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Dental procedures often produce aerosols and spatter, which have the potential to transmit pathogens such as severe acute respiratory syndrome coronavirus 2. The existing literature is limited.Aerosols and spatter were generated from an ultrasonic scaling procedure on a dental manikin and characterized via 2 optical imaging methods: digital inline holography and laser sheet imaging. Capture efficiencies of various aerosol mitigation devices were evaluated and compared.The ultrasonic scaling procedure generated a wide size range of aerosols (up to a few hundred μm) and occasional large spatter, which emit at low velocity (mostly3 m/s). Use of a saliva ejector and high-volume evacuator (HVE) resulted in overall reductions of 63% and 88%, respectively, whereas an extraoral local extractor (ELE) resulted in a reduction of 96% at the nominal design flow setting.The study results showed that the use of ELE or HVE significantly reduced aerosol and spatter emission. The use of HVE generally requires an additional person to assist a dental hygienist, whereas an ELE can be operated hands free when a dental hygienist is performing ultrasonic scaling and other operations.An ELE aids in the reduction of aerosols and spatters during ultrasonic scaling procedures, potentially reducing transmission of oral or respiratory pathogens like severe acute respiratory syndrome coronavirus 2. Position and airflow of the device are important to effective aerosol mitigation.

Influenza is a common and contagious respiratory disease caused by influenza A, B, C, and D viruses (IAV, IBV, ICV, and IDV). A multiplex real-time RT-PCR assay was developed for simultaneous detection of IAV, IBV, ICV, and IDV. The assay was designed to target unique sequences in the matrix gene of IBV and ICV, the RNA polymerase subunit PB1 of IDV, and combined with USDA and CDC IAV assays, both target the matrix gene. The host 18S rRNA gene was included as an internal control. In silico analyses indicated high strain coverages: 97.9% for IBV, 99.5% for ICV, and 100% for IDV. Transcribed RNA, viral isolates and clinical samples were used for validation. The assay specifically detected target viruses without cross-reactivity, nor detection of other common pathogens. The limit of detection was approximately 30 copies for each viral RNA template, which was equivalent to a threshold cycle value of ~37.

Highlights • A multiplex real-time PCR assay was developed for PCV2 and PCV3 detection and differentiation. • Sensitivity was determined by standard curves with viral isolates and positive diagnostic samples. • Specificity was determined by testing common swine viruses. • The assay was validated on 336 swine diagnostic samples. • Limited data indicated that PCV3 has higher prevalence than PCV2 in the US., A multiplex quantitative real-time polymerase chain reaction (mqPCR) assay was developed and validated for the detection and differentiation of porcine circovirus type 3 (PCV3) and type 2 (PCV2) strains. The assay coverage was 97.9% (184/188) for PCV3 and 99.1% (1889/1907) for PCV2 sequences that were available from the current GenBank database. The PCR amplification efficiencies were 98–99% for plasmids, and 92–96% for diagnostic samples, with correlation coefficients all greater than 0.99. The limit of detection (LOD) determined as plasmid copies per reaction was 17 for PCV3 and 14 for PCV2. The assay specifically detected the targeted viruses without cross reacting to each other or to other common porcine viruses. Among 336 swine clinical samples collected in 2018, 101 (30.1%) were PCV3 positive, 56 (16.7%) were PCV2 positive and 18 (5.4%) were co-positives. Sixty selected PCV3 positives were confirmed by Sanger sequencing, and 53 of the 56 PCV2 positive samples were tested positive by another validated PCR assay.

Objective Seamless access to information about the individuals and organizations involved in the care of a specific patient (“care teams”) is crucial to effective and efficient care coordination. This is especially true for vulnerable and complex patient populations such as pediatric patients with special needs. Despite wide adoption of electronic health records (EHR), current EHR systems do not adequately support the visualization and management of care teams within and across health care organizations. Electronic health information exchange has the potential to address this issue. In the present study, we assessed the adequacy of available health information exchange data standards to support the information needs related to care coordination of complex pediatric patients. Methods We derived data elements from the information needs of clinicians and parents to support patient care teams; and mapped them to data elements in the Health Level Seven (HL7) Consolidated Clinical Document Architecture (C-CDA) standard and in the HL7 Fast Healthcare Interoperability Resources (FHIR) standard. We also identified additional C-CDA data elements and FHIR resources that include patients’ care team members. Results Information about care team members involved in patient care is generally well-represented in the C-CDA and FHIR specifications. However, there are gaps related to patients’ non-clinical events and care team actions. In addition, there is no single place to find information about care team members; rather, information about practitioners and organizations may be available in several different types of C-CDA data elements and FHIR resources. Conclusion Through standards-based electronic health information exchange, it appears to be feasible to build patient care team representations irrespective of the location of patient care. In order to gather care team information across disparate systems, exchange of multiple C-CDA documents and/or execution of multiple FHIR queries will be necessary. This approach has the potential to enable comprehensive patient care team views that may help improve care coordination.

This experiment was designed to study mechanisms affecting growth of in vivo-derived (IVD) and in vitro-produced (IVP) fetuses of cattle. Day-7 IVD or IVP cattle blastocysts were transferred to recipients, with pregnant females being slaughtered on Days 90 or 180 of gestation or allowed to undergo parturition. Uteri and contents were dissected and physically measured, and maternal and fetal plasma and amniotic and allantoic fluids were collected for IGF-1 and IGF-2 determinations, and IGFBP profile characterization. Transcripts for IGF-1 and IGF-2 mRNA in placental and fetal tissues, and IGF-1r and IGF-2r in placentomes were determined. There was a greater fetal weight in the IVP group, which was associated with greater IGF-1 and IGF-2 concentrations in maternal circulation, and changes in IGFBP profiles within fetal fluids. Day-90 IVP-derived fetuses were longer, had greater organ weights, larger placentomes, less placentome IGF-2r mRNA transcript, and greater maternal IGF-1 and IGF-2 concentrations than controls. On Day 180 and at parturition tissues from IVP-derived fetuses/calves were from larger uteri, with larger placentomes/fetal membranes, fetuses/calves weighed more, had greater fetal hepatic IGF-2 mRNA transcript, had less fetal plasma IGF-1 and greater allantoic IGF-2 concentrations, greater and lesser IGFBP activities in the allantoic and amniotic fluids, respectively, and greater glucose and fructose accumulation in fetal fluids. Components of the IGF system were differentially regulated not only according to the gestation period (Days 90 or 180) and fluid type (maternal or fetal plasma, amniotic or allantoic fluids), but also based on conceptus origin (IVP or IVD) in cattle.

Due to the adverse effects of fossil fuel use, it is becoming increasingly important to produce next-generation biofuels from renewable, sustainable sources. Filamentous N2-fixing strains of cyanobacteria have emerged as promising industrial microorganisms capable of producing a range biofuels and chemicals using CO2, water, and sunlight. In this study, a life cycle analysis (LCA) was conducted on a hypothetical production facility that uses a genetically engineered strain of filamentous cyanobacteria to produce the cyclic hydrocarbon limonene. Two scenarios were evaluated in which the only difference between the scenarios was the limonene productivity of the engineered cyanobacteria strain. In Scenario 1, the cyanobacterium was assumed to produce limonene at a rate of 1.8 mg/L/h, resulting in an annual production of 32,727 L/yr of limonene. In Scenario 2, limonene productivity was 55.5 mg/L/h, resulting in annual production of 1,000,000 L/yr. Both scenarios were assumed to produce the same amount of cellular biomass, that was converted to biogas by anaerobic digestion and the biogas was converted by gas turbines into electricity to power the facility. Excess electricity was assumed to be sold to the grid. The major environmental burdens of the facility, which were measured in eco-points and calculated based on the Eco-indicator 99 method, were the cyanobacteria nutrient supply (especially sodium nitrate) and the photobioreactor (PBR) electrical requirements. The lower output of limonene in Scenario 1 meant that less energy was required for product recovery, leaving more electricity for sale to the grid. Even though a higher limonene productivity will worsen the environmental profile of the process, both scenarios described in this study have less of a negative environmental impact than biodiesel production. This study strongly suggests both scenarios of the theoretical limonene production facility described herein holds great potential as a future solution for producing next-generation biofuels directly from solar energy.

Seneca Valley virus 1 (SVV-1) can cause vesicular disease that is clinically indistinguishable from foot-and-mouth disease, vesicular stomatitis and swine vesicular disease. SVV-1-associated disease has been identified in pigs in several countries, namely USA, Canada, Brazil and China. Diagnostic tests are required to reliably detect this emerging virus, and this report describes the development and evaluation of a novel real-time (r) reverse-transcription (RT) PCR assay (rRT-PCR), targeting the viral polymerase gene (3D) of SVV-1. This new assay detected all historical and contemporary SVV-1 isolates examined (n=8), while no cross-reactivity was observed with nucleic acid templates prepared from other vesicular disease viruses or common swine pathogens. The analytical sensitivity of the rRT-PCR was 0.79 TCID50/ml and the limit of detection was equivalent using two different rRT-PCR master-mixes. The performance of the test was further evaluated using pig nasal (n=25) and rectal swab samples (n=25), where concordant results compared to virus sequencing were generated for 43/50 samples. The availability of this assay, will enable laboratories to rapidly detect SVV-1 in cases of vesicular disease in pigs, negated for notifiable diseases, and could enable existing knowledge gaps to be investigated surrounding the natural epidemiology of SVV-1.

Porcine respirovirus 1 (PRV1) was first reported in the pig nasopharyngeal samples in Hong Kong in 2013. It has been widespread in US swine herds. Recently, PRV1 was also detected in South America and European countries. Currently, there is no validated diagnostic assay available for the detection of this virus. In this study, we developed a real-time reverse transcriptase quantitative PCR (RT-qPCR) assay targeting the hemagglutinin-neuraminidase (HN) gene for molecular diagnosis. The analytical sensitivity of this RT-qPCR assay was evaluated using in vitro transcribed RNA standard, and the limit of detection was 10 copies of viral RNA in a 20 μl reaction. No cross-reactivity was observed with nucleic acid prepared from common swine respiratory pathogens. The diagnostic performance of this assay was determined with 114 pig nasal swabs and 19 oral fluid samples with known PRV1 infection status. The RT-qPCR results were consistent with conventional RT-PCR and DNA sequencing of the HN gene, demonstrating a 100 % sensitivity and 100 % specificity. This assay was further applied to field samples. Among 310 nasal swab samples that were tested, 201 samples from 8 swine farms were PRV1 positive. No viremia was detected in PRV1 infected pigs using the available field samples. Nasal swab and oral fluid samples appear to be reliable for PRV1 detection with the RT-qPCR assay. Taken together, we developed and validated an RT-qPCR assay for accurate detection of PRV1 in nasal swab and oral fluid samples. It will be a useful tool for the rapid diagnosis of PRV1 infection and in aid of PRV1 epidemiological surveillance.

Pluripotent stem cells including embryonic stem cells (ESCs), embryonic germ cells (EGCs), and induced pluripotent stem cells (iPSCs) are capable of self-renew and limitlessly proliferating in vitro with undifferentiated characteristics. They are able to differentiate in vitro, spontaneously or responding to suitable signals, into cells of all three primary germ layers. Consequently, these pluripotent stem cells will be valuable sources for cell replacement therapy in numerous disorders. However, the promise of human ESCs and EGCs is cramped by the ethical argument about destroying embryos and fetuses for cell line creation. Moreover, there are still carcinogenic risks existing toward the goal of clinical application for human ESCs, EGCs, and iPSCs. Therefore, a suitable animal model for stem cell research will benefit the further development of human stem cell technology. The pigs, on the basis of their similarity in anatomy, immunology, physiology, and biochemical properties, have been wide used as model animals in the study of various human diseases. The development of porcine pluripotent stem cell lines will hold the opportunity to provide an excellent material for human counterpart to the transplantation in biomedical research and further development of cell-based therapeutic strategy.

The development of easily accessible tools for human immunophenotyping to classify patients into discrete disease endotypes is advancing personalized therapy. However, no systematic approach has been developed for the study of inflammatory lung diseases with often complex and highly heterogeneous disease etiologies. We have devised an internally standardized flow cytometry approach that can identify parallel inflammatory alveolar macrophage phenotypes in both the mouse and human lungs. In mice, lung innate immune cell alterations during endotoxin challenge, influenza virus infection, and in two genetic models of chronic obstructive lung disease could be segregated based on the presence or absence of CD11b alveolar macrophage upregulation and lung eosinophilia. Additionally, heightened alveolar macrophage CD11b expression was a novel feature of acute lung exacerbations in the SHIP-1−/− model of chronic obstructive lung disease, and anti-CD11b antibody administration selectively blocked inflammatory CD11bpos but not homeostatic CD11bneg alveolar macrophages in vivo. The identification of analogous profiles in respiratory disease patients highlights this approach as a translational avenue for lung disease endotyping and suggests that heterogeneous innate immune cell phenotypes are an underappreciated component of the human lung disease microenvironment. Supplementary information The online version of this article (doi:10.1038/mi.2015.84) contains supplementary material, which is available to authorized users.

Highlights • Bovine anaplasmosis has a wide distribution across the State of Kansas. • Several commonly promoted management practices associated with anaplasmosis were found to be associated with herd infection status. • Many commonly promoted management practices were found to not be associated with herd infection status., Bovine anaplasmosis is a hemolytic disease of cattle caused by Anaplasma marginale which can cause anemia, adult mortality, abortion, and performance reduction. The objectives of this study were to estimate herd-level infection prevalence of bovine anaplasmosis in Kansas cow-calf herds and assess management practices associated with herd infection status. Licensed Kansas veterinarians were randomly selected and provided clientele to generate randomly selected participant herds. Blood samples were collected from 10 mature cows during processing of 925 herds between October 1, 2016 and March 1, 2017. A management survey was completed by 780 herd-owners. Sample status was determined by competitive enzyme-linked immunosorbent assay (cELISA); operations indicating vaccination for anaplasmosis were tested with A.marginale-specific polymerase chain reaction (PCR). Survey data underwent logistic regression analysis for calculation of odds ratios and confidence intervals. The herd-level prevalence was 52.5 % of cow-calf herds. Prevalence ranged from 19.1 % of herds in Western Kansas to 87.3 % of herds in Eastern Kansas. Vaccinated herds were more likely (OR = 2.38; CI = 1.16–4.85; p = 0.02) to be positive compared to non-vaccinated herds, and herds that utilized insecticide ear-tags were more likely to be positive (OR = 1.9; CI = 1.42–2.55; p < 0.01) compared to herds which do not. Operations that prescribe-burned 21–50 % and >50 % of their pastures were more likely to be test positive, OR = 5.74 (CI = 3 .14–10.51; p < 0.01) and OR = 4.78 (CI = 2.33–10.17; p < 0.01), respectively, than operations that prescribe-burned

Distillers dried grains with soluble (DDGS) is the main byproduct of the corn-based ethanol industry. DDGS could be used as a renewable energy source through pyrolysis or gasification. However, biochar, the main byproduct, does not have commercial value. DDGS biochar can be utilized in supercapacitors when it is converted to activated carbon. Activated carbon samples were prepared at 950 and 1050 °C with KOH/ bio-char loading of 0.075 mol g −1 . Symmetrical supercapacitors were assembled using the activated carbon samples as electrode materials and 1 mol L −1 tetraethylammonium tetrafluoroborate (TEA BF 4 ) in acetonitrile as the electrolyte. The 950 °C activated carbon presented higher specific capacitance than the 1050 °C sample (150 F g −1 verse 70 F g −1 ). The capacitance was stable after 1000 cycles. More importantly, the capacitive performances of these activated carbons are comparable to ordered mesoporous carbons and commercial graphene. Thus, we highlight the success of preparing high performance supercapacitor electrode material from renewable biomass as well as the potential for generating advanced carbon materials from pyrolysis and gasification byproduct.

Neutrophilic inflammation persists in COPD despite best current therapies and it is particularly resistant to inhaled glucocorticosteroids. Persistent neutrophil activation not only contributes to matrix breakdown, but can maintain inflammation through the release of endogenous damage associated molecule patterns (DAMPs). Inhibiting excessive neutrophilic inflammation is challenging as many pathogen recognition receptors can initiate migration and the targeting of downstream signaling molecules may compromise essential host defense mechanisms. Here, we discuss new strategies to combat this inflammation in COPD by focusing on the anti-inflammatory role of ALX/FPR2 receptors. ALX/FPR2 is a promiscuous G-protein coupled receptor (GPCR) responding to lipid and peptide agonists that can either switch on acute inflammation or promote resolution of inflammation. We highlight this receptor as an emerging target in the pathogenesis of COPD because known ALX/FPR2 endogenous agonists are enriched in COPD. Serum Amyloid A (SAA) has recently been discovered to be abundantly expressed in COPD and is a potent ALX/FPR2 agonist that unlike almost all other inflammatory chemoattractants, is induced by glucocorticosteroids. SAA not only initiates lung inflammation via ALX/FPR2 but can allosterically modify this receptor so that it no longer transduces pro-resolving signals from endogenous lipoxins that would otherwise promote tissue healing. We propose that there is an imbalance in endogenous and microbial ALX/FPR2 receptor agonists in the inflamed COPD lung environment that oppose protective anti-inflammatory and pro-resolution pathways. These insights open the possibility of targeting ALX/FPR2 receptors using synthetic agonists to resolve persistent neutrophilic inflammation without compromising essential host defense mechanisms.

A series of N-aryl pyridinone inhibitors of p38 mitogen activated protein (MAP) kinase were designed and prepared based on the screening hit SC-25028 (1) and structural comparisons to VX-745 (5). The focus of the investigation targeted the dependence of potency and metabolic stability on the benzyloxy connectivity, the role of the C-6 position and the substitution pattern on the N-phenyl ring. Further optimization produced the highly selective and potent pyridinones 2 and 3. These inhibitors exhibited activity in both acute and chronic models of inflammation.

Background Ischemia/reperfusion (IR) injury is an unavoidable consequence of tissue transplantation or replantation that often leads to inflammation and cell death. Excessive complement activation following IR induces endothelial cell injury, altering vascular and endothelial barrier function causing tissue dysfunction. To mitigate the IR response, various systemic anti-complement therapies have been tried. Recently, we developed a localized therapy that uses biotinylated fusogenic lipid vesicles (BioFLVs) to first incorporate biotin tethers onto cell membranes, which are then used to bind therapeutic fusion proteins containing streptavidin (SA) resulting in the decoration of cell membranes. The therapy is applied in two steps using solutions delivered intra-arterially. Materials and Methods Alteration of formulation, concentration and duration of incubation of BioFLVs were conducted to demonstrate the ability of the system to modulate biotin tether incorporation in cultured cells. Using a rat hind limb model, the ability of BioFLVs to decorate endothelium of femoral vessels with FITC-labeled SA for 48 h of reperfusion was demonstrated. The feasibility of a BioFLV-based anti-complement therapy was tested in cultured cells using SA fused with vaccinia virus complement control protein (SA-VCP), a C3 convertase inhibitor. Human ovarian carcinoma (SKOV-3) cells were incubated with BioFLVs first and then with SA-VCP. To activate complement the cells were treated with a SKOV-3-specific antibody (trastuzumab) and incubated in human serum. Results Decoration of cells with SA-VCP effectively reduced complement deposition. Conclusions We conclude that BioFLV-mediated decoration of cell membranes with anti-complement proteins reduces complement activation and deposition in vitro and has the potential for application against inappropropriate complement activation in vivo .

Bronchiolitis obliterans syndrome (BOS) is the main long-term complication after lung transplantation. Previous studies indicate that neutrophil mobilization causes high protease concentrations in the lung allograft during BOS. This study assessed net protease activity and the functional aspect of proteases in BOS.The net gelatinase and net serine protease activity was assessed in bronchoalveolar lavage (BAL) fluid from 12 pairs of 24 lung allograft recipients with and without BOS, carefully selected from a larger cohort that was otherwise clinically matched. We determined the identity and total activity of gelatinases and concentrations of matrix metalloproteinases (MMP) 2 and 9, as well as the concentration of serine protease, neutrophil elastase (NE), and one major antiprotease, secretory leukocyte protease inhibitor (SLPI).Net gelatinase activity was substantially increased in BOS (n = 12), with total MMP-9 activity exceeding total MMP-2 activity (p0.01). Correspondingly, the total mean (interquartile range) concentration of MMP-9 was increased in BOS (62 [160] ng/ml) vs non-BOS (20 [24] ng/ml; p0.05), but not MMP-2 (BOS: 0.6 [0.7]; non-BOS: 0.6 [0.8] ng/ml, p = 0.23). Notably, net gelatinase activity correlated with MMP-9 (rho = 0.9, p0.01) and percentage of neutrophils (rho = 0.8, p0.01). Despite increased levels of NE and unaltered levels of SLPI, net serine protease levels remained unaltered, suggesting that NE does not contribute to BOS pathology.Our study supports that there is an unopposed increase in gelatinase activity in BOS, which in part is likely to be accounted for by MMP-9 from local neutrophils. No corresponding evidence was found for serine protease activity.

Interleukin (IL)-17 may play a critical role for the innate immune response in mammals. However, little is known about its production in T lymphocytes in comparison with other cells, in lung tissue and in the bronchoalveolar space in vivo. Even less is known about the effects of anti-inflammatory pharmacotherapy on this IL-17 production. In this study on mice we show that one single, intranasal exposure to endotoxin from Escherichia coli increases extracellular IL-17 protein in bronchoalveolar (BAL) samples during 3 days, and is accompanied by a local increase in neutrophils and other inflammatory cells. This endotoxin exposure also elevates IL-17 mRNA in lung tissue samples. Moreover, after endotoxin exposure, the absolute number of CD3-positive cells containing intracellular IL-17 protein is increased as well; from a moderate cell number in lung tissue samples and from virtually none in BAL samples; with the number in lung tissue exceeding that observed in BAL samples. Notably, we also demonstrate that among the cells that contain intracellular IL-17 protein after endotoxin exposure, the percentage of CD3-positive cells is similar to that of CD3-negative cells in lung tissue. In contrast, CD3-negative cells dominate among IL-17-containing cells in BAL samples. A high systemic dose of a glucocorticoid receptor agonist attenuates the endotoxin-induced increase in extracellular IL-17 protein in BAL samples, IL-17 mRNA in lung tissue samples, and in IL-17-containing CD3-positive cells in BAL and lung tissue samples. This is also true for the endotoxin-induced accumulation of neutrophils and other inflammatory BAL cells in vivo. A systemic dose of a calcineurin phosphatase inhibitor exerts a less complete and more selective effect on the endotoxin-induced increase in extracellular IL-17 protein and on neutrophils in BAL samples. In vitro, endotoxin also increases extracellular IL-17 protein in a co-culture of CD3-positive spleen cells and adherent mononuclear BAL cells; an increase that was inhibited by a glucocorticoid as well as by a calcineurin phosphatase inhibitor. In conclusion, endotoxin-induced IL-17 production and release from T lymphocytes originates from cells that reside in lung tissue and from cells that have been recruited to the bronchoalveolar space. In both compartments, there is also a substantial number of cells other than T lymphocytes that contain IL-17 after endotoxin exposure. The sustained IL-17 production from T lymphocytes and the associated neutrophil accumulation may be inhibited non-selectively through glucocorticoid receptor stimulation and more selectively through calcineurin phosphatase inhibition.

Many resources and methods are available for dissemination and promotion of the principles and value of evidence-based dentistry among colleagues.

The 5-lipoxygenase (5-LOX) pathway has been associated with a variety of inflammatory diseases including asthma, atherosclerosis, rheumatoid arthritis, pain, cancer and liver fibrosis. Several classes of 5-LOX inhibitors have been identified, but only one drug, zileuton, a redox inhibitor of 5-LOX, has been approved for clinical use. To better evaluate the efficacy of 5-LOX inhibitors for pharmacological intervention, a rat model was modified to test the in vivo efficacy of 5-LOX inhibitors. Inflammation was produced by adding carrageenan into a newly formed air pouch and prostaglandins produced. While macrophages and neutrophils are present in the inflamed pouch, little 5-LOX products are formed. Cellular 5-LOX activation was obtained by adding calcium ionophore (A23187) into the pouch thus providing a novel model to evaluate the efficacy and selectivity of 5-LOX inhibitors. Also, we described modifications to the in vitro 5-LOX enzyme and cell assays. These assays included a newly developed fluorescence-based enzyme assay, a 5-LOX redox assay, an ex vivo human whole blood assay and an IgE-stimulated rat mast cell assay, all designed for maximal production of leukotrienes. Zileuton and CJ-13,610, a competitive, non-redox inhibitor of 5-LOX, were evaluated for their pharmacological properties using these assays. Although both compounds achieved dose-dependent inhibition of 5-LOX enzyme activity, CJ-13,610 was 3-4 fold more potent than zileuton in all-assays. Evaluation of 5-LOX metabolites-by LC/MS/MS and ELISA confirmed that both compounds selectively inhibited all products downstream of 5-hydroperoxy eicosatetraenoic acid (5-HPETE), including 5-oxo-6,8,11,14-eicosatetraenoic acid (5-oxoETE), without inhibition of 12-lipoxygenase (12-LOX), 15-lipoxygenase (15-LOX), or cyclooxygenase (COX) products. In the rat air pouch model, oral dosing of CJ-13,610 and zileuton resulted in selective inhibition 5-LOX activity from pouch exudate and ex vivo rat whole blood with similar potency to in vitro assay. These data show that the rat air pouch model is a reliable and useful tool for evaluating in vivo efficacy of 5-LOX inhibitors and may aid in the development of the next generation of 5-LOX inhibitors, such as the non-redox inhibitors similar to CJ-13,610.

SummaryRespiratory viruses are associated with severe acute exacerbations of chronic obstructive pulmonary disease (COPD) in hospitalized patients. However, exacerbations are increasingly managed in the community, where the role of viruses is unclear. In community exacerbations, the causal association between viruses and exacerbation maybe confounded by random fluctuations in the prevalence of circulating respiratory viruses.Therefore, to determine whether viral respiratory tract infections are causally associated with community exacerbations, a time-matched case-control study was performed. Ninety-two subjects (mean age 72yrs), with moderate to severe COPD, (mean FEV1 40% predicted), were enrolled. Nasopharyngeal swabs for viral multiplex polymerase chain reaction and atypical pneumonia serology were obtained at exacerbation onset. Control samples were collected in synchrony, from a randomly selected stable patient drawn from the same cohort.In 99 weeks of surveillance, there were 148 exacerbations. Odds of viral isolation were 11 times higher in cases, than their time-matched controls (34 discordant case-control pairs; in 31 pairs only the case had virus and in three pairs only control). Picornavirus (26), influenza A (3), parainfluenza 1,2,3 (2), respiratory syncytial virus (1), and adenovirus (1) were detected in cases while adenovirus (1) and picornavirus (2) were detected in controls. In patients with moderate or severe COPD the presence of a virus in upper airway secretions is strongly associated with the development of COPD exacerbations. These data support the causative role of viruses in triggering COPD exacerbations in the community.

The term severe refractory asthma (SRA) in adults applies to patients who remain difficult to control despite extensive re-evaluation of diagnosis and management following an observational period of at least 6 months by a specialist. Factors that influence asthma control should be recognized and adequately addressed prior to confirming the diagnosis of SRA. This report presents statements according to the literature defining SRA in order address the important questions. Phenotyping SRA will improve our understanding of mechanisms, natural history, and prognosis. Female gender, obesity, and smoking are associated with SRA. Atopy is less frequent in SRA, but occupational sensitizers are common inducers of new-onset SRA. Viruses contribute to severe exacerbations and can persist in the airways for long periods. Inflammatory cells are in the airways of the majority of patients with SRA and persist despite steroid therapy. The T(H)2 immune process alone is inadequate to explain SRA. Reduced responsiveness to corticosteroids is common, and epithelial cell and smooth muscle abnormalities are found, contributing to airway narrowing. Large and small airway wall thickening is observed, but parenchymal abnormalities may influence airway limitation. Inhaled corticosteroids and bronchodilators are the mainstay of treatment, but patients with SRA remain uncontrolled, indicating a need for new therapies.

Non-homologous end joining (NHEJ) is the major DNA double-strand break (DSB) repair pathway in mammalian cells and is likely responsible for the non-homologous integration of transgenes. In higher eukaryotes, this pathway predominates over the homologous recombination (HR) pathway and therefore may account for the low level of HR events that occur in mammalian cells. We evaluated the effects of transient RNAi-induced down-regulation of key components of the NHEJ pathway in human HCT116 cells. Treatment with siRNA targeting Ku70 and Xrcc4 reduced corresponding protein levels by 80-90% 48h after transfection, with a return to normal levels by 96h. Additionally, down-regulation of Ku70 and Xrcc4 resulted in a concomitant depletion of both Ku70 and Ku86 proteins. Biological consequences of transient RNAi-mediated depletion of Ku70 and Xrcc4 included sensitization to gamma radiation and a significant decrease in the expression of a linear GFP reporter gene. The results highlight the possibility of a successful means to manipulate the NHEJ pathway by RNAi.

Appetite is regulated by a number of hypothalamic neuropeptides including neuropeptide Y (NPY), a powerful feeding stimulator that responds to feeding status, and drugs such as nicotine and cannabis. There is debate regarding the extent of the influence of obesity on hypothalamic NPY. We measured hypothalamic NPY in male Sprague-Dawley rats after short or long term exposure to cafeteria-style high fat diet (32% energy as fat) or laboratory chow (12% fat). Caloric intake and body weight were increased in the high fat diet group, and brown fat and white fat masses were significantly increased after 2 weeks. Hypothalamic NPY concentration was only significantly decreased after long term consumption of the high fat diet. Nicotine decreases food intake and body weight, with conflicting effects on hypothalamic NPY reported. Body weight, plasma hormones and brain NPY were investigated in male Balb/c mice exposed to cigarette smoke for 4 days, 4 and 12 weeks. Food intake was significantly decreased by smoke exposure (2.32+/-0.03g/24h versus 2.71+/-0.04g/24h in control mice (non-smoke exposed) at 12 weeks). Relative to control mice, smoke exposure led to greater weight loss, while pair-feeding the equivalent amount of chow caused an intermediate weight loss. Chronic smoke exposure, but not pair-feeding, was associated with decreased hypothalamic NPY concentration, suggesting an inhibitory effect of cigarette smoking on brain NPY levels. Thus, consumption of a high fat diet and smoke exposure reprogram hypothalamic NPY. Reduced NPY may contribute to the anorexic effect of smoke exposure.

An estimated 2,500 cases of listeriosis occur annually in the United States. Listeriosis is particularly severe among pregnant women and immunocompromised individuals. Little is known regarding the effect of the food matrix on the ability of L. monocytogenes to survive in the gastrointestinal tract and cause systemic infection. Mice were inoculated with various doses of L. monocytogenes in skim milk, Half & Half, or whipping cream to determine whether differences in milk fat content influence the ability of L. monocytogenes to survive passage through the gut and infect the liver or spleen. The number of fecal samples positive for L. monocytogenes increased with increasing doses of L. monocytogenes for all three vehicles. The number of L. monocytogenes cells isolated from liver or spleen of mice dosed with L. monocytogenes was not significantly different among treatment vehicles. Dose-response models revealed that as the dosage of L. monocytogenes was increased in different milk vehicles, the number of L. monocytogenes cells in liver or spleen also increased. Although fat content of food had no dose-dependent effect on L. monocytogenes infection in the murine gastrointestinal tract, we cannot discount the possibility that it may be a factor in L. monocytogenes infections of humans because of differences in the physiology of gastrointestinal tracts of mice and humans.

The ability of Listeria monocytogenes to survive and grow at refrigeration temperature in some ready to eat (RTE) poultry products is a public health concern. The inhibitory effect of clove oil (1% and 2%, v/w) applied to the surface of RTE chicken frankfurters was determined on seven strains of L. monocytogenes inoculated at low (102–103 cfu/g) or high cell numbers (104–106 cfu/g), and stored at 5 °C for 2 weeks or at 15 °C for 1 week. All strains of L. monocytogenes survived and grew on control frankfurters at 5 °C and 15 °C but growth was inhibited under both storage conditions in the presence of either 1% or 2% clove oil. Depending on the sensory considerations, the addition of clove oil to frankfurters may be an effective strategy to control L. monocytogenes in chicken frankfurters.

Chronic obstructive pulmonary disease (COPD) is characterised by persistent airflow limitation, neutrophilic inflammation, macrophage accumulation, and the production of cytokines, chemokines and proteases. Cigarette smoking is the major cause of COPD and there is currently no satisfactory therapy to help treat individuals with this disease. A better understanding of the cellular and molecular responses triggered by cigarette smoke may provide new molecular targets for the development of therapeutic agents. This brief review highlights some of the mouse models used to define the cellular, molecular and pathological consequences of cigarette smoke exposure.

Chronic obstructive pulmonary disease (COPD) is an incurable group of lung diseases characterised by progressive airflow limitation and loss of lung function, which lead to profound disability. It is mostly caused by cigarette smoke. Although COPD is one of the most prevalent diseases worldwide and its incidence is increasing, current therapies do little to improve the condition. Much current research focuses on strategies to halt the accelerated rate of decline in lung function that occurs in the disease. However, as most symptoms occur when the lungs are already extensively and irreversibly damaged, it is uncertain whether an agent able to slow or halt decline in lung function would actually provide relief to COPD patients. As lung function worsens, systemic comorbidities contribute markedly to disability. Loss of lean body mass (skeletal muscle) has recently been identified as a major determinant of disability in COPD and an independent predictor of mortality. In contrast to lung structure damage, skeletal muscle retains regenerative capacity in COPD. In this review, we discuss mechanisms of wasting in COPD, focusing on therapeutic strategies that might improve the health and productive life expectancy of COPD patients by improving skeletal muscle mass and function. Single or combination approaches exploiting the suppression of procatabolic inflammatory mediators, inhibition of ubiquitin ligases, repletion of anabolic hormones and growth factors, inhibition of myoblast apoptosis, remediation of systemic oxidative stress and promotion of repair, and regeneration via stimulation of satellite cell differentiation hold considerable therapeutic promise.

A study was done to determine if a free-living, bacterivorous nematode, Caenorhabditis elegans, migrates to bovine manure, turkey manure, composted bovine manure, composted turkey manure, and manure-amended soil inoculated with Salmonella Newport. Movement of the worm to lettuce, strawberries, and carrots was also studied. C. elegans moved most rapidly to turkey manure and strawberries, with 35% and 60% of worms, respectively, associating with samples within 30 min. Survival and reproduction of C. elegans in test materials were not affected by the presence of S. newport. Bovine manure and bovine manure compost inoculated with S. newport (8.6 log10 CFU/g) were separately placed in the bottom of a glass jar and covered with a layer of soil (5 cm) inoculated (50 worms/g) or not inoculated with C. elegans. A piece of lettuce, strawberry, or carrot was placed on top of the soil before jars were sealed and held at 20 -C for up to 10 days. In the system using soil inoculated with C. elegans, S. newport initially in bovine manure was detected on the surface of lettuce, strawberry, and carrot samples within 3, 1, and 1 days, respectively. The pathogen was detected on lettuce, strawberry, and carrot within 1, 7, and 1 days, respectively, when initially present in bovine manure compost. With one exception, the pathogen was not detected on the produce over the 10-day incubation period when C. elegans was not present in the soil. Results indicate that C. elegans has the potential for transporting S. newport in soil to the surface of preharvest fruits and vegetables in contact with soil. D 2005 Elsevier B.V. All rights reserved.

A study was undertaken to determine the persistence of Escherichia coli O157:H7 and salmonellae in the gut of a free-living nematode, Caenorhabditis elegans, as affected by temperature and relative humidity and to determine if infected worms transmit Salmonella enterica serotype Newport to progeny and uninfected worms. Worms were fed cells of a non-pathogenic strain of E. coli (OP50), E. coli O157:H7, S. enterica serotype Newport, and S. enterica serotype Poona, followed by incubating at 4, 20, or 37 degrees C for up to 5 days. Initial populations of ingested pathogens significantly increased by up to 2.93 log(10) cfu/worm within 1 day at 20 degrees C on K agar and remained constant for an additional 4 days. When worms were placed on Bacto agar, populations of ingested pathogens remained constant at 4 degrees C, decreased significantly at 20 degrees C, and increased significantly at 37 degrees C within 3 days. Worms fed E. coli OP50 or S. Newport were incubated at 4 or 20 degrees C at relative humidities of 33%, 75%, or 98% to determine survival characteristics of ingested bacteria. Fewer cells of the pathogens survived incubation at 33% relative humidity compared to higher relative humidities. Populations of ingested E. coli OP50 and S. Newport decreased by up to 1.65 and 3.44 log(10) cfu/worm, respectively, in worms incubated at 20 degrees C and 33% relative humidity. Placement together on K agar of adult worms, labeled with green fluorescent protein (gfp) in the pharynx area, that had ingested gfp-labeled S. Newport and uninfected wild type worms resulted in transfer of the pathogen to gut of wild type worms. S. Newport was isolated from C. elegans two generations removed from exposure to the pathogen. Results of these studies show that C. elegans may serve as a temporary reservoir of foodborne pathogens, and could perhaps be a vector for contaminating preharvest fruits and vegetables, thus potentially increasing the risk of enteric infections associated with consumption of raw produce.

Porcine embryonic germ (EG) cells share common features with porcine embryonic stem (ES) cells, including morphology, alkaline phosphatase activity and capacity for in vitro differentiation. Porcine EG cells are also capable of in vivo development by producing chimeras after blastocyst injection; however, the proportion of injected embryos that yield a chimera and the proportion of cells contributed by the cultured cells in each chimera are too low for practical use in genetic manipulation. Moreover, somatic, but not germ-line chimerism, has been reported from blastocyst injection using porcine ES or EG cells. To test whether efficiency of chimera production from blastocyst injection can be improved upon by changing the host embryo, we used as host embryos four groups according to developmental stage or length in culture: fresh 4-cell and 8-cell stage embryos subsequently cultured into blastocysts, fresh morulae, fresh blastocysts, and cultured blastocysts. Injection and embryo transfer of fresh and cultured blastocysts produced similar percentages of live piglets (17% versus 19%). Four piglets were judged to have a small degree of pigmentation chimerism, but microsatellite analysis failed to confirm chimerism in these or other piglets. Polymerase chain reaction analysis for detection of the porcine SRY gene in female piglets born from embryos injected with male EG cells identified six chimeras, at least one, but not more than two, from each treatment. Chimerism was confirmed in two putative pigmentation chimeras and in four piglets without overt signs of chimerism. The low percentage of injected embryos that yielded a chimera and the small contribution by EG cells to development of each confirmed chimera indicated that procedural changes in how EG cells were combined with host embryos were unsuccessful in increasing the likelihood that porcine EG cells will participate in embryonic development. Alternatively, our results suggested that improvements are needed in EG cell isolation and culture procedures to ensure in vitro maintenance of EG cell developmental capacity.

Fifteen organic phosphate pesticides were tested by computer tracking for their acute behavioral toxicity with the nematode Caenorhabditis elegans. Thirteen of these 15 chemicals are used as insecticides and are anticholesterase agents. The other two chemicals are used as herbicides. EC50 values for each chemical were compared to the corresponding LD50 acute lethality value in rats and mice. Order of toxicity was found to be significantly correlated in comparisons of C. elegans to both rats and mice. Mechanistic investigations were conducted by assaying 8 of the 15 chemicals for anticholinesterase activity in C. elegans. Significant cholinesterase inhibition was confirmed for five chemicals that had displayed high behavioral toxicity, while three chemicals of low behavioral toxicity showed no significant decrease in cholinesterase activity. Toxicity for two chemicals that do not inhibit cholinesterase in mammals was linked to pH effects. Detailed comparison of individual chemicals and metabolic issues are discussed. These results have positive implications for the use of C. elegans as a mammalian neurological model and support the use of C. elegans in early rounds of chemical toxicity screening.

Free-living nematodes may harbor, protect, and disperse bacteria, including those ingested and passed in viable form in feces. These nematodes are potential vectors for human pathogens and may play a role in foodborne diseases associated with fruits and vegetables eaten raw. In this study, we evaluated the associations between a free-living soil nematode, Caenorhabditis elegans, and Escherichia coli, an avirulent strain of Salmonella Typhimurium, Listeria welshimeri, and Bacillus cereus. On an agar medium, young adult worms quickly moved toward colonies of all four bacteria; over 90% of 3-day-old adult worms entered colonies within 16 min after inoculation. After 48 h, worms moved in and out of colonies of L. welshimeri and B. cereus but remained associated with E. coli and Salmonella Typhimurium colonies for at least 96 h. Young adult worms fed on cells of the four bacteria suspended in K medium. Worms survived and reproduced with the use of nutrients derived from all test bacteria, as determined for eggs laid by second-generation worms after culturing for 96 h. Development was slightly slower for worms fed gram-positive bacteria than for worms fed gram-negative bacteria. Worms that fed for 24 h on bacterial lawns formed on tryptic soy agar dispersed bacteria over a 3-h period when they were transferred to a bacteria-free agar surface. The results of this study suggest that C. elegans and perhaps other free-living nematodes are potential vectors for both gram-positive and gram-negative bacteria, including foodborne pathogens in soil.

The lung innate immune response to lipopolysaccharide (LPS) coordinates cellular inflammation, mediator, and protease release essential for host defense but deleterious in asthma, chronic obstructive pulmonary disease, and cystic fibrosis. In vitro, LPS signals to the transcription factors NFkappaB via TLR4, MyD88, and IL-1R-associated kinase (IRAK), to AP-1 by mitogen-activated protein (MAP) kinases, and via an alternate route in IRAK-deficient mice, but the in vivo lung signaling pathway(s) are not understood. We investigated the role of Akt and Erk1/2 as LPS intensely stimulates granulocyte/macrophage-colony-stimulating factor (GM-CSF) release, and neutralizing GM-CSF profoundly suppressed LPS-induced inflammation, suppressed expression and activity of lung proteases, significantly reduced GM-CSF and tumor necrosis factor alpha (TNFalpha) mRNA expression, and dampened nuclear localization of both NFkappaB (p50/65) and AP-1. LPS markedly activated Akt and Erk1/2, but not p38, in a GM-CSF-dependent manner in direct temporal association with NFkappaB and AP-1 activation. Pharmacological inhibition of Akt or Erk activation in LPS-treated tracheal explants ex vivo inhibited the release of GM-CSF. These data implicate GM-CSF-dependent activation of Akt in the amplification of this response and demonstrate the role of Erks rather than p38 in lung LPS inflammatory responses. Inhibition of GM-CSF may be of therapeutic benefit in inflammatory diseases in which LPS contributes to lung damage.

Wood shavings used for poultry bedding may become increasingly difficult or expensive to obtain. A post-consumer paper product, derived of pelletized newspaper, was evaluated to determine the feasibility of its use as an alternative type of turkey bedding. Results of these trials indicate that a pelletized newspaper product can be used as successfully as pine shavings for bedding under brooding turkeys. Turkeys brooded on the newspaper bedding had weight gains equal to their shavings-brooded counterparts. Mortality was reduced by using the pelletized newspaper.

This study was designed to characterize conceptus development based on pre- and postnatal measurements of in vivo- and in vitro-derived bovine pregnancies. In vivo-produced embryos were obtained after superovulation, whereas in vitro-produced embryos were derived from established procedures for bovine IVM, IVF and IVC. Blastocysts were transferred to recipients to obtain pregnancies of single (in vivo/singleton or in vitro/singleton groups) or twin fetuses (in vitro/twins group). Ultrasonographic examinations were performed weekly, from Day 30 of gestation through term. Videotaped images were digitized, and still-frames were used for the measurement of conceptus traits. Calves and fetal membranes (FM) were examined and measured upon delivery. In vitro-produced fetuses were smaller than in vivo controls (P < 0.05) during early pregnancy (Day 37 to Day 58), but in vitro/singletons presented significantly higher weights at birth than in vivo/control and in vitro/twin calves (P < 0.05). From late first trimester of pregnancy (Day 72 to Day 93), placentomes surrounding in vitro-derived singleton fetuses were longer and thinner than controls (P < 0.05). At term, the presence of giant cotyledons in the fetal membranes in the in vitro group was associated with a larger cotyledonary surface area in the fetal horn (P < 0.05). The biphasic growth pattern seen in in vitro-produced pregnancies was characterized by conceptus growth retardation during early pregnancy, followed by changes in the development of the placental tissue. Resulting high birth weights may be a consequence of aberrant placental development due to the disruption of the placental restraint on fetal growth toward the end of pregnancy.

Introduction. We hypothesized that the late phase of microvascular protection induced by ischemic preconditioning or by adenosine is protein kinase C (PKC) dependent. Materials and methods. The cremaster muscle of male Sprague–Dawley rats underwent 45 min of ischemic preconditioning and, 24 h later, 4 h of warm ischemia followed by 60 min of reperfusion. To mimic the effects of IPC, adenosine (ADO; an adenosine receptor agonist) or 4-phorbol 12-myristate 13-acetate (PMA; a PKC activator) was delivered to the vascular network of the cremaster 24 h before the prolonged ischemia via local intra-arterial infusion. To block the microvascular protection induced by ADO or IPC, chelerythrine (CHE; a PKC blocker) was given by local intra-arterial infusion prior to the administration of ADO or the initiation of IPC. Microvascular responses in the cremaster muscle to ischemic preconditioning or pharmacological preconditioning were determined by measuring terminal arteriole diameter and capillary perfusion using intravital microscopy and by the evaluation of the endothelium-dependent nitric oxide system in terminal arterioles. Results. Blockade of PKC using CHE on day 1 eliminated both ADO- and IPC-induced microvascular protections seen on day 2. However, the microvascular protection induced by the administration of PMA (without IPC) that was given 24 h before the 4 h of warm ischemia/reperfusion was significantly better than the control group response (sham IPC), but was not as good as the protection induced by IPC or ADO alone. Conclusion. The overall results from these studies suggest that ischemic or ADO preconditioning induces late-phase microvascular protection in skeletal muscle by a PKC-dependent mechanism.

Based on the structure of N -[( R,R )-( E )-1-(4-chlorobenzyl)-3-(2-oxoazepan-3-yl)carbamoyl]allyl- N -methyl-3,5-bis(trifluoromethyl)benzamide ( 1 ), attempts to improve the NK 2 affinity have resulted in the discovery of N -[( R,R )-( E )-1-(3,4-dichlorobenzyl)-3-(2-oxoazepan-3-yl)carbamoyl]allyl- N -methyl-3,5-bis(trifluoromethyl)benzamide ( 9 , DNK333 ) exhibiting a 5-fold improved affinity to the NK 2 receptor in comparison to 1 . Simplification of the structure via elimination of a chiral centre led to 3-[ N ′-3,5-bis(trifluoromethyl)benzoyl- N -(3,4-dichlorobenzyl)- N ′-methylhydrazino]- N -[( R )-2-oxo-azepan-3-yl]propionamide ( 22 ), a potent and fairly balanced NK 1 /NK 2 antagonist.

Ischemia and reperfusion in skeletal muscle is unavoidable during many reconstructive surgeries. Typical examples include replantation, transplantation, and free muscle transfer. One important complication during or after surgery is arterial insufficiency or a no-reflow phenomenon. The microcirculation is a primary target of ischemia and reperfusion injury. Vasoconstriction, poor blood flow, and capillary no-reflow, are the prominent features in the microcirculation seen during reperfusion. Currently, extensive efforts have focused on the theory that reactive oxygen species induce endothelial dysfunction in the microcirculation during reperfusion. Some intervention approaches, including ischemic preconditioning, are developing to interfere with or modulate the pathophysiological processes that are set in motion during ischemia and reperfusion.

Purpose. The goal of these studies was to determine the initiating factors for late preconditioning in the microcirculation of skeletal muscle. Materials and methods. The cremaster muscle of male Sprague–Dawley rats underwent 4 h of ischemia and then 60 min of reperfusion. Ischemic preconditioning (IPC) consisted of 45 min of ischemia but was done 24 h before the 4 h of ischemia. To mimic the effects of IPC in the late phase, adenosine (ADO) or sodium nitroprusside (SNP) was given 24 h before the prolonged ischemia via local intraarterial infusion. To block the effects of IPC in the late phase, 8-sulfophenyl-theophylline (a nonspecific ADO receptor blocker) or NW-nitro- l -arginine (a nonselective nitric oxide synthase antagonist) was given prior to IPC. Microvascular response to IPC and pharmacological preconditioning were determined by measuring arteriole diameters and capillary perfusion using intravital microscopy. Results. Administration of ADO or SNP on day 1 without IPC produced a similar microvascular protection against prolonged ischemia/reperfusion on day 2 as that induced by IPC alone. In contrast, blocking ADO receptors or nitric oxide synthase on day 1 just prior to IPC eliminated the IPC-induced microvascular protection seen on day 2. In addition, inhibition of nitric oxide synthase on day 1 diminished the protection induced by ADO, but blocking ADO receptors on day 1 did not compromise the protection induced by SNP. Conclusion. The results from these studies suggest that up regulation of ADO is the initiating factor with secondary up regulation of nitric oxide in late preconditioning. Both ADO and nitric oxide contribute to initiating microvascular protection in the late phase of IPC.

The failure of interspecies and hybrid pregnancies between the domestic sheep (Ovis aries) and goat (Capra hircus) is not completely understood. The sheep-goat hematopoietic chimera is a unique model for studying the role of the maternal immune response in failure of interspecies and hybrid pregnancies between these species. Hematopoietic chimeras were created by in utero transplantation of sheep fetal liver cells into goat fetuses. The resulting chimeric females were recipients of sheep demi-embryos genetically identical to their sheep cells and/or were bred to a ram to create a hybrid pregnancy. Pregnancy sera were analyzed for the presence of anti-species antibodies (Ab) using a lymphocyte microcytotoxicity assay. None of the concepti survived to term. Gross and histological evaluations of two interspecies sheep concepti revealed abnormal placentome formation. The humoral immune response of several hematopoietic chimeras to the challenging concepti differed from control animals. We observed delayed onset of Ab production, low and absent titers, and persistent Ab titers with delayed fetal death. Ultrasonography typically revealed normal fetal development associated with high volumes of placental fluids and retarded placentome development. We conclude that fetal death was associated with abnormal placental development that was not the result of maternal humoral immune attack.

The production of antibodies during pregnancy or after parturition is a natural occurrence in many mammalian species. Fetal cells have been detected in the peripheral blood of women and mice and are thought to be the immune stimulus for antibody production. The aim of this study was to investigate if the production of maternal anti-fetal antibodies during ruminant pregnancy is the result of fetal leukocyte trafficking across the placenta. Maternal pregnancy serum was collected from 94 does whose fetuses received sheep hematopoietic stem cells via in utero transplantation at 49 to 62 d of gestation. Serum samples were collected before surgery and at weekly intervals throughout gestation. A lymphocyte microcytotoxicity assay was used to screen the serum samples from does that carried chimeric fetuses to term (n = 75). Of these 75 does, 28 parous does had presurgery serum that contained alloreactive antibodies. Nine of the 75 does had nonreactive presurgery serum, but they produced alloreactive antibody titers during gestation. Xenoreactive antibodies were detected in the pregnancy sera from 2 of the 75 does tested. Hemolytic assays confirmed the species-specificity of the xenoreactive serum from these 2 does. In view of the fact that hematopoietic cells were the only source of anti-sheep antibody stimulation in this model, we propose that fetal leukocyte trafficking does take place across the caprine placenta.

Both allogeneic and xenogeneic hematopoietic chimera models have been developed, including fetal sheep models that demonstrated high levels of stable, multilineage engraftment created by in utero hematopoietic stem cell transplantation. The aim of this study was to test the efficacy of in utero transplantation to create xenogeneic sheep-goat hematopoietic chimeras. Fetal liver cells and T-cell-depleted adult bone marrow were tested as sources of hematopoietic stem cells. Donor cells were injected intraperitoneally into 130 recipient fetuses between 49 and 62 days of gestation. Groups 1 and 2 received crude fetal liver cell preparations. Group 3 received fetal liver cells that were incubated overnight in a phytohemagglutinin-stimulated lymphocyte-conditioned medium (PHA-LCM). In Group 4, hematopoietic stem cells were concentrated by using additional density separations. Group 5 fetal recipients received low-density, T-cell-depleted adult bone marrow cells. In Group 1, fetuses were accessed via hysterotomy. Hematopoietic stem cells were injected into Groups 2, 3, 4, and 5 without cutting through the uterine wall. Fetal survival in the five groups ranged from 56 to 100%. The percentage of chimeras from injected fetuses ranged from 43 to 92% by FACS and PCR analyses; however, levels of chimerism were low (

To the extent that alternative medicine offers a substitute for mainstream physician services, physicians’ incomes are reduced by the incursion of alternative providers into the medical marketplace. State regulations restricting the practice of alternative medicine create rents for physicians whose incomes are protected from competition with alternative providers. Focusing on homeopathy as representative of an alternative therapeutic that potentially substitutes for conventional medicine, a cross-state empirical analysis reveals that mainstream physicians’ incomes are higher in states with more restrictive regulations governing the practice of homeopathy. This finding suggests that regulatory barriers to alternative medicine are motivated more by the interests of orthodox physicians in seeking protection from competition than by the interests of consumers in quality assurance.

Statement of Problem. Several methods have been used to determine the surface characteristics of resin composites in vivo and compare composite wear rates with enamel wear rates. Purpose. This pilot study describes the surface characteristics of resin composites and the wear of resin composites and enamel during 1 year of in vivo service. Material and Methods. A single Class II posterior resin composite restoration (Z100) was placed in 10 patients. Restored teeth and unrestored adjacent control teeth were measured for wear 4 times within the first year. A null point contact stylus profiler and fitting software were used to measure epoxy casts. Maximum depth of wear, average depth of wear, and characteristics of the restoration margin were determined. Paired t tests were used to compare the control and restored teeth, and ANOVA was used to assess the progression of wear over time ( P Results. After 1 year, maximum depth of wear over the entire preparation region was on average 204.8 μm (± 129.8), significantly greater than the 36.8 μm (± 10.1) average maximum depth of wear of enamel at occlusal contact areas on control teeth ( P =.009). Maximum depth of wear progressed over time ( P =.009). Fracture of excess composite, commonly called flash fracture, occurred in 50% of the restored teeth extending over the preparation margin. Conclusion. Composite restorations wore significantly faster than enamel contact areas on control teeth. Also of concern were the marginal flash fractures that could facilitate secondary caries. (J Prosthet Dent 2000;84:353-9.)

Background. The purpose of the present study was to determine if platelet-activating factor is an important mediator that produces vasospasm during reperfusion after ischemia in skeletal muscle. Materials and Methods. A vascular isolated cremaster muscle in male Sprague–Dawley rats was coupled with local intraarterial drug infusion as a model to study microcirculation responses to ischemia/reperfusion injury. Arteriole diameters and capillary perfusion were measured using intravital microscopy. Group 1: platelet-activating factor dose response. Group 2: Effects of a cyclooxygenase inhibitor; indomethacin, and a thromboxane synthetase inhibitor, imidazole, on the response to platelet-activating factor. Group 3: Effects of nitric oxide synthesis inhibitor; N ω -nitro- l -arginine methyl ester, on the response to platelet-activating factor. Group 4: Effects of a platelet-activating factor receptor antagonist, CV-3988, indomethacin, and imidazole after 4 h of warm ischemia and reperfusion. Results. Intraarterial infusion of platelet-activating factor produced a dose-related but mild vasoconstriction. Pretreatment with indomethacin or imidazole resulted in significant vasodilation actually emanating from platelet-activating factor infusion. Nitric oxide inhibition (with N ω -nitro- l -arginine methyl ester) enhanced the vasoconstriction produced by platelet-activating factor. Pretreatment with CV-3988, indomethacin, or imidazole significantly attenuated ischemia/reperfusion-induced vasospasm and capillary no-reflow in the cremaster muscles. Conclusions. Ischemia/reperfusion-induced vasoconstriction is at least in part mediated by platelet-activating factor and thromboxane A 2 .

Preparation, structure and reactions of glycosylmanganese pentacarbonyl complexes are discussed. Anomerically pure complexes of pyranosyl and furanosyl complexes were prepared in excellent yield. The conformations of the anomeric glucosyl and mannosyl complexes were derived from detailed analysis of their 1D and 2D 1H- and 13C-NMR spectra including NOE data. The complexes are further characterized by 55Mn-NMR chemical shifts and 55Mn, 13C one-bond coupling constants. These compounds undergo various migratory insertion reactions resulting in formation of C-glycosyl derivatives. Applications of this technology to the synthesis of C-glycosyl and C-aryl glycosidic systems are discussed.