Ninety-Day Mortality: Redefining the Perioperative Period After Lung Resection.

Operative mortality is an important outcome for patients, surgeons, healthcare institutions, and policy makers. Although measures of perioperative mortality have conventionally been limited to in-hospital and 30-day mortality (or a composite endpoint combining both), there is a large body of evidence emerging to support the extension of the perioperative period after lung resection to a minimum of 90 days after surgery. Several large-volume studies from centers across the world have reported that 90-day mortality after lung resection is double 30-day mortality. Hence, true perioperative mortality after lung resection is likely to be significantly higher than what is currently reported. In the contemporary era, where new treatment modalities such as stereotactic ablative body radiotherapy are emerging as viable nonsurgical alternatives for the treatment of lung cancer, accurate estimation of perioperative risk and reliable reporting of perioperative mortality are of particular importance. It is likely that shifting the discussion from 30-day to 90-day mortality will lead to altered decision making, particularly for specific patient subgroups at an increased risk of 90-day mortality. We believe that 90-day mortality should be adopted as the standard measure of perioperative mortality after lung resection and that strategies to reduce the risk of mortality within 90 days of surgery should be investigated.

Evaluation of a Powered Vascular Stapler in Video-Assisted Thoracic Surgery Lobectomy.

BACKGROUND: A narrow-profile powered vascular stapler (PVS) was developed to provide superior access and precise staple placement in thoracic procedures. The objective of this study was to determine if the PVS would yield an equivalent rate of hemostatic interventions compared with standard of care (SOC) staplers in video-assisted thoracoscopic surgery lobectomy. MATERIALS AND METHODS: A randomized, controlled, multicenter study was conducted comparing PVS with SOC staplers in lobectomies performed for non-small cell lung cancer. The primary performance endpoint was the incidence of intraoperative hemostatic interventions, and the primary safety endpoint was the frequency of postoperative bleeding-related interventions. RESULTS: A total of 98 subjects participated in the SOC group and 103 in the PVS group. Rates of intraoperative hemostatic interventions were 5.3% and 8.3% for the SOC and PVS groups, respectively. These rates were not statistically different (P = 0.137), although the upper bound of the 95% confidence interval for the difference in intervention rates between PVC and SOC exceeded a predefined 3% criterion for equivalence. Simple compressions were performed more frequently in the PVS subjects, which accounted for the higher intervention rate in this group. Postoperative interventions for bleeding were required in one SOC subject (1.0%) and one subject from the PVS group (0.9%). Procedure-related adverse events occurred in 21 (21.9%) SOC subjects and 23 (21.9%) PVS subjects, with no adverse events related to use of the study devices. CONCLUSIONS: The PVS exhibited similar overall safety and effectiveness to SOC staplers in video-assisted thoracoscopic surgery lobectomy.

Does induction chemoradiotherapy increase survival in patients with Pancoast tumour?

A best evidence topic in cardiothoracic surgery was written according to a structured protocol. The question addressed was whether induction (neoadjuvant) chemoradiotherapy (iCRT) compared with other therapeutic strategies improves survival in patients with Pancoast tumours. Altogether 248 papers were identified using the below-mentioned search. Ten of them represented the best evidence to answer the clinical question. The author, journal, date and country of publication, patient group, relevant outcomes and weaknesses were tabulated. Four studies were retrospective comparative studies of induction chemoradiotherapy and surgery (trimodal therapy) versus other therapeutic strategies. Two studies were retrospective and four were prospective investigating trimodal therapy. These papers comprised a total of 550 patients. The overall survival was better with trimodal therapy compared with RT (radiotherapy) followed by surgery group in all three comparative studies. The 2-year survival varied in the trimodal therapy group from 70 to 93%, in comparison to RT group where variation was from 22 to 49%. Five-year survival for trimodal therapy varied between 36.4 and 84% in the results of two comparative studies, compared with 11 and 49% for RT and surgery, respectively. One paper looked at survival in patients who underwent surgery alone [30% at 2-year and 20% at 4-year overall survival (OS)]. The 5-year OS in the retrospective group varied between 38 and 59%. Similar results were reported for the prospective group with 5-year OS between 44 and 56%. Despite a large variation in pCR (complete pathological response) (15-93%) and R0 (77-100%) reported, both represented a positive prognostic factor for survival. Three papers looked at the impact of staging following induction chemoradiotherapy. The majority of patients had T3 disease. An advantage in survival was seen in patients with early disease compared with advanced stage. No randomized controlled trials were identified. All the 10 articles suggested there was a benefit in trimodal therapy with improvement in overall survival. We conclude that combining induction chemoradiotherapy with surgery for Pancoast tumour may offer a survival benefit compared with radiotherapy with surgery or surgery alone.

Metastatic index of non-small-cell lung cancer and long-term survival.

AIM: We aimed to determine whether metastatic index is a factor determining long-term survival in patients undergoing curative resection for non-small-cell lung cancer. METHODS: There were 2695 consecutive pulmonary resections performed between October 2001 and September 2011 in our institution; 1795 were potentially curative resections for non-small-cell lung cancer with bronchial margin length data available. Benchmarking against the International Association for the Study of Lung Cancer data set was performed. Cox multivariate analysis was undertaken. Metastatic index was defined as N stage× bronchial resection margin length. RESULTS: Benchmarking failed to reveal any significant differences between our data and the International Association for the Study of Lung Cancer data set. Univariate analysis identified metastatic index as a significant factor determining long-term survival (p = 0.04). Cox regression demonstrated that metastatic index (hazard ratio 1.29, p = 0.0002), age (hazard ratio 1.02, p < 0.0001), body mass index (hazard ratio 0.98, p = 0.006), female sex (hazard ratio 0.65, p < 0.0001), T1 stage (hazard ratio 0.67, p < 0.0001), T2 stage (hazard ratio 2.13, p < 0.0001), T3 stage (hazard ratio 1.59, p = 0.03), forced expiratory volume in 1 s (hazard ratio 0.70, p < 0.0001), pneumonectomy (hazard ratio 1.43, p = 0.001), histology subtype adenosquamous (hazard ratio 3.77, p = 0.01) and squamous (hazard ratio 0.83, p = 0.03) were all significant determinants of long-term survival. CONCLUSION: Metastatic index is a significant factor determining long-term survival in patents with adenocarcinoma undergoing potentially curative surgery with a lobectomy.

Prediction of in-hospital mortality following pulmonary resections: improving on current risk models.

OBJECTIVES: Using a large, prospectively collected and independently validated thoracic database, we created a risk-prediction tool for in-hospital mortality with the aim of improving on the accuracy of Thoracoscore. METHODS: A prospectively collected and independently validated database containing lung resections was utilized, N = 2574. Logistic regression analysis with bootstrapping, and by the use of a random training and test set was utilized. Comparisons against the Thoracoscore, ESOS.01 and the Society of Thoracic Surgeons (STS) models were performed. RESULTS: A logistic model identified age [odds ratio (OR) 1.1, 95% confidence interval (CI) 1.0-1.2, P = 0.0002], sex (OR 0.34, 95% CI 0.14-0.83, P = 0.02), predicted postoperative FEV1 (OR 0.96, 95% CI 0.94-0.99, P = 0.002), emphysema (OR 3.2, 95% CI 1.0-9.9, P = 0.04), excess alcohol consumption (OR 1.0, 95% CI 1.0-1.0, P = 0.04), pre-existing renal disease (OR 4.3, 95% CI 1.1-17.1, P = 0.04), predicted in-hospital mortality with an receiver operating curve (ROC) of 0.81 and a Hosmer-Lemeshow test of 0.9. Bootstrap analysis confirmed the above risk factors (ROC 0.82 and Hosmer-Lemeshow 0.2). Comparisons between Thoracoscore, ESOS.01 and the STS risk models demonstrated that none was very accurate, as all had low ROC values of 0.69, 0.70 and 0.61, respectively. The STS risk model does not apply to our population (ROC 0.61, Hosmer-Lemeshow, P = 0.004), and the ESOS.01 has poor predictive power (Hosmer-Lemeshow, P < 0.0001). CONCLUSIONS: Logistic regression based on age, sex, predicted postoperative FEV1, alcohol consumption and pre-existing renal disease predicts in-hospital mortality with improved accuracy compared with the use of Thoracoscore, ESOS.01 and the STS risk model.

Lung cancer staging: a physiological update.

The tumour-node metastasis (TNM) classification system is anatomically based. We investigated whether the addition of simple physiological variables, age and body mass index (BMI), would affect survival curves, i.e. a composite anatomical and physiological staging system. We retrospectively analysed a prospectively validated thoracic surgery database (n = 1981). Cox multivariate analysis was performed to determine possible significant factors. Kaplan-Meier survival curves were constructed with combined anatomical and physiological factors. Cox multivariate analysis revealed age (P < 0.001) and BMI (P = 0.01) as significant factors affecting survival. Receiver operating curve analysis determined cut-off levels for age of 67 and BMI of 27.6. A composite anatomical and physiological survival curve based on TNM for BMI > 27.6 and age < 67 was produced. Age and BMI criteria resulted in significantly different survival curves, for stage I (P < 0.0001) and stage II (P = 0.0032), but not for stage III (P = 0.06). Neural network analysis confirmed the importance of BMI and age above cancer stage with regard to long-term survival. Combining age < 67, BMI > 27.6 and TNM anatomical classification results in very different estimated survival curves from the usual TNM system. Patients from stages I, II and III may have survival equivalent to a stage higher or lower depending on their age and BMI.

The Norwood procedure using a right ventricle-pulmonary artery conduit: comparison of the right-sided versus left-sided conduit position.

OBJECTIVE: We proposed that a right-sided right ventricle-pulmonary artery conduit during the stage I Norwood procedure would facilitate pulmonary artery reconstruction during the stage II procedure. METHODS: Between 2002 and 2006, 153 patients underwent Norwood stage I reconstruction with a right ventricle-pulmonary artery conduit (125 in the right-sided group and 28 in the left-sided group). The previous 150 consecutive classic Norwood procedures (1997-2002) were used as a control group. Outcomes from stages I and II were analyzed, including ventricular function and pulmonary artery morphology. RESULTS: The 30-day survival was 88% (110/125) in the right-sided group, 75% (21/28) in the left-sided group, and 70% (105/150) in the control group (P < .001, right-sided vs control groups). The conduit length was 35 +/- 9 mm in the right-sided group and 26 +/- 8 mm in the left-sided group (P = .001). Survival at 6 months demonstrated a significant survival benefit in the right-sided right ventricle-pulmonary artery conduit group over the control group (P = .009, log-rank test). There was no difference in ventricular function between the groups and no regional dyskinesia associated with the right ventricle-pulmonary artery conduit. Despite larger branch pulmonary artery size in the right ventricle-pulmonary artery conduit groups (compared with the control group), central pulmonary artery stenoses were common (62% in the right conduit and 80% in the left conduit). Bypass and ischemic times at stage II were 49 +/- 10 and 23 +/- 13 minutes in the right-sided group compared with 61.5 +/- 9.5 and 31 +/- 14 minutes in the left-sided group (P < .001 and P = .03, respectively). The 30-day mortality after the stage II procedure was 1.3% (1/76) in the right-sided group, 0% (0/18) in the left-sided group, and 3.3% (3/90) in the control group. CONCLUSION: The right-sided conduit is a safe technique and has improved 30-day and overall post-stage II survival compared with that seen with the classic Norwood procedure. The right ventricle-pulmonary artery conduit is associated with central pulmonary artery stenosis but good development of the branch pulmonary arteries and preservation of ventricular function. The right-sided conduit significantly reduces cardiopulmonary bypass times at stage II.

Alveolar macrophage secretory products effect type 2 pneumocytes undergoing hypoxia-reoxygenation.

BACKGROUND: Activation of the alveolar macrophage is centrally important to the development of lung ischemia reperfusion injury. Alveolar macrophages and type 2 pneumocytes secrete a variety of proinflammatory mediators in response to oxidative stress. The manner in which they interact and how the macrophage may influence pneumocyte responses in lung ischemia reperfusion injury is unknown. Utilizing an in vitro model of hypoxia and reoxygenation, we sought to determine if the proinflammatory response of type 2 pneumocytes to oxidative stress would be amplified by alveolar macrophage secretory products. METHODS: Cultured pneumocytes were exposed to control media or media from cultured macrophages exposed to hypoxia and reoxygenation. Pneumocytes were subsequently subjected to hypoxia and reoxygenation and assessed for both nuclear translocation of nuclear factor kappa B and inflammatory cytokine and chemokine secretion. To examine for any reciprocal interactions, we reversed the experiment, exposing macrophages to conditioned pneumocyte media. RESULTS: In the presence of media from stimulated macrophages, production of proinflammatory mediators by type 2 pneumocytes was dramatically enhanced. In contrast, exposure of the macrophage to conditioned pneumocyte media had an inhibitory effect on macrophage responses subsequently exposed to hypoxia and reoxygenation. CONCLUSIONS: The alveolar macrophage drives the development of lung reperfusion injury in part through amplification of the inflammatory response of type 2 pneumocytes subjected to hypoxia and reoxygenation.

A comparison of phylogenetic network methods using computer simulation.

BACKGROUND: We present a series of simulation studies that explore the relative performance of several phylogenetic network approaches (statistical parsimony, split decomposition, union of maximum parsimony trees, neighbor-net, simulated history recombination upper bound, median-joining, reduced median joining and minimum spanning network) compared to standard tree approaches, (neighbor-joining and maximum parsimony) in the presence and absence of recombination. PRINCIPAL FINDINGS: In the absence of recombination, all methods recovered the correct topology and branch lengths nearly all of the time when the substitution rate was low, except for minimum spanning networks, which did considerably worse. At a higher substitution rate, maximum parsimony and union of maximum parsimony trees were the most accurate. With recombination, the ability to infer the correct topology was halved for all methods and no method could accurately estimate branch lengths. CONCLUSIONS: Our results highlight the need for more accurate phylogenetic network methods and the importance of detecting and accounting for recombination in phylogenetic studies. Furthermore, we provide useful information for choosing a network algorithm and a framework in which to evaluate improvements to existing methods and novel algorithms developed in the future.

Poly (ADP) ribose synthetase inhibition in alveolar macrophages undergoing hypoxia and reoxygenation.

BACKGROUND: Inhibition of the nuclear enzyme poly ribose synthetase (PARS) protects against in vivo lung ischemia reperfusion injury (LIRI). The effectiveness of intratracheal treatment suggests that PARS inhibition may primarily modulate alveolar macrophage (AM) activation. These studies attempted to characterize the effects of PARS on AM activation in response to oxidative stress. METHODS: Primary cultures of AM were rendered hypoxic for 2 h and reoxygenated for up to 4 h. Cells were preincubated with INO-1001, a specific PARS inhibitor 1 h prior to hypoxia. Gel shift assays characterized nuclear factor kappa B (NFkappaB), and enzyme linked immunosorbent assay quantitated chemokine/cytokine protein secretion. RESULTS: Hypoxia and reoxygenation resulted in an increase in the early nuclear translocation of NFkappaB, and an increase in the secretion of the cytokine tumor necrosis factor-alpha (TNF-alpha), chemokines macrophage inflammatory protein (MIP-1alpha), monocyte chemoattractant protein one (MCP-1) and cytokine induced neutrophil chemoattractant (CINC). Pretreatment of AM with INO-1001 decreased both the early translocation of NFkappaB and the production of TNF-alpha (p<0.05) and MIP-1alpha p=0.02, but did not affect CINC or MCP-1 production. CONCLUSIONS: These findings indicate that PARS inhibition in the AM blunts their response to oxidative stress and may help explain the protective effects of intratracheal PARS inhibition in LIRI.

Poly (ADP) ribose polymerase inhibition improves rat cardiac allograft survival.

BACKGROUND: Heart transplantation is an accepted treatment modality for end-stage heart failure. However, acute cellular rejection (ACR) continues to be a morbid complication. Recently a novel mechanism of inflammatory allograft injury has been characterized which involves overactivation of the nuclear enzyme poly (ADP-ribose) polymerase (PARP). In the present studies, we compared the efficacy of INO-1001, a novel, potent PARP inhibitor, in limiting ACR with and without adjuvant low-dose cyclosporine (CSA). METHODS: Heterotopic heart transplantation was performed utilizing Brown-Norway strains as donors and Lewis rats as recipients. Groups received daily intraperitoneal injections of: vehicle, low-dose CSA, low-dose INO-1001, high-dose INO-1001, and low-dose CSA combined with high-dose INO-1001. Additional animals were sacrificed on postoperative Day 5 for histologic assessments of allograft inflammation, including immunohistochemistry for nitrotyrosine and poly (ADP-ribose) (the product of PARP) staining. RESULTS: PARP inhibition significantly prolonged allograft survival relative to vehicle controls. The combination of low-dose CSA and INO-1001 resulted in a marked increase in allograft survival and significant reductions in allograft rejection scores. This was associated with decreased nitrotyrosine and PAR staining in transplanted cardiac allografts. CONCLUSIONS: Pharmacologic inhibition of INO-1001 prolongs allograft survival in a dose-dependent fashion in a rodent model of heart transplantation. PARP inhibitors may permit reductions in the dose of CSA needed for adequate immunosuppression after heart transplantation.

Role of poly (ADP) ribose synthetase in lung ischemia-reperfusion injury.

BACKGROUND: The activation of poly (adenosine diphosphate) ribose synthetase (PARS) is known to be important in the cellular response to oxidative stress. Previous studies have reported that PARS inhibition confers protection in models of endotoxic shock and ischemia-reperfusion. The purpose of this study was to determine the role of PARS inhibition in lung ischemia-reperfusion injury (LIRI). METHODS: Left lungs of Long-Evans rats were rendered ischemic for 90 minutes and reperfused for up to 4 hours. Treated animals received 3 mg/kg of INO-1001 (a PARS inhibitor) intravenously 30 minutes before ischemia. Injury was quantitated in terms of tissue myeloperoxidase (MPO) content, vascular permeability ((125)I radiolabeled bovine serum albumin extravasation) and bronchoalveolar lavage (BAL) leukocyte content. BAL fluid was assessed for cytokine and chemokine content by enzyme-linked immunoassay. Further samples were processed for nuclear protein analysis by electromobility shift assay (EMSA) and cellular death by terminal deoxyribonucleotidyl transferase-mediated d-UTP biotin nick-end labeling (TUNEL) assay and caspase-3 staining. RESULTS: Lung vascular permeability was reduced in treated animals by 73% compared with positive controls (p < 0.009). The protective effects of PARS inhibition correlated with a 46% reduction in tissue MPO content (p < 0.008) and marked reductions in BAL leukocyte accumulation. This positively correlated with the diminished expression of pro-inflammatory mediators and nuclear transcription factors, as well as decreased levels of cellular death. CONCLUSIONS: The deleterious effects of LIRI are in part mediated by the formation of free radicals and superoxides, which lead to DNA single-strand breaks. This leads to activation of PARS, which causes rapid cellular energy depletion and cell death. PARS inhibition is protective against this and represents a potentially useful therapeutic tool in the prevention of LIRI.

FR167653 reduces obliterative airway disease in rats.

BACKGROUND: Obliterative bronchiolitis (OB) is the main cause of late mortality among long-term survivors of lung transplantation. Although p38 kinase is functional in multiple acute inflammatory injury models, its role in chronic lung rejection is undefined. p38 regulates the expression of the cytokines tumor necrosis (TNF)-alpha and interleukin (IL)-1beta, 2 mediators involved in the development of OB in a tracheal transplant model. These studies assessed whether specific inhibition of p38 with FR167653 (FR) protects against the development of OB in rat tracheal allografts. METHODS: Rat airways were heterotopically transplanted from Brown-Norway donors into Lewis recipients, and animals were sacrificed on day 14 (6 per group). Treated animals received 10 mg/kg daily of FR intraperitoneally beginning either immediately or at day 7 after transplant. Allografts were assessed by computerized morphometry, and tracheas were processed for TNF-alpha mRNA and protein expression by quantitative reverse transcriptase-polymerase chain reaction and immunohistochemistry, respectively. Electrophoretic mobility shift assays evaluated nuclear factor kappa beta (NFkappaB) transactivation. RESULTS: Control allografts averaged 61% occlusion and 98% loss of epithelium at 14 days, whereas FR administration reduced luminal occlusion to 28% (p < 0.001) and epithelial loss to 71% (p < 0.001). Delayed treatment beginning on day 7 slowed progression of disease, as tracheal occlusion averaged 44% and epithelial loss averaged 80%, both of which were significant (p < 0.05) improvements relative to 14-day controls. NFkappa transactivation (p < 0.004) and TNF-alpha mRNA and protein expression were reduced dramatically by FR at 14 days. CONCLUSIONS: A specific p38 inhibitor, FR 167653, ameliorates obliterative airway disease in rat tracheal allografts via attenuated NFkappaB transactivation, which ultimately results in diminished TNF-alpha mRNA and protein expression.

Poly (ADP) ribose synthetase inhibition reduces obliterative airway disease in rat tracheal allografts.

BACKGROUND: Obliterative bronchiolitis (OB) is the major long-term complication affecting lung transplant recipients, and is characterized pathologically by chronic inflammatory and fibroproliferative airway disease. Based on studies revealing anti-inflammatory and anti-apoptotic properties of poly (ADP)-ribose synthetase (PARS) inhibitors, we hypothesized that their administration would be protective in a heterotopic model of experimental OB. METHODS: We transplanted rat tracheas from Brown-Norway donors into Lewis recipients, and treated 2 groups with a novel PARS inhibitor, INO-1001. One group received 14 days of treatment, whereas a second received delayed treatment beginning on Day 7 post-transplant. Tracheas were analyzed by light microscopy and computerized morphometry. Effects on cytokine transcription, nuclear transcription factor activation and cellular death were assessed by in situ hybridization for tumor necrosis factor-alpha (TNF-alpha), electromobility shift assays for nuclear factor-kappaB (NF-kappaB) and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assays, respectively. RESULTS: PARS inhibition significantly decreased luminal obstruction (p < 0.001) and enhanced preservation of epithelial lining (p < 0.001) at 14 days post-transplant. Day 7 controls confirmed the development of an obstructive lesion in the lumen, averaging 28% occlusion. Delayed treatment (beginning on Day 7) arrested (p < 0.001) progression of the established lesion. Allograft airways treated with INO-1001 demonstrated attenuated NF-kappaB nuclear translocation, reduced transcription of TNF-alpha mRNA, and decreased cellular death on TUNEL and caspase 3 staining. CONCLUSIONS: PARS inhibition is anti-inflammatory, protects against experimental OB, and is associated with enhanced preservation of respiratory epithelium and decreased cellular death. Delayed treatment with INO-1001 arrests progression of the lesion developed by Day 7. These studies suggest that activation of PARS plays a critical role in the development of airway obliterative disease.

Intratracheal poly (ADP) ribose synthetase inhibition ameliorates lung ischemia reperfusion injury.

BACKGROUND: We previously demonstrated that intravenous poly (ADP) ribose synthetase (PARS) inhibition protects against experimental lung ischemia reperfusion injury (LIRI) in an in situ, hilar occlusion model. This study determined its efficacy when administered intratracheally (IT). METHODS: Left lungs of rats were rendered ischemic for 90 minutes, and reperfused for up to 4 hours. Treated animals received INO-1001, a PARS inhibitor, intratracheally 30 minutes before ischemia, while controls were given IT vehicle at equivalent volumes. All groups contained at least 4 animals. Lung injury was quantitated utilizing vascular permeability to radiolabeled albumin, tissue myeloperoxidase (MPO) content, alveolar leukocyte cell counts, and arterial pO(2) at 4 hours of reperfusion. Electrophoretic mobility shift assays (EMSA) assessed the nuclear translocation of NFkappaB and AP-1 in injured left lungs, while ELISAs quantitated secreted cytokine induced neutrophil chemoattractant (CINC) and MCP-1 protein in bronchoalveolar lavage fluid. RESULTS: Intratracheal PARS inhibition was 73% (p < 0.0001) and 87% (p < 0.0001) protective against increases in vascular permeability and alveolar leukocyte accumulation, respectively, and improved arterial pO(2) (p < 0.0004) at 4 hours of reperfusion. Myeloperoxidase (MPO) activity in treated lungs was reduced by 70% (p < 0.02). The nuclear translocation of NFkappaB and AP-1 was attenuated at 15 minutes of reperfusion, and the secretion of CINC and MCP-1 (p < 0.05) protein into the alveolus was diminished at 4 hours of reperfusion. CONCLUSIONS: Intratracheal INO-1001 protects against experimental LIRI. The reduction in secreted chemokine protein at 4 hours of reperfusion appears to be mediated at the pretranscriptional level through attenuated NFkappaB and AP-1 activation. This route may optimize future donor organ management and improve lung recipient outcomes.

Alpha chemokines regulate direct lung ischemia-reperfusion injury.

BACKGROUND: Alpha chemokines function predominantly to recruit and activate neutrophils, which are important effectors of acute lung injury. This study evaluated whether blockade of 2 potent alpha chemokines, macrophage inflammatory protein-2 (MIP-2) and cytokine-induced neutrophil chemoattractant (CINC), is protective against lung ischemia-reperfusion injury in a warm in situ hilar clamp model. METHODS: Left lungs of Long-Evans rats underwent normothermic ischemia for 90 minutes and reperfusion for up to 4 hours. Treated animals received antibodies to MIP-2 or CINC immediately prior to reperfusion. Lung injury was quantitated by vascular permeability to (125)I-radiolabeled bovine serum albumin, lung tissue neutrophil sequestration (myeloperoxidase [MPO] content), and alveolar leukocyte content in bronchoalveolar lavage (BAL) fluid. CINC and MIP-2 mRNA expression were assessed by northern blot, while ribonuclease protection assays were performed to evaluate mRNA expression for a number of early response cytokines. MIP-2 and CINC protein expression in injured lungs was determined by immunoblotting. RESULTS: Treatment with antibodies to CINC or MIP-2 was associated with significant protection against increases in vascular permeability, MPO content and alveolar leukocyte sequestration in injured lungs. Expression of CINC and MIP-2 mRNA peaked after 2 hours of reperfusion in injured lungs, and protein levels were evident on immunoblotting after 3 hours of reperfusion. Neither CINC nor MIP-2 blockade appeared to modulate cytokine mRNA expression. CONCLUSIONS: CINC and MIP-2 are important mediators involved in direct lung ischemia-reperfusion injury. They appear to function by modulating neutrophil recruitment, but not inflammatory cytokine release.

Early tumor necrosis factor-alpha release from the pulmonary macrophage in lung ischemia-reperfusion injury.

OBJECTIVE: Tumor necrosis factor-alpha is a proinflammatory mediator required for the development of experimental lung ischemia-reperfusion injury. The alveolar macrophage is a rich source of tumor necrosis factor-alpha in multiple models of acute lung injury. The present study was undertaken to determine whether the alveolar macrophage is an important source of tumor necrosis factor-alpha in lung ischemia-reperfusion injury and whether suppression of its function protects against injury. METHODS: Left lungs of Long-Evans rats underwent normothermic ischemia for 90 minutes and reperfusion for up to 4 hours. Treated animals received gadolinium chloride, a rare earth metal that inhibits macrophage function. Injury was quantitated via lung tissue neutrophil accumulation (myeloperoxidase content), lung vascular permeability, and bronchoalveolar lavage fluid leukocyte, cytokine, and chemokine content. Separate samples were generated for immunohistochemistry. RESULTS: Tumor necrosis factor-alpha secretion occurred at 15 minutes of reperfusion and was localized to the alveolar macrophage by immunohistochemistry. In gadolinium-treated animals, lung vascular permeability was reduced by 66% at 15 minutes (P <.03) of reperfusion and by 34% at 4 hours (P <.02) of reperfusion. Suppression of macrophage function resulted in a 35% reduction in lung myeloperoxidase content (P <.03) and similar reductions in bronchoalveolar lavage leukocyte accumulation. Tumor necrosis factor-alpha and microphage inflammatory protein-1alpha protein levels were markedly reduced in the bronchoalveolar lavage of gadolinium-treated animals by enzyme-linked immunosorbent assay. CONCLUSIONS: The alveolar macrophage secretes tumor necrosis factor-alpha protein by 15 minutes of reperfusion, which orchestrates the early events that eventually result in lung ischemia-reperfusion injury at 4 hours. Gadolinium pretreatment markedly reduces tumor necrosis factor-alpha elaboration, resulting in significant protection against lung ischemia-reperfusion injury.

Beta-chemokine function in experimental lung ischemia-reperfusion injury.

BACKGROUND: Although chemokines are functionally important in models of ischemia-reperfusion injury, little is known about their role in lung ischemia-reperfusion injury (LIRI). This study examined the role of the beta-chemokines, macrophage inflammatory protein (MIP)-1alpha, monocyte chemoattractant protein (MCP)-1, and regulated upon activation normal T cells expressed and secreted (RANTES) in LIRI. METHODS: Left lungs of Long-Evans rats underwent normothermic ischemia for 90 minutes and reperfusion for up to 4 hours. Treated animals received anti-MIP-1alpha, anti-MCP-1, or anti-RANTES antibodies before reperfusion. Changes in lung vascular permeability were measured with iodine 125-labeled bovine serum albumin. Neutrophil accumulation in the lung parenchyma was determined by myeloperoxidase activity, and bronchoalveolar lavage was performed to measure leukocyte cell counts. Western blots, Northern blots, and ribonuclease protection assays assessed beta-chemokine messenger RNA and protein levels. RESULTS: Animals receiving anti-MIP-1alpha demonstrated reduced vascular permeability compared with controls (p < 0.001). Attenuation of permeability was less dramatic in animals treated with anti-MCP-1 and anti-RANTES antibody, which demonstrated permeability decreases of 15% and 16%, respectively (p < 0.02). Lung neutrophil accumulation was reduced in animals receiving anti-MIP-1alpha antibody (p < 0.005) but was unchanged in animals receiving either anti-MCP-1 or anti-RANTES. Bronchoalveolar lavage leukocyte content was also reduced by treatment with anti-MIP-1alpha (p < 0.003) and was unchanged in anti-MCP-1-treated and anti-RANTES-treated animals. MIP-1alpha treatment decreased tumor necrosis factor-alpha messenger RNA in injured left lungs. CONCLUSIONS: MIP-1alpha is functionally significant in the development of LIRI. It likely exerts its effects in part by mediating the expression of proinflammatory and antiinflammatory cytokines and influencing tissue neutrophil recruitment. MCP-1 and RANTES seem to play relatively minor roles in the development of direct LIRI.

Proinflammatory response of alveolar type II pneumocytes to in vitro hypoxia and reoxygenation.

Type II pneumocytes (T2P) are integral in preserving the integrity of the alveolar space by modulating the fluid composition surrounding the alveolar epithelium. There is also mounting evidence supporting their contribution to the development of acute inflammatory lung injury subsequent to oxidative stress. This study characterized the response of T2P to in vitro hypoxia and reoxygenation (H&R). Rat T2P from a cultured cell line (RLE-6TN) were rendered hypoxic for 2 h, and reoxygenated for up to 6 h. Activation of signaling kinases, the nuclear translocation of proinflammatory transcription factors, and quantification of secreted cytokine and chemokine protein content were assessed. Type II pneumocytes expressed activated extracellular signal regulated kinase (ERK) 1/2 maximally at 15 min of reoxygenation. C-jun n-terminal kinase (JNK) and p38 activation was minimal at all time points studied. The nuclear translocation of nuclear factor kappa B (NFkappaB) and activator protein (AP)-1 were dramatic after 15 min of reoxygenation. There was a significant increase in the protein secretion of CINC (p = 0.03), IL-1beta (p = 0.02), and monocyte chemoattractant protein-1 (p < 0.001) at 6 h of reoxygenation. Type II pneumocytes respond directly to H&R. ERK 1/2 activity peaks at 15 min of reoxygenation, and correlates temporally with the nuclear translocation of NFkappaB and AP-1. These signaling cascades likely promote the elaboration of proinflammatory mediators.