) that is one of the primary innate immune cell types in the respiratory tract and may contribute to protective responses against RSV infection. As an effective strategy for enhancing anti-viral function of AM, this study suggests the intranasal administration of Bacillus subtilis spore which induces expansion of AM in the lung with activation and enhanced production of inflammatory cytokines along with several genes associated with M1 macrophage differentiation. Such effect by spore on AM was largely dependent on TLR-MyD88 signaling and, most importantly, resulted in a profound reduction of viral titers and pathological lung injury upon RSV infection. Taken together, our results suggest a protective role of AM in RSV infection and its functional modulation by B. subtilis spore, which may be a useful and potential therapeutic approach against RSV.

Alveolar Macrophage Phagocytosis and Bacteria Clearance in Mice.

Alveolar macrophages (AMs) guard the alveolar space of the lung. Phagocytosis by AMs plays a critical role in the defense against invading pathogens, the removal of dead cells or foreign particles, and in the resolution of inflammatory responses and tissue remodeling, processes that are mediated by various surface receptors of the AMs. Here, we report methods for the analysis of the phagocytic function of AMs using in vitro and in vivo assays and experimental strategies to differentiate between the pattern recognition receptor-, complement receptor-, and Fc gamma receptor-mediated phagocytosis. Finally, we discuss a method to establish and characterize a P. aeruginosa pneumonia model in mice to assess bacterial clearance in vivo. These assays represent the most common methods to evaluate AM functions and can also be used to study macrophage function and bacterial clearance in other organs.

GTS-21 Reduces Inflammation in Acute Lung Injury by Regulating M1 Polarization and Function of Alveolar Macrophages.

Acute lung injury (ALI) is a severe outcome of sepsis. Alveolar macrophages (AMs) play key roles in defense, resolution in ALI. The polarization of AMs is dependent on micro environmental stimuli and might influence the progression of ALI. Gainesville Tokushima scientists (GTS)-21, a selective α7 nicotinic acetylcholine receptor agonist of the cholinergic anti-inflammatory pathway (CAP), has recently been established to be promising in the treatment of ALI. However, the molecular mechanism underlying the GTS-21-mediated suppression of inflammatory responses has been explored only partially. In this study, we examined the relation between GTS-21 and AM polarization in ALI. The adoptive transfer of M1 (classically activated) and M2 (alternatively activated)-polarized AMs was performed to AM-depleted ALI mice, along with the administration of GTS-21 in a murine model of lipopolysaccharide (LPS)-induced ALI and in isolated AMs that had been stimulated by LPS in vitro. The adoptive transfer of M1-polarized AMs aggravated the inflammatory response in the lung in contrast to the adoptive transfer of M2-polarized AMs. GTS-21 protected the lung from the effect of LPS, preventing injury and decreasing the number of AMs, AM-related pro-inflammatory cytokine levels, high mobility group box 1 expression levels in AMs. In addition, GTS-21 significantly diminished the number of M1-polarized AM and increased the number of M2-polarized AM, by flow cytometry, RT-PCR, enzyme-linked immunosorbent assay, and the Arg1 and iNOS activity assays. The GTS-21 substantially ameliorates LPS-induced ALI. This protection is predominantly associated with the inhibition of pulmonary AM M1 polarization and alteration in AM function.

Upregulation of Chitinase 1 in Alveolar Macrophages of HIV-Infected Smokers.

Recent studies suggest that HIV infection is an independent risk factor for the development of chronic obstructive pulmonary disease (COPD). We hypothesized that HIV infection and cigarette smoking synergize to alter the function of alveolar macrophages (AMs). To test this hypothesis, global transcriptome analysis was performed on purified AMs from 20 individuals split evenly between HIV-uninfected nonsmokers and smokers and untreated HIV-infected nonsmokers and smokers. Differential expression analysis identified 143 genes significantly altered by the combination of HIV infection and smoking. Of the differentially expressed genes, chitinase 1 (CHIT1) and cytochrome P450 family 1 subfamily B member 1 (CYP1B1), both previously associated with COPD, were among the most upregulated genes (5- and 26-fold, respectively) in the untreated HIV-infected smoker cohort compared with HIV-uninfected nonsmokers. Expression of CHIT1 and CYP1B1 correlated with the expression of genes involved in extracellular matrix organization, oxidative stress, immune response, and cell death. Using time-of-flight mass cytometry to characterize AMs, a significantly decreased expression of CD163, an M2 marker, was seen in HIV-infected subjects, and CD163 inversely correlated with CYP1B1 expression in AMs. CHIT1 protein levels were significantly upregulated in bronchoalveolar lavage fluid from HIV-infected smokers, and increased CHIT1 levels negatively correlated with lung function measurements. Overall, these findings raise the possibility that elevated CHIT1 and CYP1B1 are early indicators of COPD development in HIV-infected smokers that may serve as biomarkers for determining this risk.

Alveolar macrophage functions during the transition phase to active immunity in calves.

The first 3 to 6 mo of the life of calves is the period during which active immunity is established. During this period, greater morbidity and mortality is caused by bronchopneumonia because of the immaturity of the pulmonary immune system or the exaggerated cytotoxic response at subsequent infection. The aim of this study was to examine the maturity of the immune system during this phase of activation of acquired immunity in calves. For this purpose, the functions of phagocytosis and the reactive oxygen species (ROS) of alveolar macrophages CD14+ were evaluated. Further, the classes of immunoglobulins and the cytokines implicated in lymphocyte response patterns Th1 and Th2 in 10 healthy Holstein calves were quantified. Samples were taken from calves every 15 d, from the third to the sixth month of life. The alveolar macrophage CD14+ functions increased progressively until 150 d of age (phagocytosis, P = 0.02, ROS, P = 0.05), IgG1 and IgG2 isotype secretion reached an equilibrium, and the cytokine profiles were compatible with the Th1 response. At 165 d of age, there was a decrease in cellular function (phagocytosis P = 0.02, ROS P = 0.04) and an increase in IgG1 titers (P = 0.005) and IL-10 mRNA expression (P = 0.09). At 180 d of life, we observed an IgG1 and IgG2 secretion balance, a decrease in IL-10 mRNA expression, and an increase in IL-12 mRNA (P = 0.04) and tumor necrosis factor (TNF)-α mRNA expressions (P = 0.0003) and alveolar macrophage oxidative metabolism were observed. These results indicate that the calves had an active immune response that was distinctive for the age group. The CD14+ response is more reactive at 150 d. A regulatory and/or humoral response begins at 165 d of life as the equilibrium of Th1 and Th2 profiles is reached at 180 d of life. This may be clinically relevant for the development of specific therapies and prophylactic measures for bronchopneumonia in calves at 135 to 180 d of life.

Vitamin D <sub>3</sub>-Vitamin D Receptor Axis Suppresses Pulmonary Emphysema by Maintaining Alveolar Macrophage Homeostasis and Function

Chronic obstructive pulmonary disease (COPD) is a complex disease with poorly understood molecular and cellular mechanisms. By using systems biology approaches and transcriptional analysis, we identify that vitamin D receptor (VDR) is a key transcription factor for regulating alveolar macrophage (AM) homeostasis and function. VDR is highly expressed in AM and in response to vitamin D3 mediates inhibition of GM-CSF-induced AM proliferation. VDR-deficient (Vdr-/-) mice progressively accumulate AM and concomitantly develop emphysema without apparent infiltration of immune cells into the lung tissue. Intratracheal transfer of Vdr-/- AM into wildtype mice readily induces emphysema. The production of reactive oxygen species (ROS) at basal level and in response to heme or lipopolysaccharide is elevated in Vdr-/- AM and suppressed by vitamin D3 in wildtype AM. These results show that vitamin D3-VDR axis regulates AM homeostasis and function and defect in this regulation results in emphysema. Our findings identify the molecular and cellular mechanisms by which vitamin D3, VDR, and AM interact to contribute to specific COPD pathology and provide new insights into modulating this pathway for inhibiting emphysema in COPD patients. Funding Statement: This work was supported in part by National Institutes of Health Grants AI69208 and CA197605, Ivan R. Cottrell Professorship and Research Fund, and the Koch Institute Support (core) Grant P30-CA14051 from the National Cancer Institute. Declaration of Interests: The authors declare no competing interests. Ethics Approval Statement: All protocols were approved by the Ethics Committee of The First Affiliated Hospital, Jilin University. All experiments were performed in accordance with the relevant guidelines and regulations. In addition, written informed consent was obtained from each subject.

Mesenchymal Stromal Cells Modulate PAF-stimulated Equine Alveolar Macrophages

Abstract Platelet-activating factor (PAF) is a potent proinflammatory mediator that is produced in increased amounts in the lungs of asthmatic humans and horses. The present pilot study, shows that mesenchymal stromal cells can modulate alveolar macrophage function in asthma, interfering in the activity of PAF, being another potential pathway for mesenchymal stromal cells benefits in asthma.

Pregnancy Induces a Steady-State Shift in Alveolar Macrophage M1/M2 Phenotype That Is Associated With a Heightened Severity of Influenza Virus Infection: Mechanistic Insight Using Mouse Models.

Influenza virus infection during pregnancy is associated with enhanced disease severity. However, the underlying mechanisms are still not fully understood. We hypothesized that normal alveolar macrophage (AM) functions, which are central to maintaining lung immune homeostasis, are altered during pregnancy and that this dysregulation contributes to the increased inflammatory response to influenza virus infection. Time-mated BALB/c mice were infected with a low dose of H1N1 influenza A virus at gestation day 9.5. Inflammatory cells in bronchoalveolar lavage (BAL) fluid were assessed by flow cytometry. Our findings confirm previous reports of increased severity of influenza virus infection in pregnant mice. The heightened inflammatory response detected in BAL fluid from infected pregnant mice was characterized by neutrophil-rich inflammation with concomitantly reduced numbers of AM, which were slower to return to baseline counts, compared with nonpregnant infected mice. The increased infection severity and inflammatory responses to influenza during pregnancy were associated with a pregnancy-induced shift in AM phenotype at homeostatic baseline, from the M1 (ie, classical activation) state toward the M2 (ie, alternative activation) state, as evidence by increased expression of CD301 and reduced levels of CCR7. These results show that pregnancy is associated with an alternatively activated phenotype of AM before infection, which may contribute to heightened disease severity.

A mouse model of binge alcohol consumption and Burkholderia infection.

BackgroundBinge drinking, an increasingly common form of alcohol consumption, is associated with increased mortality and morbidity; yet, its effects on the immune system’s ability to defend against infectious agents are poorly understood. Burkholderia pseudomallei, the causative agent of melioidosis can occur in healthy humans, yet binge alcohol use is progressively being recognized as a major risk factor. Although our previous studies demonstrated that binge alcohol exposure results in reduced alveolar macrophage function and increased Burkholderia virulence in vitro, no experimental studies have investigated the outcomes of binge alcohol on Burkholderia spp. infection in vivo.Principal findingsIn this study, we used the close genetic relatives of B. pseudomallei, B. thailandensis E264 and B. vietnamiensis, as useful BSL-2 model systems. Eight-week-old female C57BL/6 mice were administered alcohol comparable to human binge drinking episodes (4.4 g/kg) or PBS intraperitoneally 30 min before a non-lethal intranasal infection. In an initial B. thailandensis infection (3 x 105), bacteria accumulated in the lungs and disseminated to the spleen in alcohol administered mice only, compared with PBS treated mice at 24 h PI. The greatest bacterial load occurred with B. vietnamiensis (1 x 106) in lungs, spleen, and brain tissue by 72 h PI. Pulmonary cytokine expression (TNF-α, GM-CSF) decreased, while splenic cytokine (IL-10) increased in binge drunk mice. Increased lung and brain permeability was observed as early as 2 h post alcohol administration in vivo. Trans-epithelial electrical resistance (TEER) was significantly decreased, while intracellular invasion of non-phagocytic cells increased with 0.2% v/v alcohol exposure in vitro.ConclusionsOur results indicate that a single binge alcohol dose suppressed innate immune functions and increased the ability of less virulent Burkholderia strains to disseminate through increased barrier permeability and intracellular invasion of non-phagocytic cells.

Effect of High Glucose on Human Alveolar Macrophage Phenotype and Phagocytosis of Mycobacteria.

Diabetes mellitus (DBM) reduces immunological activity and increases susceptibility to various infections, including tuberculosis (TB). Human alveolar macrophage (hAM) functions are altered in DBM. To mimic hyperglycemic conditions in the lung alveolus, we co-cultured a hAM cell line (Daisy cell line) with human umbilical vein endothelial cells for 48h in the presence of culture media alone, normal glucose (5mM), and high glucose (22mM). Using flow cytometry, immunophenotype characterization included cell surface markers CD 11c, CD14, CD16, CD86, CD163, CD169, CD206, CX3CR-1, CSF-1R, and matrix metalloproteinase-9 (MMP9). Phagocytic function was measured by immunofluorescence microscopy at 24h after inoculation of cells with GFP-expressing Mycobacterium smegmatis. Direct exposure of AMs to high glucose and exposure in the co-culture system yield different results for the same phenotypic markers. MMP9 expression was increased under both conditions. CD169 and CX3CR1 expressions were decreased when AMs were exposed directly to high glucose but increased under co-culture. Immunofluorescence assay revealed that phagocytosis decreased in AMs when directly exposed to increased glucose levels from 2.5mM to normal glucose (5mM), yet AMs under co-culture did not show decreased phagocytosis until concentrations were raised to 25mM. Alteration in the expression of certain receptors may contribute to defective sentinel function of AMs, promoting susceptibility to TB in a diabetic host. Variability in cell surface marker expression under direct glucose exposure compared to exposure via co-culture reveals that cell signaling between endothelial cells and AMs may play a crucial role in the phenotypic expression of AMs.