To Tube or Not to Tube ... That Continues to Be the Question.

In the prehospital setting, "to tube, or not to tube" will persist as a probing question - long after this article is published. It is the hope of the authors simply to position a compilation of thoughts to consider in regards to alternate airways vs. endotracheal intubation. Ultimately, it's all about the right care, for the right patient, at the right time!

High prevalence of deleterious mutations in concomitant nonsyndromic cleft and outflow tract heart defects.

Our previous work demonstrating enrichment of outflow tract (OFT) congenital heart disease (CHD) in children with cleft lip and/or palate (CL/P) suggests derangements in common underlying developmental pathways. The current pilot study examines the underlying genetics of concomitant nonsyndromic CL/P and OFT CHD phenotype. Of 575 patients who underwent CL/P surgery at Children's Hospital Los Angeles, seven with OFT CHD, negative chromosomal microarray analysis, and no recognizable syndromic association were recruited with their parents (as available). Whole genome sequencing of blood samples paired with whole-blood-based RNA sequencing for probands was performed. A pathogenic or potentially pathogenic variant was identified in 6/7 (85.7%) probands. A total of seven candidate genes were mutated (CHD7, SMARCA4, MED12, APOB, RNF213, SETX, and JAG1). Gene ontology analysis of variants predicted involvement in binding (100%), regulation of transcription (42.9%), and helicase activity (42.9%). Four patients (57.1%) expressed gene variants (CHD7, SMARCA4, MED12, and RNF213) previously involved in the Wnt signaling pathway. Our pilot analysis of a small cohort of patients with combined CL/P and OFT CHD phenotype suggests a potentially significant prevalence of deleterious mutations. In our cohort, an overrepresentation of mutations in molecules associated with Wnt-signaling was found. These variants may represent an expanded phenotypic heterogeneity within known monogenic disease genes or provide novel evidence of shared developmental pathways. The mechanistic implications of these mutations and subsequent developmental derangements resulting in the CL/P and OFT CHD phenotype require further analysis in a larger cohort of patients.

A syndemic effect: Interrelationships between systemic racism, health disparities, and COVID-19.

ABSTRACT: This article discusses the interconnection between the syndemic effect of racial inequities and disparities as well as the impact of the COVID-19 pandemic on Black Americans. It also highlights meaningful reforms and priorities to achieve health equity in Black communities.

Splice-Break: exploiting an RNA-seq splice junction algorithm to discover mitochondrial DNA deletion breakpoints and analyses of psychiatric disorders.

Deletions in the 16.6 kb mitochondrial genome have been implicated in numerous disorders that often display muscular and/or neurological symptoms due to the high-energy demands of these tissues. We describe a catalogue of 4489 putative mitochondrial DNA (mtDNA) deletions, including their frequency and relative read rate, using a combinatorial approach of mitochondria-targeted PCR, next-generation sequencing, bioinformatics, post-hoc filtering, annotation, and validation steps. Our bioinformatics pipeline uses MapSplice, an RNA-seq splice junction detection algorithm, to detect and quantify mtDNA deletion breakpoints rather than mRNA splices. Analyses of 93 samples from postmortem brain and blood found (i) the 4977 bp 'common deletion' was neither the most frequent deletion nor the most abundant; (ii) brain contained significantly more deletions than blood; (iii) many high frequency deletions were previously reported in MitoBreak, suggesting they are present at low levels in metabolically active tissues and are not exclusive to individuals with diagnosed mitochondrial pathologies; (iv) many individual deletions (and cumulative metrics) had significant and positive correlations with age and (v) the highest deletion burdens were observed in major depressive disorder brain, at levels greater than Kearns-Sayre Syndrome muscle. Collectively, these data suggest the Splice-Break pipeline can detect and quantify mtDNA deletions at a high level of resolution.

A Team-Based Approach to Delivering Person-Centered Care at the End of Life.

The use of team-based models of care is widely regarded as a mechanism for enhancing the delivery of high-quality care, especially at the end of life. Active collaboration to promote effective coordination and delivery of person-centered care is an integral part of the team-based model that is the focus of this article.

Phospho-dependent Regulation of SAMHD1 Oligomerisation Couples Catalysis and Restriction.

SAMHD1 restricts HIV-1 infection of myeloid-lineage and resting CD4+ T-cells. Most likely this occurs through deoxynucleoside triphosphate triphosphohydrolase activity that reduces cellular dNTP to a level where reverse transcriptase cannot function, although alternative mechanisms have been proposed recently. Here, we present combined structural and virological data demonstrating that in addition to allosteric activation and triphosphohydrolase activity, restriction correlates with the capacity of SAMHD1 to form "long-lived" enzymatically competent tetramers. Tetramer disruption invariably abolishes restriction but has varied effects on in vitro triphosphohydrolase activity. SAMHD1 phosphorylation also ablates restriction and tetramer formation but without affecting triphosphohydrolase steady-state kinetics. However phospho-SAMHD1 is unable to catalyse dNTP turnover under conditions of nucleotide depletion. Based on our findings we propose a model for phosphorylation-dependent regulation of SAMHD1 activity where dephosphorylation switches housekeeping SAMHD1 found in cycling cells to a high-activity stable tetrameric form that depletes and maintains low levels of dNTPs in differentiated cells.

HIV-1 restriction factor SAMHD1 is a deoxynucleoside triphosphate triphosphohydrolase.

SAMHD1, an analogue of the murine interferon (IFN)-γ-induced gene Mg11 (ref. 1), has recently been identified as a human immunodeficiency virus-1 (HIV-1) restriction factor that blocks early-stage virus replication in dendritic and other myeloid cells and is the target of the lentiviral protein Vpx, which can relieve HIV-1 restriction. SAMHD1 is also associated with Aicardi-Goutières syndrome (AGS), an inflammatory encephalopathy characterized by chronic cerebrospinal fluid lymphocytosis and elevated levels of the antiviral cytokine IFN-α. The pathology associated with AGS resembles congenital viral infection, such as transplacentally acquired HIV. Here we show that human SAMHD1 is a potent dGTP-stimulated triphosphohydrolase that converts deoxynucleoside triphosphates to the constituent deoxynucleoside and inorganic triphosphate. The crystal structure of the catalytic core of SAMHD1 reveals that the protein is dimeric and indicates a molecular basis for dGTP stimulation of catalytic activity against dNTPs. We propose that SAMHD1, which is highly expressed in dendritic cells, restricts HIV-1 replication by hydrolysing the majority of cellular dNTPs, thus inhibiting reverse transcription and viral complementary DNA (cDNA) synthesis.

The eukaryotic elongation factor eEF1A1 interacts with SAMHD1.

Mutations in SAMHD1 cause Aicardi-Goutières syndrome (AGS), a Mendelian inflammatory disease which displays remarkable clinical and biochemical overlap with congenital viral infection. SAMHD1 (SAM domain and HD domain-containing protein 1) has also been defined as an HIV-1 restriction-factor that, through a novel triphosphohydrolase activity, inhibits early stage HIV-1 replication in myeloid-derived dendritic cells (MDDCs), macrophages and resting CD4+ T-cells. The potent activity of SAMHD1 is likely to be the subject of a variety of regulatory mechanisms. Knowledge of proteins that interact with SAMHD1 may not only enhance our understanding of the pathogenesis of AGS, but may also provide further details on the link between the regulation of cellular dNTPs and HIV-1 restriction. In the present study, we used a yeast two-hybrid screen and pull-down analysis followed by MS to identify the eukaryotic elongation factor 1A1 (eEF1A1) as a potential interaction partner of SAMHD1. This interaction was confirmed by unbiased co-immunoprecipitation and demonstrated in situ by a proximity ligation assay (PLA). We show that this interaction is enhanced in mutant SAMHD1 cell lines and suggest that eEF1A1 may mediate SAMHD1 turnover by targeting it to the proteosome for degradation through association with Cullin4A and Rbx1.

Interactive hazards education program for youth in a low SES community: a quasi-experimental pilot study.

A pilot study of an interactive hazards education program was carried out in Canberra (Australia), with direct input from youth participants. Effects were evaluated in relation to youths' interest in disasters, motivation to prepare, risk awareness, knowledge indicators, perceived preparedness levels, planning and practice for emergencies, and fear and anxiety indicators. Parents also provided ratings, including of actual home-based preparedness activities. Using a single group pretest-posttest with benchmarking design, a sample of 20 youths and their parents from a low SES community participated. Findings indicated beneficial changes on a number of indicators. Preparedness indicators increased significantly from pre- to posttest on both youth (p < 0.01) and parent ratings (p < 0.01). Parent ratings reflected an increase of just under six home-based preparedness activities. Youth knowledge about disaster mitigation also was seen to increase significantly (p < 0.001), increasing 39% from pretest levels. While personalized risk perceptions significantly increased (p < 0.01), anxiety and worry levels were seen either not to change (generalized anxiety, p > 0.05) or to reduce between pre- and posttest (hazards-specific fears, worry, and distress, ps ranged from p < 0.05 to < 0.001). In terms of predictors of preparedness, a number of variables were found to predict posttest preparedness levels, including information searching done by participants between education sessions. These pilot findings are the first to reflect quasi-experimental outcomes for a youth hazards education program carried out in a setting other than a school that focused on a sample of youth from a low SES community.

Strategies for Developing a Nurturing Environment for the next Generation of Nurses.

With the introduction of more complex health conditions and the changing landscape of the healthcare infrastructure, burnout is increasingly becoming a crisis for the nursing profession and for the public. Recruitment in nursing must consider the concept of a nurturing environment as a key driver of sustainability within the profession. Human beings cannot flourish in hostile and unwelcoming environments. Failure to thrive in nursing is a real phenomenon that is driven by multiple factors, including incivility, workplace bullying, and lack of support. Mitigation requires intentional, strategic interventions toward building nurturing environments in education and practice for the next generation of nurses.

Evaluation of a Seizure Action Plan in an Adult Epilepsy Center.

Background: Seizure action plans (SAPs) provide valuable information for patients to manage seizure emergencies, but are underutilized in adult epilepsy centers. The purpose of this project was to implement a structured SAP for adult patients with epilepsy. Methods: A pre/postimplementation design was used. Provider SAP utilization rates were analyzed over a 16-week period. A pre and postimplementation survey assessed participant perceived impact of the SAP on knowledge and comfort associated with managing seizure emergencies. Provider barriers and facilitators were also assessed. Results: Average provider SAP utilization rate was 51.45%. A total of 204 participants completed the surveys, which showed a significant increase in knowledge and comfort for all items, p < 0.001. At postsurvey analysis, 98% of participants felt that all patients with epilepsy should have a SAP regardless of seizure burden. Discussion: Implementing a structured SAP increased provider utilization and patient and care partner knowledge and comfort of managing seizure emergencies.

Enhancing Spatial Transcriptomics Analysis by Integrating Image-Aware Deep Learning Methods.

Spatial transcriptomics (ST) represents a pivotal advancement in biomedical research, enabling the transcriptional profiling of cells within their morphological context and providing a pivotal tool for understanding spatial heterogeneity in cancer tissues. However, current analytical approaches, akin to single-cell analysis, largely depend on gene expression, underutilizing the rich morphological information inherent in the tissue. We present a novel method integrating spatial transcriptomics and histopathological image data to better capture biologically meaningful patterns in patient data, focusing on aggressive cancer types such as glioblastoma and triple-negative breast cancer. We used a ResNet-based deep learning model to extract key morphological features from high-resolution whole-slide histology images. Spot-level PCA-reduced vectors of both the ResNet-50 analysis of the histological image and the spatial gene expression data were used in Louvain clustering to enable image-aware feature discovery. Assessment of features from image-aware clustering successfully pinpointed key biological features identified by manual histopathology, such as for regions of fibrosis and necrosis, as well as improved edge definition in EGFR-rich areas. Importantly, our combinatorial approach revealed crucial characteristics seen in histopathology that gene-expression-only analysis had missed.Supplemental Material: https://github.com/davcraig75/song_psb2014/blob/main/SupplementaryData.pdf.

Profiling the molecular and clinical landscape of glioblastoma utilizing the Oncology Research Information Exchange Network brain cancer database.

Background: Glioblastoma exhibits aggressive growth and poor outcomes despite treatment, and its marked variability renders therapeutic design and prognostication challenging. The Oncology Research Information Exchange Network (ORIEN) database contains complementary clinical, genomic, and transcriptomic profiling of 206 glioblastoma patients, providing opportunities to identify novel associations between molecular features and clinical outcomes. Methods: Survival analyses were performed using the Logrank test, and clinical features were evaluated using Wilcoxon and chi-squared tests with q-values derived via Benjamini-Hochberg correction. Mutational analyses utilized sample-level enrichments from whole exome sequencing data, and statistical tests were performed using the one-sided Fisher Exact test with Benjamini-Hochberg correction. Transcriptomic analyses utilized a student's t-test with Benjamini-Hochberg correction. Expression fold changes were processed with Ingenuity Pathway Analysis to determine pathway-level alterations between groups. Results: Key findings include an association of MUC17, SYNE1, and TENM1 mutations with prolonged overall survival (OS); decreased OS associated with higher epithelial growth factor receptor (EGFR) mRNA expression, but not with EGFR amplification or mutation; a 14-transcript signature associated with OS > 2 years; and 2 transcripts associated with OS < 1 year. Conclusions: Herein, we report the first clinical, genomic, and transcriptomic analysis of ORIEN glioblastoma cases, incorporating sample reclassification under updated 2021 diagnostic criteria. These findings create multiple avenues for further investigation and reinforce the value of multi-institutional consortia such as ORIEN in deepening our knowledge of intractable diseases such as glioblastoma.

Common mitochondrial deletions in RNA-Seq: evaluation of bulk, single-cell, and spatial transcriptomic datasets.

Common mitochondrial DNA (mtDNA) deletions are large structural variants in the mitochondrial genome that accumulate in metabolically active tissues with age and have been investigated in various diseases. We applied the Splice-Break2 pipeline (designed for high-throughput quantification of mtDNA deletions) to human RNA-Seq datasets and describe the methodological considerations for evaluating common deletions in bulk, single-cell, and spatial transcriptomics datasets. A robust evaluation of 1570 samples from 14 RNA-Seq studies showed: (i) the abundance of some common deletions detected in PCR-amplified mtDNA correlates with levels observed in RNA-Seq data; (ii) RNA-Seq library preparation method has a strong effect on deletion detection; (iii) deletions had a significant, positive correlation with age in brain and muscle; (iv) deletions were enriched in cortical grey matter, specifically in layers 3 and 5; and (v) brain regions with dopaminergic neurons (i.e., substantia nigra, ventral tegmental area, and caudate nucleus) had remarkable enrichment of common mtDNA deletions.

Debakarn Koorliny Wangkiny: steady walking and talking using first nations-led participatory action research methodologies to build relationships.

Aboriginal participatory action research (APAR) has an ethical focus that corrects the imbalances of colonisation through participation and shared decision-making to position people, place, and intention at the centre of research. APAR supports researchers to respond to the community's local rhythms and culture. APAR supports researchers to respond to the community's local rhythms and culture. First Nations scholars and their allies do this in a way that decolonises mainstream approaches in research to disrupt its cherished ideals and endeavours. How these knowledges are co-created and translated is also critically scrutinised. We are a team of intercultural researchers working with community and mainstream health service providers to improve service access, responsiveness, and Aboriginal client outcomes. Our article begins with an overview of the APAR literature and pays homage to the decolonising scholarship that champions Aboriginal ways of knowing, being, and doing. We present a research program where Aboriginal Elders, as cultural guides, hold the research through storying and cultural experiences that have deepened relationships between services and the local Aboriginal community. We conclude with implications of a community-led engagement framework underpinned by a relational methodology that reflects the nuances of knowledge translation through a co-creation of new knowledge and knowledge exchange.

Spatial Transcriptomic Analysis of a Diverse Patient Cohort Reveals a Conserved Architecture in Triple-Negative Breast Cancer.

Triple-negative breast cancer (TNBC) is an aggressive disease that disproportionately affects African American (AA) women. Limited targeted therapeutic options exist for patients with TNBC. Here, we employ spatial transcriptomics to interrogate tissue from a racially diverse TNBC cohort to comprehensively annotate the transcriptional states of spatially resolved cellular populations. A total of 38,706 spatial features from a cohort of 28 sections from 14 patients were analyzed. Intratumoral analysis of spatial features from individual sections revealed heterogeneous transcriptional substructures. However, integrated analysis of all samples resulted in nine transcriptionally distinct clusters that mapped across all individual sections. Furthermore, novel use of join count analysis demonstrated nonrandom directional spatial dependencies of the transcriptionally defined shared clusters, supporting a conserved spatio-transcriptional architecture in TNBC. These findings were substantiated in an independent validation cohort comprising 17,861 spatial features representing 15 samples from 8 patients. Stratification of samples by race revealed race-associated differences in hypoxic tumor content and regions of immune-rich infiltrate. Overall, this study combined spatial and functional molecular analyses to define the tumor architecture of TNBC, with potential implications in understanding TNBC disparities. SIGNIFICANCE: Spatial transcriptomics profiling of a diverse cohort of triple-negative breast cancers and innovative informatics approaches reveal a conserved cellular architecture across cancers and identify proportional differences in tumor cell composition by race.

The Foundational Data Initiative for Parkinson Disease: Enabling efficient translation from genetic maps to mechanism.

The Foundational Data Initiative for Parkinson Disease (FOUNDIN-PD) is an international collaboration producing fundamental resources for Parkinson disease (PD). FOUNDIN-PD generated a multi-layered molecular dataset in a cohort of induced pluripotent stem cell (iPSC) lines differentiated to dopaminergic (DA) neurons, a major affected cell type in PD. The lines were derived from the Parkinson's Progression Markers Initiative study, which included participants with PD carrying monogenic PD variants, variants with intermediate effects, and variants identified by genome-wide association studies and unaffected individuals. We generated genetic, epigenetic, regulatory, transcriptomic, and longitudinal cellular imaging data from iPSC-derived DA neurons to understand molecular relationships between disease-associated genetic variation and proximate molecular events. These data reveal that iPSC-derived DA neurons provide a valuable cellular context and foundational atlas for modeling PD genetic risk. We have integrated these data into a FOUNDIN-PD data browser as a resource for understanding the molecular pathogenesis of PD.

A guide for functional analysis of BRCA1 variants of uncertain significance.

Germline mutations in the tumor suppressor gene BRCA1 confer an estimated lifetime risk of 56-80% for breast cancer and 15-60% for ovarian cancer. Since the mid 1990s when BRCA1 was identified, genetic testing has revealed over 1,500 unique germline variants. However, for a significant number of these variants, the effect on protein function is unknown making it difficult to infer the consequences on risks of breast and ovarian cancers. Thus, many individuals undergoing genetic testing for BRCA1 mutations receive test results reporting a variant of uncertain clinical significance (VUS), leading to issues in risk assessment, counseling, and preventive care. Here, we describe functional assays for BRCA1 to directly or indirectly assess the impact of a variant on protein conformation or function and how these results can be used to complement genetic data to classify a VUS as to its clinical significance. Importantly, these methods may provide a framework for genome-wide pathogenicity assignment.

Outcomes in pediatric atypical spitz tumors treated without sentinel lymph node biopsy.

The diagnosis of atypical Spitz tumor (AST) in a pediatric patient conveys an uncertain potential for malignancy. Although pediatric melanoma is rare, AST may be treated aggressively with sentinel lymph node biopsy (SLNB) and subsequent completion lymphadenectomy. These procedures have unclear therapeutic benefit and potential morbidity. We aimed to collect outcomes on children with AST treated with excision alone to assist in the management of these lesions. We queried our institution's pathology database for AST specimens submitted between 1994 and 2009. A dermatopathologist reviewed pathology slides to confirm AST diagnosis. Clinical information was obtained from medical records, and outcomes surveys were administered to children with AST. Twenty-nine patients met AST diagnostic criteria and were included in this study. Mean age at first excision was 9.0 ± 4.2 (range 2.3-17.5), and 19 patients underwent more than one excision procedure to achieve clear margins. No patient had SLNB. Fourteen patients (48%) with mean follow-up time of 8.4 years (range 3.5-15.8) completed clinical outcomes surveys. Outcomes with mean follow-up time of 2.8 years (range 0.02-8.1 years) were obtained for 10 additional patients from medical records. There were no reports of recurrence, additional lesions, or metastases in these 24 patients. We report one of the largest series of children with AST treated using excision alone and who remain disease free after a significant follow-up period. Our data suggest that SLNB is not warranted in the routine management of pediatric AST. We recommend complete excision with clear margins and careful clinical follow-up.