Comparison of Coagulation Parameters in Arterial and Venous Blood in Cardiac Surgery Measured Using the Quantra System.

OBJECTIVE: Perioperative coagulation testing often is performed with arterial samples even though device reference ranges typically are established in venous samples. Although limited studies exist comparing coagulation parameters across sampling sites, viscoelastic testing devices have demonstrated some differences. The objective of this study was to compare coagulation parameters determined using the Quantra System for venous and arterial samples. DESIGN: Prospective, observational study. SETTING: Tertiary care university hospital. PARTICIPANTS: The study comprised 30 adult patients undergoing cardiac surgery. INTERVENTIONS: Paired arterial and venous samples were obtained at 2 of the following time points: baseline, during bypass, or after protamine reversal of heparin. Quantra measurements included Clot Time (CT), Heparinase Clot Time (CTH), Clot Time Ratio (CTR), Clot Stiffness (CS), and Fibrinogen (FCS) and Platelet (PCS) Contributions to clot stiffness. MEASUREMENTS AND MAIN RESULTS: The relationship and agreement between matched data pairs were established and statistical analysis was performed via paired t tests. CTR, CS, FCS, and PCS were unaffected by the sampling site, whereas CT and CTH demonstrated statistically significant differences between arterial and venous samples (p < 0.001). Arterial clot times were prolonged relative to the venous ones with a mean percent error of 14.2 % and 11.9 %, respectively. These results are in general agreement with those reported for other viscoelastic testing devices. CONCLUSIONS: This study demonstrates that Quantra clot stiffness-based parameters (CS, FCS, PCS) are unaffected by sampling site, whereas the clot time parameters (CT and CTH) show good correlation in the presence of a bias. CTR, a ratio of CT and CTH, also is unaffected.

Comparison of SEER Sonorheometry With Rotational Thromboelastometry and Laboratory Parameters in Cardiac Surgery.

BACKGROUND: The Quantra Hemostasis Analyzer is a novel diagnostic device that uses an ultrasound-based technology, sonic estimation of elasticity via resonance (SEER) sonorheometry, to characterize the dynamic changes in viscoelastic properties of a blood sample during coagulation. Cardiac surgery utilizing cardiopulmonary bypass (CPB) is associated with a significant impact on the coagulation system and can result in perioperative coagulopathy. The aim of this study was to correlate SEER sonorheometry results to corresponding rotational thromboelastometry (ROTEM) and laboratory parameters obtained before, during, and after CPB. METHODS: The Quantra uses a multiwell cartridge that performs 4 independent measurements with different combination of reagents. The output test results include Clot Time, Clot Stiffness, Fibrinogen and Platelet Contribution, Clot Time Ratio, and Heparinase Clot Time. Clot Time was compared with ROTEM INTEM clotting time and the adjusted partial thromboplastin time. Clot Stiffness was compared with ROTEM EXTEM. The Fibrinogen Contribution to the Clot Stiffness was correlated to ROTEM FIBTEM as well as fibrinogen concentration by the Clauss method. The Platelet Contribution to Clot Stiffness was compared with absolute platelet count and ROTEM-determined clot elasticity attributable to platelets. RESULTS: Fifty-five patients undergoing elective cardiac surgery were enrolled in this prospective observational study. Clot Time exhibited good correlation with ROTEM INTEM clotting time (pre-CPB r = 0.84, post-CPB r = 0.65) and adjusted partial thromboplastin time (pre-CPB r = 0.72, post-CPB r = 0.89); however, the majority of values were within a narrow normal range. Clot Stiffness exhibited significant correlation with ROTEM EXTEM A10 throughout the course of the study in all samples (r = 0.84). Fibrinogen Contribution correlated strongly with FIBTEM A10 (r = 0.85), and moderately with the fibrinogen concentration (r = 0.73) determined with the Clauss assay. The Platelet Contribution to Clot Stiffness showed moderate correlation to absolute platelet counts (r = 0.48). However, the correlation between Platelet Contribution and ROTEM-determined clot elasticity attributable to platelets was stronger (r = 0.78) than platelet number. All of the correlation coefficients were statistically significant with P < .001. CONCLUSIONS: SEER sonorheometry demonstrates significant correlation with ROTEM for determining Clot Stiffness and assessing Fibrinogen Contribution. SEER sonorheometry results can provide valuable information about the coagulation status in patients undergoing cardiac surgery using CPB.

Influence of catastrophizing, anxiety, and depression on in-hospital opioid consumption, pain, and quality of recovery after adult spine surgery.

OBJECTIVE Perception of perioperative pain is influenced by various psychological factors. The aim of this study was to determine the impact of catastrophizing, anxiety, and depression on in-hospital opioid consumption, pain scores, and quality of recovery in adults who underwent spine surgery. METHODS Patients undergoing spine surgery were enrolled in this study, and the preoperatively completed questionnaires included the verbal rating scale (VRS), Pain Catastrophizing Scale (PCS), Hospital Anxiety and Depression Scale (HADS), and Oswestry Disability Index (ODI). Quality of recovery was assessed using the 40-item Quality of Recovery questionnaire (QoR40). Opioid consumption and pain scores according to the VRS were recorded daily until discharge. RESULTS One hundred thirty-nine patients were recruited for the study, and 101 completed the QoR40 assessment postoperatively. Patients with higher catastrophizing scores were more likely to have higher maximum pain scores postoperatively (estimate: 0.03, SE: 0.01, p = 0.02), without increased opioid use (estimate: 0.44, SE: 0.27, p = 0.11). Preoperative anxiety (estimate: 1.18, SE: 0.65, p = 0.07) and depression scores (estimate: 1.06, SE: 0.71, p = 0.14) did not correlate with increased postoperative opioid use; however, patients with higher preoperative depression scores had lower quality of recovery after surgery (estimate: -1.9, SE: 0.56, p < 0.001). CONCLUSIONS Catastrophizing, anxiety, and depression play important roles in modulating postoperative pain. Preoperative evaluation of these factors, utilizing a validated tool, helps to identify patients at risk. This might allow for earlier psychological intervention that could reduce pain severity and improve the quality of recovery.

Vascular growth and remodeling in compensatory lung growth following right lobectomy.

Studies in animal models have shown that, following lobectomy (LBX), there is compensatory growth in the remaining lung. The vascular growth response following right LBX (R-LBX) is poorly understood. To test the hypothesis that arterial growth and remodeling occur in response to LBX, in proportion to the amount of right lung tissue removed, two (24% of lung mass; R-LBX2 group) or three right lobes (52% of lung mass; R-LBX3 group) were removed via thoracotomy from adult rats. Sham control animals underwent thoracotomy only. Arteriograms were generated 3 wk after surgery. The areas of the left lung arteriogram, arterial branching, length of arterial branches, arterial density, and arterial-to-alveolar ratios were measured. To determine whether R-LBX causes vascular remodeling and pulmonary hypertension, muscularization of arterioles and right ventricular hypertrophy were assessed. Lung weight and volume indexes were greater in R-LBX3. Arterial area of the left lung increased 26% in R-LBX2 and 47% in R-LBX3. The length of large arteries increased in R-LBX3 and to a lesser extent in R-LBX2. The ratio of distal pulmonary arteries to alveoli was similar after R-LBX2 compared with sham but was 30% lower in R-LBX3. Muscularization of arterioles increased after R-LBX3, but not in R-LBX2. Right ventricular hypertrophy increased 50-70% in R-LBX3, but not in R-LBX2. Whereas removal of three right lung lobes induced arterial growth in the left lungs of adult rats, which was proportionate to the number of lobes removed, the ratio of distal pulmonary arteries to alveoli was not normal, and vascular remodeling and pulmonary hypertension developed.

Adenosine A1 receptor activation attenuates lung ischemia-reperfusion injury.

OBJECTIVES: Ischemia-reperfusion injury contributes significantly to morbidity and mortality in lung transplant patients. Currently, no therapeutic agents are clinically available to prevent ischemia-reperfusion injury, and treatment strategies are limited to maintaining oxygenation and lung function. Adenosine can modulate inflammatory activity and injury by binding to various adenosine receptors; however, the role of the adenosine A1 receptor in ischemia-reperfusion injury and inflammation is not well understood. The present study tested the hypothesis that selective, exogenous activation of the A1 receptor would be anti-inflammatory and attenuate lung ischemia-reperfusion injury. METHODS: Wild-type and A1 receptor knockout mice underwent 1 hour of left lung ischemia and 2 hours of reperfusion using an in vivo hilar clamp model. An A1 receptor agonist, 2-chloro-N6-cyclopentyladenosine, was administered 5 minutes before ischemia. After reperfusion, lung function was evaluated by measuring airway resistance, pulmonary compliance, and pulmonary artery pressure. The wet/dry weight ratio was used to assess edema. The myeloperoxidase and cytokine levels in bronchoalveolar lavage fluid were measured to determine the presence of neutrophil infiltration and inflammation. RESULTS: In the wild-type mice, 2-chloro-N6-cyclopentyladenosine significantly improved lung function and attenuated edema, cytokine expression, and myeloperoxidase levels compared with the vehicle-treated mice after ischemia-reperfusion. The incidence of lung ischemia-reperfusion injury was similar in the A1 receptor knockout and wild-type mice; and 2-chloro-N6-cyclopentyladenosine had no effects in the A1 receptor knockout mice. In vitro treatment of neutrophils with 2-chloro-N6-cyclopentyladenosine significantly reduced chemotaxis. CONCLUSIONS: Exogenous A1 receptor activation improves lung function and decreases inflammation, edema, and neutrophil chemotaxis after ischemia and reperfusion. These results suggest a potential therapeutic application for A1 receptor agonists for the prevention of lung ischemia-reperfusion injury after transplantation.

Rapamycin blocks fibrocyte migration and attenuates bronchiolitis obliterans in a murine model.

BACKGROUND: Fibrocytes are integral in the development of fibroproliferative disease. The CXCL12/CXCR4 chemokine axis has been shown to play a central role in fibrocyte migration and the development of bronchiolitis obliterans (BO) after lung transplantation. Inhibition of the mammalian target of rapamycin (mTOR) pathway with rapamycin has been shown to decrease expression of both CXCR4 and its receptor agonist CXCL12. Thus, we hypothesized that rapamycin treatment would decrease fibrocyte trafficking into tracheal allografts and prevent BO. METHODS: A total alloantigenic mismatch murine heterotopic tracheal transplant (HTT) model of BO was used. Animals were either treated with rapamycin or dimethyl sulfoxide (DMSO) for 14 days after tracheal transplantation. Fibrocyte levels were assessed by flow cytometry, and allograft neutrophil, CD3(+) T-cell, macrophage, and smooth muscle actin (SMA) levels were assessed by immunohistochemistry. Tracheal luminal obliteration was assessed on hematoxylin and eosin (H&E) stains. RESULTS: Compared with DMSO-treated controls, rapamycin-treated mice showed a significant decrease in fibrocyte levels in tracheal allografts. Fibrocyte levels in recipient blood showed a similar pattern, although it was not statistically significant. Furthermore, animals treated with rapamycin showed a significant decrease in tracheal allograft luminal obliteration compared with controls. Based on immunohistochemical analyses, populations of α-SMA-positive (α-SMA(+)) cells, neutrophils, CD3(+) T cells, and macrophages were all decreased in rapamycin-treated allografts versus DMSO controls. CONCLUSIONS: Rapamycin effectively reduces recruitment of fibrocytes into tracheal allografts and mitigates development of tracheal luminal fibrosis. Further studies are needed to determine the cellular and molecular mechanisms that mediate the protective effect of rapamycin against BO.

Adenosine A3 receptor activation attenuates lung ischemia-reperfusion injury.

BACKGROUND: Severe ischemia-reperfusion (IR) injury leads to primary graft dysfunction after lung transplantation. Adenosine receptors modulate inflammation after IR, and the adenosine A3 receptor (A3R) is expressed in lung tissue and inflammatory cells. This study tests the hypothesis that A3R agonism attenuates lung IR injury by a neutrophil-dependent mechanism. METHODS: Wild-type and A3R knockout (A3R-/-) mice underwent 1-hour left lung ischemia followed by 2-hours reperfusion (IR). A selective A3R agonist, Cl-IB-MECA, was administered (100 µg/kg intravenously) 5 minutes prior to ischemia. Study groups included sham, IR, and IR+Cl-IB-MECA (n = 6/group). Lung injury was assessed by measuring lung function, pulmonary edema, histopathology, and proinflammatory cytokines, and myeloperoxidase levels in bronchoalveolar lavage fluid. Parallel in vitro experiments were performed to evaluate neutrophil chemotaxis, and neutrophil activation was measured after exposure to acute hypoxia and reoxygenation. RESULTS: Treatment of wild-type mice with Cl-IB-MECA significantly improved lung function and decreased edema, cytokine expression, and neutrophil infiltration after IR. The Cl-IB-MECA had no effects in A3R-/- mice; Cl-IB-MECA significantly decreased activation of wild-type, but not A3R-/-, neutrophils after acute hypoxia and reoxygenation and inhibited chemotaxis of wild-type neutrophils. CONCLUSIONS: Exogenous activation of A3R by Cl-IB-MECA attenuates lung dysfunction, inflammation, and neutrophil infiltration after IR in wild-type but not A3R-/- mice. Results with isolated neutrophils suggest that the protective effects of Cl-IB-MECA are due, in part, to the prevention of neutrophil activation and chemotaxis. The use of A3R agonists may be a novel therapeutic strategy to prevent lung IR injury and primary graft dysfunction after transplantation.

Tissue-derived proinflammatory effect of adenosine A2B receptor in lung ischemia-reperfusion injury.

OBJECTIVE: Ischemia-reperfusion injury after lung transplantation remains a major source of morbidity and mortality. Adenosine receptors have been implicated in both pro- and anti-inflammatory roles in ischemia-reperfusion injury. This study tests the hypothesis that the adenosine A(2B) receptor exacerbates the proinflammatory response to lung ischemia-reperfusion injury. METHODS: An in vivo left lung hilar clamp model of ischemia-reperfusion was used in wild-type C57BL6 and adenosine A(2B) receptor knockout mice, and in chimeras created by bone marrow transplantation between wild-type and adenosine A(2B) receptor knockout mice. Mice underwent sham surgery or lung ischemia-reperfusion (1 hour ischemia and 2 hours reperfusion). At the end of reperfusion, lung function was assessed using an isolated buffer-perfused lung system. Lung inflammation was assessed by measuring proinflammatory cytokine levels in bronchoalveolar lavage fluid, and neutrophil infiltration was assessed via myeloperoxidase levels in lung tissue. RESULTS: Compared with wild-type mice, lungs of adenosine A(2B) receptor knockout mice were significantly protected after ischemia-reperfusion, as evidenced by significantly reduced pulmonary artery pressure, increased lung compliance, decreased myeloperoxidase, and reduced proinflammatory cytokine levels (tumor necrosis factor-α; interleukin-6; keratinocyte chemoattractant; regulated on activation, normal T-cell expressed and secreted; and monocyte chemotactic protein-1). Adenosine A(2B) receptor knockout → adenosine A(2B) receptor knockout (donor → recipient) and wild-type → adenosine A(2B) receptor knockout, but not adenosine A(2B) receptor knockout → wild-type, chimeras showed significantly improved lung function after ischemia-reperfusion. CONCLUSIONS: These results suggest that the adenosine A(2B) receptor plays an important role in mediating lung inflammation after ischemia-reperfusion by stimulating cytokine production and neutrophil chemotaxis. The proinflammatory effects of adenosine A(2B) receptor seem to be derived by adenosine A(2B) receptor activation primarily on resident pulmonary cells and not bone marrow-derived cells. Adenosine A(2B) receptor may provide a therapeutic target for prevention of ischemia-reperfusion-related graft dysfunction in lung transplant recipients.

Inflammatory lung injury after cardiopulmonary bypass is attenuated by adenosine A(2A) receptor activation.

OBJECTIVE: Cardiopulmonary bypass has been shown to exert an inflammatory response within the lung, often resulting in postoperative pulmonary dysfunction. Several studies have shown that adenosine A(2A) receptor activation attenuates lung ischemia-reperfusion injury; however, the effect of adenosine A(2A) receptor activation on cardiopulmonary bypass-induced lung injury has not been studied. We hypothesized that specific adenosine A(2A) receptor activation by ATL313 would attenuate inflammatory lung injury after cardiopulmonary bypass. METHODS: Adult male Sprague-Dawley rats were randomly divided into 3 groups: 1) SHAM group (underwent cannulation + heparinization only); 2) CONTROL group (underwent 90 minutes of normothermic cardiopulmonary bypass with normal whole-blood priming solution; and 3) ATL group (underwent 90 minutes of normothermic cardiopulmonary bypass with ATL313 added to the normal priming solution). RESULTS: There was significantly less pulmonary edema and lung injury in the ATL group compared with the CONTROL group. The ATL group had significant reductions in bronchoalveolar lavage interleukin-1, interleukin-6, interferon-gamma, and myeloperoxidase levels compared with the CONTROL group. Similarly, lung tissue interleukin-6, tumor necrosis factor-alpha, and interferon-gamma were significantly decreased in the ATL group compared with the CONTROL group. There was no significant difference between the SHAM and ATL groups in the amount of pulmonary edema, lung injury, or levels of proinflammatory cytokines. CONCLUSION: The addition of a potent adenosine A(2A) receptor agonist to the normal priming solution before the initiation of cardiopulmonary bypass significantly protects the lung from the inflammatory effects of cardiopulmonary bypass and reduces the amount of lung injury. Adenosine A(2A) receptor agonists could represent a new therapeutic strategy for reducing the potentially devastating consequences of the inflammatory response associated with cardiopulmonary bypass.

Proinflammatory response of alveolar epithelial cells is enhanced by alveolar macrophage-produced TNF-alpha during pulmonary ischemia-reperfusion injury.

Pulmonary ischemia-reperfusion (IR) injury entails acute activation of alveolar macrophages followed by neutrophil sequestration. Although proinflammatory cytokines and chemokines such as TNF-alpha and monocyte chemoattractant protein-1 (MCP-1) from macrophages are known to modulate acute IR injury, the contribution of alveolar epithelial cells to IR injury and their intercellular interactions with other cell types such as alveolar macrophages and neutrophils remain unclear. In this study, we tested the hypothesis that following IR, alveolar macrophage-produced TNF-alpha further induces alveolar epithelial cells to produce key chemokines that could then contribute to subsequent lung injury through the recruitment of neutrophils. Cultured RAW264.7 macrophages and MLE-12 alveolar epithelial cells were subjected to acute hypoxia-reoxygenation (H/R) as an in vitro model of pulmonary IR. H/R (3 h/1 h) significantly induced KC, MCP-1, macrophage inflammatory protein-2 (MIP-2), RANTES, and IL-6 (but not TNF-alpha) by MLE-12 cells, whereas H/R induced TNF-alpha, MCP-1, RANTES, MIP-1alpha, and MIP-2 (but not KC) by RAW264.7 cells. These results were confirmed using primary murine alveolar macrophages and primary alveolar type II cells. Importantly, using macrophage and epithelial coculture methods, the specific production of TNF-alpha by H/R-exposed RAW264.7 cells significantly induced proinflammatory cytokine/chemokine expression (KC, MCP-1, MIP-2, RANTES, and IL-6) by MLE-12 cells. Collectively, these results demonstrate that alveolar type II cells, in conjunction with alveolar macrophage-produced TNF-alpha, contribute to the initiation of acute pulmonary IR injury via a proinflammatory cascade. The release of key chemokines, such as KC and MIP-2, by activated type II cells may thus significantly contribute to neutrophil sequestration during IR injury.

Reinitiation of compensatory lung growth after subsequent lung resection.

OBJECTIVE: In experimental animals, pneumonectomy results in rapid, hyperplastic compensatory growth of the remaining lung. The limits of this induced growth are unknown. We tested the hypothesis that compensatory growth can be reinitiated in the same lung after subsequent lung resection. METHODS: A left thoracotomy (Sham group) or left pneumonectomy (PNX group) was performed in Sprague-Dawley rats. A third group underwent left pneumonectomy followed 4 weeks later by a bilobectomy of the right upper and middle lobes (PNX+LBX group). Four weeks after bilobectomy in the PNX+LBX group (8 weeks in the Sham and PNX groups), right ventricular pressures were measured by using the open chest technique, and total lung weight and lower plus cardiac lobe weight indices were measured. Lungs were inflation fixed at 25 cm H2O to measure lobe volume index and to perform morphometric measurements on lung sections. Right ventricle/left ventricle plus septum weight index was measured as another index of pulmonary hypertension. RESULTS: Total lung weight index was similar in all groups. Pneumonectomy resulted in increased lower plus cardiac lobe weight and volume indices, which were significantly augmented in the PNX+LBX group. The PNX+LBX group underwent a significant increase in total volume of respiratory region, airspace, and tissue and a decrease in alveolar surface density versus the PNX group. The PNX+LBX group also had significantly increased right ventricular systolic pressure and right ventricle/left ventricle plus septum index. CONCLUSION: These results demonstrate that compensatory growth can be reinitiated in lungs that had previously undergone postpneumonectomy compensatory growth. This subsequent growth, however, is more hypertrophic, and pulmonary hypertension develops despite subsequent compensatory growth.

Differential vascular growth in postpneumonectomy compensatory lung growth.

OBJECTIVE: After pneumonectomy, compensatory growth occurs in the remaining lung. The vascular response during this growth and how individual lobes of the lung respond are not well understood. The aim of our study was to characterize vascular growth among individual lobes of the lung after pneumonectomy and determine whether changes in relative blood flow correlate with growth. METHODS: Rats underwent left pneumonectomy, and lobe weights and volumes of the right lung were measured 21 days later. Arterial growth was quantitated from arteriograms of each lobe after barium perfusion. Changes in relative blood flow were assessed by using radiolabeled microspheres. Expression of proliferating cell nuclear antigen was measured by means of Western blot analysis. RESULTS: After pneumonectomy, weight and volume indices of all lobes were significantly increased compared with those seen in sham control animals. Arterial growth occurred in all lobes after pneumonectomy, with the greatest increases occurring in the upper and middle lobes. In addition, a differential distribution of blood flow was observed where the upper and middle lobes contained the highest degree of relative flow. Pneumonectomy produced hyperplasic growth in all lobes, as indicated by significantly increased proliferating cell nuclear antigen expression. Proliferating cell nuclear antigen expression correlated with arterial growth in that increased and prolonged expression occurred in the upper lobe. CONCLUSIONS: These results show that left pneumonectomy induces significant, nonuniform, compensatory growth in all lobes of the right lung. Arterial growth occurred in each lobe after pneumonectomy, but preferentially higher vascular growth and cell proliferation in the upper lobe positively correlated with higher relative blood flow in this lobe.

Pulmonary macrophage inhibition and inhaled nitric oxide attenuate lung ischemia-reperfusion injury.

BACKGROUND: Lung ischemia-reperfusion injury (LIRI) is postulated to occur biphasically. Donor pulmonary macrophages mediate early injury, and neutrophil-dependent injury predominates in the later phase of LIRI. We hypothesized that the biphasic response to LIRI would be attenuated by the administration of gadolinium, a known pulmonary macrophage inhibitor, and inhaled nitric oxide (NO), a pulmonary vasodilator that also interferes with neutrophil chemotaxis. METHODS: Using our isolated, ventilated, blood-perfused rabbit lung model, study groups (n = 10 per group) underwent two hours of reperfusion after 18 hours of cold ischemia (4 degrees C). Lungs received gadolinium alone, or inhaled NO in the presence or absence of macrophage inhibition with gadolinium. RESULTS: Compared with control animals, pulmonary macrophage inhibition with the concurrent administration of inhaled NO increased lung compliance (p < 0.01) and oxygenation (p = 0.03), while also decreasing pulmonary artery pressure (p < 0.01), myeloperoxidase content by 63% (p < 0.01), wet to dry ratios by 23% (p < 0.01), and lung tissue (p < 0.01) and bronchoalveolar lavage tumor necrosis factor-alpha (TNF-alpha) protein levels (p < 0.01). CONCLUSIONS: The severity of LIRI was most significantly reduced by the inhibition of pulmonary macrophages and the concomitant use of inhaled NO. Pulmonary macrophages, likely through the elaboration of proinflammatory cytokines such as TNF-alpha, not only cause early injury themselves but also prime cells such as neutrophils to injure lungs in the later stages of LIRI. The LIRI was effectively blunted by the reduction of macrophage-dependent injury by gadolinium while inhaled NO also attenuated injury by reducing pulmonary hypertension and minimizing neutrophil sequestration.

Endothelial nitric oxide synthase is essential for postpneumonectomy compensatory vasodilation.

BACKGROUND: After pneumonectomy, the remaining lung vasculature must vasodilate to compensate for increased blood volume. We hypothesized that endothelial nitric oxide synthase (eNOS) is essential for compensatory vasodilation after pneumonectomy. METHODS: Adult, wild-type C57BL6 (WT) and eNOS knockout (eNOS-/-) mice underwent left pneumonectomy and recovered under normoxic conditions. Animals were lightly anesthetized at 1, 3, 7, or 14 days after pneumonectomy, and closed chest, systolic right ventricular pressure (RVP) was recorded using fine-needle cannulation. The right ventricle to left ventricle plus septum weight ratios were measured as an index of right ventricular hypertrophy. Two additional groups of mice (WT and eNOS-/-) were recovered after pneumonectomy in inhaled nitric oxide (iNO, 10 ppm), and RVP was measured on day 7. RESULTS: The eNOS-/- mice had significantly higher preoperative RVP than did WT (17.1 +/- 0.4 versus 14.2 +/- 0.2 cmH2O, p = 0.001). Both groups exhibited transient periods of pulmonary hypertension after pneumonectomy. On day 1, RVP was 80% above baseline in eNOS-/- mice (30.7 +/- 0.8 cmH2O) versus 42% in WT mice (20.2 +/- 0.7 cmH2O, p = 0.0001). The RVP returned to baseline in WT mice (16.3 +/- 0.2 cmH2O) but remained significantly elevated in eNOS-/- mice (28.6 +/- 0.9 cmH2O) at day 3 and at each time thereafter (p = 0.0001). The iNO significantly reduced RVP in eNOS-/- animals to 15.2 +/- 0.3 cmH2O (p = 0.0001) while having no effect in WT animals. Right ventricular hypertrophy was not observed in any group. CONCLUSIONS: Pneumonectomy results in a transient increase in RVP. Under normal circumstances, these pressures return to baseline within 3 days. The eNOS-/- mice failed to display compensatory vasodilation yet could be rescued with iNO. These results suggest that eNOS is essential for postpneumonectomy compensatory vasodilation.

Transdifferentiation of ciliated cells during repair of the respiratory epithelium.

Since the lung is repeatedly subjected to injury by pathogens and toxicants, maintenance of pulmonary homeostasis requires rapid repair of its epithelial surfaces. Ciliated bronchiolar epithelial cells, previously considered as terminally differentiated, underwent squamous cell metaplasia within hours after bronchiolar injury with naphthalene. Expression of transcription factors active in morphogenesis and differentiation of the embryonic lung, including beta-catenin, Foxa2, Foxj1, and Sox family members (Sox17 and Sox2), was dynamically regulated during repair and redifferentiation of the bronchiolar epithelium after naphthalene injury. Squamous cells derived from ciliated cells spread beneath injured Clara cells within 6-12 h after injury, maintaining the integrity of the epithelium. Dynamic changes in cell shape and gene expression, indicating cell plasticity, accompanied the transition from squamous to cuboidal to columnar cell types as differentiation-specific cell markers typical of the mature airway were restored. Similar dynamic changes in the expression of these transcription factors occurred in ciliated and Clara cells during regeneration of the lung after unilateral pneumonectomy. Taken together, these findings demonstrate that ciliated epithelial cells spread and transdifferentiate into distinct epithelial cell types to repair the airway epithelium.

Alveolar macrophage activation is a key initiation signal for acute lung ischemia-reperfusion injury.

Lung ischemia-reperfusion (I/R) injury is a biphasic inflammatory process. Previous studies indicate that the later phase is neutrophil-dependent and that alveolar macrophages (AMs) likely contribute to the acute phase of lung I/R injury. However, the mechanism is unclear. AMs become activated and produce various cytokines and chemokines in many inflammatory responses, including transplantation. We hypothesize that AMs respond to I/R by producing key cytokines and chemokines and that depletion of AMs would reduce cytokine/chemokine expression and lung injury after I/R. To test this, using a buffer-perfused, isolated mouse lung model, we studied the impact of AM depletion by liposome-clodronate on I/R-induced lung dysfunction/injury and expression of cytokines/chemokines. I/R caused a significant increase in pulmonary artery pressure, wet-to-dry weight ratio, vascular permeability, tumor necrosis factor (TNF)-alpha, monocyte chemoattractant protein (MCP)-1, and macrophage inflammatory protein (MIP)-2 expression, as well as decreased pulmonary compliance, when compared with sham lungs. After AM depletion, the changes in each of these parameters between I/R and sham groups were significantly attenuated. Thus AM depletion protects the lungs from I/R-induced dysfunction and injury and significantly reduces cytokine/chemokine production. Protein expression of TNF-alpha and MCP-1 are positively correlated to I/R-induced lung injury, and AMs are a major producer/initiator of TNF-alpha, MCP-1, and MIP-2. We conclude that AMs are an essential player in the initiation of acute lung I/R injury.

Upregulation of hypoxia-induced mitogenic factor in compensatory lung growth after pneumonectomy.

After pneumonectomy, the remaining lung increases in size. This process is referred to as compensatory lung growth. Various pathways likely play important roles in this growth response. The molecular mechanisms involved in compensatory lung growth, however, remain poorly understood. Hypoxia-induced mitogenic factor (HIMF), also called FIZZ1 or RELM-alpha, possesses mitogenic, vasoconstrictive, angiogenic, and antiapoptotic effects. In this study, we examined the expression of HIMF in mouse lung after pneumonectomy to test the hypothesis that HIMF expression is upregulated during compensatory lung growth. Results showed that HIMF is upregulated from Day 1 after pneumonectomy and peaking at Day 7 in the lung. HIMF upregulation is temporally and spatially related to lung cell proliferation, as demonstrated by expression of proliferating cell nuclear antigen. Immunohistochemical staining and in situ hybridization showed that upregulated HIMF protein and mRNA are mainly distributed in airway epithelium, alveolar type II cells, and endothelial cells of the pulmonary vessels. Intratracheal instillation of recombinant HIMF resulted in widespread cell proliferation, including airway epithelium, alveolar type II cells, and cells in the alveolar septa. These results indicate a new role for HIMF in compensatory lung growth, which is that HIMF may act as a lung-specific growth factor and participate in lung regeneration after pneumonectomy.