Isoflurane and Pentobarbital Anesthesia for Pulmonary Studies Requiring Prolonged Mechanical Ventilation in Mice.

Mechanical ventilation can be used in mice to support high-risk anesthesia or to create clinically relevant, intensive care models. However, the choice of anesthetic and inspired oxygen concentration for prolonged procedures may affect basic physiology and lung inflammation. To characterize the effects of anesthetics and oxygen concentration in mice experiencing mechanical ventilation, mice were anesthetized with either isoflurane or pentobarbital for tracheostomy followed by mechanical ventilation with either 100% or 21% oxygen. Body temperature, oxygen saturation, and pulse rate were monitored continuously. After 6 h, mice were euthanized for collection of blood and bronchoalveolar lavage fluid for evaluation of biomarkers of inflammation and lung injury, including cell counts and cytokine levels. Overall, both isoflurane and pentobarbital provided suitable anesthesia for 6 h of mechanical ventilation with either 21% or 100% oxygen. We found no differences in lung inflammation biomarkers attributable to either oxygen concentration or the anesthetic. However, the combination of pentobarbital and 100% oxygen resulted in a significantly higher concentration of a biomarker for lung epithelial cell injury. This study demonstrates that the combination of anesthetic agent, mechanical ventilation, and inspired oxygen concentrations can alter vital signs and lung injury biomarkers during prolonged procedures. Their combined impact may influence model development and the interpretation of research results, warranting the need for preliminary evaluation to establish the baseline effects.

Breath analysis for detection and trajectory monitoring of acute respiratory distress syndrome in swine.

Despite the enormous impact on human health, acute respiratory distress syndrome (ARDS) is poorly defined, and its timely diagnosis is difficult, as is tracking the course of the syndrome. The objective of this pilot study was to explore the utility of breath collection and analysis methodologies to detect ARDS through changes in the volatile organic compound (VOC) profiles present in breath. Five male Yorkshire mix swine were studied and ARDS was induced using both direct and indirect lung injury. An automated portable gas chromatography device developed in-house was used for point of care breath analysis and to monitor swine breath hourly, starting from initiation of the experiment until the development of ARDS, which was adjudicated based on the Berlin criteria at the breath sampling points and confirmed by lung biopsy at the end of the experiment. A total of 67 breath samples (chromatograms) were collected and analysed. Through machine learning, principal component analysis and linear discrimination analysis, seven VOC biomarkers were identified that distinguished ARDS. These represent seven of the nine biomarkers found in our breath analysis study of human ARDS, corroborating our findings. We also demonstrated that breath analysis detects changes 1-6 h earlier than the clinical adjudication based on the Berlin criteria. The findings provide proof of concept that breath analysis can be used to identify early changes associated with ARDS pathogenesis in swine. Its clinical application could provide intensive care clinicians with a noninvasive diagnostic tool for early detection and continuous monitoring of ARDS.

Analgesia and Humane Endpoints for Rodents in Sepsis Research.

Numerous regulatory bodies around the world require analgesics for rodents undergoing surgery to induce sepsis. Well-controlled pain will decrease morbidity. Options for analgesics include NSAIDs, local analgesics, and opioids. Supportive care can also decrease stress to post-operative animals. As well, humane endpoints should be agreed upon before the study commences so as to alleviate unnecessary pain and distress.

A novel swine model of the acute respiratory distress syndrome using clinically relevant injury exposures.

To date, existing animal models of the acute respiratory distress syndrome (ARDS) have failed to translate preclinical discoveries into effective pharmacotherapy or diagnostic biomarkers. To address this translational gap, we developed a high-fidelity swine model of ARDS utilizing clinically relevant lung injury exposures. Fourteen male swine were anesthetized, mechanically ventilated, and surgically instrumented for hemodynamic monitoring, blood, and tissue sampling. Animals were allocated to one of three groups: (1) Indirect lung injury only: animals were inoculated by direct injection of Escherichia coli into the kidney parenchyma, provoking systemic inflammation and distributive shock physiology; (2) Direct lung injury only: animals received volutrauma, hyperoxia, and bronchoscope-delivered gastric particles; (3) Combined indirect and direct lung injury: animals were administered both above-described indirect and direct lung injury exposures. Animals were monitored for up to 12 h, with serial collection of physiologic data, blood samples, and radiographic imaging. Lung tissue was acquired postmortem for pathological examination. In contrast to indirect lung injury only and direct lung injury only groups, animals in the combined indirect and direct lung injury group exhibited all of the physiological, radiographic, and histopathologic hallmarks of human ARDS: impaired gas exchange (mean PaO2 /FiO2 ratio 124.8 ± 63.8), diffuse bilateral opacities on chest radiographs, and extensive pathologic evidence of diffuse alveolar damage. Our novel porcine model of ARDS, built on clinically relevant lung injury exposures, faithfully recapitulates the physiologic, radiographic, and histopathologic features of human ARDS and fills a crucial gap in the translational study of human lung injury.

A comprehensive assessment of multi-system responses to a renal inoculation of uropathogenic E. coli in swine.

BACKGROUND: The systemic responses to infection and its progression to sepsis remains poorly understood. Progress in the field has been stifled by the shortcomings of experimental models which include poor replication of the human condition. To address these challenges, we developed and piloted a novel large animal model of severe infection that is capable of generating multi-system clinically relevant data. METHODS: Male swine (n = 5) were anesthetized, mechanically ventilated, and surgically instrumented for continuous hemodynamic monitoring and serial blood sampling. Animals were inoculated with uropathogenic E. coli by direct injection into the renal parenchyma and were maintained until a priori endpoints were met. The natural history of the infection was studied. Animals were not resuscitated. Multi-system data were collected hourly to 6 hours; all animals were euthanized at predetermined physiologic endpoints. RESULTS: Core body temperature progressively increased from mean (SD) 37.9(0.8)°C at baseline to 43.0(1.2)°C at experiment termination (p = 0.006). Mean arterial pressure did not begin to decline until 6h post inoculation, dropping from 86(9) mmHg at baseline to 28(5) mmHg (p = 0.005) at termination. Blood glucose progressively declined but lactate levels did not elevate until the last hours of the experiment. There were also temporal changes in whole blood concentrations of a number of metabolites including increases in the catecholamine precursors, tyrosine (p = 0.005) and phenylalanine (p = 0.005). Lung, liver, and kidney function parameters worsened as infection progressed and at study termination there was histopathological evidence of injury in these end-organs. CONCLUSION: We demonstrate a versatile, multi-system, longitudinal, swine model of infection that could be used to further our understanding of the mechanisms that underlie infection-induced multi-organ dysfunction and failure, optimize resuscitation protocols and test therapeutic interventions. Such a model could improve translation of findings from the bench to the bedside, circumventing a significant obstacle in sepsis research.

Thermoneutral Housing Temperature Improves Survival in a Murine Model of Polymicrobial Peritonitis.

Regulatory guidelines mandate housing for laboratory mice at temperatures below their thermoneutral zone, creating chronic cold stress. However, increases in housing temperature could alter immune responses. We hypothesized housing mice at temperatures within their thermoneutral zone would improve sepsis survival and alter immune responses. Male C57BL/6 mice were housed at 22°C or 30°C after cecal ligation and puncture (CLP) for 10 days. Survival of mice housed at 30°C (78%) after CLP was significantly increased compared with mice housed at 22°C (40%). Experimental groups were repeated with mice euthanized at 0, 12, 24, and 48 h post-surgery to examine select immune parameters. Raising housing temperature minimally altered systemic, peritoneal, or splenic cell counts. However, IL-6 levels in plasma and peritoneal lavage fluid were significantly lower at 12 h post-surgery in mice housed at 30°C compared with 22°C. Bacterial colony counts from peritoneal lavage fluid were significantly lower in mice housed at 30°C and in vivo studies suggested this was the result of increased phagocytosis by neutrophils. As previously demonstrated, adoptive transfer of fibrocytes significantly increased sepsis survival compared with saline at 22°C. However, there was no additive effect when adoptive transfer was performed at 30°C. Overall, the results demonstrated that thermoneutral housing improves survival after CLP by increasing local phagocytic activity and technical revisions may be necessary to standardize the severity of the model across different housing temperatures. These findings stress the pronounced impact housing temperature has on the CLP model and the importance of reporting housing temperature.

The Influence of Pain and Analgesia in Rodent Models of Sepsis.

Sepsis is a multifaceted host response to infection that dramatically affects patient outcomes and the cost of health care. Animal models are necessary to replicate the complexity and heterogeneity of clinical sepsis. However, these models entail a high risk of pain and distress due to tissue trauma, inflammation, endotoxin-mediated hyperalgesia, and other mechanisms. Several recent studies and initiatives address the need to improve the welfare of animals through analgesics and standardize the models used in preclinical sepsis research. Ultimately, the goal is to provide high-fidelity, humane animal models that better replicate the clinical course of sepsis, to provide more effective translation and advance therapeutic discovery. The purpose of this review is to discuss the current understanding of the roles of pain and analgesia in rodent models of sepsis. The current definitions of sepsis along with an overview of pain in human sepsis are described. Finally, welfare concerns associated with animal models of sepsis and the most recent considerations for relief of pain and distress are reviewed.

Phagocytosis by Fibrocytes as a Mechanism to Decrease Bacterial Burden and Increase Survival in Sepsis.

Fibrocytes are unique cells with innate and adaptive immune functions, but these mechanisms have not been fully explored. The aim of this study was to explain the mechanism by which adoptive transfer of exogenous fibrocytes improved bacterial clearance and increased sepsis survival. Initial flow cytometry-based, in vitro assays demonstrated phagocytosis by fibrocytes and intracellular bacterial killing was confirmed by direct plating of cell lysates after exposure to live bacteria. Intravenous adoptive transfer of fibrocytes at the time of cecal ligation and puncture (CLP) or 2 h after CLP in mice increased survivability. Decreased intraperitoneal bacterial burden was also observed. Quantification of peritoneal cell populations using flow cytometry demonstrated transferred and endogenous fibrocytes were significantly increased after CLP, while macrophage and neutrophil numbers were unchanged. To determine the impact in vivo, fluorescently labeled, killed bacteria were injected i.p. into mice 10 h after CLP or sham surgery ±â€Šadoptive transfer. Two hours later, flow cytometry of peritoneal cell populations after CLP alone revealed increased phagocytosis by macrophages, neutrophils, and endogenous fibrocytes. Transferred fibrocytes had significantly increased phagocytic activity in the septic peritoneum compared with sham and greater activity than any other cell type. Therefore, adoptive transfer may enhance bacterial clearance in early sepsis through the cumulative effects of endogenous and transferred fibrocytes rather than modulating the function of other endogenous phagocytes. Direct phagocytic activity coupled with previously described influences on T cell responses may explain the benefits of fibrocyte transfer in sepsis.

Pharmacokinetics of a Transdermal Fentanyl Solution in Suffolk Sheep (Ovis aries).

Sheep used as surgical models require appropriate pain management, and the commonly used transdermal fentanyl patches require a long predosing period to achieve adequate plasma concentrations. The aim of this study was to assess the pharmacokinetic parameters of an FDA-approved transdermal fentanyl solution (TFS) that has yet to be tested in sheep. In this study, we compared TFS at 2.7 mg/kg (n = 2), 1.7 mg/kg (n = 3), and 0.5 mg/kg (n = 3) with the control fentanyl patch at 2 µg/kg/h (n = 1); both products were applied topically to the intrascapular region. Plasma concentrations showed significant interanimal variability. Severe adverse effects occurred at both 2.7 and 1.7 mg/kg TFS and mild to moderate adverse effects were noted at 0.5 mg/kg. At all 3 doses, TFS had greater maximal concentration, clearance rate, and volume of distribution; shorter time to maximal concentration; and similar half-lives to those of the patch. In addition, we validated the use of a commercial human fentanyl ELISA kit, which positively correlated with the liquid chromatography-mass spectroscopy data, but absolute values did not match. Overall, at all 3 dosages tested (0.5, 1.7, and 2.7 mg/kg), TFS delivered fentanyl plasma concentrations that exceeded the minimal effective concentration; however, adverse effects were noted at all 3 dosages. Caution and further study are required before the use of TFS in sheep can be recommended fully.

Evaluation of Pain Assessment Techniques and Analgesia Efficacy in a Female Guinea Pig (Cavia porcellus) Model of Surgical Pain.

Guinea pigs (Cavia porcellus) are a frequently used species in research, often involving potentially painful procedures. Therefore, evidence-based recommendations regarding analgesia are critically needed to optimize their wellbeing. Our laboratory examined the efficacy of carprofen and extended-release (ER) buprenorphine, alone and as a multimodal combination, for relieving postsurgical pain in guinea pigs. Animals were assessed by using evoked (mechanical hypersensitivity), nonevoked (video ethogram, cageside ethogram, time-to-consumption test), and clinical (weight loss) measurements for 96 h during baseline, anesthesia-analgesia, and hysterectomy conditions. In addition, ER buprenorphine was evaluated pharmacologically. Guinea pigs treated with a single analgesic showed increased mechanical sensitivity for at least 96 h and indices of pain according to the video ethogram for as long as 8 h, compared with levels recorded during anesthesia-analgesia. In contrast, animals given both analgesics demonstrated increased mechanical sensitivity and behavioral evidence of pain for only 2 h after surgery compared with anesthesia-analgesia. The cageside ethogram and time-to-consumption tests failed to identify differences between conditions or treatment groups, highlighting the difficulty of identifying pain in guinea pigs without remote observation. Guinea pigs treated with multimodal analgesia or ER buprenorphine lost at least 10% of their baseline weights, whereas weight loss in carprofen animals was significantly lower (3%). Plasma levels for ER buprenorphine exceeded 0.9 ng/mL from 8 to 96 h after injection. Of the 3 analgesia regimens evaluated, multimodal analgesia provided the most effective pain control in guinea pigs. However the weight loss in the ER buprenorphine-treated animals may need to be considered during analgesia selection.

Benefits of 21% Oxygen Compared with 100% Oxygen for Delivery of Isoflurane to Mice (Mus musculus) and Rats (Rattus norvegicus).

At research institutions, isoflurane delivered by precision vaporizer to a face mask is the standard for rodent surgery and for procedures with durations that exceed a few minutes. Pure oxygen is often used as the carrier gas for isoflurane anesthesia, despite documented complications from long-term 100% oxygen use in humans and known occupational safety risks. We therefore examined the effect of anesthetic delivery gas on physiologic variables in mice and rats. Rodents were anesthetized for 60 min with isoflurane delivered in either 21% or 100% oxygen by means of a nose cone. We noted no difference between carrier gasses in physiologic variables in mice, including body temperature, respiratory rate, mean arterial pressure, surgical recovery time, pH, or PaCO2. However, blood gas analysis revealed evidence of a ventilation-perfusion mismatch in the 100% oxygen group. Pressure-volume hysteresis and histomorphometric analyses confirmed the presence of increased atelectasis in mice that received 100% oxygen. Unlike mice, rats that received isoflurane in 100% oxygen had acute respiratory acidosis and elevated mean arterial pressure, but atelectasis was similar between carrier gasses. Our data suggest that both 100% and 21% oxygen are acceptable for the delivery of isoflurane to mice. However, mice anesthetized for studies focused on lung physiology or architecture would benefit from the delivery of isoflurane in 21% oxygen to reduce absorption atelectasis and the potential associated downstream inflammatory effects. For rats, delivery of isoflurane in 21% and 100% oxygen both caused perturbations in physiologic variables, and choosing a carrier gas is not straightforward.

Bayesian network analysis of multi-compartmentalized immune responses in a murine model of sepsis and direct lung injury.

BACKGROUND: Inflammatory disease processes involve complex and interrelated systems of mediators. Determining the causal relationships among these mediators becomes more complicated when two, concurrent inflammatory conditions occur. In those cases, the outcome may also be dependent upon the timing, severity and compartmentalization of the insults. Unfortunately, standard methods of experimentation and analysis of data sets may investigate a single scenario without uncovering many potential associations among mediators. However, Bayesian network analysis is able to model linear, nonlinear, combinatorial, and stochastic relationships among variables to explore complex inflammatory disease systems. In these studies, we modeled the development of acute lung injury from an indirect insult (sepsis induced by cecal ligation and puncture) complicated by a direct lung insult (aspiration). To replicate multiple clinical situations, the aspiration injury was delivered at different severities and at different time intervals relative to the septic insult. For each scenario, we measured numerous inflammatory cell types and cytokines in samples from the local compartments (peritoneal and bronchoalveolar lavage fluids) and the systemic compartment (plasma). We then analyzed these data by Bayesian networks and standard methods. RESULTS: Standard data analysis demonstrated that the lung injury was actually reduced when two insults were involved as compared to one lung injury alone. Bayesian network analysis determined that both the severity of lung insult and presence of sepsis influenced neutrophil recruitment and the amount of injury to the lung. However, the levels of chemoattractant cytokines responsible for neutrophil recruitment were more strongly linked to the timing and severity of the lung insult compared to the presence of sepsis. This suggests that something other than sepsis-driven exacerbation of chemokine levels was influencing the lung injury, contrary to previous theories. CONCLUSIONS: To our knowledge, these studies are the first to use Bayesian networks together with experimental studies to examine the pathogenesis of sepsis-associated lung injury. Compared to standard statistical analysis and inference, these analyses elucidated more intricate relationships among the mediators, immune cells and insult-related variables (timing, compartmentalization and severity) that cause lung injury. Bayesian networks are an effective tool for evaluating complex models of inflammation.

Use of Femoral Head and Neck Ostectomy and Physical Therapy to Manage Osteoarthritis in a Rhesus Macaque (Macaca mulatta).

Osteoarthritis is associated with pain and immobility in both humans and animals. However, available resources for osteoarthritis management in captive NHP are limited. This case report describes a novel management strategy for a 10-y-old male macaque with unilateral hindlimb lameness, prominent muscle wasting, and severely limited range of motion. Radiographs of the affected limb showed lytic lesions of the femoral head. To relieve pain and improve mobility, femoral head and neck ostectomy (FHO) was performed, and multiple pharmacotherapies were initiated. The macaque also received a unique method of physical therapy that required no sedation, acted as enrichment, and was implemented by using a conventional caging system. The response to therapy was monitored by measuring thigh circumference in the operated and nonoperated limbs, which demonstrated improvement in both legs. The unique physical therapy in conjunction with surgery and pharmacotherapy benefited the macaque with osteoarthritis by reducing discomfort and improving mobility.

Adoptive transfer of fibrocytes enhances splenic T-cell numbers and survival in septic peritonitis.

Fibrocytes are unique, fibroblast-like cells with diverse functions and the potential for immunomodulation, which prompted investigation of their previously unexplored role in sepsis. Specifically, the study goals were to determine if adoptive transfer of fibrocytes would affect outcome in sepsis and to define relevant immunopathologic changes associated with the outcomes. Initial in vitro studies demonstrated that naive T-cell proliferation was significantly increased in cocultures with tissue-derived fibrocytes as compared with culture either alone or with fibroblasts. In vivo, the adoptive transfer of fibrocytes at the time of cecal ligation and puncture significantly improved survival of mice compared with transfer of fibroblasts or saline. Septic mice had lower blood levels of interleukin 6 (IL-6) and markers of organ injury after fibrocyte transfer as well as a reduced bacterial burden. Locally, peritoneal lavage fluid yielded lower bacterial counts, lower IL-6, and reduced inflammatory cell counts when fibrocyte transfer was compared with saline. This was also accompanied by significant increases in splenic CD4(+) and CD8(+) T cells. In vitro stimulation of the splenic T cells demonstrated that, after cecal ligation and puncture and adoptive transfer, the percentages of both CD4(+) and CD8(+) T cells with intracellular interferon γ were increased, whereas those with IL-4 remained similar between the groups. Therefore, it appears the adoptive transfer of fibrocytes improves sepsis survival, lowers bacterial burden, and promotes the proliferation of splenic T cells with a T(H)1 phenotype. These results confirm the immunomodulatory effects of exogenous, tissue-derived fibrocytes in sepsis and suggest their potential in cell therapy.

Effects of tramadol and buprenorphine on select immunologic factors in a cecal ligation and puncture model.

Sepsis research relies on animal models. The models that most closely resemble clinical disease, such as cecal ligation and puncture, require surgery. After surgery, analgesics may not be included in experimental protocols because of concern over effects on inflammatory responses. This often generates animal welfare controversies within institutions; however, there are no scientific studies directly addressing the effects of analgesics on surgical models of sepsis. The purpose of this study was to characterize the effects of opioids on key parameters used in sepsis research.Female ICR mice were divided into four treatment groups (Ringer's lactate solution, high- or low-dose tramadol,buprenorphine) for 3-week mortality studies (n = 12 per treatment). Experimental groups were then repeated, and mice were killed at 12, 24, and 48 h postsurgery for cell counts, differentials, and cytokine levels in blood, peritoneum, and airways. Mortality studies demonstrated no significant differences between controls and any treatment group. However,significant differences were noted between buprenorphine and high-dose tramadol, revealing more and later deaths with tramadol. For comparison of immune parameters, Mann-Whitney U or Student t test was performed, emphasizing comparisons between treatment and control. Although several results were significant, comparisons between control and any treatment group yielded no differences that remained consistently apparent during the observation period. Again,differences were observed between the treatments. The results suggest that judicious and limited use of some analgesics may not dramatically affect the outcome of similarly conducted cecal ligation and puncture studies when compared with those not using analgesics. However, when different analgesics are used, comparisons between studies may be complicated.

High-mobility group box 1 as a surrogate prognostic marker in dogs with systemic inflammatory response syndrome.

OBJECTIVE: To evaluate various surrogate markers associated with the inflammatory and counter-inflammatory responses with respect to mortality in dogs with systemic inflammatory response syndrome (SIRS). DESIGN: Prospective observational study. SETTING: Veterinary Teaching Hospital. ANIMALS: Twenty-eight dogs with naturally occurring diseases and SIRS from January 2007 to May 2009. INTERVENTIONS: Upon admission to the veterinary hospital, history and baseline data from the physical examination, including parameters previously defined for meeting SIRS criteria, were documented. Heparinized blood samples were collected and plasma cytokines interleukin-6 (IL-6), IL-10, and high-mobility group box 1 (HMGB1) were measured by sandwich ELISA. MEASUREMENTS AND MAIN RESULTS: In nonsurvivors, median plasma HMGB1 concentrations (0.718 microg/L, interquartile range [IQR]; 0.300-1.626 microg/L) and the ratio of HMGB1 to IL-10 (2.236, IQR; 0.972-5.367) were significantly increased as compared with those found in survivors (0.300 microg/L, IQR; 0.300-0.312 microg/L for HMGB1; 1.017, IQR; 0.862-1.126 for the ratio of HMGB1 to IL-10, P=0.007 and 0.024, respectively). Plasma IL-6, IL-10, and the ratio of IL-6 to IL-10 were not significantly different between groups. Among the parameters studied, HMGB1 and the ratio of HMGB1 to IL-10 performed the best in discriminating outcome in dogs with SIRS according to receiver operator characteristic curve analysis. CONCLUSIONS: Increases in plasma HMGB1 concentration and the ratio of HMGB1 to IL-10 may predict poorer outcomes in dogs with SIRS. The approach described may lead to reliable prognostic biomarkers and new therapeutic concepts in the study of SIRS in dogs.

Functional contribution of CXCR2 to lung injury after aspiration of acid and gastric particulates.

The effects of individual ELR+ CXC chemokines have been documented in experimental models of acid aspiration. However, aspiration lung injury would be influenced by the combined effects of these chemokines and other factors related to their function. Therefore, the role of the chemokine receptor CXCR2 was examined in lung injury induced by aspiration of acid and acid with gastric particulates. Anesthetized mice were given intratracheal injections of saline, acid solution, or acid containing gastric particles. Within 6 h, bronchoalveolar lavage fluid neutrophils and albumin increased relative to the severity of the insult. Immunohistochemistry and RT-PCR demonstrated striking increases in pulmonary expression of CXCR2 after aspiration. In CXCR2-deficient mice, neutrophil recruitment to airways was significantly reduced after aspiration of either acid or acid with particles. However, lung injury scores were unaffected in Ccr2-/- mice in the acid + particles group. Esterase-stained lung tissue demonstrated that focal aggregates of inflammatory cells contained neutrophils in the Ccr2-/- mice. These studies suggest CXCR2 and its ligands are dominant mediators of neutrophil recruitment to airways after aspiration. However, CXCR2-independent mechanisms recruit neutrophils into areas of cellular aggregation after aspiration of acidified gastric particulates.

Pulmonary inflammation and airway hyperresponsiveness in a mouse model of asthma complicated by acid aspiration.

Several studies have indicated a strong association between asthma and aspiration of stomach contents. However, the complex association between these inflammatory processes has not been studied extensively in animal models. In the present study, we developed an animal model to evaluate the inflammatory cell, chemokine, and airway responses to asthma complicated by aspiration. The model was produced by sensitizing mice to cockroach allergens from house-dust extracts. Mice with asthma-like airway responses then were inoculated intratracheally with either an acidic solution or saline. Acid aspiration increased airway hyperresponsiveness in mice with asthma for at least 8 h. After 6 h, the combined injury caused an additive, not synergistic, increase in airway hyperresponsiveness and neutrophil recruitment to the airways. Although cysteinyl leukotrienes in bronchoalveolar lavage fluid were higher after acid aspiration, treatment with a receptor antagonist before aspiration did not diminish airway hyperresponsiveness. Vagal mechanisms reportedly mediate airway responses in acid aspiration; however, pretreatment with an anticholinergic agent did not reduce airway responses to acid. These results are consistent with an effective model of the acute effects of aspiration on the allergic lung. Further studies could examine how various forms of aspiration influence the severity of asthma.

Modeling sepsis in the laboratory: merging sound science with animal well-being.

Despite impressive advances in biomedical research, few noteworthy breakthroughs have been made in the treatment of sepsis during the past several decades. This stalemate is primarily due to the intricate and heterogenic nature of the systemic immune responses characterized as the sepsis syndrome. In general, such complexity must be approached with in vivo models. Several animal models have been described, suggesting that none adequately address all of the pressing needs in sepsis research. The most clinically applicable models involve a localized infection, such as surgically induced polymicrobial sepsis, that gradually propagates a systemic immune response. Because relevant models must mimic a severe and chronic syndrome, animal well-being is often a concern in sepsis research. A balance between the needs of sepsis research and animal welfare can only be achieved through knowledge of the strengths and weaknesses of and alternatives to in vivo sepsis models.

Evaluation of saphenous venipuncture and modified tail-clip blood collection in mice.

The purpose of this study was to evaluate the effects of 2 methods of blood collection in unanesthetized mice. The saphenous venipuncture method was compared with a modified tail-clip technique that requires minimal restraint. Mice were evaluated through behavioral observation and plasma corticosterone levels. The results showed that the 2 methods produced similar corticosterone responses and that the tail-clip method produced fewer behavioral reactions. In addition, the effects of saphenous venipuncture method appeared to be dependent on the handler's technical expertise. When a series of 4 blood collections were performed over 1 wk, the 2 methods yielded similar corticosterone levels that did not increase over time. Some of the behavioral signs appeared to increase over the series of blood collections obtained by the saphenous venipuncture method. Serial complete blood counts showed that the tail vessels yielded higher total white blood cell, neutrophil, and lymphocyte counts than did the saphenous vein. Neither method appeared to cause stress-associated changes in the leukogram after serial blood collection. Overall, the effects of modified tail-clip method were similar to those of the saphenous venipuncture method in unanesthetized mice.