Reuse of Disposable Isolation Gowns in Rodent Facilities during a Pandemic.

Reuse of disposable personal protective equipment is traditionally discouraged, yet in times of heightened medical applications such as the SARS CoV-2 pandemic, it can be difficult to obtain. In this article we examine the reuse of disposable gowns with respect to still providing personnel protection. XR7, a fluorescent powder, was used to track contamination of gowns after manipulation of rodent cages. Mouse cages were treated with XR7 prior to manipulations. Disposable gowns were labeled for single person use and hung in common procedure spaces within the vivarium between usages. A simulated rack change of 140 cages was completed using XR7-treated cages. One individual changed all cages with a break occurring after the first 70 cages, requiring the gown to be removed and reused once. To simulate research activities, 5 individuals accessed 3 XR7-treated cages daily for 5 d. Each mouse in the XR7-treated cages was manipulated at least once before returning cages to the housing room. Disposable gowns were reused 5 times per individual. Gowns, gloves, clothing, bare arms, and hands were scanned for fluorescence before and after removing PPE. Fluorescence was localized to gloves and gown sleeves in closest contact with animals and caging. No fluorescence was detected on underlying clothing, or bare arms and hands after removing PPE. Fluorescence was not detected in procedure spaces where gowns were hung. The lack of fluorescence on personnel or surfaces indicate that gowns can be reused 1 time for routine husbandry tasks and up to 5 times for research personnel. A method for decontamination of used gowns using Vaporized Hydrogen Peroxide (VHP) was also validated for use in areas where animals are considered high risk such as quarantine, or for fragile immunocompromised rodent colonies.

Metabolic shifts modulate lung injury caused by infection with H1N1 influenza A virus.

Influenza A virus (IAV) infection alters lung epithelial cell metabolism in vitro by promoting a glycolytic shift. We hypothesized that this shift benefits the virus rather than the host and that inhibition of glycolysis would improve infection outcomes. A/WSN/33 IAV-inoculated C57BL/6 mice were treated daily from 1 day post-inoculation (d.p.i.) with 2-deoxy-d-glucose (2-DG) to inhibit glycolysis and with the pyruvate dehydrogenase kinase (PDK) inhibitor dichloroacetate (DCA) to promote flux through the TCA cycle. To block OXPHOS, mice were treated every other day from 1 d.p.i. with the Complex I inhibitor rotenone (ROT). 2-DG significantly decreased nocturnal activity, reduced respiratory exchange ratios, worsened hypoxemia, exacerbated lung dysfunction, and increased humoral inflammation at 6 d.p.i. DCA and ROT treatment normalized oxygenation and airway resistance and attenuated IAV-induced pulmonary edema, histopathology, and nitrotyrosine formation. None of the treatments altered viral replication. These data suggest that a shift to glycolysis is host-protective in influenza.

Postexposure Liponucleotide Prophylaxis and Treatment Attenuates Acute Respiratory Distress Syndrome in Influenza-infected Mice.

There is an urgent need for new drugs for patients with acute respiratory distress syndrome (ARDS), including those with coronavirus disease (COVID-19). ARDS in influenza-infected mice is associated with reduced concentrations of liponucleotides (essential precursors for de novo phospholipid synthesis) in alveolar type II (ATII) epithelial cells. Because surfactant phospholipid synthesis is a primary function of ATII cells, we hypothesized that disrupting this process could contribute significantly to the pathogenesis of influenza-induced ARDS. The goal of this study was to determine whether parenteral liponucleotide supplementation can attenuate ARDS. C57BL/6 mice inoculated intranasally with 10,000 plaque-forming units/mouse of H1N1 influenza A/WSN/33 virus were treated with CDP (cytidine 5'-diphospho)-choline (100 µg/mouse i.p.) ± CDP -diacylglycerol 16:0/16:0 (10 µg/mouse i.p.) once daily from 1 to 5 days after inoculation (to model postexposure influenza prophylaxis) or as a single dose on Day 5 (to model treatment of patients with ongoing influenza-induced ARDS). Daily postexposure prophylaxis with CDP-choline attenuated influenza-induced hypoxemia, pulmonary edema, alterations in lung mechanics, impairment of alveolar fluid clearance, and pulmonary inflammation without altering viral replication. These effects were not recapitulated by the daily administration of CTP (cytidine triphosphate) and/or choline. Daily coadministration of CDP-diacylglycerol significantly enhanced the beneficial effects of CDP-choline and also modified the ATII cell lipidome, reversing the infection-induced decrease in phosphatidylcholine and increasing concentrations of most other lipid classes in ATII cells. Single-dose treatment with both liponucleotides at 5 days after inoculation also attenuated hypoxemia, altered lung mechanics, and inflammation. Overall, our data show that liponucleotides act rapidly to reduce disease severity in mice with severe influenza-induced ARDS.

ATP catabolism by tissue nonspecific alkaline phosphatase contributes to development of ARDS in influenza-infected mice.

Influenza A viruses are highly contagious respiratory pathogens that are responsible for significant morbidity and mortality worldwide on an annual basis. We have shown previously that influenza infection of mice leads to increased ATP and adenosine accumulation in the airway lumen. Moreover, we demonstrated that A1-adenosine receptor activation contributes significantly to influenza-induced acute respiratory distress syndrome (ARDS). However, we found that development of ARDS in influenza-infected mice does not require catabolism of ATP to adenosine by ecto-5'-nucleotidase (CD73). Hence, we hypothesized that increased adenosine generation in response to infection is mediated by tissue nonspecific alkaline phosphatase (TNAP), which is a low-affinity, high-capacity enzyme that catabolizes nucleotides in a nonspecific manner. In the current study, we found that whole lung and BALF TNAP expression and alkaline phosphatase enzymatic activity increased as early as 2 days postinfection (dpi) of C57BL/6 mice with 10,000 pfu/mouse of influenza A/WSN/33 (H1N1). Treatment at 2 and 4 dpi with a highly specific quinolinyl-benzenesulfonamide TNAP inhibitor (TNAPi) significantly reduced whole lung alkaline phosphatase activity at 6 dpi but did not alter TNAP gene or protein expression. TNAPi treatment attenuated hypoxemia, lung dysfunction, histopathology, and pulmonary edema at 6 dpi without impacting viral replication or BALF adenosine. Treatment also improved epithelial barrier function and attenuated cellular and humoral immune responses to influenza infection. These data indicate that TNAP inhibition can attenuate influenza-induced ARDS by reducing inflammation and fluid accumulation within the lung. They also further emphasize the importance of adenosine generation for development of ARDS in influenza-infected mice.

Can images of pain enhance patient-clinician rapport in pain consultations?

A variety of treatment outcomes in chronic pain are influenced by patient-clinician rapport. Patients often report finding it difficult to explain their pain, and this potential obstacle to mutual understanding may impede patient-clinician rapport. Previous research has argued that the communication of both patients and clinicians is facilitated by the use of pain-related images in pain assessments. This study investigated whether introducing pain-related images into pain assessments would strengthen various components of patient-clinician rapport, including relative levels of affiliation and dominance, and interpersonal coordination between patient and clinician behaviour. Videos of 35 pain assessments in which pain images were present or absent were used to code behavioural displays of patient and clinician rapport at fixed intervals across the course of the assessment. Mixed modelling was used to examine patterns of patient and clinician affiliation and dominance with consultation type (Image vs Control) as a moderator. When pain images were present, clinicians showed more affiliation behaviour over the course of the consultation and there was greater correspondence between the affiliation behaviour of patient and clinician. However, relative levels of patient and clinician dominance were unaffected by the presence of pain images in consultations. Additional analyses revealed that clinicians responded directly to patients' use of pain images with displays of affiliation. Based on the results of this study, we recommend further investigation into the utility and feasibility of incorporating pain images into pain assessments to enhance patient-clinician communication.

Effect of Ventilated Caging on Water Intake and Loss in 4 Strains of Laboratory Mice.

Food availability, temperature, humidity, strain, and caging type all affect water consumption by mice. Measurement of transepidermal water loss (TEWL) is a new technique for the quantification of water turnover in mice. To understand water turnover in common strains of adult mice, male and female SCID, SKH, C57BL/6, and FVB mice were housed in same-sex groups of 5 animals in static cages or IVC. Body weight, TEWL, urine osmolality, and water consumption of mice and intracage temperature and humidity were measured every 48 h for comparison. Static cages were monitored for 7 d and IVC for 14 d before cage change. Female SCID, FVB, and C57 mice drank less water than did their male counterparts. Male and female SCID, SKH, and FVB mice in IVC drank less water and had higher urine osmolality than did those in static cages. In SCID and SKH mice, TEWL paralleled water consumption. C57 mice in static cages drank less water, had lower urine osmolality, and had less TEWL than did those in IVC. Temperature and humidity within the cage was higher than the macroenvironmental levels for all housing conditions, mouse strains, and sexes. Temperatures within IVC ranged from 76.6 to 81.4 °F compared with 69±0.4 °F in the room. Humidity within IVC ranged from 68% to 79% compared with 27.o%±2.7% within the room. These data demonstrate that mouse strain and housing conditions significantly influence water balance and indicate that macroenvironmental measurements do not always reflect the intracage environment.

Institutional Animal Care and Use Committee Considerations Regarding the Use of Virus-Induced Carcinogenesis and Oncolytic Viral Models.

The use of virus-induced carcinogenesis and oncologic experimental animal models is essential in understanding the mechanisms of cancer development to advance prevention, diagnosis, and treatment methods. The Institutional Animal Care and Use Committee (IACUC) is responsible for both the complex philosophical and practical considerations associated with animal models of cancer. Animal models of cancer carry their own unique issues that require special consideration from the IACUC. Many of the considerations to be discussed apply to cancer models in general; specific issues related to viral carcinogenesis or oncolytic viruses will be specifically discussed as they arise. Responsible animal use integrates good science, humane care, and regulatory compliance. To meet those standards, the IACUC, in conjunction with the research investigator and attending veterinarian, must address a wide range of issues, including animal model selection, cancer model selection, humane end point considerations, experimental considerations, postapproval monitoring, reporting requirements, and animal management and personnel safety considerations.

Implications of natural occlusion of ventilated racks on ammonia and sanitation practices.

Examination of ventilated rat racks prior to semiannual sanitation revealed silicone nozzles and ventilation ports that were partially or completely occluded with granular debris. We subsequently sought to document performance standards for rack sanitation and investigate the effect of ventilation port occlusion on rack function and animal husbandry practices. We hypothesized that individually ventilated cages with occluded airflow would require more frequent cage changes, comparable to those for static cages (that is, every 3 to 4 d). Sprague-Dawley rats were housed under one of 4 conditions: no airflow occlusion, occluded air-supply inlet, occluded air-exhaust outlet, and occlusion of both inlet and outlet. Cages were changed when daily ammonia concentration exceeded 20 ppm or after 14 d had elapsed. Most cages with unoccluded or partial airflow occlusion remained below the 20 ppm limit until day 12 or 13. Cages with occlusion of both inlet and outlet exceeded 20 ppm ammonia by as early as day 5. Airflow was significantly lower in cages with occlusion of both inlet and outlet airflow. Weekly inspection revealed that occlusion of ventilation ports was detectable by 3 mo after semiannual sanitation. This study demonstrates that silicone nozzles should be removed prior to rack sanitation to improve the effectiveness of cleaning ventilation ports and nozzles. While the rack is in use, silicone nozzles and ventilation ports should be inspected regularly to identify occlusion that is likely to diminish environmental quality in the cage. Intracage ammonia levels are significantly higher when both inlet and outlet airflow are occluded.

Heterozygosity for the F508del mutation in the cystic fibrosis transmembrane conductance regulator anion channel attenuates influenza severity.

BACKGROUND: Seasonal and pandemic influenza are significant public health concerns. Influenza stimulates respiratory epithelial Cl(-) secretion via the cystic fibrosis transmembrane conductance regulator (CFTR). The purpose of this study was to determine the contribution of this effect to influenza pathogenesis in mice with reduced CFTR activity. METHODS: C57BL/6-congenic mice heterozygous for the F508del CFTR mutation (HET) and wild-type (WT) controls were infected intranasally with 10 000 focus-forming units of influenza A/WSN/33 (H1N1) per mouse. Body weight, arterial O2 saturation, and heart rate were monitored daily. Pulmonary edema and lung function parameters were derived from ratios of wet weight to dry weight and the forced-oscillation technique, respectively. Levels of cytokines and chemokines in bronchoalveolar lavage fluid were measured by enzyme-linked immunosorbent assay. RESULTS: Relative to WT mice, influenza virus-infected HET mice showed significantly delayed mortality, which was accompanied by attenuated hypoxemia, cardiopulmonary dysfunction, and pulmonary edema. However, viral replication and weight loss did not differ. The protective HET phenotype was correlated with exaggerated alveolar macrophage and interleukin 6 responses to infection and was abrogated by alveolar macrophage depletion, using clodronate liposomes. CONCLUSIONS: Reduced CFTR expression modulates the innate immune response to influenza and alters disease pathogenesis. CFTR-mediated Cl(-) secretion is therefore an important host determinant of disease, and CFTR inhibition may be of therapeutic benefit in influenza.

Processing and treatment of corncob bedding affects cage-change frequency for C57BL/6 mice.

The goal of this study was to evaluate the effectiveness of a new proprietary processed corncob bedding material (PCC)compared with standard corncob in ventilated and static mouse housing systems. Intracage ammonia levels, bacterial growth, and absorptive capacity of bedding were measured for cages of C57BL/6 mice under nonautoclaved and autoclaved conditions on static and ventilated racks in a barrier facility. Ammonia concentration was measured daily, and cages were removed from the study when measurements reached or exceeded 25 ppm. Bacterial growth in bedding was quantified and speciated before exposure to mice and at the time of cage removal. The absorptive capacity of all bedding material was determined under autoclaved and nonautoclaved conditions. Ventilated cages with PCC or autoclaved corncob took longer to reach ammonia concentrations of 25 ppm than did those with corncob or autoclaved PCC; PCC-filled cages remained below 25 ppm NH3 for at least 3 wk. The type of bedding material did not affect the number of days required to reach 25 ppm in static cages. Compared with other bedding types in the absence of mice, 1/4-in. PCC had a lower and 1/8-in. corncob a higher bacterial load. Autoclaving altered the absorptive capacity of 1/4-in. bedding materials, and for 1/8-in. bedding, corncob was more absorptive than PCC regardless of autoclaving. The results of this study indicate that PCC is comparable to autoclaved corncob in controlling intracage ammonia levels, and a cage-change interval of 3 wk is possible when ventilated cages are used with this bedding.

Effectiveness of shoe covers for bioexclusion within an animal facility.

The personal protective equipment (PPE) required for entry into rodent barrier rooms often includes a hair bonnet, face mask, disposable gown, gloves, and shoe covers. Traditionally, shoe covers have been considered essential PPE for maintaining a 'clean' animal room. The introduction of microisolation caging and ventilated rack housing prompted us to reevaluate the contribution of shoe covers to bioexclusion. Contamination powder that fluoresces under black light was to track particle dispersal on the floor and personnel. The test mouse room contained a ventilated microisolation rack and biosafety cabinet. Powder was applied directly inside or outside the animal room doorway. PPE with or without shoe covers was donned outside of the animal room doorway and discarded on exiting. Participants either were scanned on entry into the room for the presence of florescence or asked to complete a simulated standard animal room activity while wearing full PPE. Animal rooms were scanned for florescence after exit of participants. All participants donning shoe covers fluoresced in multiple areas, primarily on gloves and gowns. Shoe covers had no effect on the spread of powder in normal traffic patterns, with no powder detected within caging. Powder also was used to determine the distance substances could be carried on the floor from building entry points. Results indicate that shoe covers do not improve (and actually may compromise) bioexclusion. Donning of shoe covers offers a potential for contamination of personnel from contact with shoe bottoms.

Comparison of methods for detection of pinworms in mice and rats.

Though pinworm infestation remains common in laboratory rodent colonies, there is little information regarding current practices for pinworm detection and their relative efficacy. The authors surveyed research institutions to find out the prevalence of pinworm infestations and the detection methods they used. They also tested mice and rats from colonies that were known to be infested with Syphacia sp. and compared the following detection methods: perianal tape test, fecal flotation, fecal concentration, cecal content examination, cecal flotation and histological examination. Though the different methods yielded comparable efficacy overall, the authors recommend using more than one type of test to increase detection potential.

Post-infection A77-1726 blocks pathophysiologic sequelae of respiratory syncytial virus infection.

Despite respiratory syncytial virus (RSV) bronchiolitis remaining the most common cause of lower respiratory tract disease in infants worldwide, treatment has progressed little in the past 30 years. The aim of our study was to determine whether post-infection administration of de novo pyrimidine synthesis inhibitors could prevent the reduction in alveolar fluid clearance (AFC) and hypoxemia that occurs at Day 2 after intranasal infection of BALB/c mice with RSV. BALB/c mice were infected intranasally with RSV strain A2. AFC was measured in anesthetized, ventilated mice after instillation of 5% bovine serum albumin into the dependent lung. Post-infection systemic treatment with leflunomide has no effect on AFC. However, when added to the AFC instillate, leflunomide's active metabolite, A77-1726, blocks RSV-mediated inhibition of AFC at Day 2. This block is reversed by uridine (which allows pyrimidine synthesis via the scavenger pathway) and not recapitulated by genistein (which mimics the tyrosine kinase inhibitor effects of A77-1726), indicating that the effect is specific for the de novo pyrimidine synthesis pathway. More importantly, when administered intranasally at Day 1, A77-1726, but not its vehicle dimethyl sulfoxide, maintains its beneficial effect on AFC and lung water content until Day 2. Intranasal instillation of A77-1726 at Day 1 also reduces bronchoalveolar lavage nucleotide levels, lung inflammation, and hypoxemia at Day 2 without impairing viral replication at Day 2 or viral clearance at Day 8. Post-infection intranasal or aerosolized treatment with pyrimidine synthesis inhibitors may provide symptomatic relief from the pathophysiologic sequelae of impaired AFC in children with RSV bronchiolitis.

The impact of organizational culture on clinical managers' organizational commitment and turnover intentions.

OBJECTIVES: The purpose was to investigate managers' perceptions of organizational culture and attitudinal and behavioral reactions during and after restructuring, and to test a model linking culture to outcome. BACKGROUND: Healthcare reform has altered the work environment, but few studies have documented the impact of system changes on managers responsible for clinical services. METHODS: Survey data were collected from clinical managers (N = 104; 99) employed by 3 institutional boards in Newfoundland and Labrador in 2000 and 2002. Response rates were 57.6% and 47.7%, respectively. RESULTS: For both periods, most variable ratings were in the low range and depicted moderately, positive intercorrelations. Select culture variables, trust, and job satisfaction emerged as significant predictors of commitment. Although culture and trust exerted inconsistent effects on intent, satisfaction remained a predictor over time. CONCLUSION: The findings support the negative impact of reform on clinical managers, and the strong link between positive ratings of culture, trust, and satisfaction, and greater commitment and intent to stay. Greater attention should focus on promoting more positive cultures and work-related attitudes, and less turnover intentions.

Respiratory syncytial virus induces insensitivity to beta-adrenergic agonists in mouse lung epithelium in vivo.

Respiratory syncytial virus (RSV) is the most common cause of bronchiolitis in infants and children worldwide. We wished to determine whether intratracheal administration of beta-agonists improved alveolar fluid clearance (AFC) across the distal respiratory epithelium of RSV-infected mice. Following intranasal infection with RSV strain A2, AFC was measured in anesthetized, ventilated BALB/c mice by instillation of 5% BSA into the dependent lung. We found that direct activation of protein kinase A by forskolin or 8-bromo-cAMP increased AFC at day 2 after infection with RSV. In contrast, short- and long-acting beta-agonists had no effect at either day 2 or day 4. Insensitivity to beta-agonists was not a result of elevated plasma catecholamines or lung epithelial cell beta-adrenergic receptor degradation. Instead, RSV-infected mice had significantly higher levels of phosphorylated PKCzeta in the membrane fractions of their lung epithelial cells. In addition, insensitivity to beta-agonists was mediated in a paracrine fashion by KC (the murine homolog of CXCL8) and reversed by inhibition of either PKCzeta or G protein-coupled receptor kinase 2 (GRK2). These results indicate that insufficient response to beta-agonists in RSV may be caused, at least in part, by impaired beta-adrenergic receptor signaling, as a consequence of GRK2-mediated uncoupling of beta-adrenergic receptors from adenylyl cyclase.