The complement cascade in lung injury and disease.

BACKGROUND: The complement system is an important arm of immune defense bringing innate and adaptive immunity. Although originally regarded as a major complementary defense mechanism against pathogens, continuously emerging evidence has uncovered a central role of this complex system in several diseases including lung pathologies. MAIN BODY: Complement factors such as anaphylatoxins C3a and C5a, their receptors C3aR, C5aR and C5aR2 as well as complement inhibitory proteins CD55, CD46 and CD59 have been implicated in pathologies such as the acute respiratory distress syndrome, pneumonia, chronic obstructive pulmonary disease, asthma, interstitial lung diseases, and lung cancer. However, the exact mechanisms by which complement factors induce these diseases remain unclear. Several complement-targeting monoclonal antibodies are reported to treat lung diseases. CONCLUSIONS: The complement system contributes to the progression of the acute and chronic lung diseases. Better understanding of the underlying mechanisms will provide groundwork to develop new strategy to target complement factors for treatment of lung diseases.

Ghrelin alterations during experimental and human sepsis.

BACKGROUND: Ghrelin is a hormone mainly produced by cells of the gastric mucosa, which has been shown to possess anti-inflammatory and immunomodulatory properties. The objective of the study was to investigate ghrelin levels during sepsis, as well as in an experimental sepsis model. METHODS: All consecutive admissions to the ICU of a tertiary hospital in Athens, Greece were screened for eligibility during the study. Thirty four non-septic patients upon ICU admission who subsequently developed sepsis were enrolled. Clinical data and scores were recorded, and blood samples were obtained at baseline (upon ICU admission), and at sepsis development. Total and active ghrelin, leptin, and cytokines were measured. Moreover, lipopolysaccharide (LPS) was administered to mice in order to induce endotoxemia and at specified time points, blood and tissue samples were collected. RESULTS: In patients, serum total and active ghrelin concentrations were significantly elevated in sepsis compared to baseline (553.8 ±â€¯213.4 vs 193.5 ±â€¯123.2, p < 0.001; 254.3 ±â€¯70.6 vs 56.49 ±â€¯16.3, p < 0.001). Active ghrelin levels at the sepsis stage were inversely correlated with SOFA score and length of stay in the ICU (p = 0.023 and p = 0.027 respectively). In the mouse endotoxemia model ghrelin levels were elevated following LPS treatment, and the same trend was observed for leptin, TNFα and IL-6. Ghrelin administration managed to reduce IL-6 levels in mouse serum and in BALF. Pulmonary expression of ghrelin and its receptor GHSR1a was found decreased in LPS-treated mice. CONCLUSIONS: In a well-defined cohort of ICU patients, we have demonstrated that active and total ghrelin increase in sepsis. The same is true for the experimental sepsis model used in the study. The inverse correlation of active ghrelin levels with SOFA score and length of ICU stay among septic patients is indicative of a potential protective role of active ghrelin during the septic process.

Validation of the new Sepsis-3 definitions: proposal for improvement in early risk identification.

OBJECTIVES: Sepsis-3 definitions generated controversies regarding their general applicability. The Sepsis-3 Task Force outlined the need for validation with emphasis on the quick Sequential Organ Failure Assessment (qSOFA) score. This was done in a prospective cohort from a different healthcare setting. METHODS: Patients with infections and at least two signs of systemic inflammatory response syndrome (SIRS) were analysed. Sepsis was defined as total SOFA ≥2 outside the intensive care unit (ICU) or as an increase of ICU admission SOFA ≥2. The primary endpoints were the sensitivity of qSOFA outside the ICU and sepsis definition both outside and within the ICU to predict mortality. RESULTS: In all, 3346 infections outside the ICU and 1058 infections in the ICU were analysed. Outside the ICU, respective mortality with ≥2 SIRS and qSOFA ≥2 was 25.3% and 41.2% (p <0.0001); the sensitivities of qSOFA and of sepsis definition to predict death were 60.8% and 87.2%, respectively. This was 95.9% for sepsis definition in the ICU. The sensitivity of qSOFA and of ≥3 SIRS criteria for organ dysfunction outside the ICU was 48.7% and 72.5%, respectively (p <0.0001). Misclassification outside the ICU with the 1991 and Sepsis-3 definitions into stages of lower severity was 21.4% and 3.7%, respectively (p <0.0001) and 14.9% and 3.7%, respectively, in the ICU (p <0.0001). Adding arterial pH ≤7.30 to qSOFA increased sensitivity for prediction of death to 67.5% (p 0.004). CONCLUSIONS: Our analysis positively validated the use of SOFA score to predict unfavourable outcome and to limit misclassification into lower severity. However, qSOFA score had inadequate sensitivity for early risk assessment.

Exhaled nitric oxide and carbon monoxide in mechanically ventilated brain-injured patients.

The inflammatory influence and biological markers of prolonged mechanical-ventilation in uninjured human lungs remains controversial. We investigated exhaled nitric oxide (NO) and carbon monoxide (CO) in mechanically-ventilated, brain-injured patients in the absence of lung injury or sepsis at two different levels of positive end-expiratory pressure (PEEP). Exhaled NO and CO were assessed in 27 patients, without lung injury or sepsis, who were ventilated with 8 ml kg(-1) tidal volumes under zero end-expiratory pressure (ZEEP group, n = 12) or 8 cm H2O PEEP (PEEP group, n = 15). Exhaled NO and CO was analysed on days 1, 3 and 5 of mechanical ventilation and correlated with previously reported markers of inflammation and gas exchange. Exhaled NO was higher on day 3 and 5 in both patient groups compared to day 1: (PEEP group: 5.8 (4.4-9.7) versus 11.7 (6.9-13.9) versus 10.7 (5.6-16.6) ppb (p < 0.05); ZEEP group: 5.3 (3.8-8.8) versus 9.8 (5.3-12.4) versus 9.6 (6.2-13.5) ppb NO peak levels for days 1, 3 and 5, respectively, p < 0.05). Exhaled CO remained stable on day 3 but significantly decreased by day 5 in the ZEEP group only (6.3 (4.3-9.0) versus 8.1 (5.8-12.1) ppm CO peak levels for day 5 versus 1, p < 0.05). The change scores for peak exhaled CO over day 1 and 5 showed significant correlations with arterial blood pH and plasma TNF levels (r s = 0.49, p = 0.02 and r s = -0.51 p = 0.02, respectively). Exhaled NO correlated with blood pH in the ZEEP group and with plasma levels of IL-6 in the PEEP group. We observed differential changes in exhaled NO and CO in mechanically-ventilated patients even in the absence of manifest lung injury or sepsis. These may suggest subtle pulmonary inflammation and support application of real time breath analysis for molecular monitoring in critically ill patients.

Elevated soluble endothelial protein C receptor levels at ICU admission are associated with sepsis development.

BACKGROUND: The endothelial protein C receptor (EPCR) is a protein that regulates the protein C anticoagulant and anti-inflammatory pathways. A soluble form of EPCR (sEPCR) circulates in plasma and inhibits activated protein C (APC) activities. The clinical impact of sEPCR and its involvement in the septic process is under investigation. In this study, we assessed the role of sEPCR levels as an early indicator of sepsis development. METHODS: Plasma sEPCR levels were measured in 59 critically-ill non-septic patients at the time of admission to the intensive care unit (ICU). Multiple logistic regression analysis was performed to identify potential risk factors for sepsis development and Cox-Regression models were fitted for variables to examine their relationship with time to sepsis development. RESULTS: Thirty patients subsequently developed sepsis and 29 did not. At ICU admission, sequential organ failure assessment (SOFA) scores were significantly higher in the subsequent sepsis group as compared to the non sepsis group (mean ± SD: 6.4±2.7 and 5±2.3, respectively, P=0.037). sEPCR levels were also higher in the patients who subsequently developed sepsis compared to the patients who did not (median and interquartile range: 173.4 [104.5-223.5] ng/mL vs. 98.3 [69.8-147.7] ng/mL, respectively; P=0.004). Cox regression analysis identified sEPCR as the only parameter related to sepsis development with time (relative risk: 1.078, 95% confidence interval: 1.016-1.144, by 10 units; P=0.013). CONCLUSION: Upon ICU admission, sEPCR levels in initially non-septic critically-ill patients appear elevated in the subjects who will subsequently become septic.

Association of autophagy-related 16-like 1 (ATG16L1) gene polymorphism with sepsis severity in patients with sepsis and ventilator-associated pneumonia.

Autophagy is a highly conserved mechanism of eukaryotic cells implicated in cell homeostasis and elimination of intracellular pathogens. Functional polymorphisms in genes encoding for autophagy have been associated with susceptibility to inflammatory and infectious diseases, but data on severe infections are missing. The aim of the present study was to assess whether polymorphisms in genes encoding proteins involved in autophagy influence susceptibility to ventilator-associated pneumonia (VAP). Mechanically ventilated patients with VAP were studied. Genotyping for autophagy-related 16-like 1 (ATG16L1, rs2241880) functional polymorphism was performed using the TaqMan single-nucleotide assay. Monocytes were isolated from patients and stimulated with lipopolysaccharide (LPS). Tumor necrosis factor-α (TNF-α) was measured in the supernatants of monocytes using an enzyme-linked immunosorbent assay. Procalcitonin (PCT) was also measured in the serum of patients by an immuno-time-resolved amplified cryptate technology assay. A total of 155 patients with VAP were enrolled in the study. Carriage of the minor A allele of ATG16L1 was associated with septic shock with at least one organ failure (odds ratio (OR): 2.40, p: 0.036). TNF-α production was significantly greater among the carriers of the polymorphism presenting with at least one organ failure (p: 0.040). PCT was increased upon worsening to septic shock and organ failure only among carriers of the minor frequency A alleles. In a homogeneous cohort of septic patients with VAP, the carriage of autophagy polymorphisms predisposes to VAP severity and septic shock development. This may be related with predisposition to immunoparalysis.

Intercomparison of three recruitment maneuvers in acute respiratory distress syndrome: the role of Body Mass Index.

BACKGROUND: The aim of this paper was to investigate the effectiveness of three different recruitment maneuvers (RMs) on respiratory and hemodynamic indexes, in patients with acute respiratory distress syndrome (ARDS), and determine the impact of patients' body mass index (BMI) on RMs efficacy. METHODS: This was a prospective clinical trial conducted in a general intensive care unit. In 25 consecutive early ARDS patients arterial blood gases, respiratory system elastance (Est,rs) and hemodynamics were monitored before (T(0)), and at 3, 15, and 30 min (T(3), T(15), T(30), respectively) after each of three different RMs techniques with same airway pressures (45 cmH(2)O), randomly applied in each patient; RM-1: pressure control ventilation for 1 min, RM-2: two hyperinflations with continuous positive airway pressure for 20 sec and 1 min interval in between, RM-3: three consecutive sighs. RESULTS: All RMs improved oxygenation and Est,rs shortly (T(3)) after their application, but only RM-2 had a prolonged beneficial effect on oxygenation (T(30)). Patients were further divided according to their BMI: in the low BMI group (<27.3 Kg/m(2)) the effectiveness of RMs was similar to the whole study population, while in the high BMI group (≥ 27.3 Kg/m(2)) no sustained effect followed any RM. CONCLUSION: In our cohort, recruitment by two sustained inflations resulted in a more persistent improvement of oxygenation as compared with recruitment by pressure controlled ventilation or consecutive sighs with the same airway pressure. BMI seems to have an impact on RMs effectiveness, most probably by altering the effective transpulmonary pressure. More studies are required to elucidate this observation.

Serum of patients with septic shock stimulates the expression of Trem-1 on U937 monocytes.

OBJECTIVES: To describe the concentrations of sTREM-1 in patients with sepsis and to explore the effects of their serum on the expression of TREM-1 on U937 monocytes. METHODS: Blood was sampled at regular time intervals in 56 patients with sepsis. Concentrations of tumour necrosis factor-alpha (TNFalpha), interleukin-1beta (IL-1alpha), IL-6, IL-8, IL-10 and IL-12p70 and sTREM-1 were measured. U937 monocytes were incubated in the presence of serum at sepsis onset. RESULTS: Median sTREM-1 concentration on day 1 for patients with septic shock was 915 pg/ml and 228.5 pg/ml for those without shock (p = 0.002). TNFalpha, IL-1alpha, IL-6, IL-8 and IL-10 did not differ between them. A positive correlation was found between changes of sTREM-1 and SOFA scores from day 1 to 7. Sera of patients with septic shock evoked a significant increase of the expression of TREM-1. The concentrations of TNFalpha and IL-8 in supernatants increased only after stimulating with sera of patients without shock, but not after stimulating with sera of patients with shock. CONCLUSIONS: Levels of sTREM-1 correlated with sepsis severity. sTREM-1 is considerably higher in patients with shock compared to patients without shock. The serum of shocked patients could stimulate the expression of TREM-1 on U937 monocytes.

Pulmonary capillary endothelial metabolic function in chronic thromboembolic pulmonary hypertension.

BACKGROUND: Chronic thromboembolic pulmonary hypertension (CTEPH) causes physical plugging of large pulmonary arteries as well as a distal micro-vasculopathy. Pulmonary endothelium is an active metabolic tissue in normal humans. The effects of CTEPH on pulmonary endothelial metabolism are unknown. OBJECTIVES: We studied pulmonary capillary endothelium-bound angiotensin converting enzyme (ACE) activity as an index of endothelial metabolism in patients with CTEPH. PATIENTS/METHODS: We measured single-pass transpulmonary per cent metabolism (%M) and hydrolysis of an ACE synthetic substrate and calculated functional capillary surface area (FCSA), normalized to body surface area (BSA), in 13 patients with CTEPH and 23 controls. RESULTS: Mean %M for CTEPH (71.6 +/- 4.0% SE) was similar to controls (74.7 +/- 2.7%). Substrate hydrolysis (v) was similar for CTEPH (1.47 +/- 0.22) and controls (1.51 +/- 0.11). However, FCSA/BSA was reduced (P < 0.01) for CTEPH (1530 +/- 218 mL min(-1)*m(-2)) as compared with controls (2948 +/- 245). CONCLUSIONS: The metabolically functional pulmonary capillary bed is reduced in CTEPH. However, because %M and hydrolysis are preserved, this points to a reduction in functional capillary surface area rather than reduced ACE activity on the pulmonary capillary endothelial cell. The reduction in functional capillary surface area may just be a result of decreased capillary recruitment because of upstream vascular plugging by chronic organized thrombus.

Leptin alterations in the course of sepsis in humans.

Neuroendocrine response to sepsis may be divided into acute and prolonged phase. As leptin is implicated in the stress response, leptin's profile during both phases, and the possible relationships between leptin and the neuroendocrine response to sepsis were investigated. Thirty adult patients with sepsis in an intensive care unit were studied. Blood samples were collected at the acute and the prolonged phases. In acute sepsis, leptin levels were higher in patients than in controls (10.2 +/- 2.5 vs. 4.1 +/- 1.2 ng/ml, p =0.01) and correlated positively with insulin levels and insulin resistance. A decline in leptin levels was found during prolonged sepsis (from 10.2 +/- 2.5 to 6.2 +/- 1.7 ng/ml, p=0.001), which was not related to survival (p=0.913). At the onset of sepsis, leptin levels increased in correlation with insulin and insulin resistance, possibly indicating a cause-effect relationship. However, the decline in leptin levels during the prolonged phase of sepsis was not related either to survival or to metabolic and hormonal changes.

Pulmonary endothelium in acute lung injury: from basic science to the critically ill.

BACKGROUND: Pulmonary endothelium is an active organ possessing numerous physiological, immunological, and metabolic functions. These functions may be altered early in acute lung injury (ALI) and further contribute to the development of acute respiratory distress syndrome (ARDS). Pulmonary endothelium is strategically located to filter the entire blood before it enters the systemic circulation; consequently its integrity is essential for the maintenance of adequate homeostasis in both the pulmonary and systemic circulations. Noxious agents that affect pulmonary endothelium induce alterations in hemodynamics and hemofluidity, promote interactions with circulating blood cells, and lead to increased vascular permeability and pulmonary edema formation. OBJECTIVE: We highlight pathogenic mechanisms of pulmonary endothelial injury and their clinical implications in ALI/ARDS patients.

Pulmonary capillary endothelial dysfunction in early systemic sclerosis.

OBJECTIVE: Pulmonary capillary endothelium-bound angiotensin-converting enzyme (PCEB-ACE) activity is a sensitive and quantifiable index of endothelial function in vivo. Systemic sclerosis (SSc) is characterized by endothelial damage and excess collagen formation, causing mainly pulmonary hypertension (PH) in the limited cutaneous SSc (lcSSc) subset and interstitial lung disease with pulmonary interstitial fibrosis (PIF) in the diffuse cutaneous SSc (dcSSc) subset. This study was undertaken to investigate the hypothesis that PCEB-ACE activity is reduced early in SSc, in the absence of PH or PIF. METHODS: Applying indicator-dilution techniques, we measured single-pass transpulmonary hydrolysis and percent metabolism (%M) of a synthetic ACE substrate and calculated functional capillary surface area (FCSA) in 25 SSc patients and 11 controls. Substrate hydrolysis and %M reflect ACE activity per capillary; FCSA reflects ACE activity per vascular bed. RESULTS: PCEB-ACE activity was decreased in both SSc subsets. Among patients without PH, substrate hydrolysis and %M were decreased in patients with lcSSc and more profoundly in those with dcSSc; loss of FCSA normalized to body surface area (FCSA/BSA) was observed in dcSSc, but not in lcSSc. High-resolution computed tomography of the lung, performed in all SSc patients, revealed no correlation between substrate %M, hydrolysis, or FCSA/BSA and the degree of PIF; 5 dcSSc and 5 lcSSc patients with no detectable PIF exhibited decreases in hydrolysis and %M, while FCSA/BSA was decreased only in dcSSc. CONCLUSION: Depression of PCEB-ACE activity, indicating pulmonary endothelial dysfunction, occurs early in SSc, in the absence of PH or PIF, and is more pronounced, at this early pulmonary disease stage, in dcSSc than in lcSSc.

Pulmonary capillary endothelium-bound angiotensin-converting enzyme activity in acute lung injury.

BACKGROUND: Pulmonary capillary endothelium-bound (PCEB) angiotensin-converting ectoenzyme (ACE) activity alteration is an early, sensitive, and quantifiable lung injury index in animal models. We hypothesized that (1) PCEB-ACE alterations can be found in patients with acute lung injury (ALI) and (2) PCEB-ACE activity correlates with the severity of lung injury and may be used as a quantifiable marker of the underlying pulmonary capillary endothelial dysfunction. METHODS AND RESULTS: Applying indicator-dilution techniques, we measured single-pass transpulmonary hydrolysis of the synthetic ACE substrate (3)H-benzoyl-Phe-Ala-Pro (BPAP) in 33 mechanically ventilated, critically ill patients with a lung injury score (LIS) ranging from 0 (no lung injury) to 3.7 (severe lung injury) and calculated the kinetic parameter A(max)/K(m). Both parameters decreased early during the ALI continuum and were inversely related to APACHE II score and LIS. Hydrolysis decreased with increasing cardiac output (CO), whereas 2 different patterns were observed between CO and A(max)/K(m). CONCLUSIONS: PCEB-ACE activity decreases early during ALI, correlates with the clinical severity of both the lung injury and the underlying disease, and may be used as a quantifiable marker of underlying pulmonary capillary endothelial dysfunction.

Reduced lung endothelial angiotensin-converting enzyme activity in Watanabe hyperlipidemic rabbits in vivo.

We investigated pulmonary endothelial function in vivo in 12- to 18-mo-old male Watanabe heritable hyperlipidemic (WHHL; n = 7) and age- and sex-matched New Zealand White (n = 8) rabbits. The animals were anesthetized and artificially ventilated, and the chest was opened and put in total heart bypass. The single-pass transpulmonary utilizations of the angiotensin-converting enzyme (ACE) substrate [(3)H]benzoyl-Phe-Ala-Pro (BPAP) and the 5'-nucleotidase (NCT) substrate [(14)C]AMP were estimated, and the first-order reaction parameter A(max)/K(m), where A(max) is the product of enzyme mass and the catalytic rate constant and K(m) is the Michaelis-Menten constant, was calculated. BPAP transpulmonary utilization and A(max)/K(m) were reduced in WHHL (1.69 +/- 0.16 vs. 2.9 +/- 0.44 and 599 +/- 69 vs. 987 +/- 153 ml/min in WHHL and control rabbits, respectively; P < 0.05 for both). No differences were observed in the AMP parameters. BPAP K(m) and A(max) values were estimated separately under mixed-order reaction conditions. No differences in K(m) values were found (9.79 +/- 1 vs. 9.9 +/- 1.31microM), whereas WHHL rabbit A(max) was significantly decreased (5.29 +/- 0.88 vs. 7. 93 +/- 0.8 micromol/min in WHHL and control rabbits, respectively; P < 0.05). We conclude that the observed pulmonary endothelial ACE activity reduction in WHHL rabbits appears related to a decrease in enzyme mass rather than to alterations in enzyme affinity.

Pulmonary capillary endothelium-bound angiotensin-converting enzyme activity in humans.

BACKGROUND: Pulmonary endothelium has metabolic functions including the conversion of angiotensin I to angiotensin II by angiotensin-converting ectoenzyme (ACE). In this study, we have validated an indicator-dilution technique that provides estimations of dynamically perfused capillary surface area (DPCSA) in humans, and we have characterized pulmonary endothelial ACE in vivo. METHODS AND RESULTS: In 12 adults, single-pass transpulmonary (one or both lungs) hydrolysis of the specific ACE substrate 3H-benzoyl-Phe-Ala-Pro (3H-BPAP) was measured and expressed as % metabolism (%M) and v=-ln(1-M). We also calculated Amax/Km, an index of DPCSA. %M (70.1+/-3.2 vs 67.9+/-3.1) and v (1.29+/-0.14 vs 1. 20+/-0.12) were similar in both lungs and the right lung, respectively, whereas Amax/Km//body surface area decreased from 2460+/-193 to 1318+/-115 mL/min per square meter. CONCLUSIONS: Pulmonary endothelial ACE activity can be assessed in humans at the bedside by means of indicator-dilution techniques. Our data suggest homogeneous pulmonary capillary ACE concentrations and capillary transit times (tc) in both human lungs, and similar tc within the normal range of cardiac index. Amax/Km in the right lung is 54% of total Amax/Km in both lungs, suggesting that Amax/Km is a reliable and quantifiable index of DPCSA in humans.

Vascular recruitment increases evidence of lung injury.

OBJECTIVE: Changes in pulmonary blood flow rate can alter the size of the perfused pulmonary capillary surface area. We tested the hypothesis that full recruitment of the pulmonary vascular bed may decrease evidence of lung injury by recruiting less injured capillaries. We also tested the hypothesis that endothelial ectoenzyme activity is an earlier indicator of lung injury than are permeability measures. DESIGN: Isolated canine lung lobes were perfused with autologous blood at constant blood flows of either 2.05+/-0.04 L/min (SEM) (high flow, full recruitment, n = 12) or 0.600 +/- 0.004 L/min (low flow, 33% full recruitment, n = 12) after lung injury to determine the effect of vascular recruitment on measures of injury. SETTING: Research laboratory at a medical university. SUBJECTS: Lung lobes were obtained from 36 mongrel dogs of either gender. INTERVENTIONS: Lung injury was induced by adding phorbol myristate acetate (PMA) to the blood perfusing the isolated lung. MEASUREMENTS AND MAIN RESULTS: Indicator dilution methods were used to measure single pass hydrolysis of 3[H]-benzoyl-Phe-Ala-Pro, a synthetic substrate for angiotensin converting enzyme, and calculate the modified first order kinetic parameter corresponding to the ratio of a normalized maximal enzymatic conversion rate (A(max)) to the Michaelis-Menten constant (K(m)), i.e., A(max)/K(m), before and after PMA. At a given flow rate, the decrease in A(max)/K(m)serves as an index of vascular injury. PMA decreased A(max)/K(m), percent metabolism, and fractional substrate utilization, and increased permeability, vascular resistance, and vascular pressures regardless of flow rate. The decrease in enzyme activity was detected earlier than the increase in permeability. CONCLUSION: The greater percentage decrease in percent metabolism and fractional substrate utilization and the earlier appearance of increased permeability during high flow indicates that increasing blood flow three-fold recruited injured vessels and/or increased vascular injury by increasing vascular perfusion pressures.

Endothelial ectoenzyme assays estimate perfused capillary surface area in the dog lung.

Whether the pulmonary vascular bed accommodates flow-induced increases in blood volume mainly through recruitment of previously unperfused capillaries or distension of already perfused vessels remains controversial. The modified first order reaction parameter of an enzyme and substrate, Amax/K(m), is, under nontoxic conditions, proportional to enzyme mass. Thus for ACE, an endothelium-bound ectoenzyme uniformly distributed along the luminal surface of the pulmonary capillary bed, Amax/K(m) is proportional to the dynamically perfused capillary surface area (PCSA). We estimated single-pass translobar hydrolysis and calculated the corresponding Amax/K(m) values of the synthetic ACE substrate 3H-benzoyl-Phe-Ala-Pro (BPAP), under first-order reaction conditions, in isolated blood-perfused dog lung lobes. We additionally studied blood flow distribution using radioactive microsphere techniques. Experiments were performed under zone III conditions over a wide range of lobar blood flow rates (Qb). As Qb was increased, Amax/K(m) rose linearly, while lobar vascular resistance (LVR) decreased, suggesting capillary recruitment rather than distension. Single pass BPAP hydrolysis (v approximately 2.9 at resting Qb) was not altered over a wide range of Qb, indicative of unchanging capillary transit times. When full capillary recruitment was achieved (at Qb > 70 ml/min/g lung wet weight), further Qb elevations failed to increase Amax/K(m), but decreased BPAP hydrolysis, denoting shorter transit times through the fully recruited capillary bed. Our data indicate that, as previously shown for rabbit lung, in this canine model, increases in pulmonary blood volume are mainly accommodated through recruitment of previously unperfused capillaries throughout the entire lung.