Burnout and Anxiety Among Trauma Nursing Specialties in a Rural Level I Trauma Center.

BACKGROUND: Burnout and anxiety compromise physical and mental well-being of nurses and jeopardize patient safety. Personal, professional, and workplace characteristics have been associated with burnout and anxiety across diverse practice settings, yet none in rural, community trauma centers. We sought to identify the severity and predictors of burnout and anxiety in the trauma nursing staff of a rural Level I trauma center. METHODS: A convenience sample of trauma nurses from the emergency department (ED), intensive care unit (ICU), and trauma ward was voluntarily surveyed using a demographic questionnaire, the Maslach Burnout Inventory (MBI) subscales: depersonalization (DP), emotional exhaustion (EE), and reduced personal accomplishment, as well as the Generalized Anxiety Disorder seven-item (GAD-7) scale. Multivariable linear regression identified the significant predictors of burnout and anxiety. RESULTS: Ninety-six nurses completed surveys (response rate: 83.5%). Married or divorced status, and ICU or trauma ward job assignments were associated with significantly lower adjusted DP scores. Thus, the model-predicted score for a single ED nurse was 15 versus a predicted score of 7 for a divorced ICU or trauma ward nurse, p < .001 for each group. The GAD-7 model demonstrated that race/ethnicity (Asian compared with White, coefficient: -5.06, p = .03), number of children (2 compared with 0, coefficient: -2.54, p = .02), and job tenure (5-10 years vs. <2, coefficient: -3.18, p = .01) were each associated with fewer GAD-7 points. CONCLUSION: Depersonalization and anxiety vary across the trauma nursing workforce based on identifiable personal and work-related risk factors. Group-specific, targeted interventions are needed to effectively reduce burnout and anxiety in trauma nursing staff.

The role of genomic imprinting in biology and disease: an expanding view.

Genomic imprinting is an epigenetic phenomenon that results in monoallelic gene expression according to parental origin. It has long been established that imprinted genes have major effects on development and placental biology before birth. More recently, it has become evident that imprinted genes also have important roles after birth. In this Review, I bring together studies of the effects of imprinted genes from the prenatal period onwards. Recent work on postnatal stages shows that imprinted genes influence an extraordinarily wide-ranging array of biological processes, the effects of which extend into adulthood, and play important parts in common diseases that range from obesity to psychiatric disorders.

Loss of Gnas imprinting differentially affects REM/NREM sleep and cognition in mice.

It has been suggested that imprinted genes are important in the regulation of sleep. However, the fundamental question of whether genomic imprinting has a role in sleep has remained elusive up to now. In this work we show that REM and NREM sleep states are differentially modulated by the maternally expressed imprinted gene Gnas. In particular, in mice with loss of imprinting of Gnas, NREM and complex cognitive processes are enhanced while REM and REM-linked behaviors are inhibited. This is the first demonstration that a specific overexpression of an imprinted gene affects sleep states and related complex behavioral traits. Furthermore, in parallel to the Gnas overexpression, we have observed an overexpression of Ucp1 in interscapular brown adipose tissue (BAT) and a significant increase in thermoregulation that may account for the REM/NREM sleep phenotypes. We conclude that there must be significant evolutionary advantages in the monoallelic expression of Gnas for REM sleep and for the consolidation of REM-dependent memories. Conversely, biallelic expression of Gnas reinforces slow wave activity in NREM sleep, and this results in a reduction of uncertainty in temporal decision-making processes.

Identification of an imprinting control region affecting the expression of all transcripts in the Gnas cluster.

Genomic imprinting results in allele-specific silencing according to parental origin. Silencing is brought about by imprinting control regions (ICRs) that are differentially marked in gametogenesis. The group of imprinted transcripts in the mouse Gnas cluster (Nesp, Nespas, Gnasxl, Exon 1A and Gnas) provides a model for analyzing the mechanisms of imprint regulation. We previously identified an ICR that specifically regulates the tissue-specific imprinted expression of the Gnas gene. Here we identify a second ICR at the Gnas cluster. We show that a paternally derived targeted deletion of the germline differentially methylated region (DMR) associated with the antisense Nespas transcript unexpectedly affects both the expression of all transcripts in the cluster and methylation of two DMRs. Our results establish that the Nespas DMR is the principal ICR at the Gnas cluster and functions bidirectionally as a switch for modulating expression of the antagonistically acting genes Gnasxl and Gnas. Uniquely, the Nespas DMR acts on the downstream ICR at exon 1A to regulate tissue-specific imprinting of the Gnas gene.

MicroRNAs 296 and 298 are imprinted and part of the GNAS/Gnas cluster and miR-296 targets IKBKE and Tmed9.

Genomic imprinting is the phenomenon whereby a subset of genes is differentially expressed according to parental origin. Imprinted genes tend to occur in clusters, and microRNAs are associated with the majority of well-defined clusters of imprinted genes. We show here that two microRNAs, miR-296 and miR-298, are part of the imprinted Gnas/GNAS clusters in both mice and humans. Both microRNAs show imprinted expression and are expressed from the paternally derived allele, but not the maternal allele. They arise from a long, noncoding antisense transcript, Nespas, with a promoter more than 27 kb away. Nespas had been shown previously to act in cis to regulate imprinted gene expression within the Gnas cluster. Using microarrays and luciferase assays, IKBKE, involved in many signaling pathways, and Tmed9, a protein transporter, were verified as new targets of miR-296. Thus, Nespas has two clear functions: as a cis-acting regulator within an imprinted gene cluster and as a precursor of microRNAs that modulate gene expression in trans. Furthermore, imprinted microRNAs, including miR-296 and miR-298, impose a parental specific modulation of gene expression of their target genes.

Uncoupling antisense-mediated silencing and DNA methylation in the imprinted Gnas cluster.

There is increasing evidence that non-coding macroRNAs are major elements for silencing imprinted genes, but their mechanism of action is poorly understood. Within the imprinted Gnas cluster on mouse chromosome 2, Nespas is a paternally expressed macroRNA that arises from an imprinting control region and runs antisense to Nesp, a paternally repressed protein coding transcript. Here we report a knock-in mouse allele that behaves as a Nespas hypomorph. The hypomorph mediates down-regulation of Nesp in cis through chromatin modification at the Nesp promoter but in the absence of somatic DNA methylation. Notably there is reduced demethylation of H3K4me3, sufficient for down-regulation of Nesp, but insufficient for DNA methylation; in addition, there is depletion of the H3K36me3 mark permissive for DNA methylation. We propose an order of events for the regulation of a somatic imprint on the wild-type allele whereby Nespas modulates demethylation of H3K4me3 resulting in repression of Nesp followed by DNA methylation. This study demonstrates that a non-coding antisense transcript or its transcription is associated with silencing an overlapping protein-coding gene by a mechanism independent of DNA methylation. These results have broad implications for understanding the hierarchy of events in epigenetic silencing by macroRNAs.

Maternal inheritance of the Gnas cluster mutation Ex1A-T affects size, implicating NESP55 in growth.

Genes subjected to genomic imprinting are often associated with prenatal and postnatal growth. Furthermore, it has been observed that maternally silenced/paternally expressed genes tend to favour offspring growth, whilst paternally silenced/maternally expressed genes will restrict growth. One imprinted cluster in which this has been shown to hold true is the Gnas cluster; of the three proteins expressed from this cluster, two, Gsα and XLαs, have been found to affect postnatal growth in a number of different mouse models. The remaining protein in this cluster, NESP55, has not yet been shown to be involved in growth. We previously described a new mutation, Ex1A-T, which upon paternal transmission resulted in postnatal growth retardation due to loss of imprinting of Gsα and loss of expression of the paternally expressed XLαs. Here we describe maternal inheritance of Ex1A-T which gives rise to a small but highly significant overgrowth phenotype which we attribute to reduction of maternally expressed NESP55.

The imprinted signaling protein XL alpha s is required for postnatal adaptation to feeding.

Genomic imprinting, by which maternal and paternal alleles of some genes have different levels of activity, has profound effects on growth and development of the mammalian fetus. The action of imprinted genes after birth, in particular while the infant is dependent on maternal provision of nutrients, is far less well understood. We disrupted a paternally expressed transcript at the Gnas locus, Gnasxl, which encodes the unusual Gs alpha isoform XL alpha s. Mice with mutations in Gnasxl have poor postnatal growth and survival and a spectrum of phenotypic effects that indicate that XL alpha s controls a number of key postnatal physiological adaptations, including suckling, blood glucose and energy homeostasis. Increased cAMP levels in brown adipose tissue of Gnasxl mutants and phenotypic comparison with Gnas mutants suggest that XL alpha s can antagonize Gs alpha-dependent signaling pathways. The opposing effects of maternally and paternally expressed products of the Gnas locus provide tangible molecular support for the parental-conflict hypothesis of imprinting.

A cis-acting control region is required exclusively for the tissue-specific imprinting of Gnas.

Genomic imprinting brings about allele-specific silencing according to parental origin. Silencing is controlled by cis-acting regulatory regions that are differentially marked during gametogenesis and can act over hundreds of kilobases to silence many genes. Two candidate imprinting control regions (ICRs) have been identified at the compact imprinted Gnas cluster on distal mouse chromosome 2, one at exon 1A upstream of Gnas itself and one covering the promoters for Gnasxl and the antisense Nespas (ref. 8). This imprinted cluster is complex, containing biallelic, maternally and paternally expressed transcripts that share exons. Gnas itself is mainly biallelically expressed but is weakly paternally repressed in specific tissues. Here we show that a paternally derived targeted deletion of the germline differentially methylated region at exon 1A abolishes tissue-specific imprinting of Gnas. This rescues the abnormal phenotype of mice with a maternally derived Gnas mutation. Imprinting of alternative transcripts, Nesp, Gnasxl and Nespas (ref. 13), in the cluster is unaffected. The results establish that the differentially methylated region at exon 1A contains an imprinting control element that specifically regulates Gnas and comprises a characterized ICR for a gene that is only weakly imprinted in a minority of tissues. There must be a second ICR regulating the alternative transcripts.

MouseBook: an integrated portal of mouse resources.

The MouseBook (http://www.mousebook.org) databases and web portal provide access to information about mutant mouse lines held as live or cryopreserved stocks at MRC Harwell. The MouseBook portal integrates curated information from the MRC Harwell stock resource, and other Harwell databases, with information from external data resources to provide value-added information above and beyond what is available through other routes such as International Mouse Stain Resource (IMSR). MouseBook can be searched either using an intuitive Google style free text search or using the Mammalian Phenotype (MP) ontology tree structure. Text searches can be on gene, allele, strain identifier (e.g. MGI ID) or phenotype term and are assisted by automatic recognition of term types and autocompletion of gene and allele names covered by the database. Results are returned in a tabbed format providing categorized results identified from each of the catalogs in MouseBook. Individual result lines from each catalog include information on gene, allele, chromosomal location and phenotype, and provide a simple click-through link to further information as well as ordering the strain. The infrastructure underlying MouseBook has been designed to be extensible, allowing additional data sources to be added and enabling other sites to make their data directly available through MouseBook.

Potential predictors of hospital length of stay and hospital charges among patients with all-terrain vehicle injuries in rural Northeast Texas.

BACKGROUND: All-Terrain Vehicles (ATVs) have become popular for recreation use in recent years. Texas has had more ATV related fatalities than any other state in the nation, with rural Northeast Texas having even higher rates of injuries. There is limited data examining the relationship between ATV injuries and the length of hospital stay, as well as hospital costs. This paper examines both issues in children as well as adults. METHODS: The regional trauma registry was analyzed for all ATV related injuries between January 2011- October 2016. Injury Severity Score, Glasgow Coma Scale and if they are seen at a Level I Trauma center are predictive for both hospital length of stay and charges. RESULTS: Length of Stay was predicted positively by Injury Severity Score, Emergency Department Respiration Rate and facility at which patients were treated and negatively by Glasgow Coma Scale. Hospital charges were predicted positively by age, Injury Severity Score, facility of treatment, means of transportation, and Emergency Department pulse and negatively by Glasgow Coma Scale. CONCLUSIONS: The study found that vital signs can be useful in predicting length of stay and hospital charges. This study not only confirms the findings of other studies regarding what predictors can be used, but expands the research into rural traumatic injuries. It is hoped that this data can help contribute to the development of algorithms to predict which patients will be most likely to require resource intensive treatment.

Alternately spliced WT1 antisense transcripts interact with WT1 sense RNA and show epigenetic and splicing defects in cancer.

Many mammalian genes contain overlapping antisense RNAs, but the functions and mechanisms of action of these transcripts are mostly unknown. WT1 is a well-characterized developmental gene that is mutated in Wilms' tumor (WT) and acute myeloid leukaemia (AML) and has an antisense transcript (WT1-AS), which we have previously found to regulate WT1 protein levels. In this study, we show that WT1-AS is present in multiple spliceoforms that are usually expressed in parallel with WT1 RNA in human and mouse tissues. We demonstrate that the expression of WT1-AS correlates with methylation of the antisense regulatory region (ARR) in WT1 intron 1, displaying imprinted monoallelic expression in normal kidney and loss of imprinting in WT. However, we find no evidence for imprinting of mouse Wt1-as. WT1-AS transcripts are exported into the cytoplasm and form heteroduplexes with WT1 mRNA in the overlapping region in WT1 exon 1. In AML, there is often abnormal splicing of WT1-AS, which may play a role in the development of this malignancy. These results show that WT1 encodes conserved antisense RNAs that may have an important regulatory role in WT1 expression via RNA:RNA interactions, and which can become deregulated by a variety of mechanisms in cancer.

Control of imprinting at the Gnas cluster.

Genomic imprinting is a form of epigenetic regulation in mammals whereby a small subset of genes is silenced according to parental origin. Early work had indicated regions of the genome that were likely to contain imprinted genes. Distal mouse chromosome 2 is one such region and is associated with devastating but ostensibly opposite phenotypes when exclusively maternally or paternally derived. Misexpression of proteins encoded at the Gnas complex, which is located within the region, can largely account for the imprinting phenotypes. Gnas is a complex locus with extraordinary transcriptional and regulatory complexity. It gives rise to alternatively spliced isoforms that show maternal-, paternal- and biallelic expression as well as a noncoding antisense transcript. The objective of our work at Harwell is to unravel mechanisms controlling the expression of these transcripts. We have performed targeted deletion analysis to test candidate regulatory regions within the Gnas complex and, unlike other imprinted domains, two major control regions have been identified. One controls the imprinted expression of a single transcript and is subsidiary to and must interact with, a principal control region that affects the expression of all transcripts. This principal region contains the promoter for the antisense transcript, expression of which may have a major role in controlling imprinting at the Gnas cluster.

Epigenetic properties and identification of an imprint mark in the Nesp-Gnasxl domain of the mouse Gnas imprinted locus.

The Gnas locus in the mouse is imprinted with a complex arrangement of alternative transcripts defined by promoters with different patterns of monoallelic expression. The Gnas transcript is subject to tissue-specific imprinted expression, Nesp is expressed only from the maternal allele, and Gnasxl is expressed only from the paternal allele. The mechanisms controlling these expression patterns are not known. To identify potential imprinting regulatory regions, particularly for the reciprocally expressed Nesp and Gnasxl promoters, we examined epigenetic properties of the locus in gametes, embryonic stem cells, and fetal and adult tissues. The Nesp and Gnasxl promoter regions are contained in extensive CpG islands with methylation of the paternal allele at Nesp and the maternal allele at Gnasxl. Parental allele-specific DNase I-hypersensitive sites were found at these regions, which correlate with hypomethylation rather than actual expression status. A germ line methylation mark was identified covering the promoters for Gnasxl and the antisense transcript Nespas. Prominent DNase I-hypersensitive sites present on paternal alleles in embryonic stem cells are contained within this mark. This is the second gametic mark identified at Gnas and suggests that the Nesp and Gnasxl promoters are under separate control from the Gnas promoter. We propose models to account for the regulation of imprinting at the locus.

FabAV antivenin use after copperhead snakebite: clinically indicated or knee-jerk reaction?

BACKGROUND: Crotalidae Polyvalent Immune Fab (Ovine) (FabAV) antivenin is commonly recommended after pit viper snakebites. Because copperhead envenomations are usually self-limited, some physicians are reluctant to use this costly treatment routinely, while others follow a more liberal approach. We hypothesized that, in practice, only patients with evidence of significant (moderate or severe) copperhead envenomation [those with snakebite severity score (SSS) > 3] receive FabAV and examined a large cohort to determine the relationship between clinical findings and FabAV administration. METHODS: All data from patients evaluated for copperhead snakebite at a rural tertiary referral center from 5/2002 to 10/2013 were compiled. Demographics, transfer status, antivenin use, and clinical findings were collected; SSS was calculated. The relationships among FabAV use, clinical findings, and SSS were analyzed using t-test, chi-square, and Pearson's coefficient (p < 0.05 was significant). RESULTS: During the study period, 318 patients were treated for copperhead snakebite; 44 (13.8 %) received antivenin. Median dose was four vials (range: 1-10; IQR: 4,6). There were no deaths. Most patients receiving FabAV (63.6 %) were admitted. With regard to demographics and symptoms, only the degree of swelling (moderate vs. none/mild; p < 0.01) and bite location (hand/arm vs. leg: p < 0.0001) were associated with FabAV use. A SSS > 3, indicating moderate or severe envenomation, was only very weakly correlated with antivenin use (r = 0.217; p < 0.0001). The majority of patients with SSS > 3 (65.8 %) did not receive antivenin while most patients who did receive antivenin (70.5 %) had SSS ≤ 3 (indicating mild envenomation). CONCLUSIONS: Considerable variation occurs in antivenin administration after copperhead snakebite. Use of FabAV appears poorly correlated with patients' symptoms. This practice may expose patients to the risks of antivenin and increasing costs of medical care without improving outcomes. Guidelines used for treating other pit viper strikes, such as rattlesnake or cottonmouth snakebite may be too liberal for copperhead envenomations. Our data suggests that most patients with mild or moderate envenomation appear to do well independent of FabAV use. We suggest, for patients with copperhead snakebite, that consideration be given to withholding FabAV for those without clinical evidence of severe envenomation until prospective randomized data are available.

Deletion of the Snord116/SNORD116 Alters Sleep in Mice and Patients with Prader-Willi Syndrome.

STUDY OBJECTIVES: Sleep-wake disturbances are often reported in Prader-Willi syndrome (PWS), a rare neurodevelopmental syndrome that is associated with paternally-expressed genomic imprinting defects within the human chromosome region 15q11-13. One of the candidate genes, prevalently expressed in the brain, is the small nucleolar ribonucleic acid-116 (SNORD116). Here we conducted a translational study into the sleep abnormalities of PWS, testing the hypothesis that SNORD116 is responsible for sleep defects that characterize the syndrome. METHODS: We studied sleep in mutant mice that carry a deletion of Snord116 at the orthologous locus (mouse chromosome 7) of the human PWS critical region (PWScr). In particular, we assessed EEG and temperature profiles, across 24-h, in PWScr (m+/p-) heterozygous mutants compared to wild-type littermates. High-resolution magnetic resonance imaging (MRI) was performed to explore morphoanatomical differences according to the genotype. Moreover, we complemented the mouse work by presenting two patients with a diagnosis of PWS and characterized by atypical small deletions of SNORD116. We compared the individual EEG parameters of patients with healthy subjects and with a cohort of obese subjects. RESULTS: By studying the mouse mutant line PWScr(m+/p-), we observed specific rapid eye movement (REM) sleep alterations including abnormal electroencephalograph (EEG) theta waves. Remarkably, we observed identical sleep/EEG defects in the two PWS cases. We report brain morphological abnormalities that are associated with the EEG alterations. In particular, mouse mutants have a bilateral reduction of the gray matter volume in the ventral hippocampus and in the septum areas, which are pivotal structures for maintaining theta rhythms throughout the brain. In PWScr(m+/p-) mice we also observed increased body temperature that is coherent with REM sleep alterations in mice and human patients. CONCLUSIONS: Our study indicates that paternally expressed Snord116 is involved in the 24-h regulation of sleep physiological measures, suggesting that it is a candidate gene for the sleep disturbances that most individuals with PWS experience.

Transcription driven somatic DNA methylation within the imprinted Gnas cluster.

Differential marking of genes in female and male gametes by DNA methylation is essential to genomic imprinting. In female gametes transcription traversing differentially methylated regions (DMRs) is a common requirement for de novo methylation at DMRs. At the imprinted Gnas cluster oocyte specific transcription of a protein-coding transcript, Nesp, is needed for methylation of two DMRs intragenic to Nesp, namely the Nespas-Gnasxl DMR and the Exon1A DMR, thereby enabling expression of the Gnas transcript and repression of the Gnasxl transcript. On the paternal allele, Nesp is repressed, the germline DMRs are unmethylated, Gnas is repressed and Gnasxl is expressed. Using mutant mouse models, we show that on the paternal allele, ectopic transcription of Nesp traversing the intragenic Exon1A DMR (which regulates Gnas expression) results in de novo methylation of the Exon1A DMR and de-repression of Gnas just as on the maternal allele. However, unlike the maternal allele, methylation on the mutant paternal allele occurs post-fertilisation, i.e. in somatic cells. This, to our knowledge is the first example of transcript/transcription driven DNA methylation of an intragenic CpG island, in somatic tissues, suggesting that transcription driven de novo methylation is not restricted to the germline in the mouse. Additionally, Gnasxl is repressed on a paternal chromosome on which Nesp is ectopically expressed. Thus, a paternally inherited Gnas cluster showing ectopic expression of Nesp is "maternalised" in terms of Gnasxl and Gnas expression. We show that these mice have a phenotype similar to mutants with two expressed doses of Gnas and none of Gnasxl.

Antisense Activity across the Nesp Promoter is Required for Nespas-Mediated Silencing in the Imprinted Gnas Cluster.

Macro long non-coding RNAs (lncRNAs) play major roles in gene silencing in inprinted gene clusters. Within the imprinted Gnas cluster, the paternally expressed Nespas lncRNA downregulates its sense counterpart Nesp. To explore the mechanism of action of Nespas, we generated two new knock-in alleles to truncate Nespas upstream and downstream of the Nesp promoter. We show that Nespas is essential for methylation of the Nesp differentially methylated region (DMR), but higher levels of Nespas are required for methylation than are needed for downregulation of Nesp. Although Nespas is transcribed for over 27 kb, only Nespas transcript/transcription across a 2.6 kb region that includes the Nesp promoter is necessary for methylation of the Nesp DMR. In both mutants, the levels of Nespas were extraordinarily high, due at least in part to increased stability, an effect not seen with other imprinted lncRNAs. However, even when levels were greatly raised, Nespas remained exclusively cis-acting. We propose Nespas regulates Nesp methylation and expression to ensure appropriate levels of expression of the protein coding transcripts Gnasxl and Gnas on the paternal chromosome. Thus, Nespas mediates paternal gene expression over the entire Gnas cluster via a single gene, Nesp.

Structural characterization of Rasgrf1 and a novel linked imprinted locus.

Imprinted genes in mammals are expressed either from the maternally or the paternally inherited allele. Previously, a genome wide scan identified novel imprinted genes based on their association with differentially methylated regions (DMRs). One of the identified genes, Rasgrf1, showed paternal expression in neonatal brain and was located on mouse chromosome 9. This gene is associated with a DMR, located about 30 kb upstream of Rasgrf1 exon 1. In order to better understand and identify novel elements involved in the regulation of this gene we have isolated and characterized genomic clones coding for mouse and human Rasgrf1 and RASGRF1, respectively. The mouse gene consists of 26 exons spanning approximately 140 kb of genomic DNA while the human gene has 28 exons. The human gene has an additional 39 bp exon inserted between exons 13 and 14 and exon 18 is split in two separate exons in human. The major transcription start site of Rasgrf1, as identified by primer extension, is 1324 bp upstream of the ATG translation start codon. Finally, a genomic region upstream of exon 1, spanning 489 bp, was determined to possess the essential promoter activity for Rasgrf1 gene. A second gene, A19, located 10 kb upstream of the DMR has been characterized. A19 is mainly expressed in testis and at lower levels in neonatal and adult brain tissue. The A19 transcript is non-coding and expressed in mouse testis and brain. A19 is imprinted with expression occurring from just the paternal allele in brain.