Lung on Chip: In vitro HGF effects on injured alveolar A549 epithelial cells in microfluidic system

Microfluidic systems have become competitive tools in the invitro modelling of diseases and promising alternatives to animal studies. They allow obtaining more invivo like conditions for cellular assays. Research in idiopathic pulmonary fibrosis could benefit from this novel methodological approach to understand the pathophysiology of the disease & develop efficient therapies. The use of hepatocyte growth factor (HGF) for alveolar reepithelisation is a promising approach. In this study, we show a new microfluidic system to analyse the effects of HGF on injured alveolar epithelial cells. Microfluidic systems in polydimethylsiloxane were fabricated by soft lithography. The alveolar A549 epithelial cells (10,000 cells) were seeded and studied in these microfluidic systems with media perfusion (1μl/30min). Injury tests were made on the cells by the perfusion with media containing H2O2 or bleomycin. The degree of injury was then assessed by a metabolic and an apoptotic assays. Wound assays were also performed with a central laminar flow of trypsin. Monitoring of wound closure with HGF vs control media was assessed. The alveolar A549 epithelial cells grew and proliferated in the microfluidic system. In the wound closure assay, the degree of wound closure after 5 hours was (53.3±1.3%) with HGF compared to (9.8±2.4%) without HGF (P <0.001). We present a novel microfluidic model that allows culture, injury and wounding of A549 epithelial cells and represents the first step towards the development of an invitro reconstitution of the alveolar-capillary interface. We were also able to confirm that HGF increased alveolar epithelial repair in this system.

Lung on a chip: Co-culture of alveolar epithelial cells and bone marrow derived stromal stem cells in a microfluidic system

Background: Microfluidics system are novel tools to study cell-cell interactions in vitro. This project focuses on the development of a new microfluidic device to co-culture alveolar epithelial cells and mesenchymal stem cells to study cellular interactions involved in healing the injured alveolar epithelium. Methods: Microfluidic systems in polydimethylsiloxane were fabricated by soft lithography. The alveolar A549 epithelial cells were seeded and injury tests were made on the cells by perfusion with media containing H2O2 or bleomycin during 6 or 18hrs. Rat Bone marrow derived stromal cells (BMSC) were then introduced into the system and cell-cell interaction was studied over 24 hrs. Results: A successful co-culture of A549 alveolar epithelial cells and BMS was achieved in the microfluidic system. The seeded alveolar epithelial cells and BMSC adhered to the bottom surface of the microfluidic device and proliferated under constant perfusion. Epithelial injury to mimic mechanisms seen in idiopathic pulmonary fibrosis was induced in the microchannels by perfusing with H2O2 or bleomycin. Migration of BMSC towards the injured epithelium was observed as well as cell-cell interaction between the two cell types was also seen. Conclusion: We demonstrate a novel microfluidic device aimed at showing interactions between different cell types on the basis of a changing microenvironment. Also we were able to confirm interaction between injured alvolar epithelium and BMSC, and showed that BMSC try to heal the injured epitelium.

Cell-specific expression of human HGF by alveolar type II cells induces remodeling of septal wall tissue in the lung: a morphometric study

Hepatocyte growth factor (HGF) is involved in development and regeneration of the lungs. Human HGF, which was expressed specifically by alveolar epithelial type II cells after gene transfer, attenuated the bleomycin-induced pulmonary fibrosis in an animal model. As there are also regions that appear morphologically unaffected in fibrosis, the effects of this gene transfer to normal lungs is of interest. In vitro studies showed that HGF inhibits the formation of the basal lamina by cultured alveolar epithelial cells. Thus we hypothesized that, in the healthy lung, cell-specific expression of HGF induces a remodeling within septal walls. Electroporation of a plasmid of human HGF gene controlled by the surfactant protein C promoter was applied for targeted gene transfer. Using design-based stereology at light and electron microscopic level, structural alterations were analyzed and compared with a control group. HGF gene transfer increased the volume of distal air spaces, as well as the surface area of the alveolar epithelium. The volume of septal walls, as well as the number of alveoli, was unchanged. Volumes per lung of collagen and elastic fibers were unaltered, but a marked reduction of the volume of residual extracellular matrix (all components other than collagen and elastic fibers) and interstitial cells was found. A correlation between the volumes of residual extracellular matrix and distal air spaces, as well as total surface area of alveolar epithelium, could be established. Cell-specific expression of HGF leads to a remodeling of the connective tissue within the septal walls in the healthy lung, which is associated with more pronounced stretching of distal air spaces at a given hydrostatic pressure during instillation fixation.

[Cardiogenic and non cardiogenic pulmonary edema: pathomechanisms and causes]

The development of pulmonary edema is divided in cardiogenic and non-cardiogenic. Cardiogenic edema pathogenically is caused by elevated hydrostatic pressure in the pulmonary capillaries due to left sided congestive heart failure. Non-cardiogenic pulmonary edema is categorized depending on the underlying pathogenesis in low-alveolar pressure, elevated permeability or neurogenic edema. Some important examples of causes are upper airway obstruction like in laryngeal paralysis or strangulation for low alveolar pressure, leptospirosis and ARDS for elevated permeability, and epilepsy, brain trauma and electrocution for neurogenic edema. The differentiation between cardiogenic versus non-cardiogenic genesis is not always straightforward, but most relevant, because treatment markedly differs between the two. Of further importance is the identification of the specific underlying cause in non-cardiogenic edema, not only for therapeutic but particularly for prognostic reasons. Depending on the cause the prognosis ranges from very poor to good chance of complete recovery.

Pulmonary surfactant coating of multi-walled carbon nanotubes (MWCNTs) influences their oxidative and pro-inflammatory potential in vitro

Background Increasing concern has been expressed regarding the potential adverse health effects that may be associated with human exposure to inhaled multi-walled carbon nanotubes (MWCNTs). Thus it is imperative that an understanding as to the underlying mechanisms and the identification of the key factors involved in adverse effects are gained. In the alveoli, MWCNTs first interact with the pulmonary surfactant. At this interface, proteins and lipids of the pulmonary surfactant bind to MWCNTs, affecting their surface characteristics. Aim of the present study was to investigate if the pre-coating of MWCNTs with pulmonary surfactant has an influence on potential adverse effects, upon both (i) human monocyte derived macrophages (MDM) monocultures, and (ii) a sophisticated in vitro model of the human epithelial airway barrier. Both in vitro systems were exposed to MWCNTs either pre-coated with a porcine pulmonary surfactant (Curosurf) or not. The effect of MWCNTs surface charge was also investigated in terms of amino (−NH2) and carboxyl (−COOH) surface modifications. Results Pre-coating of MWCNTs with Curosurf affects their oxidative potential by increasing the reactive oxygen species levels and decreasing intracellular glutathione depletion in MDM as well as decreases the release of Tumour necrosis factor alpha (TNF-α). In addition, an induction of apoptosis was observed after exposure to Curosurf pre-coated MWCNTs. In triple cell-co cultures the release of Interleukin-8 (IL-8) was increased after exposure to Curosurf pre-coated MWCNTs. Effects of the MWCNTs functionalizations were minor in both MDM and triple cell co-cultures. Conclusions The present study clearly indicates that the pre-coating of MWCNTs with pulmonary surfactant more than the functionalization of the tubes is a key factor in determining their ability to cause oxidative stress, cytokine/chemokine release and apoptosis. Thus the coating of nano-objects with pulmonary surfactant should be considered for future lung in vitro risk assessment studies. Keywords: Multi-walled carbon nanotubes (MWCNTs); Pulmonary surfactant (Curosurf); Macrophages; Epithelial cells; Dendritic cells; Triple cell co-culture; Pro-inflammatory and oxidative reactions

Intraperitoneal Echinococcus multilocularis infection in C57BL/6 mice affects CD40 and B7 costimulator expression on peritoneal macrophages and impairs peritoneal T cell activation

One of the most important immunopathological consequence of intraperitoneal alveolar echinococcosis (AE) in the mouse is suppression of T cell-mediated immune responses. We investigated whether and how intraperitoneal macrophages (MØs) are, respectively, implicated as antigen-presenting cells (APCs). In a first step we showed that peritoneal MØs from infected mice (AE-MØs) exhibited a reduced ability to present a conventional antigen (chicken ovalbumin, C-Ova) to specific responder lymph node T cells. In a subsequent step, AE-MØs as well as naïve MØs (positive control) proved their ability to uptake and process C-Ova fluorescein isthiocyanate (FITC). Furthermore, in comparison with naïve MØs, the surface expression of Ia molecules was up-regulated on AE-MØs at the early stage of infection, suggesting that AE-MØs provide the first signal via the antigen-Ia complex. To study the accessory activity of MØs, AE-MØs obtained at the early and late stages of infection were found to decrease Con A-induced proliferation of peritoneal naïve T cells as well as of AE-sensitized peritoneal T cells, in contrast to stimulation with naïve MØs. The status of accessory molecules was assessed by analysing the expression level of costimulatory molecules on AE-MØs, with naïve MØs as controls. It was found that B7-1 (CD80) and B7-2 (CD86) expression remained unchanged, whereas CD40 was down-regulated and CD54 (= ICAM-1) was slightly up-regulated. In a leucocyte reaction of AE-MØs with naïve or AE-T cells, both types of T cells increased their proliferative response when CD28 - the ligand of B7 receptors - was exposed to anti-CD28 in cultures. Conversely to naïve MØs, pulsing of AE-MØs with agonistic anti-CD40 did not even partially restore their costimulatory activity and failed to increase naïve or AE-T cell proliferation. Neutralizing anti-B7-1, in combination with anti-B7-2, reduced naïve and AE-T cell proliferation, whereas anti-CD40 treatment of naïve MØs increased their proliferative response to Con A. These results point at the key role of B7 receptors as accessory molecules and the necessity of the integrity of CD40-expression by naïve MØs to improve their accessory activity. Taken together, the obstructed presenting-activity of AE-MØs appeared to trigger an unresponsiveness of T cells, contributing to the suppression of their clonal expansion during the chronic phase of AE-infection.

Increased uptake of 111In-octreotide in idiopathic pulmonary fibrosis

Idiopathic pulmonary fibrosis (IPF) is characterized by an uncontrolled accumulation and activation of lung fibroblasts. A modulation of fibroblast activation has been observed in various systems with octreotide, a synthetic somatostatin analog with strong affinity for the somatostatin receptor subtype 2 (sst2). One aim of our study was to evaluate the expression of somatostatin receptors in the lungs of patients with IPF. A second aim was to evaluate the relationship between 111In-octreotide uptake and the effect of pulmonary fibrosis as assessed by lung function tests and parameters and by radiologic findings. METHODS: We investigated 11 patients with IPF, 6 patients with pulmonary fibrosis associated with systemic sclerosis (SSc), and 19 patients with disease not of the lung (control patients). The expression of somatostatin receptors was evaluated in vivo using 111In-octreotide scintigraphy. We evaluated the relationship between 111In-octreotide uptake and the activity of pulmonary fibrosis as assessed by lung function tests, bronchoalveolar lavage (BAL) cellularity, and high-resolution CT (HRCT) of the chest. Planar images and thoracic SPECT (24 h) were performed after injection of 222 MBq of 111In-octreotide. Lung uptake was quantified using the lung-to-background ratio (L/B). In addition, the expression of sst2 was evaluated in vitro, in frozen lung-tissue samples using autoradiography, and in human cultures of lung fibroblasts using a ligand-binding assay. RESULTS: Compared with lung uptake in control patients (median L/B, 1.25; range, 1.14-1.49), lung uptake was increased in all 11 IPF patients (median L/B, 2.63; range, 1.59-3.13; P < 0.001) and in 4 of 6 SSc patients (median L/B, 1.68; range, 1.42-2.16). The L/B was lower in SSc patients than in IPF patients (P = 0.011). Increased uptake correlated with the alteration of lung function (carbon monoxide diffusing capacity [rho = -0.655; P = 0.038], diffusing capacity for carbon monoxide and alveolar volume ratio [rho = -0.627; P = 0.047], vital capacity [rho = -0.609; P = 0.054], and total lung capacity [rho = -0.598; P = 0.058]) and with the intensity of alveolitis (total BAL cellularity [rho = 0.756; P = 0.045], neutrophil counts [rho = 0.738; P = 0.05]), and HRCT fibrosis score (rho = 0.673; P = 0.007). Autoradiography suggested that vascular structures were a prominent binding site. Lung fibroblasts expressed somatostatin receptors in vitro as measured by binding assay. CONCLUSION: Our preliminary results identified an increased expression of sst2 in (mainly idiopathic) pulmonary fibrosis. Lung uptake correlates with the alteration of lung function and with the intensity of alveolitis.

Effects of an alveolar recruitment maneuver on cardiovascular and respiratory parameters during total intravenous anesthesia in ponies

OBJECTIVE: To evaluate pulmonary and cardiovascular effects of a recruitment maneuver (RM) combined with positive end-expiratory pressure (PEEP) during total intravenous anesthesia in ponies. ANIMALS: 6 healthy adult Shetland ponies. PROCEDURE: After premedication with detomidine (10 microg/kg, IV), anesthesia was induced with climazolam (0.06 mg/kg, IV) and ketamine (2.2 mg/kg, IV) and maintained with a constant rate infusion of detomidine (0.024 mg/kg/h), climazolam (0.036 mg/kg/h), and ketamine (2.4 mg/kg/h). The RM was preceded by an incremental PEEP titration and followed by a decremental PEEP titration, both at a constant airway pressure difference (deltaP) of 20 cm H2O. The RM consisted of a stepwise increase in deltaP by 25, 30, and 35 cm H2O obtained by increasing peak inspiratory pressure (PIP) to 45, 50, and 55 cm H2O, while maintaining PEEP at 20 cm H2O. Hemodynamic and pulmonary variables were analyzed at every step of the PEEP titration-RM. RESULTS: During the PEEP titration-RM, there was a significant increase in PaO 2 (+12%), dynamic compliance (+ 62%), and heart rate (+17%) and a decrease in shunt (-19%) and mean arterial blood pressure (-21%) was recorded. Cardiac output remained stable. CONCLUSIONS AND CLINICAL RELEVANCE: Although baseline oxygenation was high, Pa(O2) and dynamic compliance further increased during the RM. Despite the use of high PIP and PEEP and a high tidal volume, limited cardiovascular compromise was detected. A PEEP titration-RM may be used to improve oxygenation in anesthetized ponies. During stable hemodynamic conditions, PEEP titration-RM can be performed with acceptable adverse cardiovascular effects.

Pathogenesis of pulmonary edema: learning from high-altitude pulmonary edema

Pulmonary edema is a problem of major clinical importance resulting from a persistent imbalance between forces that drive water into the airspace of the lung and the biological mechanisms for its removal. Here, we will review the fundamental mechanisms implicated in the regulation of alveolar fluid homeostasis. We will then describe the perturbations of pulmonary fluid homeostasis implicated in the pathogenesis of pulmonary edema in conditions associated with increased pulmonary capillary pressure, namely cardiogenic pulmonary edema and high-altitude pulmonary edema (HAPE), with particular emphasis on the latter that has provided important new insight into underlying mechanisms of pulmonary edema. We will provide evidence that impaired pulmonary endothelial and epithelial nitric oxide synthesis and/or bioavailability may represent a central underlying defect predisposing to exaggerated hypoxic pulmonary vasoconstriction, and, in turn, capillary stress failure and alveolar fluid flooding. We will then demonstrate that exaggerated pulmonary hypertension, while possibly a prerequisite, may not always be sufficient to cause HAPE, and how defective alveolar fluid clearance may represent a second important pathogenic mechanism. Finally, we will outline, how this new insight gained from studies in HAPE, may be translated into the management of pulmonary edema and hypoxemia related disease states in general.

[High altitude pulmonary edema. An experiment of nature to study the underlying mechanisms of hypoxic pulmonary hypertension and pulmonary edema in humans]

High altitude constitutes an exciting natural laboratory for medical research. Over the past decade, it has become clear that the results of high-altitude research may have important implications not only for the understanding of diseases in the millions of people living permanently at high altitude, but also for the treatment of hypoxemia-related disease states in patients living at low altitude. High-altitude pulmonary edema (HAPE) is a life-threatening condition occurring in predisposed, but otherwise healthy subjects, and, therefore, allows to study underlying mechanisms of pulmonary edema in humans, in the absence of confounding factors. Over the past decade, evidence has accumulated that HAPE results from the conjunction of two major defects, augmented alveolar fluid flooding resulting from exaggerated hypoxic pulmonary hypertension, and impaired alveolar fluid clearance related to defective respiratory transepithelial sodium transport. Here, after a brief presentation of the clinical features of HAPE, we review this novel concept. We provide experimental evidence for the novel concept that impaired pulmonary endothelial and epithelial nitric oxide synthesis and/or bioavailability may represent the central underlying defect predisposing to exaggerated hypoxic pulmonary vasoconstriction and alveolar fluid flooding. We demonstrate that exaggerated pulmonary hypertension, while possibly a condition sine qua non, may not be sufficient to cause HAPE, and how defective alveolar fluid clearance may represent a second important pathogenic mechanism. Finally, we outline how this insight gained from studies in HAPE may be translated into the management of hypoxemia related disease states in general.

Dendritic cells and macrophages form a transepithelial network against foreign particulate antigens

Fine particles (0.1-2.5 microm in diameter) may cause increased pulmonary morbidity and mortality. We demonstrate with a cell culture model of the human epithelial airway wall that dendritic cells extend processes between epithelial cells through the tight junctions to collect particles in the "luminal space" and to transport them through cytoplasmic processes between epithelial cells across the epithelium or to transmigrate through the epithelium to take up particles on the epithelial surface. Furthermore, dendritic cells interacted with particle-loaded macrophages on top of the epithelium and with other dendritic cells within or beneath the epithelium to take over particles. By comparing the cellular interplay of dendritic cells and macrophages across epithelial monolayers of different transepithelial electrical resistance, we found that more dendritic cells were involved in particle uptake in A549 cultures showing a low transepithelial electrical resistance compared with dendritic cells in16HBE14o cultures showing a high transepithelial electrical resistance 10 min (23.9% versus 9.5%) and 4 h (42.1% versus 14.6%) after particle exposition. In contrast, the macrophages in A549 co-cultures showed a significantly lower involvement in particle uptake compared with 16HBE14o co-cultures 10 min (12.8% versus 42.8%) and 4 h (57.4% versus 82.7%) after particle exposition. Hence we postulate that the epithelial integrity influences the particle uptake by dendritic cells, and that these two cell types collaborate as sentinels against foreign particulate antigen by building a transepithelial interacting cellular network.

Methylprednisolone Fails to Preserve Pulmonary Surfactant and Blood-Air Barrier Integrity in a Porcine Cardiopulmonary Bypass Model

BACKGROUND: Pulmonary inflammation after cardiac surgery with cardiopulmonary bypass (CPB) has been linked to respiratory dysfunction and ultrastructural injury. Whether pretreatment with methylprednisolone (MP) can preserve pulmonary surfactant and blood-air barrier, thereby improving pulmonary function, was tested in a porcine CPB-model. MATERIALS AND METHODS: After randomizing pigs to placebo (PLA; n = 5) or MP (30 mg/kg, MP; n = 5), animals were subjected to 3 h of CPB with 1 h of cardioplegic cardiac arrest. Hemodynamic data, plasma tumor necrosis factor-alpha (TNF-alpha, ELISA), and pulmonary function parameters were assessed before, 15 min after CPB, and 8 h after CPB. Lung biopsies were analyzed for TNF-alpha (Western blot) or blood-air barrier and surfactant morphology (electron microscopy, stereology). RESULTS: Systemic TNF-alpha increased and cardiac index decreased at 8 h after CPB in PLA (P < 0.05 versus pre-CPB), but not in MP (P < 0.05 versus PLA). In both groups, at 8 h after CPB, PaO(2) and PaO(2)/FiO(2) were decreased and arterio-alveolar oxygen difference and pulmonary vascular resistance were increased (P < 0.05 versus baseline). Postoperative pulmonary TNF-alpha remained unchanged in both groups, but tended to be higher in PLA (P = 0.06 versus MP). The volume fraction of inactivated intra-alveolar surfactant was increased in PLA (58 +/- 17% versus 83 +/- 6%) and MP (55 +/- 18% versus 80 +/- 17%) after CPB (P < 0.05 versus baseline for both groups). Profound blood-air barrier injury was present in both groups at 8 h as indicated by an increased blood-air barrier integrity score (PLA: 1.28 +/- 0.03 versus 1.70 +/- 0.1; MP: 1.27 +/- 0.08 versus 1.81 +/- 0.1; P < 0.05). CONCLUSION: Despite reduction of the systemic inflammatory response and pulmonary TNF-alpha generation, methylprednisolone fails to decrease pulmonary TNF-alpha and to preserve pulmonary surfactant morphology, blood-air barrier integrity, and pulmonary function after CPB.

Exogenous surfactant application in a rat lung ischemia reperfusion injury model: effects on edema formation and alveolar type II cells

BACKGROUND: Prophylactic exogenous surfactant therapy is a promising way to attenuate the ischemia and reperfusion (I/R) injury associated with lung transplantation and thereby to decrease the clinical occurrence of acute lung injury and acute respiratory distress syndrome. However, there is little information on the mode by which exogenous surfactant attenuates I/R injury of the lung. We hypothesized that exogenous surfactant may act by limiting pulmonary edema formation and by enhancing alveolar type II cell and lamellar body preservation. Therefore, we investigated the effect of exogenous surfactant therapy on the formation of pulmonary edema in different lung compartments and on the ultrastructure of the surfactant producing alveolar epithelial type II cells. METHODS: Rats were randomly assigned to a control, Celsior (CE) or Celsior + surfactant (CE+S) group (n = 5 each). In both Celsior groups, the lungs were flush-perfused with Celsior and subsequently exposed to 4 h of extracorporeal ischemia at 4 degrees C and 50 min of reperfusion at 37 degrees C. The CE+S group received an intratracheal bolus of a modified natural bovine surfactant at a dosage of 50 mg/kg body weight before flush perfusion. After reperfusion (Celsior groups) or immediately after sacrifice (Control), the lungs were fixed by vascular perfusion and processed for light and electron microscopy. Stereology was used to quantify edematous changes as well as alterations of the alveolar epithelial type II cells. RESULTS: Surfactant treatment decreased the intraalveolar edema formation (mean (coefficient of variation): CE: 160 mm3 (0.61) vs. CE+S: 4 mm3 (0.75); p < 0.05) and the development of atelectases (CE: 342 mm3 (0.90) vs. CE+S: 0 mm3; p < 0.05) but led to a higher degree of peribronchovascular edema (CE: 89 mm3 (0.39) vs. CE+S: 268 mm3 (0.43); p < 0.05). Alveolar type II cells were similarly swollen in CE (423 microm3(0.10)) and CE+S (481 microm3(0.10)) compared with controls (323 microm3(0.07); p < 0.05 vs. CE and CE+S). The number of lamellar bodies was increased and the mean lamellar body volume was decreased in both CE groups compared with the control group (p < 0.05). CONCLUSION: Intratracheal surfactant application before I/R significantly reduces the intraalveolar edema formation and development of atelectases but leads to an increased development of peribronchovascular edema. Morphological changes of alveolar type II cells due to I/R are not affected by surfactant treatment. The beneficial effects of exogenous surfactant therapy are related to the intraalveolar activity of the exogenous surfactant.

[Unusual pulmonary presentation of systemic Langerhans cell histiocytosis]

An 80-year-old nonsmoking man was referred to our hospital with bilateral perihilar pulmonary opacities. He had a history of epilepsy, sclerosing cholangitis, cutaneous lesions previously diagnosed as localised Langerhans cell histiocytosis. Symptoms included dry cough and dyspnea. Chest CT showed bilateral perihilar alveolar consolidation with bronchiectasis. Histological examination of a lung biopsy showed typical features of Langerhans cell granulomatosis. Investigations revealed anterior and posterior hypopituitarism. An important improvement occurred with corticosteroid and vinblastine treatment.

Circulating plasma surfactant protein type B as biological marker of alveolar-capillary barrier damage in chronic heart failure

BACKGROUND: Surfactant protein type B (SPB) is needed for alveolar gas exchange. SPB is increased in the plasma of patients with heart failure (HF), with a concentration that is higher when HF severity is highest. The aim of this study was to evaluate the relationship between plasma SPB and both alveolar-capillary diffusion at rest and ventilation versus carbon dioxide production during exercise. METHODS AND RESULTS: Eighty patients with chronic HF and 20 healthy controls were evaluated consecutively, but the required quality for procedures was only reached by 71 patients with HF and 19 healthy controls. Each subject underwent pulmonary function measurements, including lung diffusion for carbon monoxide and membrane diffusion capacity, and maximal cardiopulmonary exercise test. Plasma SPB was measured by immunoblotting. In patients with HF, SPB values were higher (4.5 [11.1] versus 1.6 [2.9], P=0.0006, median and 25th to 75th interquartile), whereas lung diffusion for carbon monoxide (19.7+/-4.5 versus 24.6+/-6.8 mL/mm Hg per min, P<0.0001, mean+/-SD) and membrane diffusion capacity (28.9+/-7.4 versus 38.7+/-14.8, P<0.0001) were lower. Peak oxygen consumption and ventilation/carbon dioxide production slope were 16.2+/-4.3 versus 26.8+/-6.2 mL/kg per min (P<0.0001) and 29.7+/-5.9 and 24.5+/-3.2 (P<0.0001) in HF and controls, respectively. In the HF population, univariate analysis showed a significant relationship between plasma SPB and lung diffusion for carbon monoxide, membrane diffusion capacity, peak oxygen consumption, and ventilation/carbon dioxide production slope (P<0.0001 for all). On multivariable logistic regression analysis, membrane diffusion capacity (beta, -0.54; SE, 0.018; P<0.0001), peak oxygen consumption (beta, -0.53; SE, 0.036; P=0.004), and ventilation/carbon dioxide production slope (beta, 0.25; SE, 0.026; P=0.034) were independently associated with SPB. CONCLUSIONS: Circulating plasma SPB levels are related to alveolar gas diffusion, overall exercise performance, and efficiency of ventilation showing a link between alveolar-capillary barrier damage, gas exchange abnormalities, and exercise performance in HF.