Engineering an fgfr4 knockout zebrafish to study its role in development and disease.

Fibroblast growth factor receptor 4 (FGFR4) has a role in many biological processes, including lipid metabolism, tissue repair, and vertebrate development. In recent years, FGFR4 overexpression and activating mutations have been associated with numerous adult and pediatric cancers. As such, FGFR4 presents an opportunity for therapeutic targeting which is being pursued in clinical trials. To understand the role of FGFR4 signaling in disease and development, we generated and characterized three alleles of fgfr4 knockout zebrafish strains using CRISPR/Cas9. To generate fgfr4 knockout crispants, we injected single-cell wildtype zebrafish embryos with fgfr4 targeting guide RNA and Cas9 proteins, identified adult founders, and outcrossed to wildtype zebrafish to create an F1 generation. The generated mutations introduce a stop codon within the second Ig-like domain of Fgfr4, resulting in a truncated 215, 223, or 228 amino acid Fgfr4 protein compared to 922 amino acids in the full-length protein. All mutant strains exhibited significantly decreased fgfr4 mRNA expression during development, providing evidence for successful knockout of fgfr4 in mutant zebrafish. We found that, consistent with other Fgfr4 knockout animal models, the fgfr4 mutant fish developed normally; however, homozygous fgfr4 mutant zebrafish were significantly smaller than wildtype fish at three months post fertilization. These fgfr4 knockout zebrafish lines are a valuable tool to study the role of FGFR4 in vertebrate development and its viability as a potential therapeutic target in pediatric and adult cancers, as well as other diseases.

Scalable plasma and digital cognitive markers for diagnosis and prognosis of Alzheimer's disease and related dementias.

INTRODUCTION: With emergence of disease-modifying therapies, efficient diagnostic pathways are critically needed to identify treatment candidates, evaluate disease severity, and support prognosis. A combination of plasma biomarkers and brief digital cognitive assessments could provide a scalable alternative to current diagnostic work-up. METHODS: We examined the accuracy of plasma biomarkers and a 10-minute supervised tablet-based cognitive assessment (Tablet-based Cognitive Assessment Tool Brain Health Assessment [TabCAT-BHA]) in predicting amyloid ß positive (Aß+) status on positron emission tomography (PET), concurrent disease severity, and functional decline in 309 older adults with subjective cognitive impairment (n = 49), mild cognitive impairment (n = 159), and dementia (n = 101). RESULTS: Combination of plasma pTau181, Aß42/40, neurofilament light (NfL), and TabCAT-BHA was optimal for predicting Aß-PET positivity (AUC = 0.962). Whereas NfL and TabCAT-BHA optimally predicted concurrent disease severity, combining these with pTau181 and glial fibrillary acidic protein was most accurate in predicting functional decline. DISCUSSION: Combinations of plasma and digital cognitive markers show promise for scalable diagnosis and prognosis of ADRD. HIGHLIGHTS: The need for cost-efficient diagnostic and prognostic markers of AD is urgent. Plasma and digital cognitive markers provide complementary diagnostic contributions. Combination of these markers holds promise for scalable diagnosis and prognosis. Future validation in community cohorts is needed to inform clinical implementation.

Investigating the Impact of Kratom (Mitragyna speciosa) Use Upon Male Sexual Health.

Purpose: Kratom (Mitragyna speciosa) exhibits µ-receptor agonism and is used as an opioid substitute. While opioids are known to inhibit sexual behavior, less is known regarding kratom. We conducted a pilot study to assess the subjective impact of kratom upon male sexual health including erectile and ejaculatory function. Patients and Methods: Twitter and Reddit (r/Kratom) were accessed to disseminate our survey featuring validated instruments (the International Index of Erectile Function, IIEF, and the premature ejaculation diagnostic tool, PEDT). Sexual health prior to and after 4 weeks of kratom use was assessed. Results: Most males surveyed (n = 165) were 18-40 years old (84.9%), with 95.8% of respondents using it at least weekly and 82.4% using kratom for ≥1 year. Reasons for use included treating pain (39.4%), and mental health conditions (63.6%). Kratom was associated with a positive (37.7%) and negative (20.5%) impact on sexual health. Kratom subjectively increased time to ejaculation in 104 (66.6%) patients, perceived as positive by 62 (59.6%). Seventy-eight patients answered questions about premature ejaculation. The median (with interquartile range, IQR, following;) pre-kratom and kratom use scores were 13.0; 8.0 and 6.5; 5.0, respectively (p < 0.001). Ejaculation before 5 minutes improved after kratom (51.3% vs 12.8%) (p < 0.0001). Following kratom use, patients reported lack of frustration with ejaculation prior to desire (21.8% vs 61.5%) (p < 0.001). The erectile function domain of the IIEF was statistically significantly different however - clinically similar pre-kratom use (29.0; 5.75) versus 27.0; 6.75 during kratom use (p = 0.037). Conclusion: Clinicians treating male sexual health should be aware of kratom and its potential effect on ejaculatory and erectile function.

VGLL2-NCOA2 leverages developmental programs for pediatric sarcomagenesis.

Clinical sequencing efforts are rapidly identifying sarcoma gene fusions that lack functional validation. An example is the fusion of transcriptional coactivators, VGLL2-NCOA2, found in infantile rhabdomyosarcoma. To delineate VGLL2-NCOA2 tumorigenic mechanisms and identify therapeutic vulnerabilities, we implement a cross-species comparative oncology approach with zebrafish, mouse allograft, and patient samples. We find that VGLL2-NCOA2 is sufficient to generate mesenchymal tumors that display features of immature skeletal muscle and recapitulate the human disease. A subset of VGLL2-NCOA2 zebrafish tumors transcriptionally cluster with embryonic somitogenesis and identify VGLL2-NCOA2 developmental programs, including a RAS family GTPase, ARF6. In VGLL2-NCOA2 zebrafish, mouse, and patient tumors, ARF6 is highly expressed. ARF6 knockout suppresses VGLL2-NCOA2 oncogenic activity in cell culture, and, more broadly, ARF6 is overexpressed in adult and pediatric sarcomas. Our data indicate that VGLL2-NCOA2 is an oncogene that leverages developmental programs for tumorigenesis and that reactivation or persistence of ARF6 could represent a therapeutic opportunity.

Zebrafish her3 knockout impacts developmental and cancer-related gene signatures.

HES3 is a basic helix-loop-helix transcription factor that regulates neural stem cell renewal during development. HES3 overexpression is predictive of reduced overall survival in patients with fusion-positive rhabdomyosarcoma, a pediatric cancer that resembles immature and undifferentiated skeletal muscle. However, the mechanisms of HES3 cooperation in fusion-positive rhabdomyosarcoma are unclear and are likely related to her3/HES3's role in neurogenesis. To investigate HES3's function during development, we generated a zebrafish CRISPR/Cas9 null mutation of her3, the zebrafish ortholog of HES3. Loss of her3 is not embryonic lethal and adults exhibit expected Mendelian ratios. Embryonic her3 zebrafish mutants exhibit dysregulated neurog1 expression, a her3 target gene, and the mutant her3 fails to bind the neurog1 promoter sequence. Further, her3 mutants are significantly smaller than wildtype and a subset present with lens defects as adults. Transcriptomic analysis of her3 mutant embryos indicates that genes involved in organ development, such as pctp and grinab, are significantly downregulated. Further, differentially expressed genes in her3 null mutant embryos are enriched for Hox and Sox10 motifs. Several cancer-related gene pathways are impacted, including the inhibition of matrix metalloproteinases. Altogether, this new model is a powerful system to study her3/HES3-mediated neural development and its misappropriation in cancer contexts.

In vivo isotope tracing reveals a requirement for the electron transport chain in glucose and glutamine metabolism by tumors.

In mice and humans with cancer, intravenous 13C-glucose infusion results in 13C labeling of tumor tricarboxylic acid (TCA) cycle intermediates, indicating that pyruvate oxidation in the TCA cycle occurs in tumors. The TCA cycle is usually coupled to the electron transport chain (ETC) because NADH generated by the cycle is reoxidized to NAD+ by the ETC. However, 13C labeling does not directly report ETC activity, and other pathways can oxidize NADH, so the ETC's role in these labeling patterns is unverified. We examined the impact of the ETC complex I inhibitor IACS-010759 on tumor 13C labeling. IACS-010759 suppresses TCA cycle labeling from glucose or lactate and increases labeling from glutamine. Cancer cells expressing yeast NADH dehydrogenase-1, which recycles NADH to NAD+ independently of complex I, display normalized labeling when complex I is inhibited, indicating that cancer cell ETC activity regulates TCA cycle metabolism and 13C labeling from multiple nutrients.

Facilitators and Barriers to Dementia Assessment and Diagnosis: Perspectives From Dementia Experts Within a Global Health Context.

Objectives: Dementia poses one of the greatest global health challenges, affecting 50 million people worldwide. With 10 million new cases each year, dementia is a growing burden, particularly in low- and middle-income countries (LMIC). This study aimed to identify the facilitators and barriers to providing quality dementia assessment and care in LMICs from a global health perspective. Methods/Design: A qualitative semi-structured interview study with 20 dementia expert healthcare providers from 19 countries. To be included, providers had to: practice dementia assessment or care in LMICs where the population over age 60 is projected to more than double by 2050 and be recognized as a leading dementia expert in the region based on position, research publications, and/or policy leadership. Interviews were analyzed by a multidisciplinary team of researchers using thematic analysis. Results: Barriers to dementia assessment and care included stigma about dementia, poor patient engagement in and access to healthcare, inadequate linguistic and cultural validation, limited dementia capable workforce, competing healthcare system priorities, and insufficient health financing. Facilitators included the rise in dementia awareness campaigns, dementia training for general practitioners, availability of family support and family caregivers, and national and international collaborations including coordinated policy efforts and involvement in international research initiatives. Conclusions: Findings from this study provide insights for prioritizing dementia assessment and care capacity-building in LMICs as a global health priority and for tailored public health approaches to strengthen dementia assessment and care at the individual, community, national, and multi-national levels.

Global Perspectives on Brief Cognitive Assessments for Dementia Diagnosis.

BACKGROUND: Timely diagnosis of dementia is a global healthcare priority, particularly in low to middle income countries where rapid increases in older adult populations are expected. OBJECTIVE: To investigate global perspectives on the role of brief cognitive assessments (BCAs) in dementia diagnosis, strengths and limitations of existing measures, and future directions and needs. METHODS: This is a qualitative study of 18 dementia experts from different areas of the world. Participants were selected using purposeful sampling based on the following criteria: 1) practicing in countries with projected growth of older adult population of over 100%by 2050; 2) expertise in dementia diagnosis and treatment; 3) involvement in clinical practice and training; and 4) recognition as a national dementia expert based on leadership positions within healthcare system, research, and/or policy work. Participants were individually interviewed in their language of choice over secure videoconference sessions. Interviews were analyzed by a multidisciplinary team using theme identification approach. RESULTS: Four domains with subthemes emerged illustrating participants' perspectives: 1) strengths of BCAs; 2) limitations of BCAs; 3) needs related to the use of BCAs; and 4) characteristics of an ideal BCA. While most experts agreed that BCAs were important and useful for dementia diagnosis, the themes emphasized the need for development and validation of novel measures that are sensitive, psychometrically sound, and culturally appropriate. CONCLUSION: BCAs are important for guiding diagnosis and care for dementia patients. Findings provide a roadmap for novel BCA development to assist in diagnostic decision making for clinicians serving a rapidly growing and diverse dementia population.

Detecting Alzheimer's disease biomarkers with a brief tablet-based cognitive battery: sensitivity to Aß and tau PET.

BACKGROUND: ß-amyloid (Aß) and tau positron emission tomography (PET) detect the pathological changes that define Alzheimer's disease (AD) in living people. Cognitive measures sensitive to Aß and tau burden may help streamline identification of cases for confirmatory AD biomarker testing. METHODS: We examined the association of Brain Health Assessment (BHA) tablet-based cognitive measures with dichotomized Aß -PET status using logistic regression models in individuals with mild cognitive impairment (MCI) or dementia (N = 140; 43 Aß-, 97 Aß+). We also investigated the relationship between the BHA tests and regional patterns of tau-PET signal using voxel-wise regression analyses in a subsample of 60 Aß+ individuals with MCI or dementia. RESULTS: Favorites (associative memory), Match (executive functions and speed), and Everyday Cognition Scale scores were significantly associated with Aß positivity (area under the curve [AUC] = 0.75 [95% CI 0.66-0.85]). We found significant associations with tau-PET signal in mesial temporal regions for Favorites, frontoparietal regions for Match, and occipitoparietal regions for Line Orientation (visuospatial skills) in a subsample of individuals with MCI and dementia. CONCLUSION: The BHA measures are significantly associated with both Aß and regional tau in vivo imaging markers and could be used for the identification of patients with suspected AD pathology in clinical practice.

A ubiquitin ligase mediates target-directed microRNA decay independently of tailing and trimming.

MicroRNAs (miRNAs) act in concert with Argonaute (AGO) proteins to repress target messenger RNAs. After AGO loading, miRNAs generally exhibit slow turnover. An important exception occurs when miRNAs encounter highly complementary targets, which can trigger a process called target-directed miRNA degradation (TDMD). During TDMD, miRNAs undergo tailing and trimming, suggesting that this is an important step in the decay mechanism. We identified a cullin-RING ubiquitin ligase (CRL), containing the substrate adaptor ZSWIM8, that mediates TDMD. The ZSWIM8 CRL interacts with AGO proteins, promotes TDMD in a tailing and trimming-independent manner, and regulates miRNA expression in multiple cell types. These findings suggest a model in which the ZSWIM8 ubiquitin ligase mediates TDMD by directing proteasomal decay of miRNA-containing complexes engaged with highly complementary targets.

BHA-CS: A novel cognitive composite for Alzheimer's disease and related disorders.

INTRODUCTION: Composite scores based on psychometrically rigorous cognitive assessments are well suited for early diagnosis and disease monitoring. METHODS: We developed and cross-validated the Brain Health Assessment-Cognitive Score (BHA-CS), based on a brief computerized battery, in 451 cognitively normal (CN) and 399 cognitively impaired (mild cognitive impairment [MCI] or dementia) older adults. We investigated its long-term reliability and reliable change indices at longitudinal follow-up (N = 340), and the association with amyloid beta (Aß) burden in the CN subgroup with Aß positron emission tomography (N = 119). RESULTS: The BHA-CS was accurate at detecting cognitive impairment and exhibited excellent long-term stability. Reliable decline over one year was detected in 75% of participants with dementia, 44% with MCI, and 3% of CN. Among CN, the Aß-positive group showed worse longitudinal performance on the BHA-CS compared to the Aß-negative group. DISCUSSION: The BHA-CS is sensitive to cognitive decline in preclinical and prodromal neurodegenerative disease.

PAX3-FOXO1 transgenic zebrafish models identify HES3 as a mediator of rhabdomyosarcoma tumorigenesis.

Alveolar rhabdomyosarcoma is a pediatric soft-tissue sarcoma caused by PAX3/7-FOXO1 fusion oncogenes and is characterized by impaired skeletal muscle development. We developed human PAX3-FOXO1 -driven zebrafish models of tumorigenesis and found that PAX3-FOXO1 exhibits discrete cell lineage susceptibility and transformation. Tumors developed by 1.6-19 months and were primitive neuroectodermal tumors or rhabdomyosarcoma. We applied this PAX3-FOXO1 transgenic zebrafish model to study how PAX3-FOXO1 leverages early developmental pathways for oncogenesis and found that her3 is a unique target. Ectopic expression of the her3 human ortholog, HES3, inhibits myogenesis in zebrafish and mammalian cells, recapitulating the arrested muscle development characteristic of rhabdomyosarcoma. In patients, HES3 is overexpressed in fusion-positive versus fusion-negative tumors. Finally, HES3 overexpression is associated with reduced survival in patients in the context of the fusion. Our novel zebrafish rhabdomyosarcoma model identifies a new PAX3-FOXO1 target, her3/HES3, that contributes to impaired myogenic differentiation and has prognostic significance in human disease.

Dust observations at orbital altitudes surrounding Mars.

Dust is common close to the martian surface, but no known process can lift appreciable concentrations of particles to altitudes above ~150 kilometers. We present observations of dust at altitudes ranging from 150 to above 1000 kilometers by the Langmuir Probe and Wave instrument on the Mars Atmosphere and Volatile Evolution spacecraft. Based on its distribution, we interpret this dust to be interplanetary in origin. A comparison with laboratory measurements indicates that the dust grain size ranges from 1 to 12 micrometers, assuming a typical grain velocity of ~18 kilometers per second. These direct observations of dust entering the martian atmosphere improve our understanding of the sources, sinks, and transport of interplanetary dust throughout the inner solar system and the associated impacts on Mars's atmosphere.

"I know if I drink I won't feel anything": substance use relapse among depressed women leaving prison.

PURPOSE: The purpose of this paper is to explore treatment needs and factors contributing to engagement in substance use and sobriety among women with co-occurring substance use and major depressive disorders (MDDs) as they return to the community from prison. DESIGN/METHODOLOGY/APPROACH: The paper used qualitative methods to evaluate the perspectives of 15 women with co-occurring substance use and MDDs on the circumstances surrounding their relapse and recovery episodes following release from a US prison. Women were recruited in prison; qualitative data were collected using semi-structured interviews conducted after prison release and were analyzed using grounded theory analysis. Survey data from 39 participants supplemented qualitative findings. FINDINGS: Results indicated that relationship, emotion, and mental health factors influenced women's first post-prison substance use. Women attributed episodes of recovery to sober and social support, treatment, and building on recovery work done in prison. However, they described a need for comprehensive pre-release planning and post-release treatment that would address mental health, family, and housing/employment and more actively assist them in overcoming barriers to care. PRACTICAL IMPLICATIONS: In-prison and aftercare treatment should help depressed, substance using women prisoners reduce or manage negative affect, improve relationships, and obtain active and comprehensive transitional support. ORIGINALITY/VALUE: Women with co-occurring mental health and substance use disorders are a high-risk population for negative post-release outcomes, but limited information exists regarding the processes by which they relapse or retain recovery after release from prison. Findings inform treatment and aftercare development efforts.

Exploring the use of mobile technologies for the acquisition of clinical skills.

Mobile learning has the potential to supplement information communication technology (ICT), online learning and the traditional teaching and learning methods to educate practitioners in the clinical practice area. Following the development of several Post Graduate modules of learning for the theory and clinical skills required to undertake the Newborn Infant Physical Examination (NIPE), a small research study was undertaken to combine mobile learning and NIPE. The research study explored the hypothesis that mobile devices could be used in pedagogically effective ways to support and enhance the learning and acquisition of clinical skills in the clinical arena. Participants in the study each received a handheld mobile device (iPod) that had been loaded with several Reusable Learning Objects (RLO) outlining each aspect of the physical examination to be performed. At the end of the module (12 weeks in duration), each participant completed an evaluation questionnaire. Participants confirmed that mobile learning afforded flexibility in time and place of learning and captured their interest in the learning material. This study reports that the use of mobile technology for skill acquisition is creative and innovative, placing learning firmly in the hands of the learner.

Structure of an exploding laser-produced plasma.

We describe the first-ever volumetric, time-resolved measurements performed with a moving probe within an expanding dense plasma, embedded in a background magnetized plasma. High-resolution probe measurements of the magnetic field and floating potential in multiple 2D cut planes combined with a 1 Hz laser system reveal complex three-dimensional current systems within the expanding plasma. Static (ωreal=0) flutelike density striations are observed at the leading edge of the plasma, which are correlated to variations in the current layer at the edge of the expanding plasma.

Two-dimensional micron-step probe drive for laboratory plasma measurement.

Laboratory measurement of small-scale ( approximately 1 mm) magnetic phenomena over an extended area is a challenge requiring precise diagnostics. We present a novel two dimensional magnetic probe platform capable of directly measuring the magnetic field over a 36 cm(2) region at spatial resolutions better than 1 mm. The platform is discussed in the context of an experiment at the Large Plasma Device facility at UCLA, designed to measure the magnetic interaction between two counterpropagating laser-produced plasmas. The use of a precise, repeatable positioning platform enables the recovery of information about the interaction using cross-correlation techniques.

ISEApeaks: an Excel platform for GeneScan and Immunoscope data retrieval, management and analysis.

ISEApeaks retrieves and handles DNA sequencer raw data (peak area and nucleotidic length). This Macintosh package efficiently interfaces GeneScan and Immunoscope softwares to Excel by extracting data from scattered files, organizing data and gathering it into a unique peak database for all samples.

Collaborative partnership for optimizing heart failure outcomes in the long-term care environment: a clinical experience.

RATIONALE: Heart failure (HF) is a devastating cardiovascular syndrome affecting more than 4.6 million Americans and resulting in a substantial economic burden. It is the number one cause of hospitalization in the older population. Because there is no cure for this costly disease, goals of therapy include; slowing progression, increasing patient survival, minimizing symptoms, improving patient functional capacity, and decreasing ER visits, hospital admissions, and readmissions. Studies have shown ACE inhibitors (ACEIs) to be extremely beneficial in reaching these goals. However, data nationwide indicate that less than half of HF patients are prescribed an ACEI. Therefore, a national long-term care pharmaceutical provider has committed to the development of a HF disease management program, which has been implemented through a collaborative partnership between medicine, nursing, and pharmacy. This report describes the experience and results of implementing this program in a large group of nursing homes. METHODS: With help from an independent health outcomes consulting firm and an expert panel, a multifaceted program was developed. The AMDA heart failure guidelines were used as a foundation to design program interventions, which included educational programs for the long-term care facility staff, implementation of a HF treatment algorithm, and an outcomes measurement program. The primary goals of this program were to increase awareness of HF and treatment options available and to decrease morbidity and mortality associated with HF through the optimization of pharmacologic management in a long-term care setting. OUTCOMES: A retrospective evaluation was conducted to assess changes in ACEI utilization, ACEI targeted dosing and HF-related hospitalizations 1 year after program implementation. A total of 510 patients followed by 23 consultants, representing 152 facilities met the inclusion criteria for the analysis. Data indicated an increase of 36% in newly enrolled HF patients utilizing ACEI after 1 year. A decreasing trend in HF hospitalizations was also noted from the first quarter, (2.2/100 patients) to the fourth (1.0/100 patients). The results of the Heart Failure-Health Outcomes Management Program (HF-HOMP) program demonstrate the opportunity existing for long-term care providers to improve patient health while minimizing expenditures. It also provides additional evidence that such programs should be implemented more broadly within this population.