Monitoring redox stress in human airway epithelial cells exposed to woodsmoke at an air-liquid interface.

Wildland fires contribute significantly to the ambient air pollution burden worldwide, causing a range of adverse health effects in exposed populations. The toxicity of woodsmoke, a complex mixture of gases, volatile organic compounds, and particulate matter, is commonly studied in vitro using isolated exposures of conventionally cultured lung cells to either resuspended particulate matter or organic solvent extracts of smoke, leading to incomplete toxicity evaluations. This study aimed to improve our understanding of the effects of woodsmoke inhalation by building an advanced in vitro exposure system that emulates human exposure of the airway epithelium. We report the development and characterization of an innovative system that permits live-cell monitoring of the intracellular redox status of differentiated primary human bronchial epithelial cells cultured at an air-liquid interface (pHBEC-ALI) as they are exposed to unfractionated woodsmoke generated in a tube furnace in real time. pHBEC-ALI exposed to freshly generated woodsmoke showed oxidative changes that were dose-dependent and reversible, and not attributable to carbon monoxide exposure. These findings show the utility of this novel system for studying the molecular initiating events underlying woodsmoke-induced toxicity in a physiologically relevant in vitro model, and its potential to provide biological plausibility for risk assessment and public health measures.

Different airborne particulates trigger distinct immune pathways leading to peanut allergy in a mouse model.

BACKGROUND: Environmental exposure to peanut through non-oral routes is a risk factor for peanut allergy. Early-life exposure to air pollutants, including particulate matter (PM), is associated with sensitization to foods through unknown mechanisms. We investigated whether PM promotes sensitization to environmental peanut and the development of peanut allergy in a mouse model. METHODS: C57BL/6J mice were co-exposed to peanut and either urban particulate matter (UPM) or diesel exhaust particles (DEP) via the airways and assessed for peanut sensitization and development of anaphylaxis following peanut challenge. Peanut-specific CD4+ T helper (Th) cell responses were characterized by flow cytometry and Th cytokine production. Mice lacking select innate immune signaling genes were used to study mechanisms of PM-induced peanut allergy. RESULTS: Airway co-exposure to peanut and either UPM- or DEP-induced systemic sensitization to peanut and anaphylaxis following peanut challenge. Exposure to UPM or DEP triggered activation and migration of lung dendritic cells to draining lymph nodes and induction of peanut-specific CD4+ Th cells. UPM- and DEP-induced distinct Th responses, but both stimulated expansion of T follicular helper (Tfh) cells essential for peanut allergy development. MyD88 signaling was critical for UPM- and DEP-induced peanut allergy, whereas TLR4 signaling was dispensable. DEP-induced peanut allergy and Tfh-cell differentiation depended on IL-1 but not IL-33 signaling, whereas neither cytokine alone was necessary for UPM-mediated sensitization. CONCLUSION: Environmental co-exposure to peanut and PM induces peanut-specific Tfh cells and peanut allergy in mice.

Cyanobacterial blooms, iron, and environmental pollutants.

Iron determines the abundance and diversity of life and controls primary production in numerous aqueous environments. Over the past decades, the availability of this metal in natural waters has decreased. Iron deficiency can apply a selective pressure on microbial aquatic communities. Each aquatic organism has their individual requirements for iron and pathways for metal acquisition, despite all having access to the common pool of iron. Cyanobacteria, a photosynthesizing bacterium that can accumulate and form so-called 'algal blooms', have evolved strategies to thrive in such iron-deficient aqueous environments where they can outcompete other organisms in iron acquisition in diverse microbial communities. Metabolic pathways for iron acquisition employed by cyanobacteria allow it to compete successfully for this essential nutrient. By competing more effectively for requisite iron, cyanobacteria can displace other species and grow to dominate the microbial population in a bloom. Aquatic resources are damaged by a diverse number of environmental pollutants that can further decrease metal availability and result in a functional deficiency of available iron. Pollutants can also increase iron demand. A pollutant-exposed microbe is compelled to acquire further metal critical to its survival. Even in pollutant-impacted waters, cyanobacteria enjoy a competitive advantage and cyanobacterial dominance can be the result. We propose that cyanobacteria have a distinct competitive advantage over many other aquatic microbes in polluted, iron-poor environments.

Peribronchiolar Metaplasia: A Marker of Cigarette Smoke-Induced Small Airway Injury in a Rural Cohort.

Background: Peribronchiolar metaplasia (PBM) is considered a reaction to injury characterized by the proliferation of bronchiolar epithelium into immediately adjacent alveolar walls. While an association of PBM with diffuse interstitial lung diseases has been recognized, the clinical significance of PBM remains uncertain. Methods: A cohort (n = 352) undergoing surgical resection of a lung nodule/mass in a rural area was retrospectively reviewed. Multivariate logistic regression analysis was performed to determine the association of PBM with clinical, physiological, radiographic, and histologic endpoints. Results: In the total study cohort, 9.1% were observed to have PBM as a histologic finding in resected lung tissue (n = 32). All but one of these patients with PBM were ever-smokers with a median of 42 pack years. Clinical COPD was diagnosed in two-thirds of patients with PBM. Comorbid gastroesophageal reflux disease (GERD) was significantly associated with PBM. All patients with PBM demonstrated radiologic and histologic evidence of emphysema. Measures of pulmonary function were not impacted by PBM. Mortality was not associated with the histologic observation of PBM. In a logistic regression model, centrilobular-ground glass opacity interstitial lung abnormality and traction bronchiectasis on the CT scan of the chest and histologic evidence of fibrosis, desquamative interstitial pneumonia and anthracosis all strongly predicted PBM in the cohort. Conclusion: A constellation of radiologic and histologic smoking-related abnormalities predicted PBM in study cohort. This confirms a co-existence of lung tissue responses to smoking including PBM, emphysema, and fibrosis. Acknowledging the physiologically "silent" nature of small airway dysfunction on pulmonary function testing, our findings support PBM as a histologic marker of small-airway injury associated with cigarette smoking.

Asbestos and Iron.

Theories of disease pathogenesis following asbestos exposure have focused on the participation of iron. After exposure, an open network of negatively charged functional groups on the fiber surface complexes host metals with a preference for iron. Competition for iron between the host and the asbestos results in a functional metal deficiency. The homeostasis of iron in the host is modified by the cell response, including increased import to correct the loss of the metal to the fiber surface. The biological effects of asbestos develop in response to and are associated with the disruption of iron homeostasis. Cell iron deficiency in the host following fiber exposure activates kinases and transcription factors, which are associated with the release of mediators coordinating both inflammatory and fibrotic responses. Relative to serpentine chrysotile, the clearance of amphiboles is incomplete, resulting in translocation to the mesothelial surface of the pleura. Since the biological effect of asbestos is dependent on retention of the fiber, the sequestration of iron by the surface, and functional iron deficiency in the cell, the greater clearance (i.e., decreased persistence) of chrysotile results in its diminished impact. An inability to clear asbestos from the lower respiratory tract initiates a host process of iron biomineralization (i.e., asbestos body formation). Host cells attempt to mobilize the metal sequestered by the fiber surface by producing superoxide at the phagosome membrane. The subsequent ferrous cation is oxidized and undergoes hydrolysis, creating poorly crystalline iron oxyhydroxide (i.e., ferrihydrite) included in the coat of the asbestos body.

Lung microhaemorrhage drives oxidative/inflammatory damage in α1-antitrypsin deficiency.

Background: Animal models using intratracheal instillation show that elastase, unopposed by α1-antitrypsin (AAT), causes alveolar damage and haemorrhage associated with emphysematous changes. The aim of the present study was to characterise any relationship between alveolar haemorrhage and human AAT deficiency (AATD) using bronchoalveolar lavage (BAL) and lung explant samples from AATD subjects. Methods: BAL samples (17 patients, 15 controls) were evaluated for free haem (iron protoporphyrin IX) and total iron concentrations. Alveolar macrophage activation patterns were assessed using RNA sequencing and validated in vitro using haem-stimulated, monocyte-derived macrophages. Lung explants (seven patients, four controls) were assessed for iron sequestration protein expression patterns using Prussian blue stain and ferritin immunohistochemistry, as well as ferritin iron imaging and elemental analysis by transmission electron microscopy. Tissue oxidative damage was assessed using 8-hydroxy-2'-deoxyguanosine immunohistochemistry. Results: BAL collected from AATD patients showed significantly elevated free haem and total iron concentrations. Alveolar and interstitial macrophages in AATD explants showed elevated iron and ferritin accumulation in large lysosomes packed by iron oxide cores with degraded ferritin protein cages. BAL macrophage RNA sequencing showed innate pro-inflammatory activation, replicated in vitro by haemin exposure, which also triggered reactive oxygen species generation. AATD explants showed massive oxidative DNA damage in both lung epithelial cells and macrophages. Conclusions: BAL and tissue markers of alveolar haemorrhage, together with molecular and cellular evidence of macrophage innate pro-inflammatory activation and oxidative damage, are consistent with free haem stimulation. Overall, this initial study provides evidence for a pathogenetic role of elastase-induced alveolar haemorrhage in AATD emphysema.

Cigarette Smoking, and Blood Monocyte Count Correlate with Chronic Lung Injuries and Mortality.

Background: Cigarette smoking (CS)-related monocytosis contributes to the development of chronic lung injuries via complex mechanisms. We aim to determine correlations between measures of CS and monocytes, their capacities to predict chronic lung diseases, and their associations with mortality. Methods: A single-center retrospective study of patients undergoing surgical resection for suspected lung nodules/masses was performed. CS was quantified as cigarettes smoked per day (CPD), duration of smoking, composite pack years (CPY), current smoking status, and smoking cessation years. A multivariate logistic regression analysis was performed. Results: Of 382 eligible patients, 88% were ever smokers. In this group, 45% were current smokers with mean CPD of 27.2±40.0. CPY and duration of smoking showed positive linear correlations with percentage monocyte count. Physiologically, CPY was associated with progressive obstruction, hyperinflation, and reduced diffusion capacity (DLCO). Across the quartiles of smoking, there was an accumulation of radiologic and histologic abnormalities. Anthracosis and emphysema were associated with CPD, while lung cancer, respiratory bronchiolitis (RB), emphysema, and honeycombing were statistically related to duration of smoking. Analysis using consecutive CPY showed associations with lung cancer (≥10 and <30), fibrosis (≥20 and <40), RB (≥50), anthracosis and emphysema (≥10 and onwards). Percentage monocytes correlated with organizing pneumonia (OP), fibrosis, and emphysema. The greater CPY increased mortality across the groups. Significant predictors of mortality included percentage monocyte, anemia, GERD, and reduced DLCO. Conclusion: Indices of CS and greater monocyte numbers were associated with endpoints of chronic lung disease suggesting a participation in pathogenesis. Application of these easily available metrics may support a chronology of CS-induced chronic lung injuries. While a relative lesser amount of smoking can be associated with lung cancer and fibrosis, greater CPY increases the risk for emphysema. Monocytosis predicted lung fibrosis and mortality. Duration of smoking may serve as a better marker of monocytosis and associated chronic lung diseases.

Mucus increases cell iron uptake to impact the release of pro-inflammatory mediators after particle exposure.

We tested the hypothesis that (1) mucus production can be included in the cell response to iron deficiency; (2) mucus binds iron and increases cell metal uptake; and subsequently (3) mucus impacts the inflammatory response to particle exposure. Using quantitative PCR, RNA for both MUC5B and MUC5AC in normal human bronchial epithelial (NHBE) cells decreased following exposures to ferric ammonium citrate (FAC). Incubation of mucus-containing material collected from the apical surface of NHBE cells grown at air-liquid interface (NHBE-MUC) and a commercially available mucin from porcine stomach (PORC-MUC) with iron demonstrated an in vitro capacity to bind metal. Inclusion of either NHBE-MUC or PORC-MUC in incubations of both BEAS-2B cells and THP1 cells increased iron uptake. Exposure to sugar acids (N-acetyl neuraminic acid, sodium alginate, sodium guluronate, and sodium hyaluronate) similarly increased cell iron uptake. Finally, increased metal transport associated with mucus was associated with a decreased release of interleukin-6 and -8, an anti-inflammatory effect, following silica exposure. We conclude that mucus production can be involved in the response to a functional iron deficiency following particle exposure and mucus can bind metal, increase cell uptake to subsequently diminish or reverse a functional iron deficiency and inflammatory response following particle exposure.

Exposure to isocyanates predicts atopic dermatitis prevalence and disrupts therapeutic pathways in commensal bacteria.

Atopic dermatitis (AD) is a chronic inflammatory skin condition increasing in industrial nations at a pace that suggests environmental drivers. We hypothesize that the dysbiosis associated with AD may signal microbial adaptations to modern pollutants. Having previously modeled the benefits of health-associated Roseomonas mucosa, we now show that R. mucosa fixes nitrogen in the production of protective glycerolipids and their ceramide by-products. Screening EPA databases against the clinical visit rates identified diisocyanates as the strongest predictor of AD. Diisocyanates disrupted the production of beneficial lipids and therapeutic modeling for isolates of R. mucosa as well as commensal Staphylococcus. Last, while topical R. mucosa failed to meet commercial end points in a placebo-controlled trial, the subgroup who completed the full protocol demonstrated sustained, clinically modest, but statistically significant clinical improvements that differed by study site diisocyanate levels. Therefore, diisocyanates show temporospatial and epidemiological association with AD while also inducing eczematous dysbiosis.

Lactate Production can Function to Increase Human Epithelial Cell Iron Concentration.

Introduction: Under conditions of limited iron availability, plants and microbes have evolved mechanisms to acquire iron. For example, metal deficiency stimulates reprogramming of carbon metabolism, increasing activity of enzymes involved in the Krebs cycle and the glycolytic pathway. Resultant carboxylates/hydroxycarboxylates then function as ligands to complex iron and facilitate solubilization and uptake, reversing the metal deficiency. Similarly, human intestinal epithelial cells may produce lactate, a hydroxycarboxylate, during absolute and functional iron deficiency to import metal to reverse limited availability. Methods: Here we investigate (1) if lactate can increase cell metal import of epithelial cells in vitro, (2) if lactate dehydrogenase (LDH) activity in and lactate production by epithelial cells correspond to metal availability, and (3) if blood concentrations of LDH in a human cohort correlate with indices of iron homeostasis. Results: Results show that exposures of human epithelial cells, Caco-2, to both sodium lactate and ferric ammonium citrate (FAC) increase metal import relative to FAC alone. Similarly, fumaric, isocitric, malic, and succinic acid coincubation with FAC increase iron import relative to FAC alone. Increased iron import following exposures to sodium lactate and FAC elevated both ferritin and metal associated with mitochondria. LDH did not change after exposure to deferoxamine but decreased with 24 h exposure to FAC. Lactate levels revealed decreased levels with FAC incubation. Review of the National Health and Nutrition Examination Survey demonstrated significant negative relationships between LDH concentrations and serum iron in human cohorts. Conclusions: Therefore, we conclude that iron import in human epithelial cells can involve lactate, LDH activity can reflect the availability of this metal, and blood LDH concentrations can correlate with indices of iron homeostasis.

Stress Drivers of Glucose Dynamics during Ozone Exposure Measured Using Radiotelemetry in Rats.

BACKGROUND: Inhaled irritant air pollutants may trigger stress-related metabolic dysfunction associated with altered circulating adrenal-derived hormones. OBJECTIVES: We used implantable telemetry in rats to assess real-time changes in circulating glucose during and after exposure to ozone and mechanistically linked responses to neuroendocrine stress hormones. METHODS: First, using a cross-over design, we monitored glucose during ozone exposures (0.0, 0.2, 0.4, and 0.8 ppm) and nonexposure periods in male Wistar Kyoto rats implanted with glucose telemeters. A second cohort of unimplanted rats was exposed to ozone (0.0, 0.4 or 0.8 ppm) for 30 min, 1 h, 2 h, or 4 h with hormones measured immediately post exposure. We assessed glucose metabolism in sham and adrenalectomized rats, with or without supplementation of adrenergic/glucocorticoid receptor agonists, and in a separate cohort, antagonists. RESULTS: Ozone (0.8 ppm) was associated with significantly higher blood glucose and lower core body temperature beginning 90 min into exposure, with reversal of effects 4-6 h post exposure. Glucose monitoring during four daily 4-h ozone exposures revealed duration of glucose increases, adaptation, and diurnal variations. Ozone-induced glucose changes were preceded by higher levels of adrenocorticotropic hormone, corticosterone, and epinephrine but lower levels of thyroid-stimulating hormone, prolactin, and luteinizing hormones. Higher glucose and glucose intolerance were inhibited in rats that were adrenalectomized or treated with adrenergic plus glucocorticoid receptor antagonists but exacerbated by agonists. DISCUSSION: We demonstrated the temporality of neuroendocrine-stress-mediated biological sequalae responsible for ozone-induced glucose metabolic dysfunction and mechanism in a rodent model. Stress hormones assessment with real-time glucose monitoring may be useful in identifying interactions among irritant pollutants and stress-related illnesses. https://doi.org/10.1289/EHP11088.

Interstitial lung abnormalities and interstitial lung diseases associated with cigarette smoking in a rural cohort undergoing surgical resection.

BACKGROUND: Cigarette smoking is a risk factor for interstitial lung abnormalities (ILAs) and interstitial lung diseases (ILDs). Investigation defining the relationships between ILAs/ILDs and clinical, radiographic, and pathologic findings in smokers have been incomplete. Employing a cohort undergoing surgical resection for lung nodules/masses, we (1) define the prevalence of ILAs/ILDs, (2) delineate their clinical, radiographic and pathologic predictors, and (3) determine their associations with mortality. METHODS: Patients undergoing resection of lung nodules/masses between 2017 and 2020 at a rural Appalachian, tertiary medical center were retrospectively investigated. Predictors for ILAs/ILDs and mortality were assessed using multivariate logistic regression analysis. RESULTS: In the total study cohort of 352 patients, radiographic ILAs and ILDs were observed in 35.2% and 17.6%, respectively. Among ILA patterns, subpleural reticular changes (14.8%), non-emphysematous cysts, centrilobular (CL) ground glass opacities (GGOs) (8% each), and mixed CL-GGO and subpleural reticular changes (7.4%) were common. ILD patterns included combined pulmonary fibrosis emphysema (CPFE) (3.1%), respiratory bronchiolitis (RB)-ILD (3.1%), organizing pneumonitis (2.8%) and unclassifiable (4.8%). The group with radiographic ILAs/ILDs had a significantly higher proportion of ever smokers (49% vs. 39.9%), pack years of smoking (44.57 ± 36.21 vs. 34.96 ± 26.22), clinical comorbidities of COPD (35% vs. 26.5%) and mildly reduced diffusion capacity (% predicated 66.29 ± 20.55 vs. 71.84 ± 23). Radiographic centrilobular and paraseptal emphysema (40% vs. 22.2% and 17.6% vs. 9.6%, respectively) and isolated traction bronchiectasis (10.2% vs. 4.2%) were associated with ILAs/ILDs. Pathological variables of emphysema (34.9% vs. 18.5%), any fibrosis (15.9% vs. 4.6%), peribronchiolar metaplasia (PBM, 8% vs. 1.1%), RB (10.3% vs. 2.5%), and anthracosis (21.6% vs. 14.5%) were associated with ILAs/ILDs. Histologic emphysema showed positive correlations with any fibrosis, RB, anthracosis and ≥ 30 pack year of smoking. The group with ILAs/ILDs had significantly higher mortality (9.1% vs. 2.2%, OR 4.13, [95% CI of 1.84-9.25]). CONCLUSIONS: In a rural cohort undergoing surgical resection, radiographic subclinical ILAs/ILDs patterns were highly prevalent and associated with ever smoking and intensity of smoking. The presence of radiographic ILA/ILD patterns and isolated honeycomb changes were associated with increased mortality. Subclinical ILAs/ILDs and histologic fibrosis correlated with clinical COPD as well as radiographic and pathologic emphysema emphasizing the co-existence of these pulmonary injuries in a heavily smoking population.

Demystifying idiopathic interstitial pneumonia: time for more etiology-focused nomenclature in interstitial lung disease.

INTRODUCTION: A major focus of interstitial lung disease (ILD) has centered on disorders termed idiopathic interstitial pneumonias (IIPs) which include, among others, idiopathic pulmonary fibrosis, idiopathic nonspecific interstitial pneumonia, cryptogenic organizing pneumonia, and respiratory bronchiolitis-interstitial lung disease. AREAS COVERED: We review the radiologic and histologic patterns for the nine disorders classified by multidisciplinary approach as IIP, and describe the remarkable amount of published epidemiologic, translational, and molecular studies demonstrating their associations with numerous yet definitive environmental exposures, occupational exposures, pulmonary diseases, systemic diseases, medication toxicities, and genetic variants. EXPERT OPINION: In the 21st century, these disorders termed IIPs are rarely idiopathic, but rather are well-described radiologic and histologic patterns of lung injury that are associated with a wide array of diverse etiologies. Accordingly, the idiopathic nomenclature is misleading and confusing, and may also promote a lack of inquisitiveness, suggesting the end rather than the beginning of a thorough diagnostic process to identify ILD etiology and initiate patient-centered management. A shift toward more etiology-focused nomenclature will be beneficial to all, including patients hoping for better life quality and disease outcome, general medicine and pulmonary physicians furthering their ILD knowledge, and expert ILD clinicians and researchers who are advancing the ILD field.

Cigarette Smoke Particle-Induced Lung Injury and Iron Homeostasis.

It is proposed that the mechanistic basis for non-neoplastic lung injury with cigarette smoking is a disruption of iron homeostasis in cells after exposure to cigarette smoke particle (CSP). Following the complexation and sequestration of intracellular iron by CSP, the host response (eg, inflammation, mucus production, and fibrosis) attempts to reverse a functional metal deficiency. Clinical manifestations of this response can present as respiratory bronchiolitis, desquamative interstitial pneumonitis, pulmonary Langerhans' cell histiocytosis, asthma, pulmonary hypertension, chronic bronchitis, and pulmonary fibrosis. If the response is unsuccessful, the functional deficiency of iron progresses to irreversible cell death evident in emphysema and bronchiectasis. The subsequent clinical and pathological presentation is a continuum of lung injuries, which overlap and coexist with one another. Designating these non-neoplastic lung injuries after smoking as distinct disease processes fails to recognize shared relationships to each other and ultimately to CSP, as well as the common mechanistic pathway (ie, disruption of iron homeostasis).

Diacetyl exposure disrupts iron homeostasis in animals and cells.

OBJECTIVE: Several mechanisms have been proposed for the biological effect of diacetyl. We tested the postulate that animal and cell exposures to diacetyl are associated with a disruption in iron homeostasis. MATERIALS AND METHODS: Male, Sprague-Dawley rats were intratracheally-instilled with either distilled water or diacetyl. Seven days after treatment, animals were euthanized and the lungs removed, fixed, and embedded. Sections were cut and stained for iron, collagen, and ferritin. Human epithelial (BEAS-2B) and monocytic (THP-1) cells were exposed in vitro to ferric ammonium citrate (FAC), diacetyl, and both FAC and diacetyl. Cell non-heme iron concentrations and ferritin levels were quantified using inductively coupled plasma optical emission spectroscopy and an immunoassay respectively. RESULTS: After exposure of animals to diacetyl, there were airway polypoid lesions which stained positively for both iron and the intracellular storage protein ferritin. Trichrome stain showed a deposition of collagen immediately adjacent to accumulated metal following diacetyl exposure. In in vitro cell exposures, FAC increased non-heme iron concentration but co-incubations of FAC and diacetyl elevated levels to significantly greater values. Levels of ferritin were increased with exposures of BEAS-2B and THP-1 cells to FAC but were similarly greater after co-exposure with FAC and diacetyl. CONCLUSIONS: Results of animal and cell studies support a disruption of iron homeostasis by diacetyl. It is proposed that, following internalization, diacetyl complexes intracellular sources of iron. The cell recognizes a loss of its requisite iron to diacetyl and imports greater concentrations of the metal.

Outcomes of Idiopathic Pulmonary Fibrosis Improve with Obesity: A Rural Appalachian Experience.

OBJECTIVES: Obesity can be an independent predictor of fibrosis in tissues, including the liver, heart, and skin. We evaluated a rural Appalachian cohort of idiopathic pulmonary fibrosis (IPF) for its relation to obesity. METHODS: Using American Thoracic Society 2018 diagnostic guidelines, an IPF cohort was systematically identified at an Appalachian academic medical center (2015-2019). The cohort was categorized in subgroups of body mass index (BMI) <30 or BMI ≥30 kg/m2. Demographics, clinical variables, and treatment details were collected retrospectively and evaluated for their associations with obesity. RESULTS: In our IPF cohort (N = 138), a usual interstitial pneumonia pattern was less prevalent in the obese group (n = 49) relative to the nonobese group (69% vs 85%, respectively). The obese group was younger (mean age 73.27 ± 9.12 vs 77.97 ± 9.59 years) and had a higher prevalence of hypertension (90% vs 72%), hyperlipidemia (83% vs 68%), diabetes mellitus (47% vs 25%), sleep-disordered breathing (47% vs 25%), chronic pain disorders (28% vs 15%), and deep vein thrombosis (19% vs 7%). An increased proportion of obese-IPF patients was seen at a tertiary or an interstitial lung disease center, with more surgical lung biopsies performed and incident diagnosis (ie, within 6 months of presentation) assigned. Only a minority of patients underwent lung transplantation (3.6%), all of them from the obese-IPF subgroup. Approximately 30% of the total IPF cohort died, with a lower mortality observed in the obese group (35% vs 20%, P = 0.017). An increasing BMI predicted a better survival in the total IPF cohort (BMI 25-29.9, 20-24.9, and <20 had mortality rates of 20%, 47%, and 75%, respectively; P < 0.001). CONCLUSIONS: Our study represents a first known effort to develop an IPF cohort in a rural Appalachian region. Although they shared an increased burden of comorbidities, the obese subgroup showed less advanced fibrosis with a lower mortality rate relative to nonobese subgroup, suggesting a potential "obesity paradox" in IPF. The study findings significantly advance our understanding of challenges posed by IPF in a rural population that also suffers from an alarming rate of obesity. We highlight the need for the multidisciplinary management of these patients and prospective studies to better define this complex relation.

Perls' Prussian Blue Stains of Lung Tissue, Bronchoalveolar Lavage, and Sputum.

Perls' Prussian blue (PPB) stain recognizes Fe3+ associated with hemosiderin. The employment of this stain in clinical medicine and research has been extensive and novel applications continue to evolve. Ferruginous bodies are intracellular structures in lung tissue, bronchoalveolar lavage (BAL), and sputum that stain with PPB. Inhaled, insoluble, biopersistent particles and fibers are phagocytosed by lung macrophages and thought to be coated, either partially or completely, with an iron-containing protein at the interface forming a ferruginous body. These structures can be categorized as ferruginous bodies having either an inorganic or a carbonaceous core (e.g., asbestos and byssinotic bodies, respectively). In lung tissue, BAL, and sputum, the only cells that stain with PPB are macrophages. These are described as iron- and hemosiderin-laden macrophages and called either siderophages or sideromacrophages. Siderophages can be observed in the lung tissue, BAL, and sputum after various exposures and can also be associated with many different pulmonary and extrapulmonary diseases.

Effects of albumin, transferrin and humic-like substances on iron-mediated OH radical formation in human lung fluids.

Inhalation of particulate matter is hypothesized to contribute to health effects by overproducing reactive oxygen species (ROS) and inducing oxidative stress. Fe(II) has been shown to contribute to ROS generation in acellular simulated lung fluids. Atmospheric humic-like substances (HULIS) have been shown to chelate Fe(II) and significantly enhance this ROS generation. Here, we investigate Fe(II)-mediated .OH generation from the iron active proteins in lung fluid, albumin and transferrin, and fulvic acid, a surrogate for HULIS, in human bronchoalveolar lavage fluid (BALF). We find that albumin enhances .OH generation from inorganic Fe(II) and that transferrin attenuates this enhancement. We estimate the rate constants for albumin-Fe(II) and fulvic acid-Fe(II) mediated O2.- reduction (1.9 ± 0.3) M-1 s-1 and (2.7 ± 0.3) M-1s-1 (pH = 5.5, T = 37 °C), 17-25 times the rate for free iron, which we measured to be (110 ± 20) × 10-3 M-1s-1, in agreement with the literature. .OH generation measured from fulvic acid-Fe(II) in BALF from 8 individuals with added fulvic acid is successfully predicted rates of .OH generation by mixtures of Fe(II), albumin, transferrin, fulvic acid, and ascorbate in saline solution. This indicates that fulvic acid enhances .OH formation in BALF, and that albumin and transferrin in BALF moderate the effect. We propose that fulvic acid, and thereby HULIS, is capable of mobilizing Fe(II) away from albumin and transferrin and this increases the formation rate of O2.- and ultimately of .OH. Furthermore, we find that albumin and transferrin have significantly different impacts on Fe(II)-mediated .OH than citrate, a common component of simulated lung fluids, a factor that should be considered carefully in the interpretation of results obtained from solutions containing citrate.