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Although obesity is implicated in numerous health complications leading to increased mortality, the relationship between obesity and outcomes for critically ill patients appears paradoxical. Recent studies have reported better outcomes and lower levels of inflammatory cytokines in obese patients with acute lung injury (ALI)/acute respiratory distress syndrome, suggesting that obesity may ameliorate the effects of this disease. We investigated the effects of obesity in leptin-resistant db/db obese and diet-induced obese mice using an inhaled LPS model of ALI. Obesity-associated effects on neutrophil chemoattractant response were examined in bone marrow neutrophils using chemotaxis and adoptive transfer; neutrophil surface levels of chemokine receptor CXCR2 were determined by flow cytometry. Airspace neutrophilia, capillary leak, and plasma IL-6 were all decreased in obese relative to lean mice in established lung injury (24 h). No difference in airspace inflammatory cytokine levels was found between obese and lean mice in both obesity models during the early phase of neutrophil recruitment (2–6 h), but early airspace neutrophilia was reduced in db/db obese mice. Neutrophils from uninjured obese mice demonstrated diminished chemotaxis to the chemokine keratinocyte cytokine compared with lean control mice, and adoptive transfer of obese mouse neutrophils into injured lean mice revealed a defect in airspace migration of these cells. Possibly contributing to this defect, neutrophil CXCR2 expression was significantly lower in obese db/db mice, and a similar but nonsignificant decrease was seen in diet-induced obese mice. ALI is attenuated in obese mice, and this blunted response is in part attributable to an obesity-associated abnormal neutrophil chemoattractant response.

Obesity is a risk factor for asthma. Studies in mice suggest that the adipokines leptin and adiponectin affect asthmatic responses. The purpose of this study was to determine if adipokines associated with obesity are (1) altered in obese women with asthma compared to controls and (2) associated with increased cytokines and chemokines involved in allergic inflammation.We performed a cross-sectional study of asthmatic and non-asthmatic obese premenopausal women. Participants answered questionnaires and performed lung function tests. Serum and peripheral blood mononuclear cells (PBMCs) were collected for analysis of cytokines and adipokines.A total of 22 asthmatic (mean body mass index 40.0 ± 5.1 kg/m(2)) and 20 non-asthmatic women (mean body mass index 41.3 ± 5.6 kg/m(2)) participated. We found no difference in serum adipokine concentrations between asthmatics and non-asthmatics. Serum adiponectin correlated positively with PBMC eotaxin (r(s) = 0.55, p = .0003) and RANTES (regulated upon activation, normal T-cell expressed, and secreted) (r(s) = 0.36, p = .03), whereas serum leptin correlated negatively with PBMC eotaxin (r(s) = -0.34, p = .04). There was a negative correlation between serum adiponectin and PBMC interferon-γ (r(s) = -0.41, p = .01).Perturbations of adipokines that occur in obesity were correlated with decreased cytokine production typically associated with allergic responses in PBMC of obese premenopausal women. This study suggests that although obese asthmatics may have elements of Th2-mediated inflammation, adipokine derangements in obesity are associated with Th1 rather than Th2 bias. Obesity has complex effects on allergic inflammation and is likely to be important modifier of the pathogenesis of airway disease in asthma.

IL-1β is a cytokine critical to several inflammatory diseases in which pathogenic Th17 responses are implicated. Activation of the NLRP3 inflammasome by microbial and environmental stimuli can enable the caspase-1–dependent processing and secretion of IL-1β. The acute-phase protein serum amyloid A (SAA) is highly induced during inflammatory responses, wherein it participates in systemic modulation of innate and adaptive immune responses. Elevated levels of IL-1β, SAA, and IL-17 are present in subjects with severe allergic asthma, yet the mechanistic relationship among these mediators has yet to be identified. In this study, we demonstrate that Saa3 is expressed in the lungs of mice exposed to several mixed Th2/Th17-polarizing allergic sensitization regimens. SAA instillation into the lungs elicits robust TLR2-, MyD88-, and IL-1–dependent pulmonary neutrophilic inflammation. Furthermore, SAA drives production of IL-1α, IL-1β, IL-6, IL-23, and PGE2, causes dendritic cell (DC) maturation, and requires TLR2, MyD88, and the NLRP3 inflammasome for secretion of IL-1β by DCs and macrophages. CD4+ T cells polyclonally stimulated in the presence of conditioned media from SAA-exposed DCs produced IL-17, and the capacity of polyclonally stimulated splenocytes to secrete IL-17 is dependent upon IL-1, TLR2, and the NLRP3 inflammasome. Additionally, in a model of allergic airway inflammation, administration of SAA to the lungs functions as an adjuvant to sensitize mice to inhaled OVA, resulting in leukocyte influx after Ag challenge and a predominance of IL-17 production from restimulated splenocytes that is dependent upon IL-1R signaling.

With the rising tide of obesity worldwide, clinicians and researchers alike find themselves struggling to understand not only what this means to human health, but also how we should best analyze and characterize weight gain (or loss) itself. Obesity as a disease modifier in vascular disease and diabetes has often been simplified to the concept of “risk factor”; yet, as we are learning in lung disease, our understanding must be far more nuanced to capture the complex effects associated with weight change. This is highlighted by the growing number of clinical “paradoxes” associated with obesity in which weight gain appears to confer not only health risks but health benefits, often in an unpredictable fashion (1). The issues that frame these paradoxes typically reflect our inability to explain these divergent associations mechanistically. Traditionally, our understanding of weight gain and its effects has centered on the concept of obesity as an inflammatory disease (2). However, although useful in the context of vascular disease, this concept often fails to predict the effects of changes in body mass index (BMI) on the lung and lung disease. For example, recent reports have demonstrated that in acute lung injury, obese patients have lower levels of circulating inflammatory cytokines (3) (and improved outcomes [4]) compared with lean patients, and in obese individuals with asthma, surgically induced weight loss is accompanied by evidence of increased airway inflammation (5, 6). These findings speak to a far more complicated relationship between weight change and lung function than can be ascribed solely to proinflammatory effects or even, for that matter, to immune modulation at all, and growing evidence suggests that an interplay of adipokine disarray (6–8), mechanical perturbations (9), and changes in muscle mass (10, 11) is likely to also influence lung disease and its manifestations. In this wider context, let us consider the interesting study by Sood and colleagues (pp. 274–281) in this issue of the Journal (12). These investigators report an examination of 1,641 current and former smokers at risk for or with mild to moderate chronic obstructive pulmonary disease (COPD), followed for a median of 6 years as part of the Lovelace Smokers’ Cohort in New Mexico, and present both cross-sectional and longitudinal data relating subjects’ BMI to spirometric variables and health status (as assessed by the St. George Respiratory Questionnaire [SGRQ]). Stratifying their cohort by World Health Organization BMI category, they found that at baseline, higher BMI in obese (BMI > 30 kg/m2) smokers was associated with worse health status and lower FEV1, yet in the lean (BMI < 25 kg/m2) smokers, the association between BMI and these variables was reversed. The authors report that these findings delineate a “U-shaped” association between BMI and the extreme weight categories, such that both the morbidly obese and the lean to underweight subsets of their cohort had the lowest FEV1 and worst health status. Even more interestingly, in longitudinal analyses, they found that these opposing relationships appear to be real since they applied to weight gain over the duration of the study as well: rising BMI was associated with worsening health status and falling FEV1 in obese smokers, yet improvement in these outcomes in lean smokers. Interestingly, the authors found no association between IL-6, IL-10, or leptin and outcomes. Do these findings present yet another “paradox” or do they underscore a complex dichotomy between BMI, lung function, and respiratory symptoms in smokers? Given the methods of analysis, these findings might be more intuitively understood as follows: weight loss is better tolerated by the obese than the lean in their cohort. The authors attempt to dissect weight gain from weight loss in their supplemental data (Table E2), which reveal roughly equal numbers of subjects gaining and losing weight within each BMI category. In the obese cohort, both weight gain and weight loss are strongly associated with inverse changes in FEV1 and SGRQ. However, in the lean cohort, although SGRQ appears similarly bidirectional in it association, only weight loss is significantly associated with FEV1, and in a markedly downward manner. These findings suggest that in smokers without severe COPD, weight change signifies something quite different in lean versus obese patients. But what? To answer that question, let’s consider the contrast between the Lovelace Smokers’ cohort findings and those of The Copenhagen City Heart Study (13). The latter study found that weight loss was associated with increased COPD-related mortality across the BMI spectrum of their cohort, whereas weight gain was protective. Further examination of this cohort and others suggests that much of this effect may be related to low or decreasing fat-free body mass—presumed here to indicate sarcopenia (10, 14, 15). Further studies by Marquis and colleagues (16) and Rutten and colleagues (11) have gone further to prove this association. It is understandable that loss of muscle mass as a consequence of worsening COPD, inflammation, poor nutrition, or other comorbidities might compromise pulmonary physiology and overall health status. The same effect is seen with falling BMI in the lean (BMI < 25) subset of the Lovelace cohort. How does this explain the findings in the rest of this cohort? Here it is worth noting that in the previously cited studies, mean BMIs of the cohorts were in the range of 24–25, whereas in the Lovelace cohort, the mean was nearly 28. Furthermore, unlike previous studies, the Lovelace cohort is predominantly women (>70%). Although the ratio of fat mass (FM) to fat-free mass (FFM) generally rises with BMI, it is well known that both sex and age significantly affect FM/FFM balance. For any given BMI, women have a significantly higher percentage of FM compared with men (Figure 1), and FM/FFM ratio rises with age for both sexes (17). Thus, the differences seen between overall BMI/outcome associations in the current study compared with previous studies, including the Copenhagen, are likely to reflect the effects of greater FM on the higher BMI subsets. What are these effects? Figure 1. Percentages of total body mass from fat mass (FM) and fat-free mass (FFM) in males and females across the body mass index (BMI) range. Data shown are for subjects 45 years of age (16). In this study, obese smokers had a markedly lower incidence of Global Initiative for Chronic Obstructive Lung Disease (GOLD) criteria–defined COPD (17% vs. 35% in lean smokers) despite having a higher incidence of wheeze and dyspnea and a nearly 20% rate of bronchodilator response at baseline. Although this may in part reflect a notable failure of the GOLD criteria to capture early COPD in the obese (likely due to obesity-driven elevation of the FEV1/FVC ratio), it may also suggest the presence of an additional, discrete syndrome in the obese (female) smoker. This is underscored by the salutary effects of weight loss in this subset—a distinctly different finding than reported for subjects in the Copenhagen Study and others. As Sood and others have shown previously, weight gain and obesity are associated with an increased incidence of airway obstruction and wheezing in the general population, particularly among women, irrespective of smoking status (18, 19), whereas weight loss improves these symptoms, particularly in the nonatopic obese (5). Although generally described as “patients with asthma,” this group of patients has recently been recognized as a distinct phenotype within this syndrome (20). The current work therefore begs the question, what is this form of obese obstructive lung disease that variously appears as difficult-to-treat asthma or the dyspneic, wheezing smoker? As the authors find no association between these findings and a (limited) panel of cytokines and adipokines, it is tempting to invoke the mechanical effects of obesity in this patient subset that hopefully future studies will delineate. The only thing we know for sure is that smoking cessation and weight loss are likely the most effective (and only?) treatments for this rapidly growing population of patients.

Obesity is a major epidemic in the United States and is associated with increased morbidity related to cardiovascular disease, diabetes, and asthma.1,2 These morbidities are thought to result, in part, from the production of pathogenic mediators by adipose tissue. Adipose tissue in the individual who is obese produces multiple proinflammatory cytokines, including tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), IL-6, and IL-8. Elevated levels of these cytokines in patients who are obese have been linked to the development of atherosclerotic cardiovascular disease, hypertension, and insulin resistance, and it is believed they may have systemic effects mediating many of the other medical complications of obesity.1,3-7 Thus, obesity represents a chronic inflammatory state. Acute lung injury (ALI) and its more severe form, acute respiratory distress syndrome, are characterized by an overexuberant inflammatory response in the lung associated with and perhaps driven by elevated plasma levels of inflammatory cytokines.8 Previous clinical investigations have demonstrated an association between increased mortality and high circulating levels of TNF-α, IL-1β, IL-6, and IL-8 in ALI.9-13 Such an association is also seen with biomarkers of epithelial injury (surfactant protein D [SP-D]) and vascular injury (soluble intracellular adhesion molecule [sICAM] and von Willebrand factor [vWF]), as well as disordered coagulation (plasminogen activator inhibitor 1 [PAI 1] and protein C).14-18 Because many of these same inflammatory cytokines are elevated at baseline in individuals who are obese,19 one might anticipate that individuals who are obese would have more severe lung injury and worse clinical outcomes from ALI. However, prior studies of patients with ALI have reported that mortality does not increase, and may even be lower with increasing BMI.20-22 Thus, an apparent paradox exists that patients who are obese, who manifest a proinflammatory state with increased circulating cytokines in their normal state, do not have increased mortality in the setting of critical illness with ALI, which is a condition driven, at least in part, by a proinflammatory cytokine milieu. Given this paradox in patients with ALI, we hypothesized that one of the following scenarios may be occurring: (1) increasing BMI is associated with decreased levels of circulating inflammatory cytokines, (2) increasing BMI is associated with more rapid attenuation of cytokine release during the course of ALI, or (3) BMI alters the relationship between baseline cytokine levels and mortality and morbidity from ALI. In order to address these hypotheses and to examine the effects of obesity on other biomarkers associated with outcomes in ALI, we analyzed data from prior National Heart, Lung, and Blood Institute (NHLBI) Acute Respiratory Distress Syndrome Network (ARDSNet) clinical trials in ALI because they present a large, well-characterized patient population with plasma biomarker measurements (at enrollment and Day 3) and information on outcomes from ALI.

To evaluate the association between plasma granulocyte colony-stimulating factor (G-CSF) levels and clinical outcomes including mortality in patients with acute lung injury (ALI), and to determine whether lower tidal volume ventilation was associated with a more rapid decrease in plasma G-CSF over time in patients with ALI.Retrospective measurement of G-CSF levels in plasma samples that were collected prospectively as part of a large multicenter clinical trial.Intensive care units in ten university centers.The study included 645 patients enrolled in the National Heart, Lung, and Blood Institute Acute Respiratory Distress Syndrome Clinical Network trial of lower tidal volumes compared with traditional tidal volumes for ALI.Baseline plasma levels of G-CSF were associated with an increased risk of death and a decrease in ventilator-free days and organ failure-free days in multivariate analyses controlling for ventilation strategy, age, and sex (Odds ratio death 1.2/log10 increment G-CSF, 95% confidence interval 1.01 to 1.4). Stratification of G-CSF levels into quartiles revealed a strong association between the highest levels of G-CSF and an increased risk of death and decreased ventilator-free days and organ failure-free days in multivariate analyses controlling for ventilation strategy, Acute Physiology and Chronic Health Evaluation III score, Pao2/Fio2 ratio, creatinine, and platelet count (p0.05). Subgroup multivariate analysis of patients with sepsis as their risk factor for ALI revealed a U-shaped association between mortality and G-CSF levels such that risk increased linearly from the second through fourth (highest) quartiles, yet also increased in the first (lowest) quartile. G-CSF levels decreased over time in both tidal volume groups, and there was no statistical difference in the extent of decrease between ventilator strategies.In patients with ALI, plasma G-CSF levels are associated with morbidity and mortality, but these levels are not influenced by tidal volume strategy. In patients with sepsis-related ALI, a bimodal association between baseline plasma G-CSF levels and subsequent morbidity and mortality from this disease was found.

Neutrophil retention in and release from the bone marrow is a critical process that remains incompletely understood. Previous work has implicated the CXCR4/stromal derived factor-1 (SDF-1) chemokine axis in the marrow retention of neutrophils, yet the adhesion pathways responsible for this retention are unknown. Because α4β1 integrin (VLA-4) and its ligand VCAM-1 play a central role in the interactions of hematopoietic stem cells, lymphocytes, and developing neutrophils in the marrow, we investigated whether this integrin might be involved in marrow neutrophil retention and release. In this study, we show that VLA-4 is expressed on murine marrow neutrophils and decreases with maturation, whereas blockade of this integrin leads to the release of marrow neutrophils. Marrow neutrophils adhere via VLA-4 to VCAM-1, which is expressed on marrow endothelium and stroma, and inhibition of VCAM-1 causes release of marrow neutrophils. Furthermore, SDF-1 (CXCL12) signaling through neutrophil CXCR4 augments VLA-4 adhesion to VCAM-1 in vitro, an effect that is blocked by preincubation with pertussis toxin. In vivo blockade of both CXCR4 and α4 causes synergistic release of marrow neutrophils, showing that cross-talk between CXCR4 and VLA-4 modulates marrow retention of these cells. Taken together, these results indicate that the VLA-4/VCAM adhesion pathway is critical in the retention and maturation-controlled release of neutrophils from the marrow, while providing an important link between the CXCR4/SDF-1 signaling axis and the adhesion events that govern this process.

Study Objectives The purpose of this study was to determine if asthma with rhinitis and asthma without rhinitis represent distinct forms of disease. Design We performed a prospective cross-sectional study. Participants The study included healthy controls, participants with asthma without rhinitis, and participants with both asthma and rhinitis. Interventions We compared lung function and airway inflammation between the three groups of participants. Results We recruited 32 participants: 12 normals, 8 asthmatics without rhinitis, and 12 with rhinitis. Compared to asthmatics with rhinitis, asthmatics without rhinitis had more severe airflow limitation (FEV1/FVC = 60.6% [IQR = 22.8] vs. 74.8% [IQR = 7.8] and fewer induced sputum eosinophils (2.8 [IQR = 5.8] and 9.6 [IQR = 23.8], respectively). Sputum interleukin-6 correlated inversely with lung function measured by postbronchodilator FEV1 in the study cohort (Spearman correlation coefficient = −0.55, p

Rationale: Recent studies have suggested that both embryonic stem cells and adult bone marrow stem cells can participate in the regeneration and repair of diseased adult organs, including the lungs. However, the extent of airway epithelial remodeling with adult marrow stem cells is low, and there are no available in vivo data with embryonic stem cells. Human umbilical cord blood contains both hematopoietic and nonhematopoietic stem cells, which have been used clinically as an alternative to bone marrow transplantation for hematologic malignancies and other diseases. Objectives: We hypothesized that human umbilical cord blood stem cells might be an effective alternative to adult bone marrow and embryonic stem cells for regeneration and repair of injured airway epithelium. Methods: Human cord blood was obtained from normal deliveries at the University of Vermont. Cultured plastic adherent cells were characterized as mesenchymal stem cells (MSCs) by flow cytometry and differentiation assays. Cord blood–derived MSCs (CB-MSCs) were cultured in specialized airway growth media or with specific growth factors, including keratinocyte growth factor and retinoic acid. mRNA and protein expression were analyzed with PCR and immunofluorescent staining. CB-MSCs were systematically administered to immunotolerant, nonobese diabetic/severe combined immunodeficiency (NOD-SCID) mice. Lungs were analyzed for presence of human cells. Measurements and Main Results: When cultured in specialized airway growth media or with specific growth factors, CB-MSCs differentially expressed Clara cell secretory protein (CCSP), cystic fibrosis transmembrane conductance regulator (CFTR), surfactant protein C, and thyroid transcription factor-1 mRNA, and CCSP and CFTR protein. Furthermore, CB-MSCs were easily transduced with recombinant lentiviral vectors to express human CFTR. After systemic administration to immunotolerant, NOD-SCID, mice, rare cells were found in the airway epithelium that had acquired cytokeratin and human CFTR expression. Conclusions: CB-MSCs appear to be comparable to MSCs obtained from adult bone marrow in ability to express phenotypic markers of airway epithelium and to participate in airway remodeling in vivo.

The severe and protracted inflammation that characterizes acute lung injury (ALI) is driven by the ongoing recruitment of neutrophils to the lung. Although much of the cytokine signaling responsible for the initial phase of ALI has been elaborated, relatively little is known about the mechanisms governing the recruitment of neutrophils from the bone marrow to the lung in the later period of this disease. Given its previously described chemoattractant effects on marrow neutrophils, we investigated whether stromal-derived factor-1 (SDF-1) (CXCL12) might participate in this later phase of recruitment. Using immunohistochemistry to examine both banked human lung specimens from patients with ALI and lungs from mice with LPS-induced pneumonitis, we found that pulmonary SDF-1 expression increases during ALI. We further determined that both lung SDF-1 protein expression and mRNA expression rise in a delayed but sustained pattern in this mouse model and that the major source of the increase in expression appears to be the lung epithelium. Lastly, we found that expression of the SDF-1 receptor CXCR4 rises in a similar temporal pattern on neutrophils in both the blood and airspace of LPS-injured mice and that Ab-mediated SDF-1 blockade significantly attenuates late but not early pulmonary neutrophilia in this model. These results implicate SDF-1 in neutrophil recruitment to the lung in the later period of acute lung injury and suggest a novel role for this cytokine in coordinating the transition from the inflammatory response to the initiation of tissue repair.

Cystic fibrosis (CF) was one of the first target diseases for lung gene therapy. Studies of lung gene transfer for CF have provided many insights into the necessary components of successful gene therapy for lung diseases. Many advancements have been achieved with promising results in vitro and in small animal models. However, studies in primate models and patients have been discouraging despite a large number of clinical trials. This reflects a number of obstacles to successful, sustained, and repeatable gene transfer in the lung. Cell-based therapy with embryonic stem cells and adult stem cells (bone marrow or cord blood), have been investigated recently and may provide a viable therapeutic approach in the future. In this article, the authors review CF pathophysiology with a focus on specific targets in the lung epithelium for gene transfer and summarize the current status and future directions of gene- and cell-based therapies.

Introduction: Obesity, a rapidly worsening epidemic in the developed world, is associated with increased risk for the development of ARDS. Alveolar macrophages are key participants in the overexuberant pulmonary inflammatory response that characterizes ARDS. We hypothesize that alveolar macrophage function and phenotype are abnormally skewed by obesity. Methods: Macrophage and neutrophil counts as well as cytokine levels were determined in bronchoalveolar lavage (BAL) fluid from lean (10% fat) and diet-induced obese (60% fat) mice at baseline. Alveolar macrophages were isolated from uninjured obese and lean mice exposed to LPS in vitro for 4h after which cytokine levels were determined in culture supernatant. Next, wild-type mice received PKH26-PCL dye via o.p. instillation where after they were placed on 10% vs. 60% fat diet for 10 weeks and BAL cell populations were analyzed by flow cytometry. Results: Alveolar macrophage numbers and MCP-1 levels were found to be elevated in BAL of uninjured obese mice. Furthermore, alveolar macrophages from the uninjured obese mice showed an exaggerated in vitro cytokine response to LPS. The proportion of PKH26-PCL positive macrophages remaining in BAL after 10 weeks of diet was greater in the 10% fat (lean) diet-fed mice compared to that found in 60% fat diet-fed mice, suggesting that obesity induces monocyte/macrophage recruitment from the circulation to the lung over time. Conclusion: Our findings indicate that obesity increases monocyte/macrophage recruitment to the lung and that alveolar macrophage function and phenotype manifest an exaggerated pro-inflammatory state at baseline. These changes may drive the increased risk of developing ARDS that manifests in obese humans and mice.

Obesity is a risk factor for the development of acute respiratory distress syndrome (ARDS) but mechanisms mediating this association are unknown. While obesity is known to impair systemic blood vessel function and predisposes to systemic vascular diseases, its effects on the pulmonary circulation are largely unknown. We hypothesized that the chronic low grade inflammation of obesity impairs pulmonary vascular homeostasis and primes the lung for acute injury. The lung endothelium from obese mice expressed higher levels of leukocyte adhesion markers and lower levels of cell-cell junctional proteins when compared to lean mice. We tested whether systemic factors are responsible for these alterations in the pulmonary endothelium; treatment of primary lung endothelial cells with obese serum enhanced the expression of adhesion proteins and reduced the expression of endothelial junctional proteins when compared to lean serum. Alterations in pulmonary endothelial cells observed in obese mice were associated with enhanced susceptibility to LPS-induced lung injury. Restoring serum adiponectin levels reversed the effects of obesity on the lung endothelium and attenuated susceptibility to acute injury. Our work indicates that obesity impairs pulmonary vascular homeostasis and enhances susceptibility to acute injury and provides mechanistic insight into the increased prevalence of ARDS in obese humans.

Acute lung injury/acute respiratory distress syndrome is a clinical syndrome that describes a single common end point: severe injury to the alveolar capillary membrane and the development of proteinaceous edema. Forty years of clinical and basic science research have elaborated many of the pathophysiologic mechanisms that link initiating insults to the development of alveolar membrane failure. A combination of genetics, comorbid conditions, inciting events, and factors yet to be discovered may uniquely alter the expression of these mechanisms in the individual patient.

Low tidal volume (Vt) ventilation is protective against ventilator-induced lung injury but can promote development of atelectasis. Periodic deep inflation (DI) can open the lung, but if delivered too frequently may cause damage via repeated overdistention. We therefore examined the effects of varying DI frequency on lung mechanics, gas exchange, and biomarkers of injury in mice. C57BL/6 males were mechanically ventilated with positive end-expiratory pressure (PEEP) of 2 cmH2O for 2 h. One high Vt group received a DI with each breath (HV). Low Vt groups received 2 DIs after each hour of ventilation (LV) or 2 DIs every minute (LVDI). Control groups included a nonventilated surgical sham and a group receiving high Vt with zero PEEP (HVZP). Respiratory impedance was measured every 4 min, from which tissue elastance ( H) and damping ( G) were derived. G and H rose progressively during LV and HVZP, but returned to baseline after hourly DI during LV. During LVDI and HV, G and H remained low and gas exchange was superior to that of LV. Bronchoalveolar lavage fluid protein was elevated in HV and HVZP but was not different between LV and LVDI. Lung tissue IL-6 and IL-1β levels were elevated in HVZP and lower in LVDI compared with LV. We conclude that frequent DI can safely improve gas exchange and lung mechanics and may confer protection from biotrauma. Differences between LVDI and HV suggest that an optimal frequency range of DI exists, within which the benefits of maintaining an open lung outweigh injury incurred from overdistention.

Adult marrow-derived stem cells can localize to lung and acquire immunophenotypic characteristics of lung epithelial cells. Lung injury increases recruitment of the marrow-derived cells. We speculated that comparing patterns of lung engraftment following different lung injuries would provide insight into potential mechanisms by which marrow-derived cells were recruited to lung. To evaluate this, adult female C57Bl/6 mice irradiated and engrafted with marrow from adult male transgenic GFP mice were exposed to either intranasal inhalation of endotoxin (25 μg/mouse) or 3 days of 25 ppm NO 2 and then compared 1 or 3 months later to transplanted but otherwise uninjured mice. In all cases, the majority of marrow-derived cells recruited to lung were CD45 + leukocytes. In lungs of transplanted but otherwise uninjured mice, small numbers of CD45 − donor-derived cells in alveolar septae stained positively for pro-surfactant protein C. Rare donor-derived cells located in the airway epithelium stained positively with cytokeratin. Subsequent exposure of engrafted mice to NO 2 or endotoxin did not significantly increase the number or pattern of donor-derived CD45 − cells found in recipient lungs. These results suggest that NO 2 or endotoxin lung injury does not result in significant engraftment of marrow-derived cells in lung.

The bone marrow is the primary site for neutrophil production and release into the circulation. Because the CXC chemokine receptor-4/stromal derived factor-1 (CXCR4/SDF-1) axis plays a central role in the interactions of hematopoietic stem cells, lymphocytes, and developing neutrophils in the marrow, we investigated whether reciprocal CXCR4-dependent mechanisms might be involved in neutrophil release and subsequent return to the marrow following circulation. Neutralizing antibody to CXCR4 reduced marrow retention of infused neutrophils (45.7% ± 0.5% to 6.9% ± 0.5%) and was found to mobilize neutrophils from marrow (34.4% ± 4.4%). Neutrophil CXCR4 expression and SDF-1-induced calcium flux decreased with maturation and activation of the cells, corresponding to the decreased marrow homing associated with these characteristics in vivo. Infusion of the inflammatory mediator and CXCR2 ligand KC led to mobilization of neutrophils from marrow by itself and was augmented 3-fold by low doses of CXCR4-blocking antibody that otherwise had no mobilizing effect. Examination of KC and SDF-1 calcium signaling demonstrated that the effect of KC may, in part, be due to heterologous desensitization to SDF-1. These results suggest that the CXCR4/SDF-1 axis is critical in circulating neutrophil homeostasis and that it may participate in the rapid release of neutrophils from the marrow during inflammation through a novel interaction with inflammatory CXC chemokines. (Blood. 2004;104:565-571)

An embryonic stem (ES) cell/OP9 coculture system for the effective production of functional neutrophils is described. A 3-step differentiation strategy was developed that uses liquid culture, enabling reliable and abundant production of neutrophils at high purity without the need of sorting for isolation of mature neutrophils. Use of the OP9 stromal cell line significantly enhances the number, percentage, and duration of differentiated neutrophils produced from embryonic stem cells. Effective and sustained differentiation of ES cells into neutrophils provides a useful model system for studying neutrophil differentiation and function and the factors that regulate them. Morphologic and functional evaluation of these ES-derived neutrophils indicates that large numbers of mature neutrophils can be produced from pluripotent ES cells in vitro. Specifically, their morphology, ability to produce superoxides, flux calcium, undergo chemotaxis in response to macrophage inflammatory protein 2 (MIP-2), stain for the granulocyte-specific marker–specific chloroacetate esterase, and contain the neutrophil-specific markers Gr-1 and the mouse neutrophil-specific antigen indicates that they are comparable with purified mouse bone marrow neutrophils. They also express gelatinase and lactoferrin granule proteins. During the differentiation of these ES-derived neutrophils, regional areas of neutrophil production can be identified that have been designated as neutrophil generating regions (NGRs).

Many of the body's tissues once thought to be only locally regenerative may, in fact, be actively replaced by circulating stem cells after hematopoietic stem cell transplantation. Localization of donor-derived cells ("chimerism") has recently been shown to occur in the lungs of mice after either hematopoietic stem cell transplantation or infusion of cultured marrow. To determine whether tissues of the human lung might be similarly derived from extrapulmonary sources, we examined lung specimens from a retrospective cohort of female allogeneic hematopoietic stem cell transplant recipients who received stem cells from male donors. Tissue samples from three such patients who had undergone diagnostic lung biopsy or autopsy were examined. Slides were stained by immunohistochemistry for cytokeratin (epithelium) and platelet endothelial cell adhesion molecule, CD31 (PECAM) (endothelium) and were imaged and then examined by fluorescent in situ hybridization analysis to identify male cells. The resulting overlapping in situ hybridization and immunohistochemistry images were examined for the presence and, if present, cell type of donor cells in the lung. We found significant rates of epithelial (2.5-8.0%) and endothelial (37.5-42.3%) chimerism. These results suggest that significant chimerism of the human lung may follow hematopoietic stem cell transplantation and that adult human stem cells could potentially play a therapeutic role in treatment of the damaged lung.

Problem Influenza infection severity may be mediated by estradiol and/or progesterone. Method of Study An exploratory study was designed to evaluate 17-β-estradiol and progesterone on influenza infection and examine immune-mediated response in a mouse model. Inoculation with placebo or mouse-adapted H1N1 influenza virus occurred. Treatment groups included 17-β-estradiol, progesterone, ovariectomy, and pregnancy. Mice were assessed for morbidity and mortality. Toll-like receptor gene studies and airspace cell differentials were performed. Results Onset of morbidity was earlier and morbidity duration greater for progesterone. Absence of morbidity/mortality and overall survival was greater for 17-β-estradiol. Airspace cell differentials suggest improved immune cell recruitment for 17-β-estradiol. Pregnant mouse data demonstrate significant mortality during the period of increased progesterone. Select immune cell markers demonstrate patterns of regulation that may promote proper immune response to influenza infection for 17-β-estradiol. Conclusion Estradiol may play a protective and progesterone a detrimental role in the pathophysiology of influenza infection.

Background The obesity has been shown to increase the severity of A/H1N1 infection and the development of acute respiratory distress syndrome (ARDS) and organ involvement. Methods Circulating levels of C-peptide, insulin, glucagon, leptin, acute phase reactants (procalcitonin, C-reactive protein, tissue plasminogen activator, and serum amyloids A and P), were measured in samples from 32 critically ill patients with A/H1N1 virus infection, 17 of whom had ARDS complicated by acute kidney injury (AKI) and 15 of whom had ARDS but did not develop AKI. Results Patients with ARDS and AKI (ARDS/AKI) had higher BMI and higher levels of C-peptide, insulin, leptin, procalcitonin and serum amyloid A compared to those ARDS patient who did not develop AKI. Adjusting for confounding variables using logistic regression analysis, higher levels of C-peptide (> 0.75 ng/mL) (OR = 64.8, 95% CI = 2.1–1980, p = 0.0006) and BMI > 30Kg/m2 (OR = 42.0, 95% CI = 1.2–1478, p = 0.04) were significantly associated with the development of AKI in ARDS patients. Conclusion High levels of C-peptide and BMI > 30 kg/m2 were associated with the development of AKI in ARDS patients due to A/H1N1 infection. These metabolic/obesity indicators, together with the profiles of pro-inflammatory acute phase proteins, may be important links between obesity and poor outcomes in A/H1N1 09 infection.

Obesity is a global epidemic, not simply a disease of the Western world (1), and this has grave implications for lung health. There is a growing appreciation that obesity is associated with asthma, but the study by Chen and colleagues (pp. 1194–1203) in this issue of the Journal drives home some particularly salient features of this epidemic, which are worthy of attention from clinicians and policymakers alike (2). Chen and colleagues present results of a study of 2,758 Taiwanese children, in which they examined the relationship between asthma, obesity, physical fitness, and sedentary time. The investigators enrolled fourth grade school children and followed them over 3 years with systematic annual evaluations for lung disease and measures of obesity. The approach taken, with multiple anthropomorphic measures and systematic annual follow-up, provides confidence in their conclusions. Central obesity, among measures of adiposity, was most strongly associated with the presence of asthma in their cohort. Of particular interest, they show data suggesting that low physical fitness and high “screen time” (television/video games) lead to central obesity, which appears to alter lung function as measured by FEV1/FVC. Physical inactivity is likely to be an important, perhaps modifiable, risk factor for the development of asthma in children. Central obesity was particularly associated with active asthma. This is a finding that is now gaining some traction in the literature, as many investigators have variously reported that central obesity is a risk factor for low lung function (3) and for the development of incident asthma (4). Central obesity clearly has mechanical effects on the thoracic cavity, and so it is perhaps self-evident that central obesity could lead to low lung volumes. Breathing at low lung volumes in normal human volunteers can induce airway reactivity (5), and may therefore contribute to the development of asthma, particularly in those with central obesity, but this is unlikely to be the whole story; airway reactivity in animal models appears not to be related to low lung volumes (6). Thus, other factors must be at play. Central obesity is produced by deposition of visceral adipose tissue, and this form of adipose tissue is far more pathogenic in terms of producing proinflammatory mediators and is more closely related to the metabolic complications of obesity than subcutaneous adipose tissue (7, 8). We have previously found that inflammation in visceral adipose tissue is associated with asthma in obesity (9). Mediators produced in abundance by visceral adipose tissue could have direct effects on the airways, or perhaps produce changes in immune responses that predispose to the development of asthma in obesity. How central obesity is related to asthma is an important scientific question that will require much additional study to understand, but there are immediate clinical implications of the current study in which central obesity was strongly associated with low physical activity and high screen time. This is an all-too-common scenario, as increasing numbers of children (and adults) all over the world live in urban environments with sedentary lifestyles and limited access to physically active recreational opportunities. Addressing this lifestyle issue is clearly one of the most pressing public health issues of our time. Pulmonologists have long focused on public health issues, such as smoking; obesity is now the pressing pandemic that needs our urgent attention. This study provides important scientific and clinical insights that suggest we need to pay far more attention to the role of ubiquitous modern lifestyle factors and the role that these factors play in the development of lung disease. Solving the obesity pandemic is clearly a challenge for the 21st century, but there are likely to be interventions that can be promoted during routine medical care. Those caring for patients with asthma routinely counsel patients about smoke and other environmental exposures. It is now time to start focusing on lifestyle factors such as diet and exercise as major modifiable risk factors contributing to their disease. This is likely to be particularly important for patients who are overweight, as a growing body of evidence suggests that obese patients with asthma do not respond as well to steroid controller therapy as lean patients, and tend to have more poorly controlled asthma (10). Promoting lifestyle changes may be particularly challenging in those with asthma, as patients with asthma may experience greater symptoms with physical activity, and so have more difficulty undertaking regular exercise. There are a few small studies that suggest that successful weight loss can be achieved in individuals with asthma through lifestyle modification, and this weight loss is associated with improvements in asthma control (11–13). By focusing on prevention of obesity in children, we are likely to prevent the development of a lifelong disorder that will produce lifelong pulmonary morbidity. Clinicians need to incorporate lifestyle counseling into the routine care of patients with asthma, and there is a pressing need to develop systematic lifestyle interventions that will work in this challenging patient population.

Hypoxia inducible factor 1α (HIF1α) is the mammalian transcriptional factor that controls metabolism, survival, and innate immunity in response to inflammation and low oxygen. Previous work established that generation of hypoxic microenvironments occurs within the lung during infection with the human fungal pathogen Aspergillus fumigatus. Here we demonstrate that A. fumigatus stabilizes HIF1α protein early after pulmonary challenge that is inhibited by treatment of mice with the steroid triamcinolone. Utilizing myeloid deficient HIF1α mice, we observed that HIF1α is required for survival and fungal clearance early following pulmonary challenge with A. fumigatus. Unlike previously reported research with bacterial pathogens, HIF1α deficient neutrophils and macrophages were surprisingly not defective in fungal conidial killing. The increase in susceptibility of the myeloid deficient HIF1α mice to A. fumigatus was in part due to decreased early production of the chemokine CXCL1 (KC) and increased neutrophil apoptosis at the site of infection, resulting in decreased neutrophil numbers in the lung. Addition of recombinant CXCL1 restored neutrophil survival and numbers, murine survival, and fungal clearance. These results suggest that there are unique HIF1α mediated mechanisms employed by the host for protection and defense against fungal pathogen growth and invasion in the lung. Additionally, this work supports the strategy of exploring HIF1α as a therapeutic target in specific immunosuppressed populations with fungal infections., Author Summary Due to the limited treatment options and severity of invasive fungal infections, a better understanding of fungal-host interactions is needed for the development of new therapies. Recent studies have implicated a role for hypoxia inducible factor 1-alpha (HIF1α) in the regulation of inflammation and host defense responses to microbial pathogens. In this study, we discover that HIF1α is required for protection and murine survival to Aspergillus fumigatus pulmonary challenge. First, we observed that nuclear HIF1α protein levels are reduced in the murine corticosteroid immunosuppressed model of invasive pulmonary aspergillosis, suggesting its involvement in disease outcome. We then tested the hypothesis that HIF1α is required by innate immune effector cells to control/prevent A. fumigatus growth and invasion. Surprisingly, we observed that the role of myeloid HIF1α is not to mediate innate effector cell A. fumigatus killing directly, but rather to induce and maintain a protective immune response that ensures proper effector cell recruitment and survival at the site of infection. These findings provide a better understanding of host mechanisms involved in thwarting fungal pathogenesis, have implications for host susceptibility, and reveal the potential for novel treatment strategies involving HIF1α mediated signaling in the lung in immune suppressed patients.

While it has long been recognized that obesity is a risk factor for diseases such as type II diabetes and steatohepatitis, it has only recently been appreciated that obesity is a major risk factor for lung diseases such as asthma, sleep apnea and likely also pulmonary hypertension. Obesity also dramatically alters the clinical course of other common lung diseases such as chronic obstructive pulmonary disease and acute lung injury. The purpose of this chapter will be first to review the effects of obesity on lung function, and then to discuss the effect of obesity on the aforementioned pulmonary diseases.

Obesity has reached alarming levels with approximately one third of the adult U.S. population now obese and projected to rise to over 50% by the year 2030 [1,2]. It is startling to realize that only a minority of the U.S. population are of normal weight. This epidemic is not at all unique to the United States; it is a global epidemic that is changing the nature of human health and disease [3]. The articles in this Special Issue of PPT focus on some of the major ways that obesity is affecting lung health. The reports highlight important new findings pertaining to the effects of obesity on lung physiology and pathology; they show how obesity is transforming the presentation of common lung diseases. Understanding the effects of obesity is now essential for anyone in the field of lung health. Furthermore, although there exists a substantial body of work detailing the cardiovascular and endocrinological effects of obesity and the metabolic syndrome, comparably little is known in the fields of pulmonary and critical care. That obesity produces major changes in lung physiology has been known for many years. How this relates to diseases such as asthma, is just beginning to be understood. The contribution by Mahadev et al. [4] highlights changes in respiratory physiology that occur with obesity and how this may lead to symptoms of asthma. Obesity increases the occurrence of expiratory flow limitation during tidal breathing in response to bronchoconstriction in both asthmatics and non-asthmatic controls. This is clearly associated with symptoms of breathlessness. Expiratory flow limitation during tidal breathing is one mechanism that could contribute to the development of severe asthmatic symptoms in the setting of obesity; this has major implications for our approach to the treatment of such patients. Mediators produced by adipose tissue, adipokines, play an important role in physiological processes in the lung tissue, and also fundamentally alter both innate and adaptive immune responses. Vernooy et al. [5] review the pleiotropic effects of leptin on such diverse processes as lung development and function, and innate and adaptive immune responses, and the potential importance of this to a number of human diseases [5]. In separate articles, Konter et al. [6] and Medoff [7] review the effects of adiponectin on the alveolar-capillary membrane and the pulmonary vasculature and inflammation. Studies of the effects of adipokines in the lung are going to be an important area of investigation in the coming years. Obesity is characterized by a number of metabolic derangements, which include abnormalities of cholesterol metabolism. Disorders of cholesterol metabolism are likely to have broad effects on innate and adaptive immunity, which in turn have important implications for many lung diseases as outlined in the paper by Gowdy and Fessler [8]. One such disease is acute lung injury. Obesity is a risk factor for acute lung injury as outlined in the paper by Konter et al. [6]. It also appears to alter the course of ALI, through apparently anti-inflammatory effects [9]. One possible mechanism for this finding is through the effects of hypercholesterolemia. Palvinskya et al. [10] show how chronic hypercholesterolemia impairs neutrophil functional responses and that these defects may be in part due to protracted signaling responses to low density lipoprotein and its oxidized forms. Thus, hypercholesterolemia may be an important factor altering the course of acute lung injury and immune responses in the lung [10]. Respiratory infections are fundamentally altered by the presence of obesity. This was highlighted by the 2009 H1N1 pandemic in which obese individuals were at high risk of severe disease. The paper by Mancuso reviews some of the recent work on obesity and both viral and bacterial pneumonia [11]. In particular, Dr. Mancuso reviews how leptin and adiponectin may be involved in both the pulmonary innate and adaptive immune response, and how the absence of or resistance to these adipokines may alter host defense and thus the susceptibility to pulmonary infections. Understanding how obesity may affect response to immunizations and common pulmonary infectious diseases is going to be critical if we are to develop a rational approach to treating patients with pulmonary infections in the coming years. The link between obesity and asthma has been recognized for at least the last decade, and there has been important progress made in understanding the link between these diseases which is addressed in papers in this issue. The publication by Williams et al. [12] demonstrates that airway reactivity in obese mice is eliminated in mice deficient in tumor necrosis factor-receptor 2, and this appears to be associated with increased levels of interleukin 17A in bronchoalveolar lavage fluid, implicating these pathways in the development of asthma in obesity. The publication by Sideleva et al. reviews the epidemiological links between obesity and asthma, and discusses the importance of the recently recognized phenotypes of asthma in obesity. These new insights are likely to drive a critical paradigm shift in our understanding of asthma in obesity, which will move forward our understanding and approach to these patients [13]. Recent studies do suggest that obesity is a risk factor for pulmonary hypertension, and work described by Medoff in this issue show that adiponectin may be a key factor involved in the pathogenesis of pulmonary hypertension in obesity [7]. Adiponectin levels decrease in the setting of obesity, and this may be important as adiponectin can reduce vascular remodeling in response to environmental insults such as inflammation and hypoxia. Interestingly, a role for circulating adiponectin as a protector of the alveolar-capillary membrane as well is suggested by Konter et al. in their review on obesity and acute lung injury in this issue [6]. Understanding the link between obesity and pulmonary vasculopathy should be an important research priority in the coming years. Obesity is a multi-faceted disease process, often both characterized and driven by physical inactivity. The publication by Stevenson & Yew-Booth provides insights into the potential importance of this in lung disease [14]. The authors review data suggesting that exercise and aerobic capacity are important predictors of outcome in many diseases, particularly in chronic obstructive pulmonary disease. Their provocative hypothesis that low aerobic capacity increases a smoker’s risk for developing chronic obstructive pulmonary disease appears to be borne out by recent animal modeling studies. Thus, understanding how physical inactivity may predispose to the development of lung disease is likely to be an important field of investigation as our population becomes increasingly obese, older and sedentary. Obesity is profoundly altering human physiology and pathology: it is changing the nature of both health and disease throughout the world. Common diseases are being profoundly altered by obesity both in their incidence and behavior. Drugs designed to work in lean individuals, on diseases that we understand in the setting of the lean state, are unlikely to work the same way in obesity. If we are to understand disease in the 21st century, we need to concentrate our research efforts on understanding the ‘new normal’ these demographic trends portend.

Severe, glucocorticoid-resistant asthma comprises 5-7% of patients with asthma. IL-17 is a biomarker of severe asthma, and the adoptive transfer of Th17 cells in mice is sufficient to induce glucocorticoid-resistant allergic airway disease. Nitrogen dioxide (NO2) is an environmental toxin that correlates with asthma severity, exacerbation, and risk of adverse outcomes. Mice that are allergically sensitized to the antigen ovalbumin by exposure to NO2 exhibit a mixed Th2/Th17 adaptive immune response and eosinophil and neutrophil recruitment to the airway following antigen challenge, a phenotype reminiscent of severe clinical asthma. Because IL-1 receptor (IL-1R) signaling is critical in the generation of the Th17 response in vivo, we hypothesized that the IL-1R/Th17 axis contributes to pulmonary inflammation and airway hyperresponsiveness (AHR) in NO2-promoted allergic airway disease and manifests in glucocorticoid-resistant cytokine production. IL-17A neutralization at the time of antigen challenge or genetic deficiency in IL-1R resulted in decreased neutrophil recruitment to the airway following antigen challenge but did not protect against the development of AHR. Instead, IL-1R-/- mice developed exacerbated AHR compared to WT mice. Lung cells from NO2-allergically inflamed mice that were treated in vitro with dexamethasone (Dex) during antigen restimulation exhibited reduced Th17 cytokine production, whereas Th17 cytokine production by lung cells from recipient mice of in vitro Th17-polarized OTII T-cells was resistant to Dex. These results demonstrate that the IL-1R/Th17 axis does not contribute to AHR development in NO2-promoted allergic airway disease, that Th17 adoptive transfer does not necessarily reflect an endogenously-generated Th17 response, and that functions of Th17 responses are contingent on the experimental conditions in which they are generated.

Acute Respiratory Distress Syndrome (ARDS) severity may be influenced by heterogeneity of neutrophil activation. Interferon-stimulated genes (ISG) are a broad gene family induced by Type I interferons, often as a response to viral infections, which evokes extensive immunomodulation. We tested the hypothesis that over- or under-expression of immunomodulatory ISG by neutrophils is associated with worse clinical outcomes in patients with ARDS. Genome-wide transcriptional profiles of circulating neutrophils isolated from patients with sepsis-induced ARDS (n = 31) and healthy controls (n = 19) were used to characterize ISG expression. Hierarchical clustering of expression identified 3 distinct subject groups with Low, Mid and High ISG expression. ISG accounting for the greatest variability in expression were identified (MX1, IFIT1, and ISG15) and used to analyze a prospective cohort at the Colorado ARDS Network site. One hundred twenty ARDS patients from four urban hospitals were enrolled within 72 hours of initiation of mechanical ventilation. Circulating neutrophils were isolated from patients and expression of ISG determined by PCR. Samples were stratified by standard deviation from the mean into High (n = 21), Mid, (n = 82) or Low (n = 17) ISG expression. Clinical outcomes were compared between patients with High or Low ISG expression to those with Mid-range expression. At enrollment, there were no differences in age, gender, co-existing medical conditions, or type of physiologic injury between cohorts. After adjusting for age, race, gender and BMI, patients with either High or Low ISG expression had significantly worse clinical outcomes than those in the Mid for number of 28-day ventilator- and ICU-free days (P = 0.0006 and 0.0004), as well as 90-day mortality and 90-day home with unassisted breathing (P = 0.02 and 0.004). These findings suggest extremes of ISG expression by circulating neutrophils from ARDS patients recovered early in the syndrome are associated with poorer clinical outcomes.

γδ T cells are an evolutionary conserved T cell subset with innate-like effector functions. We had previously identified a locus on chromosome 1 encompassing the Slam gene family that regulates the innate-like T cell compartment. To characterize this region further, we generated a B6.129 congenic mouse strain in which a 1.1 Mbp region (chr. 1: 171.05–172.12) from the 129X1/SvJ strain was introgressed onto the C57BL/6J background, and evaluated its effect on the host immune response in influenza-infected mice. Upon infection, we observed a significant and sustained increase in the number of lung γδ T cells in B6.129c2 congenic mice. A comparison of γδ T cell function revealed that a significantly greater fraction of B6.129c2 γδ T cells were producing IL-17A versus their B6 counterparts. These differences were subset-specific as they were observed in Vγ4 and at least one other Vγ subset, but not in Vγ1 T cells. Those IL-17-producing Vγ subsets accounted for most of the increase in B6.129c2 lung γδ T cell numbers and they displayed an attenuated cell surface expression profile of Slam family receptors. Finally, since enhanced IL-17 responses have been associated with immunopathology after influenza infection, we assessed the effect of the congenic interval on survival. We found that the survival of B6.129c2 congenic mice is significantly lower compared with B6 mice after influenza infection. Taken together, our data suggest that natural genetic variations at this locus significantly impact both IL-17A production by lung γδ T cells and mortality after influenza infection.

Delayed pulmonary toxicity syndrome, characterized by interstitial pneumonia and pulmonary fibrosis, is common following high-dose bischloroethylnitrosourea (BCNU) (carmustine, [1,3-bis (2-chloroethyl)-1-nitrosourea]) containing chemotherapeutic regimens. Depending upon the treatment protocol, it may develop in over 70% of patients. Early and aggressive corticosteroid treatment leads to improvement in the majority of patients. However, up to 8% of affected patients may fail to respond to corticosteroids and develop progressive respiratory failure leading to death. No alternatives to corticosteroids have thus far been shown useful. We report the symptomatic and physiological improvement of a patient with severe steroid-resistant delayed pulmonary toxicity syndrome, following treatment with interferon-gamma.

Infection with pandemic (pdm) A/H1N1 virus induces high levels of pro-inflammatory mediators in blood and lungs of experimental animals and humans.To compare the involvement of seasonal A/PR/8/34 and pdm A/H1N1 virus strains in the regulation of inflammatory responses, we analyzed the changes in the whole-genome expression induced by these strains in macrophages and A549 epithelial cells. We also focused on the functional implications (cytokine production) of the differential induction of suppressors of cytokine signaling (SOCS)-1, SOCS-3, retinoid-inducible gene (RIG)-I and interferon receptor 1 (IFNAR1) genes by these viral strains in early stages of the infection.We identified 130 genes differentially expressed by pdm A/H1N1 and A/PR/8/34 infections in macrophages. mRNA levels of SOCS-1 and RIG-I were up-regulated in macrophages infected with the A/PR/8/34 but not with pdm A/H1N1 virus. mRNA levels of SOCS-3 and IFNAR1 induced by A/PR/8/34 and pdm A/H1N1 strains in macrophages, as well as in A549 cells were similar. We found higher levels of IL-6, TNF-α, IL-10, CCL3, CCL5, CCL4 and CXCL8 (p0.05) in supernatants from cultures of macrophages infected with the pdm A/H1N1 virus compared to those infected with the A/PR/8/34 strain, coincident with the lack of SOCS-1 and RIG-I expression. In contrast, levels of INF-α were higher in cultures of macrophages 48h after infection with the A/PR/8/34 strain than with the pdm A/H1N1 virus.These findings suggest that factors inherent to the pdm A/H1N1 viral strain may increase the production of inflammatory mediators by inhibiting SOCS-1 and modifying the expression of antiviral immunity-related genes, including RIG-I, in human macrophages.

Mounting evidence suggests that obesity and the metabolic syndrome have significant but often divergent effects on the innate immune system. These effects have been best established in monocytes and macrophages, particularly as a consequence of the hypercholesterolemic state. We have recently described defects in neutrophil function in the setting of both obesity and hypercholesterolemia, and hypothesized that exposure to elevated levels of lipoproteins, particularly LDL its oxidized forms, contributed to these defects. As a model of chronic cholesterol exposure, we examined functional responses of bone marrow neutrophils isolated from non-obese mice with diet-induced hypercholesterolemia compared to normal cholesterol controls. Chemotaxis, calcium flux, CD11b display, and F-actin polymerization were assayed in response to several chemoattractants, while neutrophil cytokine transcriptional response was determined to LPS. Following this, the acute effects of isolated LDL and its oxidized forms on normal neutrophils were assayed using the same functional assays. We found that neutrophils from non-obese hypercholesterolemic mice had blunted chemotaxis, altered calcium flux, and normal to augmented CD11b display with prolonged actin polymerization in response to stimuli. In response to acute exposure to lipoproteins, neutrophils showed chemotaxis to LDL which increased with the degree of LDL oxidation. Paradoxically, LDL oxidation yielded the opposite effect on LDL-induced CD11b display and actin polymerization, and both native and oxidized LDL were found to induce neutrophil transcription of the monocyte chemoattractant MCP-1. Together these findings suggest that chronic hypercholesterolemia impairs neutrophil functional responses, and these defects may be in part due to protracted signaling responses to LDL and its oxidized forms.

Leptin is an adipocyte-derived hormone, recognized as a critical mediator of the balance between food intake and energy expenditure by signalling through its functional receptor (Ob-Rb) in the hypothalamus. Structurally, leptin belongs to the long-chain helical cytokine family, and is now known to have pleiotropic functions in both innate and adaptive immunity. The presence of the functional leptin receptor in the lung together with evidence of increased airspace leptin levels arising during pulmonary inflammation, suggests an important role for leptin in lung development, respiratory immune responses and eventually pathogenesis of inflammatory respiratory diseases. The purpose of this article is to review our current understanding of leptin and its functional role on the different resident cell types of the lung in health as well as in the context of three major respiratory conditions being chronic obstructive pulmonary disease (COPD), asthma, and pneumonia.

Acute lung injury (ALI) is a common clinical inflammatory syndrome ­associated with severe acute hypoxemic respiratory with a case fatality of 30–40%. Although counterintuitive, several studies now suggest that the survival of obese and extremely obese critically ill patients with ALI (as well as obese critically ill patients in general) is equal to or better than their lean critically ill counterparts. The mechanisms underlying these findings are not yet clear but may be related to obesity-associated alterations of the host inflammatory response or to decreased susceptibility to ventilator-induced lung injury in these patients. Despite possible survival benefits afforded by obesity in ALI, studies do suggest that obese patients may have longer durations of ventilation and ICU stays. This information is important for clinicians to consider when discussing prognosis and expectations with critically ill patients and their families.

Influenza virus infection is considered a major worldwide public health problem. Seasonal infections with the most common influenza virus strains (e.g., H1N1) can usually be resolved, but they still cause a high rate of mortality. The factors that influence the outcome of the infection remain unclear. Here, we show that deficiency of interleukin (IL)-6 or IL-6 receptor is sufficient for normally sublethal doses of H1N1 influenza A virus to cause death in mice. IL-6 is necessary for resolution of influenza infection by protecting neutrophils from virus-induced death in the lung and by promoting neutrophil-mediated viral clearance. Loss of IL-6 results in persistence of the influenza virus in the lung leading to pronounced lung damage and, ultimately, death. Thus, we demonstrate that IL-6 is a vital innate immune cytokine in providing protection against influenza A infection. Genetic or environmental factors that impair IL-6 production or signaling could increase mortality to influenza virus infection.

Background Recent literature suggests that obese critically ill patients do not have worse outcomes than patients who are normal weight. However, outcomes in extreme obesity (BMI ≥ 40 kg/m2) are unclear. We sought to determine the association between extreme obesity and ICU outcomes. Methods We analyzed data from a multicenter international observational study of ICU nutrition practices that occurred in 355 ICUs in 33 countries from 2007 to 2009. Included patients were mechanically ventilated adults ≥ 18 years old who remained in the ICU for > 72 h. Using generalized estimating equations and Cox proportional hazard modeling with clustering by ICU and adjusting for potential confounders, we compared extremely obese to normal-weight patients in terms of duration of mechanical ventilation (DMV), ICU length of stay (LOS), hospital LOS, and 60-day mortality. Results Of the 8,813 patients included in this analysis, 3,490 were normal weight (BMI 18.5-24.9 kg/m2), 348 had BMI 40 to 49.9 kg/m2, 118 had BMI 50 to 59.9 kg/m2, and 58 had BMI ≥ 60 kg/m2. Unadjusted analyses suggested that extremely obese critically ill patients have improved mortality (OR for death, 0.77; 95% CI, 0.62-0.94), but this association was not significant after adjustment for confounders. However, an adjusted analysis of survivors found that extremely obese patients have a longer DMV and ICU LOS, with the most obese patients (BMI ≥ 60 kg/m2) also having longer hospital LOS. Conclusions During critical illness, extreme obesity is not associated with a worse survival advantage compared with normal weight. However, among survivors, BMI ≥ 40 kg/m2 is associated with longer time on mechanical ventilation and in the ICU. These results may have prognostic implications for extremely obese critically ill patients.

Dyslipidemia influences innate immune responses in the bloodstream, but whether and how pulmonary innate immunity is sensitive to circulating lipoproteins is largely unknown. In order to define whether dyslipidemia impacts responses to bacteria in the airspace and, if so, whether differently from its effects in other tissues, airspace, bloodstream, and intraperitoneal responses to lipopolysaccharide and Klebsiella pneumoniae were investigated using murine models of dyslipidemia. Dyslipidemia reduced neutrophil recruitment to the airspace in response to lipopolysaccharide and K. pneumoniae by impairing both chemokine induction in the airspace and neutrophil chemotaxis, thereby compromising pulmonary bacterial clearance. Paradoxically, bacteria were cleared more effectively from the bloodstream during dyslipidemia. This enhanced systemic response was due, at least in part, to basal circulating neutrophilia and basal Toll like Receptor 4 (TLR4)/Myeloid Differentiation Primary Response Protein 88-dependent serum cytokine induction, and enhanced serum cytokine responses to systemically administered TLR ligands. Dyslipidemia did not globally impair neutrophil transvascular trafficking to, and host defense within all loci, as neutrophilia, cytokine induction, and bacterial clearance were enhanced within the infected peritoneum. Peritoneal macrophages from dyslipidemic animals were primed for more robust TLR responses, reflecting increased lipid rafts and increased TLR4 expression, whereas macrophages from the airspace, in which cholesterol was maintained constant during dyslipidemia, had normal responses and rafts. Dyslipidemia thus imparts opposing effects upon intra- and extrapulmonary host defense by inducing tissue-divergent TLR response phenotypes and dysregulating airspace:blood compartmental levels of neutrophils and cytokines. We propose that the airspace is a ‘privileged’ site, thereby uniquely sensitive to dyslipidemia.

CD1d-restricted NKT cells make up an innate-like T cell subset that plays a role in amplifying the response of innate immune leukocytes to TLR ligands. The Slam locus contains genes that have been implicated in innate and adaptive immune responses. In this study, we demonstrate that divergent Slam locus haplotypes modulate the response of macrophages to the TLR4 ligand LPS through their control of NKT cell number and function. In response to LPS challenge in vivo, macrophage TNF production in Slam haplotype-2+ 129S1/SvImJ and 129X1/SvJ mice was significantly impaired in comparison with macrophage TNF production in Slam haplotype-1+ C57BL/6J mice. Although no cell-intrinsic differences in macrophage responses to LPS were observed between strains, 129 mice were found to be deficient in liver NKT cell number, in NKT cell cytokine production in response to the CD1d ligand α-galactosylceramide, and in NKT cell IFN-γ production after LPS challenge in vivo. Using B6.129c1 congenic mice and adoptive transfer, we found that divergent Slam haplotypes controlled the response to LPS in vivo, as well as the diminished NKT cell number and function, and that these phenotypes were associated with differential expression of signaling lymphocytic activation molecule family receptors on NKT cells. These data suggest that the polymorphisms that distinguish two Slam haplotypes significantly modulate the innate immune response in vivo through their effect on NKT cells.

A 48-year-old male smoker was referred for the evaluation of symptoms of cough, weight-loss, and chest radiograph abnormality. The patient had a medical history significant for chronic bronchitis and had been seen by his primary care physician 2