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To facilitate computational toxicology, we developed an approach for generating high-resolution lung-anatomy and particle-deposition mouse models. Major processing steps of our method include mouse preparation, serial block-face cryomicrotome imaging, and highly automated image analysis for generating three-dimensional (3D) mesh-based models and volume-based models of lung anatomy (airways, lobes, sublobes, and near-acini structures) that are linked to local particle-deposition measurements. Analysis resulted in 34 mouse models covering 4 different mouse strains (B6C3F1: 8, BALB/C: 11, C57Bl/6: 8, and CD-1: 7) as well as both sexes (16 male and 18 female) and different particle sizes [2 μm ( n = 15), 1 μm ( n = 16), and 0.5 μm ( n = 3)]. On average, resulting mouse airway models had 1,616.9 ± 298.1 segments, a centerline length of 597.6 ± 59.8 mm, and 1,968.9 ± 296.3 outlet regions. In addition to 3D geometric lung models, matching detailed relative particle-deposition measurements are provided. All data sets are available online in the lapdMouse archive for download. The presented approach enables linking relative particle deposition to anatomical structures like airways. This will in turn improve the understanding of site-specific airflows and how they affect drug, environmental, or biological aerosol deposition. NEW & NOTEWORTHY Computer simulations of particle deposition in mouse lungs play an important role in computational toxicology. Until now, a limiting factor was the lack of high-resolution mouse lung models and measured local particle-deposition information, which are required for developing accurate modeling approaches (e.g., computational fluid dynamics). With the developed imaging and analysis approach, we address this issue and provide all of the raw and processed data in a publicly accessible repository.

Despite substantial development of sophisticated subject-specific computational models of aerosol transport and deposition in human lungs, experimental validation of predictions from these new models is sparse. We collected aerosol retention and exhalation profiles in seven healthy volunteers and six subjects with mild-to-moderate COPD (FEV1=50–80%predicted) in the supine posture. Total deposition was measured during continuous breathing of 1 and 2.9 µm-diameter particles (tidal volume of 1 L, flow rate of 0.3 L/s and 0.75 L/s). Bolus inhalations of 1 µm particles were performed to penetration volumes of 200, 500 and 800 mL (flow rate of 0.5 L/s). Aerosol bolus dispersion (H), deposition, and mode shift (MS) were calculated from these data. There was no significant difference in total deposition between healthy subjects and those with COPD. Total deposition increased with increasing particle size and also with increasing flow rate. Similarly, there was no significant difference in aerosol bolus deposition between subject groups. Yet, the rate of increase in dispersion and of decrease in MS with increasing penetration volume was higher in subjects with COPD than in healthy volunteers (H: 0.798±0.205 vs. 0.527±0.122 mL/mL, p=0.01; MS: −0.271±0.129 vs. −0.145 ±0.076 mL/mL, p=0.05) indicating larger ventilation inhomogeneities (based on H) and increased flow sequencing (based on MS) in the COPD than in the healthy group. In conclusion, in the supine posture, deposition appears to lack sensitivity for assessing the effect of lung morphology and/or ventilation distribution alteration induced by mild-to-moderate lung disease on the fate of inhaled aerosols. However, other parameters such as aerosol bolus dispersion and mode shift may be more sensitive parameters for evaluating models of lungs with moderate disease.

Computer models for inhalation toxicology and drug-aerosol delivery studies rely on ventilation pattern inputs for predictions of particle deposition and vapor uptake. However, changes in lung mechanics due to disease can impact airflow dynamics and model results. It has been demonstrated that non-invasive, in vivo, 4DCT imaging (3D imaging at multiple time points in the breathing cycle) can be used to map heterogeneities in ventilation patterns under healthy and disease conditions. The purpose of this study was to validate ventilation patterns measured from CT imaging by exposing the same rats to an aerosol of fluorescent microspheres (FMS) and examining particle deposition patterns using cryomicrotome imaging.Six male Sprague-Dawley rats were intratracheally instilled with elastase to a single lobe to induce a heterogeneous disease. After four weeks, rats were imaged over the breathing cycle by CT then immediately exposed to an aerosol of ∼ 1 μm FMS for ∼ 5 minutes. After the exposure, the lungs were excised and prepared for cryomicrotome imaging, where a 3D image of FMS deposition was acquired using serial sectioning. Cryomicrotome images were spatially registered to match the live CT images to facilitate direct quantitative comparisons of FMS signal intensity with the CT-based ventilation maps.Comparisons of fractional ventilation in contiguous, non-overlapping, 3D regions between CT-based ventilation maps and FMS images showed strong correlations in fractional ventilation (r = 0.888, p0.0001).We conclude that ventilation maps derived from CT imaging are predictive of the 1 μm aerosol deposition used in ventilation-perfusion heterogeneity inhalation studies.

A highly automated method for the segmentation of airways in the serial block-face cryomicrotome images of rat lungs is presented. First, a point inside of the trachea is manually specified. Then, a set of candidate airway centerline points is automatically identified. By utilizing a novel path extraction method, a centerline path between the root of the airway tree and each point in the set of candidate centerline points is obtained. Local disturbances are robustly handled by a novel path extraction approach, which avoids the shortcut problem of standard minimum cost path algorithms. The union of all centerline paths is utilized to generate an initial airway tree structure, and a pruning algorithm is applied to automatically remove erroneous subtrees or branches. Finally, a surface segmentation method is used to obtain the airway lumen. The method was validated on five image volumes of Sprague-Dawley rats. Based on an expert-generated independent standard, an assessment of airway identification and lumen segmentation performance was conducted. The average of airway detection sensitivity was 87.4% with a 95% confidence interval (CI) of (84.9, 88.6)%. A plot of sensitivity as a function of airway radius is provided. The combined estimate of airway detection specificity was 100% with a 95% CI of (99.4, 100)%. The average number and diameter of terminal airway branches was 1179 and 159 μm, respectively. Segmentation results include airways up to 31 generations. The regression intercept and slope of airway radius measurements derived from final segmentations were estimated to be 7.22 μm and 1.005, respectively. The developed approach enables the quantitative studies of physiology and lung diseases in rats, requiring detailed geometric airway models.

We aimed to investigate the role of endothelin-mediated vasoconstriction following acute pulmonary thromboembolism (APTE). Thirteen anesthetized piglets (~25 kg) were ventilated with 0 PEEP. Cardiac output (Qt) and wedge pressure (Pw) were measured by a Swan Ganz catheter, along with arterial and venous blood gases. APTE was induced by autologous blood clots (~0.8 g/kg, 12-16 pieces) via a jugular venous catheter at time = 0 minutes until the mean pulmonary arterial pressure (Ppa) was about 2.5 times the baseline at 30 minutes. Eight control animals (Group 1) received only normal saline afterward, while the remaining five (Group 2) received at time = 40-minute saline plus Tezosentan, a nonspecific endothelin antagonist. The drug was initially given as an intravenous bolus (10 mg/kg), followed by an infusion (2 mg/min) until the end of the experiment at 2 hours. Hemodynamic data were measured before APTE and then at 30-minute intervals. Pulmonary vascular resistance index (PVRI) was calculated as (Ppa-Pw)/CI, where CI was cardiac index or Qt/W (body weight). Fluorescent microspheres (FMS) were used to mark regional blood flows and ventilation for cluster analysis. PVRI acutely increased within minutes and remained high despite some recovery over time. With Tezosentan treatment, the results showed that endothelin-mediated vasoconstriction persisted significantly up to 2 hours and accounted for about 25% of the increase in PVRI while clot obstruction accounted for the remaining 75%. CI remained relatively constant throughout. Tezosentan also affected PVRI indirectly by mitigating the shift of regional blood flow back to the embolized areas over time, possibly by attenuating vasoconstriction in the nonembolized areas. We conclude that following APTE, although the increased PVRI is mostly due to mechanical embolic obstruction, secondary factors such as vasoconstriction and pattern of regional blood flow over time also play important roles.

Long-term survival after lung transplantation is limited by acute and chronic graft rejection. Induction of immune tolerance by first establishing mixed hematopoietic chimerism (MC) is a promising strategy to improve outcomes. In a preclinical canine model, stable MC was established in recipients after reduced-intensity conditioning and hematopoietic cell transplantation from a DLA-identical donor. Delayed lung transplantation was performed from the stem cell donor without pharmacological immunosuppression. Lung graft survival without loss of function was prolonged in chimeric (n = 5) vs. nonchimeric (n = 7) recipients (por = 0.05, Fisher's test). There were histological changes consistent with low-grade rejection in 3/5 of the lung grafts in chimeric recipients ator =1 year. Chimeric recipients after lung transplantation had a normal immune response to a T-dependent antigen. Compared to normal dogs, there were significant increases of CD4+INFgamma+, CD4+IL-4+ and CD8+ INFgamma+ T-cell subsets in the blood (p0.0001 for each of the three T-cell subsets). Markers for regulatory T-cell subsets including foxP3, IL10 and TGFbeta were also increased in CD3+ T cells from the blood and peripheral tissues of chimeric recipients after lung transplantation. Establishing MC is immunomodulatory and observed changes were consistent with activation of both the effector and regulatory immune response.

Exhaled acetone is measured to estimate exposure or monitor diabetes and congestive heart failure. Interpreting this measurement depends critically on where acetone exchanges in the lung. Health professionals assume exhaled acetone originates from alveolar gas exchange, but experimental data and theoretical predictions suggest that acetone comes predominantly from airway gas exchange. We measured endogenous acetone in the exhaled breath to evaluate acetone exchange in the lung. The acetone concentration in the exhalate of healthy human subjects was measured dynamically with a quadrupole mass spectrometer and was plotted against exhaled volume. Each subject performed a series of breathing maneuvers in which the steady exhaled flow rate was the only variable. Acetone phase III had a positive slope (0.054 ± 0.016 liter−1) that was statistically independent of flow rate. Exhaled acetone concentration was normalized by acetone concentration in the alveolar air, as estimated by isothermal rebreathing. Acetone concentration in the rebreathed breath ranged from 0.8 to 2.0 parts per million. Normalized end-exhaled acetone concentration was dependent on flow and was 0.79 ± 0.04 and 0.85 ± 0.04 for the slow and fast exhalation rates, respectively. A mathematical model of airway and alveolar gas exchange was used to evaluate acetone transport in the lung. By doubling the connective tissue (epithelium + mucosal tissue) thickness, this model predicted accurately ( R2 = 0.94 ± 0.05) the experimentally measured expirograms and demonstrated that most acetone exchange occurred in the airways of the lung. Therefore, assays using exhaled acetone measurements need to be reevaluated because they may underestimate blood levels.

Hypoxic pulmonary vasoconstriction (HPV) is known to affect regional pulmonary blood flow distribution. It is unknown whether lungs with well-matched ventilation (V̇)/perfusion (Q̇) have regional differences in the HPV response. Five prone pigs were anesthetized and mechanically ventilated (positive end-expiratory pressure = 2 cmH2O). Two hypoxic preconditions [inspired oxygen fraction (FiO2) = 0.13] were completed to stabilize the animal's hypoxic response. Regional pulmonary blood Q̇ and V̇ distribution was determined at various FiO2 (0.21, 0.15, 0.13, 0.11, 0.09) using the fluorescent microsphere technique. Q̇ and V̇ in the lungs were quantified within 2-cm3 lung pieces. Pieces were grouped, or clustered, based on the changes in blood flow when subjected to increasing hypoxia. Unique patterns of Q̇ response to hypoxia were seen within and across animals. The three main patterns (clusters) showed little initial difference in V̇/Q̇ matching at room air where the mean V̇/Q̇ range was 0.92–1.06. The clusters were spatially located in cranial, central, and caudal portions of the lung. With decreasing FiO2, blood flow shifted from the cranial to caudal regions. We determined that pulmonary blood flow changes, caused by HPV, produced distinct response patterns that were seen in similar regions across our prone porcine model.

We studied the spatial distribution of the abnormal ventilation-perfusion (V̇a/Q̇) units in a porcine model of acute pulmonary thromboembolism (APTE), using the fluorescent microsphere (FMS) technique. Four piglets (∼22 kg) were anesthetized and ventilated with room air in the prone position. Each received ∼20 g of preformed blood clots at time t = 0 min via a large-bore central venous catheter, until the mean pulmonary arterial pressure reached 2.5 times baseline. The distributions of regional V̇a and blood flow (Q̇) at five time points ( t = −30, −5, 30, 60, 120 min) were mapped by FMS of 10 distinct colors, i.e., aerosolization of 1-μm FMS for labeling V̇a and intravenous injection of 15-μm FMS for labeling Q̇. Our results showed that, at t = 30 min following APTE, mean V̇a/Q̇ (V̇a/Q̇ = 2.48 ± 1.12) and V̇a/Q̇ heterogeneity (log SD V̇a/Q̇ = 1.76 ± 0.23) were significantly increased. There were also significant increases in physiological dead space (11.2 ± 12.7% at 60 min), but the shunt fraction (V̇a/Q̇ = 0) remained minimal. Cluster analyses showed that the low V̇a/Q̇ units were mainly seen in the least embolized regions, whereas the high V̇a/Q̇ units and dead space were found in the peripheral subpleural regions distal to the clots. At 60 and 120 min, there were modest recoveries in the hemodynamics and gas exchange toward baseline. Redistribution pattern was mostly seen in regional Q̇, whereas V̇a remained relatively unchanged. We concluded that the hypoxemia seen after APTE could be explained by the mechanical diversion of Q̇ to the less embolized regions because of the vascular obstruction by clots elsewhere. These low V̇a/Q̇ units created by high flow, rather than low V̇a, accounted for most of the resultant hypoxemia.

Hypoxic pulmonary vasoconstriction (HPV) serves to maintain optimal gas exchange by decreasing perfusion to hypoxic regions. However, global hypoxia and nonuniform HPV may result in overperfusion of poorly constricted regions leading to local edema seen in high-altitude pulmonary edema. To quantify the spatial distribution of HPV and its response to regional Po2 (PrO2) among small lung regions, five pigs were anesthetized and mechanically ventilated in the supine posture. The animals were ventilated with an inspired O2 fraction (FiO2) of 0.50 and 0.21 and then (in random order) 0.15, 0.12, and 0.09. Regional blood flow (Q̇) and alveolar ventilation (V̇a) were measured by using intravenous infusion of 15 μm and inhalation of 1-μm fluorescent microspheres, respectively. PrO2 was calculated for each piece at each FiO2. Lung pieces differed in their Q̇ response to hypoxia in a manner related to their initial V̇a/Q̇ with FiO2 = 0.21. Reducing FiO2 < 0.15 decreased Q̇ to the initially high V̇a/Q̇ (higher PrO2) regions and forced Q̇ into the low V̇a/Q̇ (dorsal-caudal) regions. Resistance increased in most lung pieces as PrO2 decreased, reaching a maximum resistance when PrO2 is between 40 and 50 Torr. Local resistance decreased at Pro2 < 40 Torr. Pieces were statistically clustered with respect to their relative Q̇ response pattern to each FiO2. Some clusters were shown to be spatially organized. We conclude that HPV is spatially heterogeneous. The heterogeneity of Q̇ response may be related, in part, to the heterogeneity of baseline V̇a/Q̇.

Pulmonary arterial occlusion pressure is not thought to reflect left atrial pressure (Pla) when alveolar pressure (PA) exceeds pulmonary venous pressure because alveolar capillaries collapse and the required continuous fluid column between the pulmonary artery and left atrium is interrupted. However, arterial-to-venous flow can occur when PA exceeds both the pulmonary arterial pressure (Ppa) and pulmonary venous pressure (i.e., in Zone 1 conditions), indicating the existence of a continuous patent vascular channel. Accordingly, Ppa should reflect Pla under these conditions. To investigate this connection cannulas were placed in the pulmonary arteries and left atria of eight excised rabbit lungs. Ppa and Pla were set 5 cm H2O above PA, which ranged from 0 to 25 cm H2O. Pla was then reduced in 2 to 4 cm H2O decrements while recording Ppa when arterial-to-venous flow ceased. At all PAs greater than 0 cm H2O, Pla was accurately reflected by the Ppa when both were exceeded by PA. The greater the PA, the lower the Ppa could track Pla below PA. Pla can be accurately measured by a pulmonary arterial catheter under Zone 1 conditions.

We propose a model to measure both regional ventilation (V˙) and perfusion (Q˙) in which the regional radiodensity (RD) in the lung during xenon (Xe) washin is a function of regional V˙ (increasing RD) and Q˙(decreasing RD). We studied five anesthetized, paralyzed, mechanically ventilated, supine sheep. Four 2.5-mm-thick computed tomography (CT) images were simultaneously acquired immediately cephalad to the diaphragm at end inspiration for each breath during 3 min of Xe breathing. Observed changes in RD during Xe washin were used to determine regional V˙ and Q˙. For 16 mm3, Q˙ displayed more variance than V˙: the coefficient of variance of Q˙ (CVQ˙) = 1.58 ± 0.23, the CV of V˙ (CVV˙) = 0.46 ± 0.07, and the ratio of CVQ˙ to CVV˙ = 3.5 ± 1.1. CVQ˙(1.21 ± 0.37) and the ratio of CVQ˙ to CVV˙ (2.4 ± 1.2) were smaller at 1,000-mm3 scale, but CVV˙ (0.53 ± 0.09) was not. V˙/Q˙ distributions also displayed scale dependence: log SD of V˙ and log SD of Q˙ were 0.79 ± 0.05 and 0.85 ± 0.10 for 16-mm3 and 0.69 ± 0.20 and 0.67 ± 0.10 for 1,000-mm3 regions of lung, respectively. V˙ and Q˙ measurements made with CT and Xe also demonstrate vertically oriented and isogravitational heterogeneity, which are described using other methodologies. Sequential images acquired by CT during Xe breathing can be used to determine both regional V˙ and Q˙ noninvasively with high spatial resolution.

Hyperventilation with mixtures of O2 and CO2 has long been known to enhance carbon monoxide (CO) elimination at low HbCO levels in animals and humans. The effect of this therapy on oxygen delivery (DO2) has not been studied. Isocapnic hyperventilation utilizing mechanical ventilation may decrease cardiac output and therefore decrease DO2 while increasing CO elimination. We studied the effects of isocapnic hyperventilation on five adult mechanically ventilated sheep exposed to multiple episodes of severe CO poisoning. Five ventilatory patterns were studied: baseline minute ventilation (RR. VT), twice (2. RR) and four times (4. RR) baseline respiratory rate, and twice (2. VT) and four times (4. VT) baseline tidal volume. The mean carboxyhemoglobin (HbCO) washout half-time (t1/2) was 14.3 +/- 1.6 min for RR. VT, decreasing to 9.5 +/- 0.9 min for 2. RR, 8.0 +/- 0.5 min for 2. VT, 6.2 +/- 0.5 min for 4. RR, and 5.2 +/- 0.5 min for 4. VT. DO2 was increased during hyperventilation compared with baseline ventilation for 2. VT, 4. RR, and 4. VT ventilatory patterns. Isocapnic hyperventilation, in our animal model, did not alter arterial or pulmonary blood pressures, arterial pH, or cardiac output. Isocapnic hyperventilation is a promising therapy for CO poisoning.

The microsphere method has been used extensively to measure regional blood flow in large laboratory animals. A fundamental premise of the method is that microspheres do not alter regional flow or vascular tone. Whereas this assumption is accepted in large animals, it may not be valid in the pulmonary circulation of smaller animals. Three studies were performed to determine the hemodynamic effects of microspheres on the rat pulmonary circulation. Increasing numbers of 15-μm-diameter microspheres were injected into a fully dilated, isolated-lung preparation. Vascular resistance increased 0.8% for every 100,000 microspheres injected. Microspheres were also injected into an isolated-lung preparation in which vascular tone was increased with hypoxia. Microspheres did not induce vasodilatation, as reported in other vascular beds. Fluorescent microspheres were injected via tail veins into awake rats, and the spatial locations of the microspheres were determined. Regional distributions remained highly correlated when microspheres of one color were injected after microspheres of another color. This indicates that the initial injection did not alter regional perfusion. We conclude that, when used in appropriate numbers, 15-μm-diameter microspheres do not alter regional flow or vascular tone in the rat pulmonary circulation.

Prior studies exploring the spatial distributions of ventilation and perfusion have partitioned the lung into discrete regions not constrained by anatomical boundaries and may blur regional differences in perfusion and ventilation. To characterize the anatomical heterogeneity of regional ventilation and perfusion, we administered fluorescent microspheres to mark regional ventilation and perfusion in five Sprague–Dawley rats and then using highly automated computer algorithms, partitioned the lungs into regions defined by anatomical structures identified in the images. The anatomical regions ranged in size from the near-acinar to the lobar level. Ventilation and perfusion were well correlated at the smallest anatomical level. Perfusion and ventilation heterogeneity were relatively less in rats compared to data previously published in larger animals. The more uniform distributions may be due to a smaller gravitational gradient and/or the fewer number of generations in the distribution trees before reaching the level of gas exchange, making regional matching of ventilation and perfusion less extensive in small animals.

Inhalation of antigen in immunized mice induces an infiltration of eosinophils into the airways and increased bronchial hyperreactivity as are observed in human asthma. We employed a model of late-phase allergic pulmonary inflammation in mice to address the role of leukotrienes (LT) in mediating airway eosinophilia and hyperreactivity to methacholine. Allergen intranasal challenge in OVA-sensitized mice induced LTB4 and LTC4 release into the airspace, widespread mucus occlusion of the airways, leukocytic infiltration of the airway tissue and broncho-alveolar lavage fluid that was predominantly eosinophils, and bronchial hyperreactivity to methacholine. Specific inhibitors of 5-lipoxygenase and 5-lipoxygenase-activating protein (FLAP) blocked airway mucus release and infiltration by eosinophils indicating a key role for leukotrienes in these features of allergic pulmonary inflammation. The role of leukotrienes or eosinophils in mediating airway hyperresponsiveness to aeroallergen could not be established, however, in this murine model.

In the current study, we used a canine model of radiation-induced lung injury to test the effect of a single i.v. infusion of 10×10(6)/kg of marrow fibroblasts on the progression of damage following 15 Gy exposure to the right lung. The fibroblasts, designated DS1 cells, are a cloned population of immortalized cells isolated from a primary culture of marrow stromal cells. DS1 cells were infused at week 5 post-irradiation when lung damage was evident by imaging with high-resolution computed tomography (CT). At 13 weeks post-irradiation we found that 4 out of 5 dogs receiving DS1 cells had significantly improved pulmonary function compared to 0 out of 5 control dogs (p = 0.047, Fisher's Exact). Pulmonary function was measured as the single breath diffusion capacity-hematocrit (DLCO-Hct), the total inspiratory capacity (IC), and the total lung capacity (TLC), which differed significantly between control and DS1-treated dogs; p = 0.002, p = 0.005, and p = 0.004, respectively. The DS1-treated dogs also had less pneumonitis detected by CT imaging and an increased number of TTF-1 (thyroid transcription factor 1, NKX2-1) positive cells in the bronchioli and alveoli compared to control dogs. Endothelial-like progenitor cells (ELC) of host origin, detected by colony assays, were found in peripheral blood after DS1 cell infusion. ELC numbers peaked one day after infusion, and were not detectable by 7 days. These data suggest that infusion of marrow fibroblasts stimulates mobilization of ELC, which is associated with a reduction in otherwise progressive radiation-induced lung injury. We hypothesize that these two observations are related, specifically that circulating ELC contribute to increased angiogenesis, which facilitates endogenous lung repair.

Real-time imaging of cellular and sub-cellular dynamics in vascularized organs requires image-resolution, image-registration, and demonstrably intact physiology to be simultaneously optimized. This problem is particularly pronounced in the lung in which cells may transit at speeds > 1 mm/sec, and in which normal respiration results in large-scale tissue movements that prevent image registration. Here, we report video-rate, two-photon imaging of a physiologically intact preparation of the mouse lung that is at once stabilizing and non-disruptive. The application of our method provides evidence for differential trapping of T cells and neutrophils in mouse pulmonary capillaries and enables observation of neutrophil mobilization and dynamic vascular leak in response to stretch and inflammatory models of lung injury in mice. The system permits physiological measurement of motility rates of > 1 mm/sec, observation of detailed cellular morphology, and could be applied to other organs and tissues while maintaining intact physiology.

Pulmonary computational fluid dynamics models require that three-dimensional images be acquired over multiple points in the dynamic breathing cycle without breath holds or changes in ventilatory mechanics. With small animals, these requirements can result in long imaging times (∼90 minutes), over which lung mechanics, such as compliance, may gradually change if not carefully monitored and controlled. These changes, caused by derecruitment of parenchymal tissue, are manifested as an upward drift in peak inspiratory pressure (PIP) or by changes in the pressure waveform and/or lung volume over the course of the experiment. We demonstrate highly repeatable mechanical ventilation in anesthetized rats over a long duration for dynamic lung x-ray computed tomography (CT) imaging. We describe significant updates to a basic commercial ventilator that was acquired for these experiments. Key to achieving consistent results was the implementation of periodic deep breaths, or sighs, of extended duration to maintain lung recruitment. In addition, continuous monitoring of breath-to-breath pressure and volume waveforms and long-term trends in PIP and flow provide diagnostics of changes in breathing mechanics.

Previous studies reported that regional CO2tension might affect regional ventilation (V̇) following acute pulmonary thromboembolism (APTE). We investigated the pathophysiology and magnitude of these changes. Eight anesthetized and ventilated piglets received autologous clots at time = 0 min until mean pulmonary artery pressure was 2.5 times baseline. The distribution of V̇ and perfusion (Q̇) at four different times (−5, 30, 60, 120 min) was mapped by fluorescent microspheres. Regional V̇ and Q̇ were examined postmortem by sectioning the air-dried lung into 900–1,000 samples of ∼2 cm3each. After the redistribution of regional Q̇ by APTE, but in the scenario assuming that no V̇ shift had yet occurred, CO2tension in different lung regions at 30 min post-APTE (PXCO2) was estimated from the V̇/Q̇ data and divided into four distinct clusters: i.e., PXCO2< 10 Torr; 10 < PXCO2< 25 Torr; 25 < PXCO2< 50 Torr; PXCO2> 50 Torr. Our data showed that the clusters in higher V̇/Q̇ regions (with a PXCO2< 25 Torr) received ∼35% less V̇ when measured within 30 min of APTE, whereas, in contrast, the lower V̇/Q̇ regions showed no statistically significant increases in their V̇. However, after 30 min, there was minimal further redistribution of V̇. We conclude that there are significant compensatory V̇ shifts out of regions of low CO2tension soon following APTE, and that these variations in regional CO2tension, which initiate CO2-dependent changes in airway resistance and lung parenchymal compliance, can lead to improved gas exchange.

Previous studies reported that the degree of hypoxemia following acute pulmonary thromboembolism (APTE) was highly variable and that its mechanism was mainly due to the creation of many high and low ventilation/perfusion (V/Q) units, as a result of the heterogeneous regional blood flow (Q) caused by embolic obstruction. We studied the effect of changing cardiac output (Qt) on gas exchange after APTE in 5 embolized piglets (23 ± 3 Kg), using Dobutamine intermittently at approximately 20 μg/kg/min for 120 minutes. The distribution of ventilation (V) and perfusion (Q) at various times was mapped using fluorescent microspheres in 941 ± 60 lung regions. After APTE, increase in Qt by Dobutamine improved venous oxygen tension (PvO2) but arterial PaO2 did not change consistently. On the other hand, cluster analysis showed that the V/Q ratio of most lung regions was lowered due to increases in Q at the same time. We concluded that the effect of changing cardiac output on gas exchange following APTE was affected by the simultaneous and varying balance between the changing V/Q mismatch and the concomitantly changing PvO2, which might explain the unpredictability of PaO2 in the clinical setting.

We studied the roles of endothelins in determining ventilation (V̇a) and perfusion (Q̇) mismatch in a porcine model of acute pulmonary thromboembolism (APTE), using a nonspecific endothelin antagonist, tezosentan. Nine anesthetized piglets (∼23 kg) received autologous clots (∼20 g) via a central venous catheter at time = 0 min. The distribution of V̇a and Q̇ at five different time points (−30, −5, 30, 60, 120 min) was mapped by fluorescent microspheres of 10 different colors. Five piglets ( group 1) received tezosentan (courtesy of Actelion) starting at time = 40 min for 2 h, and four piglets ( group 2) received only saline and served as control. Our results showed that, in all of the animals at 30 min following APTE but before tezosentan, the mean V̇a/Q̇ was increased, as was V̇a/Q̇ heterogeneity (log SD V̇a/Q̇), which represented a widening of its main peak. Afterwards, tezosentan attenuated the pulmonary hypertension in group 1 but also produced moderate systemic hypotension. However, it did not improve arterial Po2 or V̇a/Q̇ mismatch. We concluded that endothelin antagonism had minimal impact on gas exchange following APTE and confirmed our earlier observation that the main mechanism for hypoxemia in APTE was due to the mechanical redistribution of pulmonary regional blood flow away from the embolized vessels, resulting in the creation of many divergent low and high V̇a/Q̇ regions.

We have shown previously that inspired CO2 (3-5%) improves ventilation-perfusion (V̇a/Q̇) matching but with the consequence of mild arterial hypercapnia and respiratory acidosis. We hypothesized that adding CO2 only late in inspiration to limit its effects to the conducting airways would enhance V̇a/Q̇ matching and improve oxygenation without arterial hypercapnia. CO2 was added in the latter half of inspiration in a volume aimed to reach a concentration of 5% in the conducting airways throughout the respiratory cycle. Ten mixed-breed dogs were anesthetized and, in a randomized order, ventilated with room air, 5% CO2 throughout inspiration, and CO2 added only to the latter half of inspiration. The multiple inert-gas elimination technique was used to assess V̇a/Q̇ heterogeneity. Late-inspired CO2 produced only very small changes in arterial pH (7.38 vs. 7.40) and arterial CO2 (40.6 vs. 39.4 Torr). Compared with baseline, late-inspired CO2 significantly improved arterial oxygenation (97.5 vs. 94.2 Torr), decreased the alveolar-arterial Po2 difference (10.4 vs. 15.7 Torr) and decreased the multiple inert-gas elimination technique-derived arterial-alveolar inert gas area difference, a global measurement of V̇a/Q̇ heterogeneity (0.36 vs. 0.22). These changes were equal to those with 5% CO2 throughout inspiration (arterial Po2, 102.5 Torr; alveolar-arterial Po2 difference, 10.1 Torr; and arterial-alveolar inert gas area difference, 0.21). In conclusion, we have established that the majority of the improvement in gas exchange efficiency with inspired CO2 can be achieved by limiting its application to the conducting airways and does not require systemic acidosis.

To investigate whether hypercapnic acidosis protects against ventilator-induced lung injury (VILI) in vivo, we subjected 12 anesthetized, paralyzed rabbits to high tidal volume ventilation (25 cc/kg) at 32 breaths per minute and zero positive end-expiratory pressure for 4 hours. Each rabbit was randomized to receive either an FI(CO(2)) to achieve eucapnia (Pa(CO(2)) approximately 40 mm Hg; n = 6) or hypercapnic acidosis (Pa(CO(2)) 80-100 mm Hg; n = 6). Injury was assessed by measuring differences between the two groups' respiratory mechanics, gas exchange, wet:dry weight, bronchoalveolar lavage fluid protein concentration and cell count, and injury score. The eucapnic group showed significantly higher plateau pressures (27.0 +/- 2.5 versus 20.9 +/- 3.0; p = 0.016), change in Pa(O(2)) (165.2 +/- 19.4 versus 77.3 +/- 87.9 mm Hg; p = 0.02), wet:dry weight (9.7 +/- 2.3 versus 6.6 +/- 1.8; p = 0.04), bronchoalveolar lavage protein concentration (1,350 +/- 228 versus 656 +/- 511 micro g/ml; p = 0.03), cell count (6.86 x 10(5) +/- 0.18 x 10(5) versus 2.84 x 10(5) +/- 0.28 x 10(5) nucleated cells/ml; p = 0.021), and injury score (7.0 +/- 3.3 versus 0.7 +/- 0.9; p < 0.0001). We conclude that hypercapnic acidosis is protective against VILI in this model.

To compare the relative contributions of gravity and vascular structure to the distribution of pulmonary blood flow, we flew with pigs on the National Aeronautics and Space Administration KC-135 aircraft. A series of parabolas created alternating weightlessness and 1.8-G conditions. Fluorescent microspheres of varying colors were injected into the pulmonary circulation to mark regional blood flow during different postural and gravitational conditions. The lungs were subsequently removed, air dried, and sectioned into ∼2 cm3 pieces. Flow to each piece was determined for the different conditions. Perfusion heterogeneity did not change significantly during weightlessness compared with normal and increased gravitational forces. Regional blood flow to each lung piece changed little despite alterations in posture and gravitational forces. With the use of multiple stepwise linear regression, the contributions of gravity and vascular structure to regional perfusion were separated. We conclude that both gravity and the geometry of the pulmonary vascular tree influence regional pulmonary blood flow. However, the structure of the vascular tree is the primary determinant of regional perfusion in these animals.

Observations made on vessels seen directly beneath the pleura may not accurately reflect what occurs in vessels located deeper in the interior of the lung. We quantified flow to subpleural and deeper, interior regions under zone 1 or 2 conditions in excised ( n = 5) and in vivo ( n = 6) rabbit lungs, in the head-up or inverted position. After infusion of radiolabeled microspheres, lungs were dried at alveolar pressure of 25 cmH2O and sliced in 1-cm sections along the gravitational plane and in three planes in the dorsal-ventral axis. Regions located

Immunized mice after inhalation of specific antigen have the following characteristic features of human asthma: airway eosinophilia, mucus and Th2 cytokine release, and hyperresponsiveness to methacholine. A model of late-phase allergic pulmonary inflammation in ovalbumin-sensitized mice was used to address the role of the alpha4 integrin (CD49d) in mediating the airway inflammation and hyperresponsiveness. Local, intrapulmonary blockade of CD49d by intranasal administration of CD49d mAb inhibited all signs of lung inflammation, IL-4 and IL-5 release, and hyperresponsiveness to methacholine. In contrast, CD49d blockade on circulating leukocytes by intraperitoneal CD49d mAb treatment only prevented the airway eosinophilia. In this asthma model, a CD49d-positive intrapulmonary leukocyte distinct from the eosinophil is the key effector cell of allergen-induced pulmonary inflammation and hyperresponsiveness.