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Co-speciation is a fundamental concept of evolutionary biology and intuitively appealing, yet in practice hard to demonstrate as it is often blurred by other evolutionary processes. We investigate the phylogeographic history of the monogenean ectoparasites Gyrodactylus teuchis and G. truttae on European salmonids of the genus Salmo. Mitochondrial cytochrome oxidase subunit 1 and the nuclear ribosomal internal transcribed spacer 2 were sequenced for 189 Gyrodactylus individuals collected from 50 localities, distributed across most major European river systems, from the Iberian- to the Balkan Peninsula. Despite both anthropogenic and naturally caused admixture of the principal host lineages among major river basins, co-phylogenetic analyses revealed significant global congruence for host and parasite phylogenies, providing firm support for co-speciation of G. teuchis and its salmonid hosts brown trout (S. trutta) and Atlantic salmon (S. salar). The major split within G. teuchis, coinciding with the initial divergence of the hosts was dated to ~1.5 My BP, using a Bayesian framework based on an indirect calibration point obtained from the host phylogeny. The presence of G. teuchis in Europe thus predates some of the major Pleistocene glaciations. In contrast, G. truttae exhibited remarkably low intraspecific genetic diversity. Given the direct life cycle and potentially high transmission potential of gyrodactylids, this finding is interpreted as indication for a recent emergence (<60 ky BP) of G. truttae via a host-switch. Our study thus suggests that instances of two fundamentally different mechanisms of speciation (co-speciation vs. host-switching) may have occurred on the same hosts in Europe within a time span of less than 1.5 My in two gyrodactylid ectoparasite species.

To improve treatment of obesity, a contributing factor to multiple systemic and metabolic diseases, a better understanding of metabolic state and environmental stress at the cellular level is essential. This work presents development of a three-dimensional (3D) in vitro model of adipose tissue displaying induced lipid accumulation as a function of fatty acid supplementation that, subsequently, investigates cellular responses to a pro-inflammatory stimulus, thereby recapitulating key stages of obesity progression. Three-dimensional spheroid organization of adipose cells was induced by culturing 3T3-L1 mouse preadipocytes on an elastin-like polypeptide-polyethyleneimine (ELP-PEI)-coated surface. Results indicate a more differentiated phenotype in 3D spheroid cultures relative to two-dimensional (2D) monolayer analogues based on triglyceride accumulation, CD36 and CD40 protein expression, and peroxisome proliferator-activated receptor-γ (PPAR-γ) and adiponectin mRNA expression. The 3T3-L1 adipocyte spheroid model was then used to test the effects of a pro-inflammatory microenvironment, namely maturation in the presence of elevated fatty acid levels followed by acute exposure to tumor necrosis factor alpha (TNF-α). Under these conditions, we demonstrate that metabolic function was reduced across all cultures exposed to TNF-α, especially so when pre-exposed to linoleic acid. Further, in response to TNF-α, enhanced lipolysis, monitored as increased extracellular glycerol and fatty acids levels, was observed in adipocytes cultured in the presence of exogenous fatty acids. Taken together, our 3D spheroid model showed enhanced adipogenic differentiation and presents a platform for elucidating the key phenotypic responses that occur in pro-inflammatory microenvironments that characterize obesogenic states.

Objectives: The objective of this study was to compare current adverse drug/allergy reaction reporting in patient electronic medical records/charts against information gathered during patient interviews in the emergency department. Our hypothesis was that current methods for allergy reporting results in significant discrepancy between what is documented and the actual allergy history upon interviewing the patient., Methods: The study was conducted between December 2011 and April 2012 in an academic emergency department. This was a convenience sample study comparing a prospective patient interview with previously documented allergy histories. Demographics for sex, age, and race were recorded. Patients to be interviewed were adults with at least one documented allergy in their chart. Descriptive statistics and percentages were used for demographic and prevalence data. Agreement between interviews and charts was assessed for both the reaction type and the reaction descriptor., Results: There were 101 patients interviewed during this 4-month period, and a total of 235 adverse drug reactions were recorded. There were 66 women and 35 men included in this study. The mean age was 51 ± 17 years. The median number of allergy instances for women was 2 (interquartile range 1-3) and for men the median number of allergy instances was 1 (interquartile range 1-2). The percentage of agreements for overall allergies was 85% and 50% for the type of reaction. Total profile agreement occurred in nine patients., Conclusions: The percentage of agreement between interviews and charting for reaction type was 50%. Even with the use of electronic medical records, better methods are needed to properly record allergies to ensure patient safety and care.

The first decade of clinical experience with transcatheter aortic valve replacement since 2002 saw the development of 2 main valve systems, namely the Edwards Sapien balloon-expandable valve series and the Medtronic self-expanding CoreValve. These 2 valve platforms now have achieved commercial approval and application worldwide in patients with severe aortic stenosis whose perioperative risk for surgical intervention is high or extreme. In the second decade of transcatheter aortic valve replacement, clinical experience and refinements in valve design have resulted in clinical drift towards lower patient risk cohorts. There are currently 2 major trials, PARTNER II and SURTAVI, that are both evaluating the role of transcatheter aortic valve replacement in intermediate-risk patient cohorts. The results from these landmark trials may usher in a new clinical paradigm for transcatheter aortic valve replacement in its second decade., (Copyright © 2015. Published by Elsevier Inc.)

The functional aortic annulus represents a sound clinical framework for understanding the components of the aortic root complex. Recent three-dimensional imaging analysis has demonstrated that the aortic annulus frequently is elliptical rather than circular. Comprehensive three-dimensional quantification of this aortic annular geometry by transesophageal echocardiography and/or multidetector computed tomography is essential to guide precise prosthesis sizing in transcatheter aortic valve replacement to minimize paravalvular leak for optimal clinical outcome. Furthermore, three-dimensional transesophageal echocardiography accurately can quantify additional parameters of the functional aortic annulus such as coronary height for complete sizing profiles for all valve types in transcatheter aortic valve replacement. Although it is maturing rapidly as a clinical imaging modality, its role in transcatheter aortic valve replacement is seen best as complementary to multidetector computed tomography in a multidisciplinary heart team model., (Copyright © 2014. Published by Elsevier Inc.)

Psychological pain is a prominent symptom of clinical depression. We asked if frontal alpha asymmetry, frontal EEG power, and frontal fractal dimension asymmetry predicted psychological pain in adults with a history of depression. Resting-state frontal EEG (F3/F4) was recorded while participants (N=35) sat upright with their eyes closed. Frontal delta power predicted psychological pain while controlling for depressive symptoms, with participants who exhibited less power experiencing greater psychological pain. Frontal fractal dimension asymmetry, a nonlinear measure of complexity, also predicted psychological pain, such that greater left than right complexity was associated with greater psychological pain. Frontal alpha asymmetry did not contribute unique variance to any regression model of psychological pain. As resting-state delta power is associated with the brain's default mode network, results suggest that the default mode network was less activated during high psychological pain. Findings are consistent with a state of arousal associated with psychological pain., (Copyright © 2015 Elsevier B.V. All rights reserved.)

The Adult Attachment Interview (AAI) and the Adult Attachment Projective (AAP) purport to measure the same attachment classifications. The aims of this study were to determine the concurrent validity of the AAI and AAP, and to compare their ability to predict indicators of risk associated with socioeconomic status (SES), depression, and 5-HTTLPR genotype. Results indicated no agreement between AAI and AAP attachment classifications in a sample of late adolescents. As predicted, individuals classified as "unresolved" with regard to loss or trauma were significantly more likely to be of lower SES, have higher levels of depression, and have the 5-HTTLPR "ss" genotype than individuals with secure, preoccupied, or dismissing attachments. These associations, however, were only significant when attachment was classified with the AAI. Results suggest that the AAI and AAP measure different facets of attachment as a result of their unique methodologies and coding criteria. Further research is needed to support their comparability before investigators can assume that the AAP is a valid substitute for the AAI.

There has been significant progress throughout 2014 in cardiothoracic and vascular anaesthesia and intensive care. There has been a revolution in the clinical approach to acute and chronic adult aortic diseases. Contemporary management of adult aortic disease is based on etiology, clinical presentation, extent, and integrated intervention with medical, endovascular and/or surgical measures. Further European guidelines have explored in depth the cardiovascular management in non-cardiac surgery with a thematic focus to reduce perioperative mortality from the leading offender, namely myocardial ischemia. Integrated guidelines address the management of myocardial revascularization including the percutaneous and surgical options. Despite 50 years since the first coronary artery bypass grafting procedure and impressive advances in interventional cardiology, surgical revascularization remains a gold standard for many patients with coronary artery disease. These advances in 2014 will likely further improve perioperative outcomes for our patients.

Though growth factors allow tissue regeneration, the trade-off between their effectiveness and adverse effects limits clinical application. The key issues in current growth factor therapy largely derive from initial burst pharmacokinetics, rapid clearance, and proteolytic cleavage resulting in clinical ineffectiveness and diverse complications. While a number of studies have focused on the development of carriers, issues arising from soluble growth factor remain. In this study, we report a prodrug of growth factors constituting a novel mode of action (MoA). To mimic endogenous protein processing in cells, we developed a recombinant BMP-2 polypeptide based on a protein transduction domain (PTD) to transduce the protein into cells followed by furin-mediated protein cleavage and secretion of active growth factor. As proof of concept, a few micrograms scale of PTD-BMP-2 polypeptide sufficed to induce bone regeneration in vivo. As a simple platform, our technique can easily be extended to delivery of BMP-7 and DKK-1 as therapeutics for TGF-β and canonical Wnt signaling, respectively, to suppress the epithelial-mesenchymal transition (EMT), which constitutes a fundamental biological mechanism of many diseases. This technique largely overcomes the limitations of current soluble growth factors and opens the door to next generation growth factor therapeutics., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

miRNAs are essential for self-renewal and differentiation of normal and malignant stem cells by regulating the expression of key stem cell regulatory genes. Here, we report evidence implicating the miR100 in self-renewal of cancer stem-like cells (CSC). We found that miR100 expression levels relate to the cellular differentiation state, with lowest expression in cells displaying stem cell markers. Utilizing a tetracycline-inducible lentivirus to elevate expression of miR100 in human cells, we found that increasing miR100 levels decreased the production of breast CSCs. This effect was correlated with an inhibition of cancer cell proliferation in vitro and in mouse tumor xenografts due to attenuated expression of the CSC regulatory genes SMARCA5, SMARCD1, and BMPR2. Furthermore, miR100 induction in breast CSCs immediately upon their orthotopic implantation or intracardiac injection completely blocked tumor growth and metastasis formation. Clinically, we observed a significant association between miR100 expression in breast cancer specimens and patient survival. Our results suggest that miR100 is required to direct CSC self-renewal and differentiation., (©2014 American Association for Cancer Research.)

Objectives: The goals of this study were to (1) identify valid variables that correlate with emergency department (ED) crowding and (2) determine a model that could be used to accurately reflect the degree of ED crowding., Methods: A site sampling form was applied to convenience sampling of 13 community hospitals in California between April 6, 2011, and May 1, 2011. The outcome variable was average perception of crowding by the ED physician and charge nurse on a 100-mm visual analog scale. We focused on 20 candidate predictor variables that represented counts and times in the ED that were collected every 4 hours. A prediction model was developed using multivariable linear regression to determine the measures that predicted ED crowding. A parsimonious model was developed to allow for a clinical useful tool that that explained a significant amount of variability predicted by the full ED crowding model., Results: A total of 2006 data sets were collected for each of the participating hospitals. A total of 1628 time entries for the hospitals were included in the study. Hospital EDs had censuses ranging from 18 000 to 98 000. Full evaluation was completed on 1489 data sets. Twenty variables were considered for the full model with 7 removed due to multicollinearity. The remaining 13 variables constituted the full model and explained 50.5% of the variability in the outcome variable. Five predictors were found to represent 92% of the variability represented by the full model., Conclusions: Five variables were highly correlated with community ED crowding and could be used to model the full set of all variables in explaining ED crowding., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Efforts to develop unbiased screens for identifying novel function-blocking monoclonal antibodies (mAbs) in human carcinomatous states have been hampered by the limited ability to design in vitro models that recapitulate tumor cell behavior in vivo. Given that only invasive carcinoma cells gain permanent access to type I collagen-rich interstitial tissues, an experimental platform was established in which human breast cancer cells were embedded in 3D aldimine cross-linked collagen matrices and used as an immunogen to generate mAb libraries. In turn, cancer-cell-reactive antibodies were screened for their ability to block carcinoma cell proliferation within collagen hydrogels that mimic the in vivo environment. As a proof of principle, a single function-blocking mAb out of 15 identified was selected for further analysis and found to be capable of halting carcinoma cell proliferation, inducing apoptosis, and exerting global changes in gene expression in vitro. The ability of this mAb to block carcinoma cell proliferation and metastatic activity was confirmed in vivo, and the target antigen was identified by mass spectroscopy as the α2 subunit of the α2β1 integrin, one of the major type I collagen-binding receptors in mammalian cells. Validating the ability of the in vitro model to predict patterns of antigen expression in the disease setting, immunohistochemical analyses of tissues from patients with breast cancer verified markedly increased expression of the α2 subunit in vivo. These results not only highlight the utility of this discovery platform for rapidly selecting and characterizing function-blocking, anticancer mAbs in an unbiased fashion, but also identify α2β1 as a potential target in human carcinomatous states.

Objectives: Inclusion of select orthopedic problems in the orthopedics observation unit (OOU) may reduce hospital admissions. Our system allows OOU status for 24 hours, but the effect on admissions is unknown. Our primary hypothesis was that we could predict which OOU patients required admission based on the presence of uncontrolled pain., Methods: Data were prospectively collected for all OOU patients in this prospective observational study, including data on extremity cellulitis, fractures, and spine injuries awaiting brace placement.The primary outcome variable was admission to the hospital versus discharge home. The a priori hypotheses were that patients with more persistent or worsening pain would require admission more often and that the OOU would result in fewer patients needing a costlier inpatient admission to the hospital. An a priori power analysis showed adequate power of 80% to detect a difference between admitted and discharged patients., Results: Data were prospectively collected from August 2011 to August 2012 for 199 consecutive OOU patients, 62% of whom were men. Diagnoses included infection (cellulitis or abscess of extremity) in 76%, fracture in 15% and other in 9% of the patients. Sixty-two patients (31%) were admitted and 7 patients (4%) made return visits for the same problem within a 30-day period. No significant relations existed between any of the independent variables and admission on bivariate analysis. Multivariable logistic regression found no significant predictors of hospital admission. Logistic regression was not performed on 30-day returns because of the low event rate (4%)., Conclusions: An OOU prevented 138 of 199 (69%) patients from being admitted to a hospital. There were no significant predictors of which patients would require admission. Pain was not a predictor of need for admission. The lack of significant predictors is important in suggesting that without the ability to predict which patients require admission, a system using an OOU can reduce admissions by more than two-thirds.

Notch1-Delta-like 4 (Dll4) signalling controls vascular development by regulating endothelial cell (EC) targets that modulate vessel wall remodelling and arterial-venous specification. The molecular effectors that modulate Notch signalling during vascular development remain largely undefined. Here we demonstrate that the transcriptional repressor, Snail1, acts as a VEGF-induced regulator of Notch1 signalling and Dll4 expression. EC-specific Snail1 loss-of-function conditional knockout mice die in utero with defects in vessel wall remodelling in association with losses in mural cell investment and disruptions in arterial-venous specification. Snail1 loss-of-function conditional knockout embryos further display upregulated Notch1 signalling and Dll4 expression that is partially reversed by inhibiting γ-secretase activity in vivo with Dll4 identified as a direct target of Snail1-mediated transcriptional repression. These results document a Snail1-Dll4/Notch1 axis that controls embryonic vascular development.

An overwhelming need for change in the U.S. healthcare delivery system, coupled with the need to improve clinical and financial outcomes, has prompted hospitals to direct renewed efforts toward achieving high quality and cost-effectiveness. Additionally, with the dawn of accountable care organizations and increasing focus on patient expectations, hospitals have begun to seek physician partners through clinical alignment. Contrary to the unsuccessful alignment strategies of the 1990s, today's efforts are more mutually beneficial, driven by the need to achieve better care coordination, increased access to infrastructure, improved quality, and lower costs. In this article, we describe a large, academic, tertiary care hospital's approach to developing and implementing alignment and integration models with its collaboration-ready physicians and physician groups. We developed four models--short of physicians' employment with the organization--tailored to meet the needs of both the physician group and the hospital: (1) medical directorship (group physicians are appointed to serve as medical directors of a clinical area), (2) professional services agreement (specific clinical services, such as overnight admissions help, are contracted), (3) co-management services agreement (one specialty group co-manages all services within the specialty service lines), and (4) lease arrangement (closest in scope to employment, in which the hospital pays all expenses and receives all revenue). Successful hospital-physician alignment requires careful planning and the early engagement of legal counsel to ensure compliance with federal statutes. Establishing an integrated system with mutually identified goals better positions hospitals to deliver cost-effective and high-quality care under the new paradigm of healthcare reform.

Psychological pain is a prominent symptom in people who experience depression, but its relation with physiological measures has not been explored. This study compared two measures of psychological pain, the Orbach & Mikulincer Mental Pain (OMMP) questionnaire and the Psychache Scale, for their relationship with resting-state heart rate variability (HRV) in 35 adults with a history of depression. Low-frequency HRV decreased significantly with increasing psychological pain, particularly in participants who did not use antidepressants, while the beat-to-beat fractal dimension decreased in participants who did use antidepressants. Neither heart rate nor high-frequency HRV was associated with psychological pain. These results suggest a state of arousal characterized by increased sympathetic activity. Results also indicate that the OMMP may be a more accurate measure of autonomic arousal associated with current psychological pain than the Psychache Scale., (Copyright © 2014 Society for Psychophysiological Research.)

Background: Cysteine and methionine are the two sulfur containing amino acids in proteins. While the roles of protein-bound cysteinyl residues as endogenous antioxidants are well appreciated, those of methionine remain largely unexplored., Scope: We summarize the key roles of methionine residues in proteins., Major Conclusion: Recent studies establish that cysteine and methionine have remarkably similar functions., General Significance: Both cysteine and methionine serve as important cellular antioxidants, stabilize the structure of proteins, and can act as regulatory switches through reversible oxidation and reduction. This article is part of a Special Issue entitled Current methods to study reactive oxygen species - pros and cons and biophysics of membrane proteins. Guest Editor: Christine Winterbourn., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Embryonic stem cells (ESCs) exhibit the dual properties of self-renewal and pluripotency as well as the ability to undergo differentiation that gives rise to all three germ layers. Wnt family members can both promote ESC maintenance and trigger differentiation while also controlling the expression of Snail1, a zinc-finger transcriptional repressor. Snail1 has been linked to events ranging from cell cycle regulation and cell survival to epithelial-mesenchymal transition (EMT) and gastrulation, but its role in self-renewal, pluripotency or lineage commitment in ESCs remains undefined. Here we demonstrate using isogenic pairs of conditional knockout mouse ESCs, that Snail1 exerts Wnt- and EMT independent control over the stem cell transcriptome without affecting self-renewal or pluripotency-associated functions. By contrast, during ESC differentiation, an endogenous Wnt-mediated burst in Snail1 expression regulates neuroectodermal fate while playing a required role in epiblast stem cell exit and the consequent lineage fate decisions that define mesoderm commitment.

Objectives: The population of the United States continues to diversify, with an increasing percentage of individuals who have limited English proficiency (LEP). A major concern facing emergency departments (EDs) around the country is increasing length of stay (LOS). Although multiple studies have shown racial and ethnic disparities in waiting time and LOS, no studies have examined specifically whether patients with LEP have a different LOS than English-speaking (ES) patients. In addition, no studies have examined whether the use of interpreters by patients with LEP has a significant impact on LOS. We hypothesized that there was a significant difference in LOS when comparing patients with LEP and ES patients and patients with LEP who used interpreters versus patients with LEP who did not., Methods: This was a prospective cohort study with LOS data collected from a level I ED patient tracking software program from October 2011 to December 2011. The primary language preferred by the patient was indicated at the time of triage and registration and the patient's use of an interpreter also was recorded. The patient's demographic data, ED visit information, and LOS were prospectively entered into an Excel spreadsheet. Percentages were compared using 95% confidence intervals and LOS was analyzed using the Student t test. With >100 subjects per group, our study had 80% power (ie, a power of 0.8) to determine a 15% difference in proportions between groups or a difference of 120 minutes (assuming a standard deviation of 300 minutes on both means)., Results: Data were collected from a total of 121 ES patients and 124 patients with LEP. In the LEP group were the languages of Spanish, Navajo, Vietnamese, Chinese, Arabic, and American Sign Language. Fifty-eight percent of patients with LEP used an interpreter. There were no differences between ES patients and patients with LEP in age, sex, mode of arrival, chief complaints, acuity, percentage admitted, percentage pediatric patients, or percentage of Medicaid/Medicare recipients. More patients with LEP were self-pay (36% vs 20%, diff 16, 95% confidence interval 2-31). There were no differences in mean LOS from time of arrival to time to being seen by a provider when comparing ES patients with patients with LEP or time of arrival to time to discharge or admission request. Comparing the patients with LEP who used interpreters with those who did not use interpreters, there was a significantly different LOS from time of arrival to time of discharge or admission request (958 ± 644 vs 628 ± 595 minutes, diff 330, 95% confidence interval 84-576)., Conclusions: There was no difference in LOS for patients with LEP; however, patients with LEP who used interpreters had a significant increase in LOS compared with those who did not use interpreters.

Hsaio and colleagues link gut microbes to autism spectrum disorders (ASD) in a mouse model. They show that ASD symptoms are triggered by compositional and structural shifts of microbes and associated metabolites, but symptoms are relieved by a Bacteroides fragilis probiotic. Thus probiotics may provide therapeutic strategies for neurodevelopmental disorders., (Copyright © 2013 Elsevier Inc. All rights reserved.)

During development, wound repair and disease-related processes, such as cancer, normal, or neoplastic cell types traffic through the extracellular matrix (ECM), the complex composite of collagens, elastin, glycoproteins, proteoglycans, and glycosaminoglycans that dictate tissue architecture. Current evidence suggests that tissue-invasive processes may proceed by protease-dependent or protease-independent strategies whose selection is not only governed by the characteristics of the motile cell population, but also by the structural properties of the intervening ECM. Herein, we review the mechanisms by which ECM dimensionality, elasticity, crosslinking, and pore size impact patterns of cell invasion. This summary should prove useful when designing new experimental approaches for interrogating invasion programs as well as identifying potential cellular targets for next-generation therapeutics., (© 2013 The Authors Journal of Microscopy © 2013 Royal Microscopical Society.)

Cell migration through 3D tissue depends on a physicochemical balance between cell deformability and physical tissue constraints. Migration rates are further governed by the capacity to degrade ECM by proteolytic enzymes, particularly matrix metalloproteinases (MMPs), and integrin- and actomyosin-mediated mechanocoupling. Yet, how these parameters cooperate when space is confined remains unclear. Using MMP-degradable collagen lattices or nondegradable substrates of varying porosity, we quantitatively identify the limits of cell migration by physical arrest. MMP-independent migration declined as linear function of pore size and with deformation of the nucleus, with arrest reached at 10% of the nuclear cross section (tumor cells, 7 µm²; T cells, 4 µm²; neutrophils, 2 µm²). Residual migration under space restriction strongly depended upon MMP-dependent ECM cleavage by enlarging matrix pore diameters, and integrin- and actomyosin-dependent force generation, which jointly propelled the nucleus. The limits of interstitial cell migration thus depend upon scaffold porosity and deformation of the nucleus, with pericellular collagenolysis and mechanocoupling as modulators.

Matrix metalloproteinases (MMPs) are zinc endopeptidases that play roles in numerous pathophysiological processes and therefore are promising drug targets. However, the large size of this family and a lack of highly selective compounds that can be used for imaging or inhibition of specific MMPs members has limited efforts to better define their biological function. Here we describe a protein engineering strategy coupled with small-molecule probe design to selectively target individual members of the MMP family. Specifically, we introduce a cysteine residue near the active-site of a selected protease that does not alter its overall activity or function but allows direct covalent modification by a small-molecule probe containing a reactive electrophile. This specific engineered interaction between the probe and the target protease provides a means to both image and inhibit the modified protease with absolute specificity. Here we demonstrate the feasibility of the approach for two distinct MMP proteases, MMP-12 and MT1-MMP (or MMP-14).

Current in vivo models for head and neck squamous cell carcinoma (HNSCC) have limitations in simulating some essential tumorigenic phenotypes, such as invasion. Most mouse models of human HNSCC are inadequate because tumor cells are injected directly into the connective tissue, thereby bypassing the basement membrane of the surface epithelium, the first barrier to invasion. In this manuscript, we establish the chick chorioallantoic membrane (CAM) assay as an in vivomodel of human HNSCC tumor progression. Using the CAM model of HNSCC, we investigated the role of enhancer of zeste homolog 2 (EZH2), a histone methyltransferase, in multiple aspects of HNSCC tumor progression. We found that knockdown of EZH2 reduced tumor size, angiogenesis, invasion, and metastasis of tumors produced by grafting human HNSCC cells onto the CAM. In addition, we demonstrate that EZH2 expression mediates a mesenchymal phenotype in HNSCC cell lines and mouse tumors. These findings demonstrate the advantages of the newly proposed CAM model of human HNSCC and highlight the emerging role of EZH2 in HSNCC tumor progression.

In vitro, topographical and biophysical cues arising from the extracellular matrix (ECM) direct skeletal stem cell (SSC) commitment and differentiation. However, the mechanisms by which the SSC-ECM interface is regulated and the outcome of such interactions on stem cell fate in vivo remain unknown. Here we demonstrate that conditional deletion of the membrane-anchored metalloproteinase MT1-MMP (Mmp14) in mesenchymal progenitors, but not in committed osteoblasts, redirects SSC fate decisions from osteogenesis to adipo- and chondrogenesis. By effecting ECM remodeling, MT1-MMP regulates stem cell shape, thereby activating a β1-integrin/RhoGTPase signaling cascade and triggering the nuclear localization of the transcriptional coactivators YAP and TAZ, which serve to control SSC lineage commitment. These data identify a critical MT1-MMP/integrin/YAP/TAZ axis operative in the stem cell niche that oversees SSC fate determination., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Using a constructivist grounded theory approach, we explored the development of father identity among boys involved in the juvenile justice system. Youth were recruited from a juvenile detention center and school district in a northern California county with a high teen birth rate. The participants were expecting a child or parenting an infant and had been arrested, incarcerated, or had committed a crime. We collected data through observations and individual interviews. Using constant comparative and dimensional analysis, we found that expectant adolescent fathers hoped for a boy and envisioned their central role as father to be making their son a man. This article contributes to greater understanding of father identity development for youth involved in the justice system. We suggest that teen parenting policies and programs include interventions sensitized by gender, accounting for the influence masculine ideals of manhood have on the development of father identity and the father-child relationship.

There are increasing numbers of mothers as well as fathers who engage in long-term migration to support their children and other family members in their home countries. In this article, the current state of the literature about children and adolescents left at home in these transnational families is surveyed and reviewed. The article reviews the effects on children of the process of separation from parents, the impact of gifts and remittances home, communication with distant parents and the quality of life with their substitute caregivers. The effects of immigration in late childhood or adolescence on these separated children are examined, as well as what is known about the processes of adaptation and family reunification, including migration traumas, impact of gender, and educational outcomes. Suggestions are given for pediatric clinicians working with reunifying families. Gaps in the literature are highlighted and the need for research into factors that promote successful family re-engagement and overall adaptation upon reunification., (Copyright © 2013 Mosby, Inc. All rights reserved.)

Research on brain areas involved in experiencing emotion and physical pain is abundant; however, psychological pain has received little attention in studies of the brain. The purpose of this systematic review was to provide an overview of studies on brain function related to psychological pain. The review was limited to studies in which participants experienced actual psychological pain or recalled a significant autobiographical event that may be assumed to have involved psychological pain. Based on results of the studies (N = 18), a tentative neural network for psychological pain is proposed that includes the thalamus, anterior and posterior cingulate cortex, the prefrontal cortex, cerebellum, and parahippocampal gyrus. Results indicated that grief may be a more accurate exemplar of psychological pain than recalled sadness, with indications of greater arousal during psychological pain. The proposed neural network for psychological pain overlaps to some extent with brain regions involved in physical pain, but results suggest a markedly reduced role for the insula, caudate, and putamen during psychological pain. Psychological pain is well known for its association with depression and as a precursor of suicidal behavior. Thus, identification of brain areas involved in psychological pain may help guide development of interventions to decrease mortality and morbidity.

Objective: Emergency department (ED) interruptions occur frequently. Recently, a hands-free communication device (HCD; Vocera) was added to the population of communications devices (a cellular telephone and a pager) in our ED. The research purpose was to determine whether this addition improved interruption times and our hypothesis was the device would decrease length of interruptions., Methods: This was a prospective cohort study of interruptions in an academic ED level 1 trauma center before and after introduction of an HCD. Interruptions were defined a priori and based on previous literature. We studied a convenience sampling of shifts. Length of interruption time was recorded. Three investigators collected interruption data, including nine different ED attendings., Results: Of 511 interruptions, there were no significant differences in types of interruptions pre- versus post-HCD. Before HCD use, we collected 40 hours of data with 65 interruptions (mean 1.6/hour). Post-HCD, 180 hours of data were collected with 446 interruptions (mean 2.5/hour). There was a significant difference in length of time of interruptions, with an average of 9 minutes pre-HCD versus 4 minutes post-HCD (P = 0.012 diff 4.9; 95% confidence interval 1.8-8.1). HCD calls were significantly shorter than non-HCD calls (1 vs 5 minutes; P < 0.001). Comparing data collectors for type of interruption during the same 4-hour shift resulted in κ = 0.73., Conclusions: The addition of Vocera HCD may improve interruptions by shortening call length.

Background: The effect of aortic valve replacement on three-dimensional mitral annular geometry has not been well described. Emerging transcatheter approaches for aortic valve replacement employ fundamentally different mechanical techniques for achieving fixation and seal of the prosthetic valve than standard surgical aortic valve replacement. This study compares the immediate impact of transcatheter aortic valve replacement (TAVR) and standard surgical aortic valve replacement (AVR) on mitral annular anatomy., Methods: Real-time three-dimensional echocardiography was performed in patients undergoing TAVR using the Edwards Sapien valve (n = 10 [Edwards Lifesciences, Irvine, CA]) or AVR (n = 10) for severe aortic stenosis. Mitral annular geometric indexes were measured using Tomtec EchoView (Tomtec Imaging Systems, Munich, Germany) to assess regional and global annular geometry., Results: Mixed between-within analysis of variance showed no differences between TAVR and AVR groups in any of the mitral annular geometric indices preoperatively. However, postoperative analysis did demonstrate an effect of AVR on geometry. Patients undergoing open AVR had significant decrease in annular height, septolateral diameter, mitral valve transverse diameter, and mitral annular area after valve replacement (p ≤ 0.006). Similar changes were not noted in the TAVR group., Conclusions: Mitral annular geometry is better preserved by TAVR than by AVR. Thus, TAVR may be a more physiologic approach to aortic replacement., (Copyright © 2013 The Society of Thoracic Surgeons. Published by Elsevier Inc. All rights reserved.)

Natural and human-mediated population structure of European grayling Thymallus thymallus was assessed in primarily Austrian reaches of the Danube River catchment. Data on phylogeographic structure based on mitochondrial DNA (mtDNA) were combined with variation found across 13 microsatellite loci to assess introgression stemming from stocking activities. Populations north of the Alps generally showed no signs of long-term introgression from allochthonous stocks, although one population comprised two distinct genotypic groups, one of which may stem from recently stocked material. South of the Alps, levels of introgression with stocks from the north were extensive and only one of six populations is believed to represent a reference state. Allelic diversity and expected heterozygosity were positively correlated with mtDNA admixture supporting more recent introgression and that there is little or no reproductive barrier between these two major lineages north and south of the Alps. A third unrelated mtDNA lineage is described from the Lafnitz, a tributary of the Raab drainage. The high lineage diversity in the upper Austrian Danube is not concordant with an existing model of T. thymallus evolutionarily significant units in Europe. Caveats in naming such units or following a strict hierarchical conservation unit structure for broadly distributed species with complex phylogeographic distributions stretching over various jurisdictions are discussed. The necessity of using both phylogeographic and population genetic approaches in evaluating the history and conservation value of populations in a conservation context is additionally highlighted., (© 2013 The Authors. Journal of Fish Biology © 2013 The Fisheries Society of the British Isles.)

The identification of relevant gene targets for engineering a desired trait is a key step in combinatorial strain engineering. Here, we applied the multi-Scalar Analysis of Library Enrichments (SCALEs) approach to map ethanol tolerance onto 1,000,000 genomic-library clones in Escherichia coli. We assigned fitness scores to each of the ∼4,300 genes in E. coli, and through follow-up confirmatory studies identified 9 novel genetic targets (12 genes total) that increase E. coli ethanol tolerance (up to 6-fold improved growth). These genetic targets are involved in the processes related to cell membrane composition, translation, serine biosynthesis, and transcription regulation. Transcriptional profiling of the ethanol stress response in 5 of these ethanol-tolerant clones revealed a total of 700 genes with significantly altered expression (mapped to 615 significantly enriched gene ontology terms) across all five clones, with similar overall changes in global gene expression between two clone clusters. All ethanol-tolerant clones analyzed shared 6% of the overexpressed genes and showed enrichment for transcription regulation-related GO terms. iTRAQ-based proteomic analysis of ethanol-tolerant strains identified upregulation of proteins related to ROS mitigation, fatty acid biosynthesis, and vitamin biosynthesis as compared to the parent strain's ethanol response. The approach we outline here will be useful for engineering a variety of other traits and further improvements in alcohol tolerance., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Background: Acute aortic dissection (AAD) is a life-threatening vascular disease without effective pharmaceutical therapy. Matrix metalloproteinases (MMPs) are implicated in the development of chronic vascular diseases including aneurysm, but the key effectors and mechanism of action remain unknown. To define further the role of MMPs in AAD, we screened circulating MMPs in AAD patients, and then generated a novel mouse model for AAD to characterize the mechanism of action., Methods and Results: MMP9 and angiotensin II were elevated significantly in blood samples from AAD patients than in those from the patients with nonruptured chronic aortic aneurysm or healthy volunteers. Based on the findings, we established a novel AAD model by infusing angiotensin II to immature mice that had been received a lysyl oxidase inhibitor, β-aminopropionitrile monofumarate. AAD was developed successfully in the thoracic aorta by angiotensin II administration to β-aminopropionitrile monofumarate-treated wild-type mice, with an incidence of 20%, 80%, and 100% after 6, 12, and 24 hours, respectively. Neutrophil infiltrations were observed in the intima of the thoracic aorta, and the overexpression of MMP9 in the aorta was demonstrated by reverse transcription polymerase chain reaction, gelatin zymography, and immunohistochemistry. The incidence of AAD was reduced significantly by 40% following the administration of an MMP inhibitor and was almost blocked completely in MMP(-/-) mice without any influence on neutrophil infiltration. Neutrophil depletion by injection of anti-granulocyte-differentiation antigen-1 (anti-Gr-1) antibody also significantly decreased the incidence of AAD., Conclusions: These data suggest that AAD is initiated by neutrophils that have infiltrated the aortic intima and released MMP9 in response to angiotensin II.

Introduction: Many traumatic pneumothoraces (PTX) are not seen on initial chest radiograph (CR) (occult PTX) but are detected only on computed tomography (CT). Although CR remains the first tool for detecting PTX, most trauma patients with significant thoracoabdominal injuries will receive both CT and CR. The primary objective of this study was to retrospectively determine the effectiveness of CR for detecting PTX in trauma patients. Our hypotheses were that CR is a sensitive indicator of PTX on CT, that chest pain and shortness of breath are good predictors of PTX on CR, and that we could determine other predictors of PTX on CR., Methods: All trauma patients presenting to our Level I trauma center with a CT-diagnosed PTX over a 2-year period who had both a CR and a chest CT were included. The CT reading was considered the gold standard for PTX diagnosis. Electronic medical records were searched using key words for diagnoses, symptoms, demographics, and radiologic results. We recorded the official radiologist readings for both CR and CT (positive or negative) and the size of the PTX on CT (large, moderate, small, or tiny). The outcome variable was dichotomized based on presence or absence of PTX detected on CR. Descriptive statistics and χ(2) tests were used for univariate analysis. A regression analysis was performed to determine characteristics predictive of a PTX on CR, and 1 variable was added to the model for every 10 positive CRs. With equal-size groups, this study has the power of 80% to detect a 10% absolute difference in single predictors of PTX on CR with 45 subjects in each group., Results: There were 134 CT-documented PTXs included in the study. Mean age was 42, and 74% were men. For 66 (49%) patients, PTX was detected on CR (sensitivity = 50%). The CR detected 30% of small PTX, 35% of moderate PTX, and 33% of large PTX. Comparing patients with and without PTX on CR, there were no significant differences in shortness of breath or chest pain. There no relationships between PTX detected on CR and age, gender, penetrating versus blunt injury, bilaterality of the PTX, or presence of lung contusion or hemothorax on CT. After adjusting for all significant variables, predictor of a PTX detected on CR was air in the tissue on CR (adjusted odds ratio [OR] = 3.8) and PTX size (compared to a tiny PTX, adjusted OR = 2.0 for a small PTX, 7.5 for a moderate PTX, and 51 for a large PTX). Chest tubes were used in 89% of patients with PTX on CR and 44% of patients with PTX only on CT (difference 45%; 95% confidence interval 30, 58)., Conclusion: Factors associated with PTX on CR included air in the soft tissue on CR and size of the PTX. Even when PTX is not apparent on CR, 44% of these PTXs received placement of a chest tube.

Slug (Snail2) plays critical roles in regulating the epithelial-mesenchymal transition (EMT) programs operative during development and disease. However, the means by which Slug activity is controlled remain unclear. Herein we identify an unrecognized canonical Wnt/GSK3β/β-Trcp1 axis that controls Slug activity. In the absence of Wnt signaling, Slug is phosphorylated by GSK3β and subsequently undergoes β-Trcp1-dependent ubiquitination and proteosomal degradation. Alternatively, in the presence of canonical Wnt ligands, GSK3β kinase activity is inhibited, nuclear Slug levels increase, and EMT programs are initiated. Consistent with recent studies describing correlative associations in basal-like breast cancers between Wnt signaling, increased Slug levels, and reduced expression of the tumor suppressor Breast Cancer 1, Early Onset (BRCA1), further studies demonstrate that Slug-as well as Snail-directly represses BRCA1 expression by recruiting the chromatin-demethylase, LSD1, and binding to a series of E-boxes located within the BRCA1 promoter. Consonant with these findings, nuclear Slug and Snail expression are increased in association with BRCA1 repression in a cohort of triple-negative breast cancer patients. Together, these findings establish unique functional links between canonical Wnt signaling, Slug expression, EMT, and BRCA1 regulation.

Background: Antiemetics have been shown to be effective in multiple hospital settings, but few studies have been done in the prehospital environment., Objectives: Our hypotheses were 1) that the amount of normal saline administered during an emergency medical services (EMS) transport was not related to a change in nausea and vomiting and 2) that the addition of the ondansetron orally disintegrating tablet (ODT) would decrease the degree of nausea., Methods: This was a pre-post study of two cohorts of consecutive patients with nausea in the prehospital setting. During phase 1 of the study, our local EMS agency adopted a protocol form to complete whenever a patient with nausea and/or vomiting was assessed and transported to one of the area hospitals. Patients were asked to rate their nausea on a visual analog scale (VAS) and a Likert scale, and saline administration and active vomiting were documented. During phase 2, our EMS system adopted the use of ondansetron ODT for nausea and continued to complete the same forms. The nausea forms completed by EMS during phase 1 (saline only) and phase 2 (ondansetron ODT) were evaluated and compared. For both phases, the primary outcome measures were the change in VAS nausea rating (0 = no nausea, 100 = most nausea imaginable) from beginning to end of the transport and the results on the Likert scale completed at the end of the transport. Relationships were considered significant if p < 0.01., Results: Data were collected from 274 transports in phase 1 and 372 transports in phase 2. The average patient age was 50 ± 12 years. In phase 1 of the study, 178 of 274 patients (65%) received normal saline (mean volume ± standard deviation = 265 ± 192 mL). There was no significant correlation between the VAS change and the amount of fluid administration in either phase of the study. Conversely, during phase 2, patients receiving ondansetron ODT showed significant improvement in both measures of nausea. The difference in nausea improvement between phase 1 and phase 2 was significant (difference in VAS change: 24.6; 95% confidence interval 20.9, 28.3)., Conclusion: There was no improvement in patient nausea related to quantity of saline alone during an EMS transport. The addition of ondansetron ODT resulted in a significant improvement in degree of nausea.

Type IV collagen, a major constituent of basement membranes, contains an unusual intermolecular sulfilimine crosslink whose route of biosynthesis has remained undefined. An oxidative triad consisting of peroxidasin, H(2)O(2) and halide is now shown to drive sulfilimine generation in vivo.

Background: A method for reducing the power of drug cues could help in treating drug abuse and addiction. Extinction has been used, with mixed success, in such an effort. Research with non-drug cues has shown that simultaneously presenting (compounding) those cues during extinction can enhance the effectiveness of extinction. The present study investigated whether this procedure could be used to similarly deepen the extinction of cocaine cues., Methods: Rats were first trained to self-administer cocaine during tone, click, and light stimuli. Then, these stimuli were subjected to extinction in an initial phase where they were presented individually. In a second extinction phase, one of the auditory stimuli (counterbalanced) was compounded with the light. The other auditory stimulus continued to be presented alone. Rats were then given a week of rest in their homecages prior to testing for spontaneous recovery of cocaine seeking., Results: The cue that was compounded with the light during the second phase of extinction training occasioned less spontaneous recovery of cocaine seeking than the cue that was always presented individually during extinction. Increasing the number of compound cue extinction sessions did not produce a greater deepened extinction effect., Conclusions: The present study showed that simultaneously presenting already-extinguished cocaine cues during additional extinction training enhanced extinction. This extends the deepened extinction effect from non-drug cues to drug cues and further confirms predictions of error-correction learning theory. Incorporating deepened extinction into extinction-based drug abuse treatments could help to reduce the power of drug cues., (Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.)

Aberrant activation of canonical Wingless-type MMTV integration site family (Wnt) signaling is pathognomonic of colorectal cancers (CRC) harboring functional mutations in either adenomatous polyposis coli or β-catenin. Coincident with Wnt cascade activation, CRCs also up-regulate the expression of Wnt pathway feedback inhibitors, particularly the putative tumor suppressor, Axin2. Because Axin2 serves as a negative regulator of canonical Wnt signaling in normal cells, recent attention has focused on the utility of increasing Axin2 levels in CRCs as a means to slow tumor progression. However, rather than functioning as a tumor suppressor, we demonstrate that Axin2 acts as a potent promoter of carcinoma behavior by up-regulating the activity of the transcriptional repressor, Snail1, inducing a functional epithelial-mesenchymal transition (EMT) program and driving metastatic activity. Silencing Axin2 expression decreases Snail1 activity, reverses EMT, and inhibits CRC invasive and metastatic activities in concert with global effects on the Wnt-regulated cancer cell transcriptome. The further identification of Axin2 and nuclear Snail1 proteins at the invasive front of human CRCs supports a revised model wherein Axin2 acts as a potent tumor promoter in vivo.

Objective: The objective of the study was to compare errors in the emergency department (ED) with pharmacists present (PPs) for resuscitations and traumas vs with pharmacists absent (PAs). Our hypothesis was that errors would be significantly fewer during PP than PA times. We also hypothesized that times with PP would affect patients greater when disposition was to more critical areas (intensive care unit, or ICUs)., Methods: The study was conducted during a 3-month period in 2009 in a level 1 trauma center with an emergency medicine residency. This was a cross-sectional cohort study comparing a prospective analysis of patients during the time (10 hour/day) with PP and a retrospective review of the time on the same days (14 hours/day) with PA. Demographics of age, race, and sex were recorded. Patient disposition was either ICU, operating room, non-ICU wards, observation unit, or discharge. Main outcome was errors recorded including medications given but not ordered, medication ordered but not given, and time delays for medications. For demographics and prevalence, descriptive statistics and percentages were used. Percent differences and 95% confidence intervals (CIs) and χ2 were derived. Logistic regression used predictor variables of age, race, sex, disposition, and presence or absence of pharmacists. An a priori power analysis was performed. The study was powered at 80% with 186 subjects per group (PP vs PA), to find a difference of 20% between the 2 groups in percent of medical errors., Results: There were 694 patients included in the 3-month period. A total of 242 presented during PP times and 452 during PA times. There were 383 (55%) male, 301 (43%) female, and 10 (2%) unknown sex. Mean age was 45±18 years in PP group and 48±20 years in PA group (P, nonsignificant). There was no difference in ethnicity between groups. There were 6 (3%) patients with errors recorded during PP times and 137 (30%) with errors recorded during PA times (difference, 27%; 95% CI, 23-32). Controlling for age, race, sex, and disposition, medical errors were 13.5 times more likely during PA than during PP times (adjusted odds ratio, 13.5; 95% CI, 5.7-31.9)., Conclusion: With pharmacists absent, over 13 times more errors are recorded in our ED than with pharmacists present. An on-site pharmacist in the ED may be helpful in reducing medical errors., (Copyright © 2012 Elsevier Inc. All rights reserved.)