Repurposing DNase I and alginate lyase to degrade the biofilm matrix of dual-species biofilms of Staphylococcus aureus and Pseudomonas aeruginosa grown in artificial sputum medium: In-vitro assessment of their activity in combination with broad-spectrum antibiotics.

BACKGROUND: Biofilm-associated pulmonary infections pose therapeutic challenges in cystic fibrosis patients, especially when involving multiple bacterial species. Enzymatic degradation of the biofilm matrix may offer a potential solution to enhance antibiotic efficacy. This study investigated the repurposing of DNase I, commonly used for its mucolytic activity in cystic fibrosis, to target extracellular DNA within biofilms, as well as potential synergies with alginate lyase and broad-spectrum antibiotics in dual-species biofilms of Pseudomonas aeruginosa and Staphylococcus aureus. METHODS: Dual-species biofilms were grown in artificial sputum medium using S. aureus and P. aeruginosa isolated by pairs from the same patients and exposed to various combinations of enzymes, meropenem, or tobramycin. Activity was assessed by measuring biofilm biomass and viable counts. Matrix degradation and decrease in bacterial load were visualized using confocal microscopy. Biofilm viscoelasticity was estimated by rheology. RESULTS: Nearly complete destruction of the biofilms was achieved only if combining the enzymatic cocktail with the two antibiotics, and if using supratherapeutic levels of DNase I and high concentrations of alginate lyase. Biofilms containing non-pigmented mucoid P. aeruginosa required higher antibiotic concentrations, despite low viscoelasticity. In contrast, for biofilms with pigmented mucoid P. aeruginosa, a correlation was observed between the efficacy of different treatments and the reduction they caused in elasticity and viscosity of the biofilm. CONCLUSIONS: In this complex, highly drug-tolerant biofilm model, enzymes prove useful adjuvants to enhance antibiotic activity. However, the necessity for high enzyme concentrations emphasizes the need for thorough concentration-response evaluations and safety assessments before considering clinical applications.

Dimethyl Sulfide (DMS) in Amarone Wines: Influence of Aging, Withering, Grape Variety, and Geographical Origin.

Occurrence of dimethyl sulfide (DMS), a potent aroma compound accumulating during aging, was investigated in commercial and experimental Amarone wines. In commercial Amarone, DMS was observed in concentrations ranging from 2.9 to 64.3 µg/L. Model aging studies on experimental wines indicated that DMS in Amarone is strongly associated with aging and that wines from different vineyards can vary significantly in their ability to accumulate DMS during aging. The capacity of certain vineyards to give wines with higher DMS-forming potential was consistent across three consecutive vintages, representing a true terroir factor to be expressed with aging. Wine content of primary amino acids (PAN), a commonly analyzed enological parameter of grape must, was shown to be positively correlated with DMS accumulation during aging. Grape withering also increased DMS-forming potential mostly due to increased PAN resulting from concentration due to water loss. Increased pH due to withering also contributed to a higher DMS content of withered wines, but to a lower extent. In certain vineyard sites, an influence of vintage conditions on DMS-forming potential was also observed.

Analysis of Adverse Drug Reactions in Pediatric Patients with Epilepsy: An Intensive Pharmacovigilance Study.

Epilepsy is a chronic neurological disease characterized by the presence of spontaneous seizures, with a higher incidence in the pediatric population. Anti-seizure medication (ASM) may produce adverse drug reactions (ADRs) with an elevated frequency and a high severity. Thus, the objective of the present study was to analyze, through intensive pharmacovigilance over 112 months, the ADRs produced by valproic acid (VPA), oxcarbazepine (OXC), phenytoin (PHT), and levetiracetam (LEV), among others, administered to monotherapy or polytherapy for Mexican hospitalized pediatric epilepsy patients. A total of 1034 patients were interviewed; 315 met the inclusion criteria, 211 patients presented ADRs, and 104 did not. A total of 548 ASM-ADRs were identified, and VPA, LEV, and PHT were the main culprit drugs. The most frequent ADRs were drowsiness, irritability, and thrombocytopenia, and the main systems affected were hematologic, nervous, and dermatologic. LEV and OXC caused more nonsevere ADRs, and PHT caused more severe ADRs. The risk analysis showed an association between belonging to the younger groups and polytherapy with ADR presence and between polytherapy and malnutrition with severe ADRs. In addition, most of the severe ADRs were preventable, and most of the nonsevere ADRs were nonpreventable.

An e-learning pediatric cardiology curriculum for Pediatric Postgraduate trainees in Rwanda: implementation and evaluation.

BACKGROUND: Access to pediatric sub-specialty training is a critical unmet need in many resource-limited settings. In Rwanda, only two pediatric cardiologists are responsible for the country's clinical care of a population of 12 million, along with the medical education of all pediatric trainees. To strengthen physician training opportunities, we developed an e-learning curriculum in pediatric cardiology. This curriculum aimed to "flip the classroom", allowing residents to learn key pediatric cardiology concepts digitally before an in-person session with the specialist, thus efficiently utilizing the specialist for additional case based and bedside teaching. METHODS: We surveyed Rwandan and US faculty and residents using a modified Delphi approach to identify key topics in pediatric cardiology. Lead authors from Rwanda and the USA collaborated with OPENPediatrics™, a free digital knowledge-sharing platform, to produce ten core topics presented in structured videos spanning 4.5 h. A mixed methods evaluation was completed with Rwandan pediatric residents, including surveys assessing knowledge, utilization, and satisfaction. Qualitative analysis of structured interviews was conducted using NVivo. RESULTS: Among the 43 residents who participated in the OPENPediatrics™ cardiology curriculum, 33 (77%) completed the curriculum assessment. Residents reported using the curriculum for a median of 8 h. Thirty-eight (88%) reported viewing the curriculum on their personal or hospital computer via pre-downloaded materials on a USB flash drive, with another seven (16%) reporting viewing it online. Twenty-seven residents viewed the course during core lecture time (63%). Commonly reported barriers to utilization included lack of time (70%), access to internet (40%) and language (24%). Scores on knowledge assessment improved from 66.2% to 76.7% upon completion of the curriculum (p < 0.001) across all levels of training, with most significant improvement in scores for PGY-1 and PGY-2 residents. Residents reported high satisfaction with the visuals, engaging presentation, and organization of the curriculum. Residents opined the need for expanded training material in cardiac electrocardiogram and echocardiogram and requested for slower narration by foreign presenters. CONCLUSION: Video-based e-learning via OPENPediatrics™ in a resource-limited setting was effective in improving resident's knowledge in pediatric cardiology with high levels of utilization and satisfaction. Expanding access to digital curriculums for other pediatric sub-specialties may be both an effective and efficient strategy for improving training in settings with limited access to subspecialist faculty.

Two Sides to One Story-Aroma Chemical and Sensory Signature of Lugana and Verdicchio Wines.

Lugana and Verdicchio are two Italian white wines with a Protected Designation of Origin (PDO) label. These two wine types are produced in different regions using the same grape variety. The aim of this work is to investigate the existence of volatile chemical markers that could help to elucidate differences between Lugana and Verdicchio wines both at chemical and sensory levels. Thirteen commercial wine samples were analyzed by Gas Chromatography-Mass Spectrometry (GC-MS), and 76 volatile compounds were identified and quantified. Verdicchio and Lugana had been differentiated on the basis of 19 free and glycosidically bound compounds belonging to the chemical classes of terpenes, benzenoids, higher alcohols, C6 alcohols and norisoprenoids. Samples were assessed by means of a sorting task sensory analysis, resulting in two clusters formed. These results suggested the existence of 2 product types with specific sensory spaces that can be related, to a good extend, to Verdicchio and Lugana wines. Cluster 1 was composed of six wines, 4 of which were Lugana, while Cluster 2 was formed of 7 wines, 5 of which were Verdicchio. The first cluster was described as "fruity", and "fresh/minty", while the second as "fermentative" and "spicy". An attempt was made to relate analytical and sensory data, the results showed that damascenone and the sum of 3 of esters the ethyl hexanoate, ethyl octanoate and isoamyl acetate, was characterizing Cluster 1. These results highlighted the primary importance of geographical origin to the volatile composition and perceived aroma of Lugana and Verdicchio wines.

A Group Visit for High-Risk Pediatric Asthma Patients: A Quality Improvement Initiative to Improve Asthma Care.

INTRODUCTION: Asthma disproportionately affects poor and minority children. Limited parental knowledge and confidence in asthma management, as well as stress from chronic illness, may contribute to poor outcomes. Novel approaches for providing care are essential for this vulnerable population. Our objective was to evaluate the feasibility and impact of an asthma group visit for high-risk children. METHODS: Our primary care practice cares for more than 2600 children with asthma. The majority have public insurance. Children classified as high risk (≥1 asthma-related emergency department visit/hospitalization in the preceding 2 years) were eligible. Children received brief physical examinations, medication review, and updated Asthma Action Plans. Educational sessions were held for children and parents. Pre and post surveys were used to assess parents' experience and changes in confidence in asthma management. RESULTS: Twenty children and their parents participated. Mean parent confidence scores (5-point Likert-type scale, 5 indicating greatest confidence) improved in managing their child's asthma symptoms (3.60, 4.40, P ≤ .005), managing their child's asthma medications (3.85, 4.30, P ≤ .005), using their child's Asthma Action Plan (3.79, 4.45, P ≤ .02), communicating with the school about their child's food allergies (4.32, 4.72, P ≤ .03), and helping their child relax to reduce emotional triggers of asthma (3.25, 4.47, P ≤ .01). All families reported that they would return to a group visit. CONCLUSION: Group visits are feasible for providing care, education, and peer support to a vulnerable population. Parents expressed satisfaction and improved confidence in aspects of asthma management. Group visits have the potential to improve asthma outcomes for high-risk families.

Short- and long-term effects of leucine and branched-chain amino acid supplementation of a protein- and energy-reduced diet on muscle protein metabolism in neonatal pigs.

The objective of this study was to determine if enteral leucine or branched-chain amino acid (BCAA) supplementation increases muscle protein synthesis in neonates who consume less than their protein and energy requirements, and whether this increase is mediated via the upregulation of the mechanistic target of rapamycin complex 1 (mTORC1) pathway or the decrease in muscle protein degradation signaling. Neonatal pigs were fed milk replacement diets containing reduced energy and protein (R), R supplemented with BCAA (RBCAA), R supplemented with leucine (RL), or complete protein and energy (CON) at 4-h intervals for 9 (n = 24) or 21 days (n = 22). On days 9 and 21, post-prandial plasma amino acids and insulin were measured at intervals for 4 h; muscle protein synthesis rate and activation of mTOR-related proteins were determined at 120 min post-feeding in muscle. For all parameters measured, the effects of diet were not different between day 9 or day 21. Compared to CON and R, plasma leucine and BCAA were higher (P ≤ 0.01) in RL- and RBCAA-fed pigs, respectively. Body weight gain, protein synthesis, and activation of S6 kinase (S6K1), 4E-binding protein (4EBP1), and eukaryotic initiation factor 4 complex (eIF4E·eIF4G) were decreased in RBCAA, RL, and R relative to CON (P < 0.01). RBCAA and RL upregulated (P ≤ 0.01) S6K1, 4EBP1, and eIF4E·eIF4G compared to R. In conclusion, when protein and energy are restricted, both leucine and BCAA supplementation increase mTOR activation, but do not enhance skeletal muscle protein synthesis and muscle growth in neonatal pigs.

Examining Cohort Effects in Developmental Trajectories of Substance Use.

The current study demonstrates the application of an analytic approach for incorporating multiple time trends in order to examine the impact of cohort effects on individual trajectories of eight drugs of abuse. Parallel analysis of two independent, longitudinal studies of high-risk youth that span ages 10 to 40 across 23 birth cohorts between 1968 and 1991 was conducted. The two studies include the Michigan Longitudinal Study (current analytic sample of n=579 over 12 cohorts between 1980-1991 and ages 10-27) and the Adolescent/Adult and Family Development Project (current analytic sample of n=849 over 11 cohorts between 1968-1978 and ages 10-40). A series of nonlinear, multi-level growth models controlled simultaneously for cohort and age trends in substance use trajectories. Evidence was found for both age and cohort effects across most outcomes as well as several significant age-by-cohort interactions. Findings suggest cohort trends in developmental trajectories of substance use are sample and drug-specific in the adolescent and early to mid-adult years. Thus, studies that do not control for both trends may confound cohort and developmental trends in substance use. For this reason, demonstration of one analytic approach that can be used to examine both time trends simultaneously is informative for future multi-cohort longitudinal studies where change over time is of interest.

Profiling families in conflict: Multigenerational continuity in conflict predicts deleterious adolescent and adult outcomes.

The present study investigated the associations between multigenerational continuity in family conflict and current psychopathology symptoms and social impairment experienced by parents and adolescents. We sampled 246 families from a multigenerational, high-risk, longitudinal study of parents (G1s) and their children (G2s), followed from adolescence (Mage = 14.3 years, 57% female, 71% Caucasian, 26% Hispanic or Latino) to adulthood as well as the children of G2 targets (G3s; Mage = 12.1 years, 47% female, 51% Caucasian, 33% Hispanic or Latino). Family conflict was measured by composite latent variables incorporating mother, father, and adolescent reports in G1-G2 families and incorporating G2 target, G2 target's spouse, and G3 adolescent report in G2-G3 families. Indicators of G2 and G3 impairment including psychopathology symptoms (e.g., internalizing, externalizing, and substance use symptoms) and social role impairment (e.g., marital satisfaction, parenting behavior) were predicted from G1-G2 family conflict, G2-G3 family conflict, and the interaction between G1-G2 and G2-G3 family conflict. Results indicate that G1-G2 family conflict uniquely predicted indicators of G2 and G3 psychopathology, as well as G2 social impairment, even after controlling for more temporally proximal G2-G3 family conflict. Results further indicate that for G2 externalizing, internalizing, and marital functioning outcomes, high G2-G3 family conflict was associated with highest G2 impairment when G1-G2 family conflict was also high. It appears that for many G2 outcomes the interactive effects of multigenerational conflict are associated with greater risk for impairment. (PsycINFO Database Record

Comparing Psychiatric Symptoms among Children of Substance-Abusing Parents With Different Treatment Histories.

It appears that no studies to date have compared the psychiatric functioning of children of substance-abusing parents (COSs) across substance abuse treatment histories (e.g., inpatient, outpatient, residential). Different treatment histories may reflect differences in the severity of drug use, degree of impairment, or drug of choice, which may result in different symptomatology among COSs. Moreover, this information may be important for family-based treatment and prevention planning. In the current study, we examine psychiatric symptoms experienced by children whose parents have different addiction treatment histories. Eighty-one parents in treatment for a substance use disorder who had a 2-8 year old child completed an anonymous, computerized assessment. Respondents reported on their child's symptomatology as well as their own treatment history and current substance use. Overall, children were remarkably similar in psychiatric functioning and demographic characteristics across parent's treatment histories. However, children whose parents had only received medication management or detoxification had significantly higher somatization scores, more social withdrawal, and greater attention problems than children whose parents received outpatient treatment (but not inpatient/residential treatment). Children whose parents had been treated in an inpatient or residential setting had marginally higher attention problems than children whose parents received outpatient treatment (but not inpatient/residential treatment). Differences across treatment histories were reflected in the severity of psychiatric symptomatology in the young children of parents in treatment. However, given the modest size of these differences, prevention and intervention programs with children of substance-abusing parents may not need to be tailored as a function of parental treatment history as recruitment from various treatment centers may provide a sample with similar characteristics and risk profiles.

Future directions in the developmental science of addictions.

This article addresses important future directions for the study of addictions, emphasizing the incorporation of developmental perspectives into how we think about substance use and disorder as unfolding processes over time and context for a heterogeneous group of individuals. These perspectives articulate complexities in the developmental processes that underlie change and continuity in human behavior over time. We consider two key developmental concepts, namely, "time" and "heterogeneity." We argue that a lack of attention to time sampling creates ambiguity in the meaning of time-linked assessments, challenges in discerning which of multiple clocks may govern behavior, and the inability in some instances to distinguish which of multiple etiological processes may be driving behavior within our samples. Moreover, artificial divisions among disorders that commonly co-occur with substance use are a barrier to the further integration of the study and treatment of addictions with that of psychopathology. Similar to recent changes in the study of psychiatric disorders more broadly, we argue that identifying common deficits among commonly comorbid disorders, rather than patterns of comorbidity per se, is key to identifying early emerging risk factors for substance use and disorder, with important implications for identifying risk populations and developmental periods as well as potentially malleable intervention targets. Attention to time sampling in theory-driven research designs and attempts to identify more homogenous groups of individuals who use and eventually abuse substances over time are two examples of ways to better understand some of the complexity underlying the development of addictions.

Advancements in using TiO2 bionanoconjugates for precision degradation of intracellular biological structures.

Due to their low cost, photocatalytic properties, and unique surface chemistry, titanium dioxide (TiO2) nanoparticles are among the most widely used nanoparticles in industry today. Over the last decade, TiO2 nanoparticles have also been chemically and biologically enhanced to create TiO2 bionanoconjugates that can be used for biological applications such as imaging and manipulating desired biological structures. This review particularly focuses on the manner in which these specific chemical and biological modifications in TiO2 bionanoconjugates alter pre and post photoexcitation events to enable precision degradation of intracellular biological structures. Secondary emphasis is given to imaging aspects when necessary to understanding the effects that targeting these bionanoconjugates has on degradation of neighboring biological structures. The advantages of TiO2 bionanoconjugates to standard techniques, as well as future research directions, are also discussed.

Propiedades antioxidantes de los frutos secos y la disminución del colesterol total y LDL-colesterol / Antioxidant properties of nuts and decreased total and LDL cholesterol

Disminuir el colesterol total y el LDL colesterol mediante el consumo de frutos secos (maní, nuez y almendras) con propiedades antioxidantes para reducir el riesgo de enfermedades cardiovasculares. Método: El presente trabajo es cuantitativo transversal, cuasi-experimental. Se tomó una muestra de 45 participantes del Instituto Interamericano de Cooperación para la Agricultura (IICA), ambos sexos, entre 18 y 65 años de edad, con hipercolesterolemia total mayor a 200 mg/dl y LDL-colesterol mayor a 100 mg/dl uno a ambos alterados. Sin plan nutricional establecido, sin antecedentes de alcohol, tabaco, alergias a frutos secos, hipotiroidismo, mantenimiento actividad física usual y no consumo de estatinas. Los frutos secos fueron maní, nueces y almendras, distribuidos en tres grupos, cada participante consumió 40 gramos diarios de un tipo de fruto seco por seis semanas. Se realizó un pre y post test de examen bioquímico de lípidos intravenoso. Instrumentos utilizados: cuestionario para recolección de datos y pesa de alimentos. Estudio realizado con una confianza estadística estimada (95 por ciento). Resultado: Todos los participantes presentan niveles altos de ambos tipos de colesterol, uno u otro. En promedio estadístico el colesterol total disminuyó 12,7 mg/dl y el LDL colesterol disminuyó 10.88 mg/dl. El colesterol total promedio estadístico disminuye 11,7 mg/dl consumiendo maní, 7,7 mg/dl, consumiendo almendras y 19.4 mg/dl consumiendo nueces. Discusión: El consumo de los frutos secos está asociado con la disminución del colesterol total y LDL colesterol, por sus propiedades antioxidantes, que tiene un afecto protector contra enfermedades cardiovasculares...

A pilot randomized controlled trial of brief cognitive-behavioral therapy for anxiety in patients with terminal cancer.

INTRODUCTION: Patients with terminal cancer often experience marked anxiety that is associated with poor quality of life. Although cognitive-behavioral therapy (CBT) is an evidence-based treatment for anxiety disorders, the approach needs to be adapted to address realistic concerns related to having cancer, such as worries about disease progression, disability, and death. In this pilot randomized controlled trial (clinicaltrials.gov identifier NCT00706290), we examined the feasibility and potential efficacy of brief CBT to reduce anxiety in patients with terminal cancer. METHODS: We adapted CBT by developing treatment modules targeting skills for relaxation, coping with cancer worries, and activity pacing. Adults with incurable malignancies and elevated anxiety based on the Hamilton Anxiety Rating Scale (HAM-A) were randomly assigned to individual CBT or a waitlist control group. Primary outcomes included the number of completed CBT visits and the change in HAM-A scores from baseline to 8-week follow-up per a treatment-blind evaluator. The feasibility criterion was 75% adherence to the intervention. RESULTS: We randomized 40 patients with terminal cancers to CBT (n = 20) or waitlist control (n = 20) groups; 70% completed posttreatment assessments. Most patients who received CBT (80%) participated in at least five of the required six therapy sessions. Analysis of covariance models, adjusted for baseline scores, showed that those assigned to CBT had greater improvements in HAM-A scores compared to the control group, with an adjusted mean difference of -5.41 (95% confidence interval: -10.78 to -0.04) and a large effect size for the intervention (Cohen's d = 0.80). CONCLUSION: Providing brief CBT tailored to the concerns of patients with terminal cancer was not only feasible but also led to significant improvements in anxiety.

Understanding the diverse needs of children whose parents abuse substances.

In this review, we consider the potential service needs of children of substance abusing parents based on what we know about the risk outcomes faced by these children and the parenting deficits often present in these families. Importantly, our review does not address the etiological role of parental substance abuse in children's negative outcomes but instead we discuss the complex inter-related risk factors that often co-occur with and exacerbate risk associated with parental alcohol and drug use. We first review studies showing the elevated risk that children of substance abusing parents face in general for poorer academic functioning; emotional, behavioral, and social problems; and an earlier onset of substance use, faster acceleration in substance use patterns, and higher rates of alcohol and drug use disorders. We then review studies showing contextual risk factors for children of substance abusing parents, including parenting deficits (less warmth, responsiveness, and physical and verbal engagement as well as harsher and more over-involved interaction styles), greater risk for child maltreatment, and less secure attachment patterns. We conclude with a discussion of future directions for research and guidelines for professionals working with children and their families where parental substance abuse is present.

Anxiety disorders in long-term survivors of adult cancers.

BACKGROUND: Little is known about the prevalence of anxiety disorders among long-term survivors of adult cancers. Using data from the National Comorbidity Survey Replication (NCS-R), we compared rates of anxiety disorders between long-term cancer survivors and individuals without a history of cancer. METHODS: A nationally representative sample of 9282 adults participated in a household survey to assess the prevalence of DSM-IV psychiatric disorders, a subset of whom also answered questions about medical comorbidities, including cancer. Long-term survivors were defined as those who received an adult cancer diagnosis at least 5 years before the survey. Multiple logistic regression analyses were used to examine associations between cancer history and anxiety disorders in the past year. RESULTS: The NCS-R sample consisted of 225 long-term cancer survivors and 5337 people without a history of cancer. Controlling for socio-demographic variables, long-term cancer survivors were more likely to have an anxiety disorder (odds ratio [OR]: 1.49, 95% confidence interval [CI]: 1.04-2.13), including specific phobia (OR: 1.59, 95% CI: 1.06-2.44) and medical phobia (OR: 3.45, 95% CI: 1.15-10.0), during the past 12 months compared with those without cancer histories. Rates for social anxiety disorder, generalized anxiety disorder, posttraumatic stress disorder, panic disorder, and agoraphobia were not significantly different between groups. CONCLUSION: Long-term survivors of adult cancers were more likely to have an anxiety disorder diagnosis, namely specific phobia, in the past 12 months compared with the general public. Further longitudinal study is needed to clarify the timing and course of anxiety relative to the cancer diagnosis.

Code status documentation in the outpatient electronic medical records of patients with metastatic cancer.

BACKGROUND: Advanced care planning (ACP) is considered an essential component of medical care in the United States, especially in patients with incurable diseases. However, little is known about clinical practices in outpatient oncology settings related to discussing end-of-life care and documenting code status preferences in ambulatory medical records. OBJECTIVE: To assess the rate of documentation of code status in the electronic longitudinal medical records (LMR) of patients with metastatic cancer. DESIGN: Retrospective review of 2,498 patients with metastatic solid tumors at an academic cancer center. An electronic patient database and the LMR were queried to identify demographic information, cancer type, number of clinic visits, and documentation of code status. PARTICIPANTS: The sample consisted of adult patients with metastatic prostate, breast, ovarian, bladder kidney, colorectal, non-colorectal gastrointestinal (GI), and lung cancers. MEASUREMENTS: Primary outcome was the percentage of documented code status in the LMR. MAIN RESULTS: Among the 2,498 patients, 20.3% had a documented code status. Code status was designated most frequently in patients with non-colorectal GI (193/609, 31.7%) and lung (179/583, 30.7%) cancers and least frequently in patients with genitourinary malignancies [bladder/kidney (4/89, 4.5%), ovarian (4/93, 4.3%), and prostate (7/365, 1.9%) cancers]. Independent predictors of having documented code status included religious affiliation, cancer type, and a greater number of visits to the cancer center. Younger patients and black patients were less likely to be designated as DNR/DNI. CONCLUSIONS: Despite the incurable nature of metastatic cancer, only a minority of patients had a code status documented in the electronic medical record.

Circadian genes are expressed during early development in Xenopus laevis.

BACKGROUND: Circadian oscillators are endogenous time-keeping mechanisms that drive twenty four hour rhythmic changes in gene expression, metabolism, hormone levels, and physical activity. We have examined the developmental expression of genes known to regulate circadian rhythms in order to better understand the ontogeny of the circadian clock in a vertebrate. METHODOLOGY/PRINCIPAL FINDINGS: In this study, genes known to function together in part of the core circadian oscillator mechanism (xPeriod1, xPeriod2, and xBmal1) as well as a rhythmic, clock-controlled gene (xNocturnin) were analyzed using in situ hybridization in embryos from neurula to late tailbud stages. Each transcript was present in the developing nervous system in the brain, eye, olfactory pit, otic vesicle and at lower levels in the spinal cord. These genes were also expressed in the developing somites and heart, but at different developmental times in peripheral tissues (pronephros, cement gland, and posterior mesoderm). No difference was observed in transcript levels or localization when similarly staged embryos maintained in cyclic light were compared at two times of day (dawn and dusk) by in situ hybridization. Quantitation of xBmal1 expression in embryonic eyes was also performed using qRT-PCR. Eyes were isolated at dawn, midday, dusk, and midnight (cylic light). No difference in expression level between time-points was found in stage 31 eyes (p = 0.176) but stage 40 eyes showed significantly increased levels of xBmal1 expression at midnight (RQ = 1.98+/-0.094) when compared to dawn (RQ = 1+/-0.133; p = 0.0004). CONCLUSIONS/SIGNIFICANCE: We hypothesize that when circadian genes are not co-expressed in the same tissue during development that it may indicate pleiotropic functions of these genes that are separate from the timing of circadian rhythm. Our results show that all circadian genes analyzed thus far are present during early brain and eye development, but rhythmic gene expression in the eye is not observed until after stage 31 of development.