Alveolar Macrophages Research Articles

Augmenting Macrophages Apoptosis Induced by Carnitine Palmitoyl Transferase 1A Inhibition via Acetyl-CoA-Associated Protein Acetylation.

Macrophage apoptosis contributes to acute lung injury (ALI). However, the relationship between cell metabolism and the apoptosis of macrophages remains unclear.In our study, murine alveolar macrophages (MH-S) were stimulated by lipopolysaccharide (LPS) to induce an apoptosis model; cell viability, mitochondrial membrane potential (MMP) and apoptosis rate were determined. TCA metabolites and fatty acids were measured; qPCR and western blot were used to detect gene and protein expressions. The LPS-induced ALI mice model was established, and pathological changes, inflammatory cytokines, and protein acetylation were evaluated.The results showed that LPS exposure impaired cell viability and increased apoptosis of alveolar macrophages (AM) in a concentration-dependent manner. LPS also downregulated the expression of the FAO rate-limiting enzyme carnitine palmitoyl transferase 1A (CPT1A), which was accompanied by suppression of fatty acid oxidation (FAO) and alterations of the fatty acid profile. CPT1A inhibitor etomoxir also promoted cell apoptosis of AM and decreased MMP. Overexpression of CPT1A ameliorated cell apoptosis of AM induced by LPS. Etomoxir and LPS decreased acetyl-CoA levels, and supplementation of acetyl-CoA prevented LPS-induced cell apoptosis. In addition, LPS led to the alteration of acetylated protein profiles. Invivo study, excessive cell apoptosis, decreased expression of proteins related to FAO, and decreased acetyl-CoA levels were detected in ALI animal models. Acetyl-CoA could relieve the apoptosis and inflammation in the lung induced by LPS. These findings suggested the essential role of CPT1A and acetyl-CoA in cell apoptosis of AM induced by LPS.

Alveolar macrophages from persons with HIV mount impaired TNF signaling networks to M. tuberculosis infection

People living with HIV (PLWH) have an increased risk for developing tuberculosis after M. tuberculosis infection, despite anti-retroviral therapy (ART). To delineate the underlying mechanisms, we conducted single cell transcriptomics on bronchoalveolar lavage cells from PLWH on ART and HIV uninfected healthy controls infected with M. tuberculosis ex vivo. We identify an M1-like proinflammatory alveolar macrophage subset that sequentially acquires TNF signaling capacity in controls but not in PLWH. Cell-cell communication analyses reveal interactions between M1-like macrophages and effector memory T cells within TNF superfamily, chemokine, and costimulatory networks in the airways of controls. These interaction networks were lacking in PLWH infected with M. tuberculosis, where anti-inflammatory M2-like alveolar macrophages and T regulatory cells dominated along with dysregulated T cell signatures. Our data support a model in which impaired TNF-TNFR signaling, M2-like alveolar macrophages and aberrant macrophage-T cell crosstalk, lead to ineffective immunity to M. tuberculosis in PLWH on ART.

Acute kidney injury triggers hypoxemia by lung intravascular neutrophil retention that reduces capillary blood flow.

Sterile acute kidney injury (AKI) is common in the clinic and frequently associated with unexplained hypoxemia that does not improve with dialysis. AKI induces remote lung inflammation with neutrophil recruitment in mice and humans, but which cellular cues establish neutrophilic inflammation and how it contributes to hypoxemia is not known. Here we report that AKI induces rapid intravascular neutrophil retention in lung alveolar capillaries without extravasation into tissue or alveoli, causing hypoxemia by reducing lung capillary blood flow in the absence of substantial lung interstitial or alveolar edema. In contrast to direct ischemic lung injury, lung neutrophil recruitment during remote lung inflammation did not require cues from intravascular non-classical monocytes or tissue-resident alveolar macrophages. Instead, lung neutrophil retention depended on neutrophil chemoattractant CXCL2 released by activated classical monocytes. Comparative single-cell RNA-sequencing analysis of direct and remote lung inflammation revealed that alveolar macrophages are highly activated and produce CXCL2 only in direct lung inflammation. Establishing a CXCL2 gradient into the alveolus by intratracheal CXCL2 administration during AKI-induced remote lung inflammation enabled neutrophils to extravasate. We thus discovered important differences in lung neutrophil recruitment in direct versus remote lung inflammation and identified lung capillary neutrophil retention that negatively affects oxygenation by causing a ventilation-perfusion mismatch as a driver of AKI-induced hypoxemia.

VASP Knockdown Ameliorates Lipopolysaccharide-Induced Acute Lung Injury with Inhibition of M1 Macrophage Polarization Through the cGMP-PKG Signaling Pathway.

Alveolar macrophage (AM) polarization plays a pivotal role in the inflammatory response during acute lung injury (ALI). As reported previously, vasodilator-stimulated phosphoprotein (VASP) may function as an anti-inflammatory agent in hepatic tissues. However, the specific role of VASP in ALI-induced macrophage polarization remains unclear. To elucidate the role of VASP in ALI, we established a lipopolysaccharide (LPS)-induced M1 polarization model of MH-S cells. RNA sequencing was performed to identify differentially expressed genes during macrophage polarization. The results revealed significant upregulation of the VASP gene. Subsequently, VASP gene knockdown in the lungs was achieved by intratracheal delivery of VASP-AAV6, and the resulting ALI symptoms and macrophage polarization were assessed. The VASP gene was also knocked down in MH-S cells; these cells were then stimulated with LPS for 24h, and polarization-related markers of macrophages were analyzed. Finally, to validate the involvement of the PKG-VASP signaling pathway, experiments were conducted with a PKG agonist (8-Br-cGMP) and inhibitor (KT5823), and the effects of modulating the PKG-VASP pathway on macrophage polarization were investigated. VASP knockdown notably ameliorated ALI symptoms in these mice with LPS-induced ALI. Additionally, in vitro experiments showed that the PKG-VASP signaling pathway plays a pivotal role in macrophage polarization. VASP knockdown protected mice from LPS-induced ALI by inhibiting M1 polarization, and its protective effects were partially mediated by the cGMP-PKG signaling pathway.

S-nitrosylation-triggered secretion of mycobacterial PknG leads to phosphorylation of SODD to prevent apoptosis of infected macrophages

The tuberculosis-causing agent Mycobacterium tuberculosis (M.tb) establishes its niche inside macrophages by secretion of several virulence factors and engaging many host factors. Mycobacterial infection of macrophages results in a proinflammatory trigger-mediated secretion of TNFα. Protein kinase G (PknG), a Serine/Threonine kinase, is essential for mycobacterial survival within the macrophage. Pathogenic mycobacteria, upon infection, can trigger the secretion of proinflammatory cytokine TNFα, but whether secreted PknG plays any role in TNFα secretion at early stages of infection remains undeciphered. Moreover, at early infection stages, prevention of macrophage apoptosis is vital to successful mycobacterial pathogenesis. Our studies show that mycobacteria-secreted PknG can dampen the expression and concomitant secretion of proinflammatory TNFα. During early infection, M.tb infection-induced generation of reactive nitrogen intermediates (RNI) leads to S-nitrosylation of PknG on Cys109, thereby enabling its secretion into macrophages. Upon M.tb infection, secreted S-nitrosylated PknG phosphorylates macrophage Silencer of Death Domains (SODD) at Thr405, as identified through our phosphoproteomic studies. Thereafter, phosphorylated SODD, through an irreversible binding with the TNFR1 death domain, prevents Caspase8 activation and concomitant extrinsic apoptotic trigger. Moreover, alveolar macrophages from mice infected with PknG-knockout M.tb also exhibited SODD phosphorylation and hindered Caspase8 activation to prevent extrinsic macrophage apoptosis. Therefore, this work exhibits S-nitrosylation-mediated secretion of PknG to induce phosphorylation of macrophage SODD, which, through irreversible interaction with TNFR1, prevented extrinsic macrophage apoptosis at the early stages of infection.

Absence of c-Maf and IL-10 enables type I IFN enhancement of innate responses to LPS in alveolar macrophages

Abstract Alveolar macrophages (AMs) are lung-resident myeloid cells and airway sentinels for inhaled pathogens and environmental particles. While AMs can be highly inflammatory in response to respiratory viruses, they do not mount proinflammatory responses to all airborne pathogens. For example, we previously showed that AMs fail to mount a robust proinflammatory response to Mycobacterium tuberculosis. Here, we address this discrepancy by investigating the capacity of murine AMs for direct innate immune sensing, using LPS as a model. Use of LPS-coated fluorescent beads enabled us to distinguish between directly exposed and bystander cells to measure transcriptional responses, by RNA-sequencing after cell sorting, and cytokine responses, by flow cytometry. We find that AMs have decreased proinflammatory responses to low-dose LPS compared to other macrophage types (bone marrow–derived macrophages, peritoneal macrophages), as measured by TNF, IL-6, Ifnb, and Ifit3. The reduced response to low-dose LPS correlates with minimal TLR4 and CD14 surface expression, despite sufficient internal expression of TLR4. We also find that AMs do not produce IL-10 in response to a variety of stimuli due to low expression of the transcription factor c-Maf, while exogenous c-Maf expression restores IL-10 production in AMs. Lastly, we show that lack of IL-10 enables type I IFN enhancement of AM responses to LPS. Overall, we demonstrate AMs have a cell-intrinsic hyporesponsiveness to LPS, which makes them uniquely tolerant to low-dose exposure. Regulation of AM innate responses by distinct CD14, c-Maf, and IL-10 expression patterns has important implications for both respiratory infections and environmental airborne exposures.

Inherent PD-L1 22C3 Expression in Alveolar Macrophages Impacts the Combined Positive Score Status in Breast Cancer With Pulmonary Metastasis.

This study aimed to determine the impact of inherent programmed death-ligand 1 (PD-L1)-expressing alveolar macrophages (AMs) on the combined positive score (CPS) of PD-L1 (22C3) in metastatic breast cancer in the lungs. A total of 87 patients with pulmonary metastases of breast cancer were included in this study. Immunohistochemical staining of various PD-L1 antibodies was performed. The CPSs and CPSs excluding the number of PD-L1 positive AMs [CPS(NAM)s] with PD-L1 (22C3) were determined and compared. Among 87 enrolled patients, 22 had luminal A breast cancer, 24 had luminal B breast cancer, 13 had HER-2-positive breast cancer, and 28 had triple-negative breast cancer (TNBC). CPSs ≥ 10 was observed only in luminal B (12.5%) and TNBC (35.7%) subtypes (p < 0.001), whereas CPS(NAM)s ≥ 10 was observed only in TNBC (14.3%) (p = 0.011). Changes from the CPS-positive to the CPS(NAM)-negative status occurred in nine cases (10.3%), with significantly higher proportions being observed in the luminal B (12.5%) and TNBC (21.4%) subtypes (p = 0.007). Tumors showing changes from the CPS-positive to the CPS(NAM)-negative status were larger (p < 0.001); similar findings were observed in the TNBC subgroup (p = 0.001). The inclusion of PD-L1 expressing AMs leads to differences in CPS positivity, especially in large TNBC subtype tumors.

Screening effective-component compatibility from Jinshui Chenfei formula for silicosis treatment by serum-pharmacochemistry and feedback system control.

Screening effective-component compatibility from Jinshui Chenfei formula for silicosis treatment by serum-pharmacochemistry and feedback system control.

Integration of single-cell RNA sequencing and network pharmacology to elucidate the effect of Yantiao Formula on alleviating ALI by regulating the polarization of alveolar macrophages.

Integration of single-cell RNA sequencing and network pharmacology to elucidate the effect of Yantiao Formula on alleviating ALI by regulating the polarization of alveolar macrophages.

Mannose-modified exosomes loaded with MiR-23b-3p target alveolar macrophages to alleviate acute lung injury in Sepsis.

Mannose-modified exosomes loaded with MiR-23b-3p target alveolar macrophages to alleviate acute lung injury in Sepsis.

Exploring the Molecular Mechanism by which Kaempferol Attenuates Sepsis-related Acute Respiratory Distress Syndrome Based on Network Pharmacology and Experimental Verification.

Sepsis-related acute respiratory distress syndrome (ARDS) is a fatal disease without effective therapy. Kaempferol is a flavonoid compound extracted from natural plant products; it exerts numerous pharmacological effects. Kaempferol attenuates sepsis-related ARDS; however, the underlying protective mechanism has not been elucidated completely. This study aimed to use network pharmacology and experimental verification to investigate the mechanisms by which kaempferol attenuates sepsis-related ARDS. We screened the targets of kaempferol by PharMapper, Swiss Target Prediction, and CTD database. We identified the targets of sepsis-related ARDS by GeneCards, DisGeNet, OMIM, and TTD. The Weishengxin platform was used to map the targets of both kaempferol and sepsis-related ARDS. We created a Venn diagram to identify the intersection targets. We constructed the "component-intersection targets-disease" network diagram using Cytoscape 3.9.1 software. The intersection targets were imported into the STRING database for developing the protein-protein interaction network. Metascape was used for the Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis. We selected the leading 20 KEGG pathways to establish the KEGG relationship network. Finally, we performed experimental verification to confirm our prediction results. Through database screening, we obtained 502, 360, and 78 kaempferol targets, disease targets of sepsis-related ARDS, and intersection targets, respectively. The core targets consisted of tumor necrosis factor-alpha (TNF-α), interleukin (IL)-6, albumin (ALB), IL-1β, and AKT serine/ threonine kinase (AKT)1. GO enrichment analysis identified 426 items, which were principally involved in response to lipopolysaccharide, regulation of inflammatory response, inflammatory response, positive regulation of cell migration, positive regulation of cell adhesion, positive regulation of protein phosphorylation, response to hormone, regulation of reactive oxygen species (ROS) metabolic process, negative regulation of apoptotic signaling pathway, and response to decreased oxygen levels. KEGG enrichment analysis identified 151 pathways. After eliminating the disease and generalized pathways, we obtained the hypoxia-inducible factor 1 (HIF-1), nuclear factor κB (NF-κB), and phosphoinositide 3-kinase (PI3K)-Akt signaling pathways. Our experimental verification confirmed that kaempferol blocked the HIF-1, NF-κB, and PI3K-Akt signaling pathways, diminished TNF-α, IL-1β, and IL-6 expressions, suppressed ROS production, and inhibited apoptosis in lipopolysaccharide (LPS)-induced murine alveolar macrophage (MH-S) cells. Kaempferol can reduce inflammatory response, ROS production, and cell apoptosis by acting on the HIF-1, NF-κB, and PI3K-Akt signaling pathways, thereby alleviating sepsis- related ARDS.

Porcine serum amyloid A3 promotes the adhesion, invasion, and proliferation of Actinobacillus pleuropneumoniae.

Porcine serum amyloid A3 promotes the adhesion, invasion, and proliferation of Actinobacillus pleuropneumoniae.

HIV alters the landscape of alveolar macrophages subpopulations in the human lung

HIV alters the landscape of alveolar macrophages subpopulations in the human lung

Biomimetic fucoidan nanoparticles with regulation of macrophage polarization for targeted therapy of acute lung injury

Biomimetic fucoidan nanoparticles with regulation of macrophage polarization for targeted therapy of acute lung injury

Differences in glycolytic metabolism between tissue-resident alveolar macrophages and recruited lung macrophages

Immunometabolism has emerged as a key area of focus in immunology and has the potential to lead to new treatments for immune-related diseases. It is well-established that glycolytic metabolism is essential for adaptation to hypoxia and for macrophage inflammatory function. Macrophages have been shown to upregulate their glycolytic metabolism in response to pathogens and pathogen-associated molecular patterns such as LPS. As a direct link to the external environment, the lungs’ distinctive nutrient composition and multiple macrophage subtypes provide a unique opportunity to study macrophage metabolism. This review aims to highlight how the steady-state airway and severely inflamed airway offer divergent environments for macrophage glycolytic metabolism. We describe the differences in glycolytic metabolism between tissue-resident alveolar macrophages, and other lung macrophages at steady-state and during inflammation/injury. We also provide an overview of experimental guidelines on how to assess metabolism at the cellular level using Seahorse-based bioenergetic analysis including a review of pharmacologic agents used to inhibit or activate glycolysis.

Recruitment of pulmonary intravascular macrophages in SARS-CoV-2 infected hamsters.

The mechanisms by which severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes severe lung inflammation and mortality remain unclear. While the role of alveolar macrophages in COVID-19 is known, data on pulmonary intravascular macrophages (PIMs) is lacking. PIMs are key inflammatory cells present in species like cattle and pigs. Though constitutively absent in humans and rodents, their recruitment in rodents triggers exaggerated inflammation. We investigated the recruitment of PIMs and other immune cells, using immunofluorescence, hematoxylin and eosin (H&E) staining, and immunogold labeling in a hamster model of SARS-CoV-2 infection. Syrian golden hamsters were divided into 6 groups: uninfected control, unvaccinated-infected at 2-, 5-, and 14-days post infection (dpi) and vaccinated-infected at 5- and 14-dpi. Lung tissues were analyzed for neutrophils (myeloperoxidase), monocytes/macrophages (CCR2, CX3CR1), macrophages (IBA-1), and T cells (CD3). Septal macrophages increased at 2-, 5-, and 14-dpi in infected animals vs. control. CX3CR1 + cells decreased at 14-dpi in unvaccinated animals, but CX3CR1/CCR2 double positive cells were higher at 5-dpi, indicating a pro-inflammatory macrophage phenotype. PIMs were confirmed by transmission electron microscopy. These are the first data showing recruitment of pro-inflammatory PIMs in SARS-CoV-2 infected lungs.

A single-cell map of patients with non-small cell lung cancer harboring rare-driver mutations after anti-PD-1 treatment.

A single-cell map of patients with non-small cell lung cancer harboring rare-driver mutations after anti-PD-1 treatment.

Identification of Escherichia coli 166 isolate as an effective inhibitor of African swine fever virus replication.

African swine fever is a lethal disease with mortality rates approaching 100% in both domestic pigs and wild boars. With no effective vaccines or treatments available, there is an urgent need for new biologics to combat the African swine fever virus (ASFV). In this study, we isolated bacteria from the intestinal contents of wild boar using culture-based methods and identified them through 16S ribosomal DNA (rDNA) sequencing. These isolates underwent high-throughput screening to evaluate their immunomodulatory effects on J774-Dual cells and their ability to inhibit ASFV replication in vitro. Among them, an Escherichia coli strain, designated as E. coli 166, exhibited strong inhibitory effects on various ASFV strains' replication, including three genotype II strains: virulent strain HLJ/18, moderately virulent strain HLJ/HRB1/20, genetically modified low-virulent strain HLJ/18-6GD, and one genotype I low-virulent strain SD/DY-I/21. Notably, this inhibition did not require direct interaction between the bacteria and porcine alveolar macrophages (PAMs). Both live and heat-inactivated E. coli 166 demonstrated a strong inhibitory effect on ASFV replication. Genetic modification of E. coli 166 did not alter its inhibitory phenotype. Further analysis revealed that PAMs pretreated with E. coli 166 showed upregulation of NF-κB and downregulation of CD163 at different time points post-infection, whereas PAMs only infected with ASFV exhibited the opposite trend. These findings suggest that E. coli 166 holds promise as a biological agent for controlling ASFV infection, through indirect mechanisms involving bacterial metabolites or lysis products. Future studies should focus on identifying the specific components responsible for its antiviral effects.IMPORTANCEThe emergence of the African swine fever virus (ASFV) as a devastating pathogen in swine populations necessitates the development of novel strategies for its control. In this study, Escherichia coli strain 166 (E. coli 166) demonstrated a remarkable ability to inhibit the replication of multiple ASFV strains in porcine alveolar macrophages (PAMs), even without direct bacterial contact. Both live and heat-inactivated E. coli 166 retained this inhibitory effect, suggesting that secreted metabolites or lysis products may play a key role. Furthermore, pretreatment of PAMs with E. coli 166 resulted in upregulated NF-κB activity and downregulated expression of the ASFV entry receptor CD163, presenting an immune-modulatory mechanism distinct from PAMs solely infected with ASFV. These findings highlight the potential of E. coli 166 as a biological agent to combat ASFV, offering a promising alternative or complementary approach to traditional antiviral strategies.

Molecular docking, free energy calculations, ADMETox studies, DFT analysis, and dynamic simulations highlighting a chromene glycoside as a potential inhibitor of PknG in Mycobacterium tuberculosis.

Tuberculosis (TB), caused by the Mycobacterium tuberculosis (M.tb), remains a serious medical concern globally. Resistant M.tb strains are emerging, partly because M.tb can survive within alveolar macrophages, resulting in persistent infection. Protein kinase G (PknG) is a mycobacterial virulence factor that promotes the survival of M.tb in macrophages. Targeting PknG could offer an opportunity to suppress the resistant M.tb strains. In the present study, multiple computational tools were adopted to screen a library of 460,000 molecules for potential inhibitors of PknG of M.tb. Seven Hits (1-7) were identified with binding affinities exceeding that of the reference compound (AX20017) towards the PknG catalytic domain. Next, the ADMETox studies were performed to identify the best hit with appropriate drug-like properties. The chromene glycoside (Hit 1) was identified as a potential PknG inhibitor with better pharmacokinetic and toxicity profiles rendering it a potential drug candidate. Furthermore, quantum computational analysis was conducted to assess the mechanical and electronic properties of Hit 1, providing guidance for further studies. Molecular dynamics simulations were also performed for Hit 1 against PknG, confirming the stability of its complex. In sum, the findings in the current study highlight Hit 1 as a lead with potential for development of drugs capable of treating resistant TB.

A progranulin variant causing childhood interstitial lung disease responsive to anti-TNF-α biologic therapy.

A progranulin variant causing childhood interstitial lung disease responsive to anti-TNF-α biologic therapy.