Development and Functions of Alveolar Macrophages

Macrophages residing in various tissue types are unique in terms of their anatomical locations, ontogenies, developmental pathways, gene expression patterns, and immunological functions. Alveolar macrophages (AMs) reside in the alveolar lumen of the lungs and serve as the first line of defense for the respiratory tract. The immunological functions of AMs are implicated in the pathogenesis of various pulmonary diseases such as allergic asthma, chronic obstructive pulmonary disorder (COPD), pulmonary alveolar proteinosis (PAP), viral infection, and bacterial infection. Thus, the molecular mechanisms driving the development and function of AMs have been extensively investigated. In this review article, we discuss the roles of granulocyte-macrophage colony-stimulating factor (GM-CSF) and transforming growth factor (TGF)-β in AM development, and provide an overview of the anti-inflammatory and pro-inflammatory functions of AMs in various contexts. Notably, we examine the relationships between the metabolic status of AMs and their development processes and functions. We hope that this review will provide new information and insight into AM development and function.     

Viral Infection of Human Lung Macrophages Increases PDL1 Expression via IFNβ

Lung macrophages are an important defence against respiratory viral infection and recent work has demonstrated that influenza-induced macrophage PDL1 expression in the murine lung leads to rapid modulation of CD8+ T cell responses via the PD1 receptor. This PD1/PDL1 pathway may downregulate acute inflammatory responses to prevent tissue damage. The aim of this study was to investigate the mechanisms of PDL1 regulation by human macrophages in response to viral infection. Ex-vivo viral infection models using influenza and RSV were established in human lung explants, isolated lung macrophages and monocyte-derived macrophages (MDM) and analysed by flow cytometry and RT-PCR. Incubation of lung explants, lung macrophages and MDM with X31 resulted in mean cellular infection rates of 18%, 18% and 29% respectively. Viral infection significantly increased cell surface expression of PDL1 on explant macrophages, lung macrophages and MDM but not explant epithelial cells. Infected MDM induced IFNγ release from autologous CD8+ T cells, an effect enhanced by PDL1 blockade. We observed increases in PDL1 mRNA and IFNβ mRNA and protein release by MDM in response to influenza infection. Knockdown of IFNβ by siRNA, resulted in a 37.5% reduction in IFNβ gene expression in response to infection, and a significant decrease in PDL1 mRNA. Furthermore, when MDM were incubated with IFNβ, this cytokine caused increased expression of PDL1 mRNA. These data indicate that human macrophage PDL1 expression modulates CD8+ cell IFNγ release in response to virus and that this expression is regulated by autologous IFNβ production.     

Selective down-regulation of alveolar macrophage oxidative response to opsonin-independent phagocytosis

We have compared the oxidative response of alveolar macrophages (AM) during opsonin-dependent and independent phagocytosis by using multiparameter flow cytometry. The respiratory burst of AM during phagocytosis was quantitated by the intracellular oxidation of the nonfluorescent precursors dichlorofluorescin diacetate (DCFH) or hydroethidine (HE, a reduced precursor of ethidium) to their fluorescent (oxidized) counterparts. After loading freshly isolated normal hamster AM with DCFH or HE, red or green fluorescent beads, respectively, were added to the shaking cell suspensions. Ingestion of opsonized particles by AM caused a marked increase in oxidation of both DCFH and HE proportional to the number of beads ingested. In contrast, uptake of one to three unopsonized particles per cell led to inhibition of oxidative activity compared to control cells incubated without particles. AM ingesting four or more unopsonized particles showed some increase in oxidative metabolism, but far less than that with identical numbers of particles in opsonin-dependent ingestion. Similar results were obtained using fluorescent labeled staphylococcal bacteria. Using three-color flow cytometry to study cells ingesting both types of particles, cells first ingesting unopsonized beads were also found to have an inhibited oxidative response to subsequently ingested opsonized particles. The mitochondrial poison antimycin inhibited most of the intracellular oxidative response to either type of phagocytosis. The remaining antimycin-insensitive, membrane derived respiratory burst of AM was also substantially diminished after phagocytosis of unopsonized particles vs similar numbers of opsonized particles. The greatly increased mitochondrial respiration in AM during phagocytosis of opsonized particles may be related to bactericidal mechanisms. Killing of ingested Staphylococcus by AM was markedly impaired in the presence of antimycin. The results suggest that AM may ingest the numerous, unopsonized inert particles that are inhaled without generation of potentially toxic oxygen metabolites, while retaining the capacity to undergo a respiratory burst after ingesting opsonized particles and bacteria. The mechanism(s) for this distinct response may include generation of an inhibitor of intracellular oxidative metabolism.     

Human adipose tissue macrophages display activation of cancer-related pathways

Obesity is associated with a significantly increased risk for cancer suggesting that adipose tissue dysfunctions might play a crucial role therein. Macrophages play important roles in adipose tissue as well as in cancers. Here, we studied whether human adipose tissue macrophages (ATM) modulate cancer cell function. Therefore, ATM were isolated and compared with monocyte-derived macrophages (MDM) from the same obese patients. ATM, but not MDM, were found to secrete factors inducing inflammation and lipid accumulation in human T47D and HT-29 cancer cells. Gene expression profile comparison of ATM and MDM revealed overexpression of functional clusters, such as cytokine-cytokine receptor interaction (especially CXC-chemokine) signaling as well as cancer-related pathways, in ATM. Comparison with gene expression profiles of human tumor-associated macrophages showed that ATM, but not MDM resemble tumor-associated macrophages. Indirect co-culture experiments demonstrated that factors secreted by preadipocytes, but not mature adipocytes, confer an ATM-like phenotype to MDM. Finally, the concentrations of ATM-secreted factors related to cancer are elevated in serum of obese subjects. In conclusion, ATM may thus modulate the cancer cell phenotype.     

TNF-alpha inhibits HIV-1 replication in peripheral blood monocytes and alveolar macrophages by inducing the production of RANTES and decreasing C-C chemokine receptor 5 (CCR5) expression.

The pathogenesis of HIV-1 infection is influenced by the immunoregulatory responses of the host. Macrophages present in the lymphoid tissue are susceptible to infection with HIV-1, but are relatively resistant to its cytopathic effects and serve as a reservoir for the virus during the course of disease. Previous investigators have demonstrated that increased serum levels of TNF-alpha contribute to the clinical symptoms of AIDS and that TNF-alpha stimulates the production of HIV-1 in chronically infected lymphocytic and monocytic cell lines by increasing HIV-1 gene expression. Although previous studies have suggested that TNF-alpha may increase HIV-1 infection of primary human mononuclear cells, some recent studies have indicated that TNF-alpha suppresses HIV-1 infection of macrophages. We now demonstrate that TNF-alpha suppresses HIV-1 replication in freshly infected peripheral blood monocytes (PBM) and alveolar macrophages (AM) in a dose-dependent manner. As TNF-alpha has been shown to increase the production of C-C chemokine receptor (CCR5)-binding chemokines under certain circumstances, we hypothesized that TNF-alpha inhibits HIV-1 replication by increasing the expression of these HIV-suppressive factors. We now show that TNF-alpha treatment of PBM and AM increases the production of the C-C chemokine, RANTES. Immunodepletion of RANTES alone or in combination with macrophage inflammatory protein-1alpha and -1beta block the ability of TNF-alpha to suppress viral replication in PBM and AM. In addition, we found that TNF-alpha treatment reduces CCR5 expression on PBM and AM. These findings suggest that TNF-alpha plays a significant role in inhibiting monocytotropic strains of HIV-1 by two distinct, but complementary, mechanisms.     

Quantification of PPAR-gamma protein in monocyte/macrophages from healthy smokers and non-smokers: a possible direct effect of nicotine

Previous observations demonstrated that Peroxisome Proliferator-Activated Receptor-gamma (PPAR-gamma), a key regulator of adipocyte differentiation, is expressed in a large variety of cells, including cells of the monocyte/macrophage lineage. This study was aimed to quantify both the constitutive and ligand-induced PPAR-gamma expression in monocytes and monocyte-derived macrophages (MDM) isolated from healthy smokers and non-smokers, and to evaluate the possible direct effect of nicotine. PPAR-gamma protein was detected by Western blot and quantification was performed by calculating the ratio between PPAR-gamma and beta-actin protein expression. Cytokine release was measured with enzyme-linked immunoassay kits. Constitutive PPAR-gamma protein was detected in human monocytes and its expression was up-regulated along with differentiation to MDM. The endogenous ligand 15-deoxy-delta(12,14)-prostaglandin J(2) and the synthetic agonist ciglitazone enhanced PPAR-gamma expression, the former being effective also at low micromolar concentrations. Both agonists significantly inhibited the basal secretion of pro-inflammatory cytokines (e.g., TNF-alpha, IL-6), ciglitazone being more potent. Monocytes and MDM from healthy smokers presented a significantly enhanced (4-fold and 2.5-fold, respectively) constitutive PPAR-gamma expression, as compared to those from healthy non-smokers. However, ligand-induced PPAR-gamma expression and inhibition of cytokine secretion were similar in healthy smokers and non-smokers. Nicotine dose-dependently enhanced PPAR-gamma expression with a maximum at 10 muM, and inhibited release of pro-inflammatory cytokines; these effects were reversed by alpha-bungarotoxin. Nicotine and PPAR-gamma agonists did not exert synergistic effects. In conclusion, monocytes and MDM from healthy smokers present a constitutively enhanced PPAR-gamma expression; this effect is reproduced, to some extent, by nicotine in vitro.