Impaired Phagocytic Function Research Articles

HIV-1 Nef mobilizes lipid rafts in macrophages through a pathway that competes with ABCA1-dependent cholesterol efflux

HIV infection, through the actions of viral accessory protein Nef, impairs activity of cholesterol transporter ABCA1, inhibiting cholesterol efflux from macrophages and elevating the risk of atherosclerosis. Nef also induces lipid raft formation. In this study, we demonstrate that these activities are tightly linked and affect macrophage function and HIV replication. Nef stimulated lipid raft formation in macrophage cell line RAW 264.7, and lipid rafts were also mobilized in HIV-1-infected human monocyte-derived macrophages. Nef-mediated transfer of cholesterol to lipid rafts competed with the ABCA1-dependent pathway of cholesterol efflux, and pharmacological inhibition of ABCA1 functionality or suppression of ABCA1 expression by RNAi increased Nef-dependent delivery of cholesterol to lipid rafts. Nef reduced cell-surface accessibility of ABCA1 and induced ABCA1 catabolism via the lysosomal pathway. Despite increasing the abundance of lipid rafts, expression of Nef impaired phagocytic functions of macrophages. The infectivity of the virus produced in natural target cells of HIV-1 negatively correlated with the level of ABCA1. These findings demonstrate that Nef-dependent inhibition of ABCA1 is an essential component of the viral replication strategy and underscore the role of ABCA1 as an innate anti-HIV factor.

Loss of Transcription Factor Nuclear Factor-Erythroid 2 (NF-E2) p45-related Factor-2 (Nrf2) Leads to Dysregulation of Immune Functions, Redox Homeostasis, and Intracellular Signaling in Dendritic Cells

Dendritic cells (DCs) are critical mediators of immunity and immune tolerance by orchestrating multiple aspects of T cell activation and function. Immature DCs (iDCs) expressing low levels of co-stimulatory receptors are highly efficient at antigen capture but are poor activators of T cells. Maturation of DCs is associated with increased expression of co-stimulatory molecules. Co-stimulatory receptor gene expression is regulated by intracellular redox, NF-κB, and MAPK pathways and by histone deacetylase (HDAC) activity. The transcription factor, Nrf2, is important for maintaining intracellular glutathione (GSH) levels and redox homeostasis and has been implicated in modulating DC co-stimulatory receptor expression. It is unclear whether Nrf2 mediates this effect by GSH-dependent mechanisms and whether it influences DC signaling pathways. Using bone marrow-derived iDCs from Nrf2(+/+) and Nrf2(-/-) mice, we demonstrate that Nrf2(-/-) iDCs have lower basal GSH levels, enhanced co-stimulatory receptor expression, impaired phagocytic functions, and increased antigen-specific CD8 T cell stimulation capacity. Interestingly, lowering GSH levels in Nrf2(+/+) iDCs did not recapitulate the Nrf2(-/-) iDC phenotype. Loss of Nrf2 resulted in elevated basal levels of reactive oxygen species but did not affect basal NF-κB activity or p38 MAPK phosphorylation. Using pharmacological inhibitors, we demonstrate that enhanced co-stimulatory receptor phenotype of Nrf2(-/-) iDC does not require ERK activity but is dependent on HDAC activity, indicating a potential interaction between Nrf2 function and HDAC. These results suggest that Nrf2 activity is required to counter rises in intracellular reactive oxygen species and to regulate pathways that control DC co-stimulatory receptor expression.

Effect of Uremia on Structure and Function of Immune System

End-stage renal disease (ESRD) is simultaneously associated with immune activation, marked by systemic inflammation, and immune deficiency. Systemic inflammation contributes to atherosclerosis, cardiovascular disease, cachexia, and anemia, whereas immune deficiency leads to impaired response to vaccination, and increased incidence and severity of microbial infections. ESRD-associated inflammation and immune deficiency are associated with the following: (a) general expansion of monocytes and elevations of their basal integrin, Toll-like receptor (TLR)-2, TLR-4 expression, cytokine production, and reactive oxygen species (ROS) generation and reduced phagocytic capacity, (b) depletion and impaired inhibitory activity of regulatory T cells, (c) spontaneous activation, degranulation, increased basal ROS production, decreased phagocytic capacity, and increased apoptosis of the circulating polymorphonuclear leukocytes, (d) upregulation of ROS production machinery and chemokine expression in the cellular constituents of various tissues, highlighting participation of nonimmune cells in the prevailing inflammatory state, (e) depletion of the antigen-presenting dendritic cells, (f) reduced CD4/CD8 T cell ratio and depletion of naïve and central memory T cells, (g) diffuse B cell lymphopenia leading to impaired humoral immunity, and (h) increased proinflammatory activity of low-density lipoprotein and reduced anti-inflammatory capacity of high-density lipoprotein. Thus, ESRD-associated inflammation is due to activation of innate immune system, orchestrated by monocytes, macrophages, granulocytes, and cellular constituents of other organs/tissues. This is coupled with immune deficiency that is caused by depletion of dendritic cells, naïve and central memory T cells and B cells, and impaired phagocytic function of polymorphonuclear leukocytes and monocytes.

OP06 Maternal obesity programs offspring innate immune dysregulation in non-alcoholic fatty liver disease

IntroductionObesity induced, non-alcoholic fatty liver disease (NAFLD), is the major cause of chronic liver dysfunction. In tandem, obesity and NAFLD rates in reproductive age women are rising. We have previously...

Endotoxaemia in the pathogenesis of cytopenias in liver cirrhosis. Could oral antibiotics raise blood counts?

Cytopenias are frequently observed in patients with cirrhosis and are associated with increased morbidity. In particular, thrombocytopenia can impact routine care of patients with cirrhosis by potentially postponing or interfering with diagnostic and therapeutic procedures including liver biopsy and medically indicated or elective surgery. The pathogenesis of cytopenias in cirrhosis remains largely unknown. Historically, the concept of hypersplenism has long been associated with the cirrhosis-related hematological disorders but was never proven. On the other hand, intestinal bacterial overgrowth and altered gut permeability in cirrhotic patients lead to increased translocation of bacteria and endotoxin into the portal circulation. The impaired phagocytic function of the reticuloendothelial system together with the portosystemic shunting allow endotoxin to reach the systemic circulation and high concentrations of circulating endotoxin are found in cirrhotic patients even with no clinical evidence of infection and correlate with the severity of liver disease. Endotoxin activates monocytes and promotes the release of proinflammatory cytokines. Indeed, serum levels of interleukin-1, interleukin-6, tumor necrosis factor-α, and interferon-γ are elevated in patients with cirrhosis in proportion to the severity of liver disease. Endotoxaemia stimulates the vascular production of nitric oxide (NO) directly or indirectly via the cytokine cascade, and correlates with serum NO metabolite levels in cirrhosis. Several lines of evidence strongly suggest that endotoxaemia may reduce peripheral blood counts either directly or through the release of cytokines and NO. Previous studies in experimental models of cirrhosis and cirrhotic patients have demonstrated that long-term administration of oral antibiotics such as trimethoprim-sulfamethoxazole, norfloxacin, and rifaximin can reduce bacterial translocation and circulating levels of endotoxin, TNF-α, IL-6, and NO. We hypothesize that endotoxaemia plays a pivotal role in the pathogenesis of cytopenias in cirrhosis and that intestinal decontamination could raise peripheral blood counts by the suppression of endotoxaemia and the inhibition of cytokine and NO production.

Effect of abrupt weaning at housing on leukocyte distribution, functional activity of neutrophils, and acute phase protein response of beef calves.

BackgroundSixteen, spring-born, single suckled, castrated male calves of Limousin × Holstein-Friesian and Simmental × Holstein-Friesian dams respectively, were used to investigate the effect of weaning on total leukocyte and differential counts, neutrophil functional activity, lymphocyte immunophenotypes, and acute phase protein response. Calves grazed with their dams until the end of the grazing season when they were housed in a slatted floor shed. On the day of housing, calves were assigned to a treatment, (i) abruptly weaned (W: n = 8) or (ii) non-weaned (controls) (C: n = 8). Weaned calves were housed in pens without their dams, whereas non-weaned (control) calves were housed with their dams. Blood was collected on day -7, 0 (housing), 2, 7, and 14 to determine total leukocyte and differential counts and concentration of fibrinogen and haptoglobin. Lymphocyte immunophenotypes were characterised using selected surface antigens (CD4+, CD8+, WC1+ (γδ T cells), MHC Class II+ lymphocytes), and the functional activities of neutrophils (surface expression of L-selectin (CD62L), phagocytic and oxidative burst activity) were investigated using flow cytometry.ResultsTreatment × sampling time interactions (P < 0.05) were detected for total leukocyte and neutrophil counts, all lymphocyte subsets, mean fluorescence intensity of CD62L+ neutrophils, and percentage neutrophils performing phagocytosis. On d 2, total leukocyte and neutrophil count increased (P < 0.001), and percentage CD4+ and CD8+ lymphocytes, percentage phagocytic neutrophils, mean fluorescence intensity of CD62L+ neutrophils decreased (P < 0.05) in W compared with baseline (d 0), whereas they were unchanged (P > 0.05) in C. On d 2, percentage WC1+ lymphocytes decreased (P < 0.05), whereas percentage MHC class II+ lymphocytes increased (P < 0.05) in W and C, however the magnitude of change was greater in W than C. There were no treatment × sampling time interactions (P > 0.05) for monocyte, eosinophil, and basophil counts, percentage G1+ neutrophils, or percentage oxidative burst positive neutrophils.ConclusionsAbrupt weaning resulted in increased neutrophil counts and impaired trafficking and phagocytic function. Together with the changes in lymphocyte subsets, the results suggest that there was a greater transitory reduction in immune function at housing in abruptly weaned than non-weaned beef calves.

Nanosized Aluminum Altered Immune Function

On the basis of their uses in jet fuels and munitions, the most likely scenario for aluminum nanoparticle (NP) exposure is inhalation. NPs have been shown to be capable of penetrating deep into the alveolar regions of the lung, and therefore human alveolar macrophages (U937) with human type II pneumocytes (A549) were cultured together and exposed to NPs dispersed in an artificial lung surfactant to more accurately mimic the lung microenvironment. Two types of NPs were evaluated: aluminum (Al) and aluminum oxide (Al2O3). Following a 24-h incubation, cell viability was assessed using MTS, and mild toxicity was observed at higher doses with the U937 cells affected more than the A549. Since the U937 cells provided protection from NP toxicity, the cocultures were exposed to a benign concentration of NPs and infected with the respiratory pathogen community-associated methicillin-resistant Staphylococcus aureus (ca-MRSA) to determine any changes in cellular function. Phagocytosis assays demonstrated that the NPs impaired phagocytic function, and bacterial growth curves confirmed that this reduction in phagocytosis was not related to NP-bacteria interactions. Furthermore, NFkappaB PCR arrays and an IL-6 and TNF-alpha real time PCR demonstrated that both types of NPs altered immune response activation. This change was confirmed by ELISA assays that evaluated the secretion of IL-6, IL-8, IL-10, IL-1beta, and TNF-alpha and illustrated that the NPs repressed secretion of these cytokines. Therefore, although the NPs were not toxic to the cells, they did impair the cell's natural ability to respond to a respiratory pathogen regardless of NP composition.

Impaired phagocytosis by alveolar macrophages from diabetic rats is related to the deficient coupling of LTs to the FcγR signaling cascade

Diabetic individuals are more susceptible to infections and this seems to be related to impaired phagocyte function. Alveolar macrophages (AMs) are the first barrier to prevent respiratory infections. Leukotrienes (LTs) increase AM phagocytic activity via FcγR. In this study, we compared AMs from diabetic and non-diabetic rats for phagocytosis via FcγR and the roles of LTs and insulin. Diabetes was induced in male Wistar rats by alloxan (42 mg/kg, i.v.); macrophages were obtained by bronchoalveolar lavage and IgG-opsonised sheep red blood cells (IgG-SRBC) were used as targets. LTs were added to the AMs 5 min before the addition of IgG-SRBC. AMs were treated with a LT synthesis inhibitor (zileuton, 10 μM), or antagonists of the LTB 4 receptor (CP105.696, 10 μM) or cys-LT receptor (MK571, 10 μM), 30 or 20 min before the addition of IgG-SRBC, respectively. We found that the phagocytosis of IgG-SRBC by AMs from diabetic rats is impaired compared with non-diabetic rats. Treatment with the LT inhibitor/antagonists significantly reduced AM phagocytosis in non-diabetic but not diabetic rats. During the phagocytosis of IgG-SRBC LTB 4 and LTC 4 were produced by AMs from both groups. The addition of exogenous LTB 4 or LTD 4 potentiated phagocytosis similarly in both groups. Phagocytosis was followed by the phosphorylation of PKC-δ, ERK and Akt. This was reduced by zileuton treatment in AMs from non-diabetic but not diabetic rats. The addition of insulin to AMs further increased the phagocytosis by increasing PKC-δ phosphorylation. These results suggest that the impaired phagocytosis found in AMs from diabetic rats is related to a deficient coupling of LTs to the FcγR signaling cascade and that insulin has a key role in this coupling. An essential role for insulin in innate immunity is suggested.

The role of Pseudomonas lipopolysaccharide in cystic fibrosis airway infection.

Pseudomonas aeruginosa (PA) is a ubiquitous environmental Gram-negative bacterium found in soil and water. This opportunistic pathogen can cause infections in individuals with impaired phagocytic function, such as those with burns, exposure to chemotherapy, or cystic fibrosis (CF). PA infects the lungs of most individuals with CF, and is associated with severe progressive pulmonary disease that is the major cause of premature death in this disorder. The specific adaptations of PA to the CF airway responsible for bacterial persistence and antibiotic tolerance are not completely understood but may include increased alginate production (i.e., mucoid phenotype), biofilm formation, and specific lipid A modifications. During adaptation to the CF airway, PA synthesizes a variety of lipid A structures that alter host innate immune responses and promote bacterial persistence and chronic infection. The synthesis of specific lipid A structures is attributable to bacterial enzymes that: (1) remove the 3OH-C10:0 acyl chain from the 3-position (PagL); (2) add a C16:0 acyl chain to the 3OH-C10:0 chain at the 3'-position (PagP); (3) add C12:0 and 2OH-C12:0 acyl chains to the 3OH-C12:0 chains at the 2- and 2'-positions (HtrB and LpxO); and (4) add aminoarabinose to phosphate groups at the 1- and 4'-positions (PmrH, PmrF, PmrI, PmrJ, PmrK, and PmrE). These lipid A modifications represent an essential aspect of PA adaptation to the CF airway.

Loops and networks in control of Francisella tularensis virulence

Francisella tularensis is a highly infectious, Gram-negative bacterium responsible for the disease tularemia in a broad variety of animals, including humans. F. tularensis intracellular multiplication occurs mainly in macrophages. However, F. tularensis is able to infect many other cell types, including other phagocytic (dendritic cells, polymorphonuclear leukocytes) and nonphagocytic (alveolar epithelial cells, hepatocytes, endothelial cells and fibroblasts) cells. The ability of professional phagocytic cells to engulf and kill microbes is an essential component of innate defense. The ability of F. tularensis to impair phagocyte function and survive in the cytosol of infected cells thus constitutes a central aspect of its virulence. The F. tularensis intracellular lifecycle relies on the tightly regulated expression of a series of genes. The unraveling secrets of the regulatory cascades governing the regulation of virulence of F. tularensis will be discussed along with future challenges yet to be solved.

Response to Inhaled Granulocyte-Macrophage Colony-Stimulating Factor in a Patient With Alveolar Proteinosis

Pulmonary alveolar proteinosis is a rare disease characterized by the accumulation of lipoproteinaceous material derived from alveolar surfactant in the alveoli, with a consequent deterioration in gas exchange. Pathogenesis is related to impaired phagocytic function of alveolar macrophages. In recent years, a new treatment for pulmonary alveolar proteinosis—consisting of subcutaneous administration of granulocytemacrophage colony-stimulating factor (GM-CSF)—has become available. The commonly accepted treatment, and the one to have shown greatest efficacy in pulmonary alveolar proteinosis, is whole lung lavage. Instead of subcutaneous administration, GM-CSF can also be inhaled as an aerosol. This route of administration of GM-CSF is safe and effective in the treatment of pulmonary alveolar proteinosis and represents an alternative to subcutaneous administration or whole lung lavage. We present a patient with pulmonary alveolar proteinosis who was treated with inhaled GM-CSF and describe her clinical and functional outcome after 1 year of treatment.

Proteinosis alveolar. Respuesta al tratamiento con factor estimulante de colonias de granulocitos y macrófagos por vía inhalada

Pulmonary alveolar proteinosis is a rare disease characterized by the accumulation of lipoproteinaceous material derived from alveolar surfactant in the alveoli, with a consequent deterioration in gas exchange. Pathogenesis is related to impaired phagocytic function of alveolar macrophages. In recent years, a new treatment for pulmonary alveolar proteinosis—consisting of subcutaneous administration of granulocyte-macrophage colony-stimulating factor (GM-CSF)—has become available. The commonly accepted treatment, and the one to have shown greatest efficacy in pulmonary alveolar proteinosis, is whole lung lavage. Instead of subcutaneous administration, GM-CSF can also be inhaled as an aerosol. This route of administration of GM-CSF is safe and effective in the treatment of pulmonary alveolar proteinosis and represents an alternative to subcutaneous administration or whole lung lavage. We present a patient with pulmonary alveolar proteinosis who was treated with inhaled GM-CSF and describe her clinical and functional outcome after 1 year of treatment.

Pseudomonas aeruginosaType III secretion system interacts with phagocytes to modulate systemic infection of zebrafish embryos

Pseudomonas aeruginosa is an opportunistic human pathogen that can cause serious infection in those with deficient or impaired phagocytes. We have developed the optically transparent and genetically tractable zebrafish embryo as a model for systemic P. aeruginosa infection. Despite lacking adaptive immunity at this developmental stage, zebrafish embryos were highly resistant to P. aeruginosa infection, but as in humans, phagocyte depletion dramatically increased their susceptibility. The virulence of an attenuated P. aeruginosa strain lacking a functional Type III secretion system was restored upon phagocyte depletion, suggesting that this system influences virulence through its effects on phagocytes. Intravital imaging revealed bacterial interactions with multiple blood cell types. Neutrophils and macrophages rapidly phagocytosed and killed P. aeruginosa, suggesting that both cell types play a role in protection against infection. Intravascular aggregation of erythrocytes and other blood cells with resultant circulatory blockage was observed immediately upon infection, which may be relevant to the pathogenesis of thrombotic complications of human P. aeruginosa infections. The real-time visualization capabilities and genetic tractability of the zebrafish infection model should enable elucidation of molecular and cellular details of P. aeruginosa pathogenesis in conditions associated with neutropenia or impaired phagocyte function.

Methicillin-resistant Staphylococcus aureus (MRSA) mitral valve acute bacterial endocarditis (ABE) in a patient with Job’s syndrome (hyperimmunoglobulin E syndrome) successfully treated with linezolid and high-dose daptomycin

Job's syndrome (hyperimmunoglobulin E syndrome) is a congenitally acquired primary immune deficiency. The primary host defense defect in Job's syndrome is impaired phagocytosis. Accordingly, patients with Job's syndrome have difficulties eradicating staphylococcal infections. A continuous, high-grade Staphylococcus aureus bacteremia with a cardiac valve vegetation is the hallmark of S. aureus acute bacterial endocarditis (ABE). ABE may be caused by methicillin-sensitive Staphylococcus aureus or methicillin-resistant Staphylococcus aureus (MRSA). We report a case of Job's syndrome MRSA mitral valve ABE. Presumably because of impaired phagocytic function, his MRSA ABE was complicated by extensive metastatic septic complications manifested as brain abscess, multiple epidural abscesses, and multifocal vertebral osteomyelitis. The patient did not respond to 5 days of appropriately dosed linezolid and daptomycin and remained bacteremic because abscess drainage was not an option in this case and the continuous, high-grade MRSA bacteremia continued despite appropriate therapy. High-dose daptomycin (12 mg/kg intravenously every 24 hours) was given, and his MRSA bacteremia was rapidly terminated. Because daptomycin does not cross the blood-brain barrier in therapeutic concentrations, linezolid was used to treat the brain abscess. The extensiveness of infection in this case is remarkable and is probably related to impaired phagocytic function from Job's syndrome. High-dose daptomycin therapy rapidly cleared the bacteremia and cured the endocarditis and epidural abscesses/vertebral osteomyelitis. The patient was treated with 8 weeks of high-dose daptomycin therapy with no adverse effects. If MRSA and methicillin-sensitive S. aureus bacteremias are unresponsive to usually effective antistaphylococcal agents, and surgical drainage of abscesses and removal of infected devices are not clinically possible, then a prolonged, high dose of daptomycin is a therapeutic alternative in such situations. To the best of our knowledge, this is the first case of MRSA mitral valve ABE complicated by extensive epidural abscesses and vertebral osteomyelitis in a patient with Job's syndrome.

Constitutive activation of SHP2 protein tyrosine phosphatase inhibits ICSBP-induced transcription of the gene encoding gp91PHOX during myeloid differentiation

The IFN consensus sequence-binding protein (ICSBP; also referred to as IFN regulatory factor 8) is a transcription factor which is expressed in myeloid and B cells. In previous studies, we found that ICSBP activated transcription of the gene encoding gp91(PHOX) (the CYBB gene), a rate-limiting component of the phagocyte respiratory burst oxidase expressed exclusively after the promyelocyte stage of myelopoiesis. Previously, we found that CYBB transcription was dependent on phosphorylation of specific ICSBP tyrosine residues. Since ICSBP is tyrosine-phosphorylated during myelopoiesis, this provided a mechanism of differentiation stage-specific CYBB transcription. In the current studies, we found that ICSBP was a substrate for Src homology-containing tyrosine phosphatase 2 (SHP2-PTP) in immature myeloid cells but not during myelopoiesis. Therefore, SHP2-PTP inhibited CYBB transcription and respiratory burst activity in myeloid progenitor cells by dephosphorylating ICSBP. In contrast, we found that ICSBP was a substrate for a leukemia-associated, constitutively active mutant form of SHP2, described previously, throughout differentiation. Consistent with this, constitutive SHP2 activation blocked ICSBP-induced CYBB transcription and respiratory burst activity in differentiating myeloid cells. ICSBP-deficiency and constitutive SHP2 activation have been described in human myelodysplastic syndromes. As these two abnormalities may coexist, our results identified a potential molecular mechanism for impaired phagocyte function in this malignant myeloid disease.

Stress-Induced Downregulation of Macrophage Phagocytic Function Is Attenuated by Exercise Training in Rats

Background/Aims: Acute restraint stress may induce impaired macrophage phagocytic function. Moderate physical training is associated with beneficial effects on immunological functions. We investigated the effects of moderate physical training on phagocytic function of alveolar macrophages in rats submitted to acute restraint stress. Methods: Thirty male Wistar rats weighing 210–226 g were randomly divided into 4 groups: nontrained rats (n = 7), nontrained rats submitted to stress (n = 8), trained rats (n = 7) and trained rats submitted to stress (n = 8). Trained rats were submitted to a program of moderate running training over a period of 8 weeks. Rats subjected to restraint stress were kept immobilized in glass cylinders (8 cm in diameter and 24 cm long) during 60 min. Phagocytosis capacity of macrophages was evaluated by either Escherichia coli orzymosan stimuli. Results: Restraint stress induced a decrease in phagocytosis of E. coli and zymosan particle stimulation by macrophages. Neither of these alterations was observed in trained animals submitted to acute restraint stress. Conclusions: Our data confirm that acute restraint stress is associated with impaired function of macrophages. Moreover, moderate physical training attenuates the effects of acute stress by a mechanism that involves an increase in tolerance of macrophages.

The Nature of Immunological Reaction in the Peripheral Airways of Cigarette Smokers

The influence of tobacco smoke on human health is still an important problem worldwide. Complex inflammatory processes and changes in the immune system are crucial in the pathogenesis of smoking related disorders like chronic obstructive lung disease (COPD), lung cancer, asthma, interstitial lung diseases (ILD) and atherosclerosis. The objective of this review is to present the alterations in the immune system in smokers. Discrete changes in peripheral blood of smokers may be found, such as leukocytosis and elevated proportion of T cells. However, the mainly affected system is the respiratory tract. In bronchial epithelium, metaplastic and dysplastic changes are accompanied by elevated expression of adhesion molecules and secretion of many proinflammatory cytokines. In the population of pulmonary macrophages, an elevated proportion of cells, changes in expression surface markers: CD14, CD54, CD11 and CD71 with impaired phagocytic and antigen presenting function were observed. The influence of tobacco smoke on the population of granulocytes results in elevated proportion of neutrophils and eosinophils and elevated concentration of the products of neutrophils: neutrophil elastase (NE), interleukin- 8 (IL-8) and tumor necrosis factor (TNFα). Tobacco smoke seems to alter the way of death of neutrophils from apoptosis to necrosis. On the other hand, an elevated expression of the receptors of apoptosis on lymphocytes was found. Cigarette smoke enhances the recirculation of lymphocytes, which results in the augmentation of activated and cytotoxic/ suppressor cells in the bronchial lumen. Smoking cessation is the most effective method of prophylaxis and treatment of diseases related to tobacco smoking. However, many immunological changes in smokers are not completely reversible after quitting smoking.

Eicosanoid regulation of pulmonary innate immunity post-hematopoietic stem cell transplantation

Hematopoietic stem cell transplantation (HSCT) is a therapeutic option for a number of malignant and inherited disorders. However, the efficacy of this therapy is limited by a number of serious infectious and noninfectious complications. Pulmonary infections represent a significant cause of morbidity and mortality post-HSCT and can occur both pre- and post-hematopoietic reconstitution. Susceptibility to Gram-negative bacterial infections despite full hematopoietic engraftment suggests that innate immunity remains impaired months to years post-HSCT. This review will describe the process and complications of HSCT and will summarize what is known about innate immune reconstitution post-HSCT. Data from the literature as well as our own laboratory will be presented to suggest that an eicosanoid imbalance characterized by over-production of prostaglandins and under-production of leukotrienes leads to impaired lung phagocyte function post-HSCT. Of therapeutic interest, strategies which limit production of prostaglandins can improve pulmonary host defense in animal HSCT models, which suggests that this may also be beneficial for human HSCT recipients.

Iron transport by Nramp2/DMT1: pH regulation of transport by 2 histidines in transmembrane domain 6

Mutations at natural resistance-associated macrophage protein 1 (Nramp1) impair phagocyte function and cause susceptibility to infections while mutations at Nramp2 (divalent metal transporter 1 [DMT1]) affect iron homeostasis and cause severe microcytic anemia. Structure-function relationships in the Nramp superfamily were studied by mutagenesis, followed by functional characterization in yeast and in mammalian cells. These studies identify 3 negatively charged and highly conserved residues in transmembrane domains (TM) 1, 4, and 7 as essential for cation transport by Nramp2/DMT1. The introduction of a charged residue (Gly185Arg) in TM4 found in the naturally occurring microcytic anemia mk (mouse) and Belgrade (rat) mutants is shown to cause a partial or complete loss of function in mammalian and yeast cells, respectively. A pair of mutation-sensitive and highly conserved histidines (His267, His272) was identified in TM6. Surprisingly, inactive His267 and His272 mutants could be rescued by lowering the pH of the transport assay. This indicates that His267/His272 are not directly involved in metal binding but, rather, play an important role in pH regulation of metal transport by Nramp proteins.

Hyperoxia impairs antibacterial function of macrophages through effects on actin.

Oxidative stress may impair alveolar macrophage function in patients with inflammatory lung diseases or those exposed to high concentrations of oxygen. We investigated putative mechanisms of injury to macrophages by oxidative stress, using RAW 264.7 cells exposed to 95% oxygen for 48 h. Hyperoxia-exposed macrophages were less able to phagocytose and kill Klebsiella pneumoniae than normoxic controls, despite increased production of nitric oxide, a free radical important in pathogen killing. Exposure of macrophages to hyperoxia had marked effects on the actin cytoskeleton, including increased actin polymerization, loss of cortical actin, formation of stress fibers, de novo synthesis of actin, and actin oxidation. Hyperoxia induced changes in cell morphology, with increased cell size and pseudopod formation. Exposure of macrophages to jasplakinolide, an agent that increases actin polymerization, also impaired their ability to phagocytose Klebsiella. Alveolar macrophages isolated from mice exposed to 100% oxygen for 84 h also demonstrated impaired phagocytic function, as well as similar effects on the actin cytoskeleton and cell morphology to macrophages exposed to hyperoxia in vitro. We conclude that oxidative stress in vitro and in vivo impairs macrophage antibacterial function through effects on actin.