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Mitochondria are central players in host immunometabolism as they function not only as metabolic hubs but also as signaling platforms regulating innate immunity. Environmental exposures to mitochondrial toxicants occur widely and are increasingly frequent. Exposures to these mitotoxicants may pose a serious threat to organismal health and the onset of diseases by disrupting immunometabolic pathways. In this study, we investigated whether the Complex I inhibitor rotenone could alter

Covid-19 raised many novel ethical issues including regarding the allocation of opportunities to participate in clinical trials during a public health emergency. In this article, we explore how hospitals that have a scarcity of trial opportunities, either overall or in a specific trial, can equitably allocate those opportunities in the context of an urgent medical need with limited therapeutic interventions. We assess the three main approaches to allocating trial opportunities discussed in the literature: patient choice, physician referral, and randomization/lottery. As, we argue, none of the three typical approaches are ethically ideal for allocating trial opportunities in the pandemic context, many hospitals have instead implemented hybrid solutions. We offer practical guidance to support those continuing to face these challenges, and we analyze options for the future.

Introduction: The purpose of this study was to evaluate the effect of collimation on image quality and radiation dose to the eye lenses of the personnel involved in computed radiography of the canine pelvis.Materials and Methods: A retrospective study of canine pelvic radiographs (N = 54) was undertaken to evaluate the relationship between image quality and the degree of field the collimation used. This was followed by a prospective cadaver study (N = 18) that assessed the effects on image quality and on scattered radiation dose of different collimation field areas and exposure parameters. All radiographs were analyzed for image quality using a Visual Grading Analysis (VGA) with three observers. Finally, the potential scattered radiation dose to the eye lens of personnel restraining a dog for pelvic radiographs was measured.Results: The retrospective study showed a slightly better (statistically non-significant) VGA score for the radiographs with optimal collimation. Spatial and contrast resolution and image sharpness showed the greatest improvement in response to minimizing the collimation field. The prospective study showed slightly better VGA scores (improved image quality) with the optimal collimation. Increasing the exposure factors especially the tube current and exposure time (mAs) resulted in improved low contrast resolution and less noise in the radiographs. The potential eye lens radiation dose increased by 14, 28, and 40% [default exposures, increased the tube peak potential (kVp), increased mAs, respectively] as a result of reduced collimation (increased beam size).Conclusion: The degree of collimation has no statistically significant on image quality in canine pelvic radiology for the range of collimation used but does have an impact on potential radiation dose to personnel in the x-ray room. With regard to radiation safety, increases in kVp are associated with less potential scatter radiation exposure compared to comparable increases in mAs.

Dopamine neuron loss is involved in the pathology of Parkinson's Disease (PD), a highly prevalent neurodegenerative disorder affecting over 10 million people worldwide. Since many details about PD etiology remain unknown, studies investigating genetic and environmental contributors to PD are needed to discover methods of prevention, management, and treatment. Proper characterization of dopaminergic neuronal loss may be relevant not only to PD research, but to other increasingly prevalent neurodegenerative disorders. There are established genetic and chemical models of dopaminergic neurodegeneration in the Caenorhabditis elegans model system, with easy visualization of neurobiology supported by the nematodes' transparency and invariant neuronal architecture. In particular, hermaphroditic C. elegans' dopaminergic neuron morphological changes can be visualized using strains with fluorescent reporters driven by cell-specific promotors such as the dat-1 dopamine transporter gene, which is expressed exclusively in their eight dopaminergic neurons. With the capabilities of this model system and the appropriate technology, many laboratories have studied dopaminergic neurodegeneration. However, there is little consistency in the way the data is analyzed and much of the present literature uses binary scoring analyses that capture the presence of degeneration but not the full details of the progression of neuron loss. Here, we introduce a universal scoring system to assess morphological changes and degeneration in C. elegans' cephalic neuron dendrites. This seven-point scale allows for analysis across a full range of dendrite morphology, ranging from healthy neurons to complete dendrite loss, and considering morphological details including kinks, branching, blebs, and breaks. With this scoring system, researchers can quantify subtle age-related changes as well as more dramatic chemical-induced changes. Finally, we provide a practice set of images with commentary that can be used to train, calibrate, and assess the scoring consistency of researchers new to this method. This should improve within- and between- laboratory consistency, increasing rigor and reproducibility.

Myeloid-derived suppressor cells (MDSCs) are a heterogeneous group of cells, which can suppress the immune response, promote tumor progression and impair the efficacy of immunotherapies. Consequently, the pharmacological targeting of MDSC is emerging as a new immunotherapeutic strategy to stimulate the natural anti-tumor immune response and potentiate the efficacy of immunotherapies. Herein, we leveraged genetically modified models and a small molecule inhibitor to validate Calcium-Calmodulin Kinase Kinase 2 (CaMKK2) as a druggable target to control MDSC accumulation in tumor-bearing mice. The results indicated that deletion of CaMKK2 in the host attenuated the growth of engrafted tumor cells, and this phenomenon was associated with increased antitumor T cell response and decreased accumulation of MDSC. The adoptive transfer of MDSC was sufficient to restore the ability of the tumor to grow in Camkk2-/- mice, confirming the key role of MDSC in the mechanism of tumor rejection. In vitro studies indicated that blocking of CaMKK2 is sufficient to impair the yield of MDSC. Surprisingly, MDSC generated from Camkk2-/- bone marrow cells also showed a higher ability to terminally differentiate toward more immunogenic cell types (e.g inflammatory macrophages and dendritic cells) compared to wild type (WT). Higher intracellular levels of reactive oxygen species (ROS) accumulated in Camkk2-/- MDSC, increasing their susceptibility to apoptosis and promoting their terminal differentiation toward more mature myeloid cells. Mechanistic studies indicated that AMP-activated protein kinase (AMPK), which is a known CaMKK2 proximal target controlling the oxidative stress response, fine-tunes ROS accumulation in MDSC. Accordingly, failure to activate the CaMKK2-AMPK axis can account for the elevated ROS levels in Camkk2-/- MDSC. These results highlight CaMKK2 as an important regulator of the MDSC lifecycle, identifying this kinase as a new druggable target to restrain MDSC expansion and enhance the efficacy of anti-tumor immunotherapy.

This study aimed to decrease surgeon exposure to ionizing radiation through a new learning technique, "deliberate practice", which consists in improving performance by setting goals with feedback. The hypothesis was that exposure to ionizing radiation during distal radius fracture surgery using the minimally invasive plate osteosynthesis (MIPO) technique decreased faster with "deliberate" practice than with "naïve" practice. Radiographic dosimetry was measured in the first 30 fractures operated on by MIPO by 6 surgeons. The first 3 surgeons operated "naively" (Group 1) and the next 3 according to the "deliberate" procedure (Group 2). Group 2 received weekly feedback (number of exposed hands, number of fluoroscopic views, exposure duration, and X-ray dose). An expert, using fluoroscopic images and surgical videos, provided suggestions for improvement. Mean number of exposed hands was 23.66 in Group 1 and 1.9 in Group 2. Mean number of fluoroscopic views was 78.31 and 35.0, respectively. Mean X-ray exposure time was 74.34 and 32.89 s, respectively. Mean dosimetry was 1.40 mGy (and 0.59 mGy, respectively. The hypothesis was thus confirmed: dosimetry decreased faster in Group 2 than in Group 1. Teaching this deliberate practice should be generalized, to decrease the growth phase and increase the plateau phase of the learning curve.

Silver nanoparticles (AgNPs) are well-proven antimicrobial nanomaterials, yet little is elucidated regarding the mechanism underlying cytotoxicity induced by these nanoparticles. Here, we tested the hypothesis that mitochondria are primary intracellular targets of two AgNPs and silver ions, in mouse hepatocytes (AML12) cultured in glucose and galactose-based media. AML12 cells were more sensitive to mitochondrial uncoupling when grown with galactose rather than glucose. However, 24 hour-treatments with 15 nm AgNPs and 6 nm GA-AgNPs (5 and 10 μg/mL), and AgNO(3) (1 and 3 μg/mL), concentrations that resulted in either 10 or 30% cytotoxicity, failed to cause more toxicity to AML12 cells grown on galactose than glucose. Furthermore, colocalization analysis and subcellular Ag quantification did not show any enrichment of silver content in mitochondria in either medium. Finally, the effects of the same exposures on mitochondrial respiration were mild or undetectable, a result inconsistent with mitochondrial toxicity causing cell death. Our results suggest that neither ionic Ag nor the AgNPs that we tested specifically target mitochondria, and are inconsistent with mitochondrial dysfunction being the primary cause of cell death after Ag exposure under these conditions.

The nicotinamide cofactor specificity of enzymes plays a key role in regulating metabolic processes and attaining cellular homeostasis. Multiple studies have used enzyme engineering tools or a directed evolution approach to switch the cofactor preference of specific oxidoreductases. However, whole-cell adaptation toward the emergence of novel cofactor regeneration routes has not been previously explored. To address this challenge, we used an Escherichia coli NADPH-auxotrophic strain. We continuously cultivated this strain under selective conditions. After 500 to 1,100 generations of adaptive evolution using different carbon sources, we isolated several strains capable of growing without an external NADPH source. Most isolated strains were found to harbor a mutated NAD+-dependent malic enzyme (MaeA). A single mutation in MaeA was found to switch cofactor specificity while lowering enzyme activity. Most mutated MaeA variants also harbored a second mutation that restored the catalytic efficiency of the enzyme. Remarkably, the best MaeA variants identified this way displayed overall superior kinetics relative to the wild-type variant with NAD+. In other evolved strains, the dihydrolipoamide dehydrogenase (Lpd) was mutated to accept NADP+, thus enabling the pyruvate dehydrogenase and 2-ketoglutarate dehydrogenase complexes to regenerate NADPH. Interestingly, no other central metabolism oxidoreductase seems to evolve toward reducing NADP+, which we attribute to several biochemical constraints, including unfavorable thermodynamics. This study demonstrates the potential and biochemical limits of evolving oxidoreductases within the cellular context toward changing cofactor specificity, further showing that long-term adaptive evolution can optimize enzyme activity beyond what is achievable via rational design or directed evolution using small libraries. IMPORTANCE In the cell, NAD(H) and NADP(H) cofactors have different functions. The former mainly accepts electrons from catabolic reactions and carries them to respiration, while the latter provides reducing power for anabolism. Correspondingly, the ratio of the reduced to the oxidized form differs for NAD+ (low) and NADP+ (high), reflecting their distinct roles. We challenged the flexibility of E. coli’s central metabolism in multiple adaptive evolution experiments using an NADPH-auxotrophic strain. We found several mutations in two enzymes, changing the cofactor preference of malic enzyme and dihydrolipoamide dehydrogenase. Upon deletion of their corresponding genes we performed additional evolution experiments which did not lead to the emergence of any additional mutants. We attribute this restricted number of mutational targets to intrinsic thermodynamic barriers; the high ratio of NADPH to NADP+ limits metabolic redox reactions that can regenerate NADPH, mainly by mass action constraints.

Over the last decades, the revolution in DNA sequencing has changed the way we understand the genetics and biology of B-cell lymphomas by uncovering a large number of recurrently mutated genes, whose aberrant function is likely to play an important role in the initiation and/or maintenance of these cancers. Dissecting how the involved genes contribute to the physiology and pathology of germinal center (GC) B cells –the origin of most B-cell lymphomas– will be key to advance our ability to diagnose and treat these patients. Genetically engineered mouse models (GEMM) that faithfully recapitulate lymphoma-associated genetic alterations offer a valuable platform to investigate the pathogenic roles of candidate oncogenes and tumor suppressors in vivo, and to pre-clinically develop new therapeutic principles in the context of an intact tumor immune microenvironment. In this review, we provide a summary of state-of-the art GEMMs obtained by accurately modelling the most common genetic alterations found in human GC B cell malignancies, with a focus on Burkitt lymphoma, follicular lymphoma, and diffuse large B-cell lymphoma, and we discuss how lessons learned from these models can help guide the design of novel therapeutic approaches for this disease.

CtIP is a DNA end resection factor widely implicated in alternative end-joining (A-EJ)–mediated translocations in cell-based reporter systems. To address the physiological role of CtIP, an essential gene, in translocation-mediated lymphomagenesis, we introduced the T855A mutation at murine CtIP to nonhomologous end-joining and Tp53 double-deficient mice that routinely succumbed to lymphomas carrying A-EJ–mediated IgH-Myc translocations. T855 of CtIP is phosphorylated by ATM or ATR kinases upon DNA damage to promote end resection. Here, we reported that the T855A mutation of CtIP compromised the neonatal development of Xrcc4(−/−)Tp53(−/−) mice and the IgH-Myc translocation-driven lymphomagenesis in DNA-PKcs(−/−)Tp53(−/−) mice. Mechanistically, the T855A mutation limits DNA end resection length without affecting hairpin opening, translocation frequency, or fork stability. Meanwhile, after radiation, CtIP-T855A mutant cells showed a consistent decreased Chk1 phosphorylation and defects in the G2/M cell cycle checkpoint. Consistent with the role of T855A mutation in lymphomagenesis beyond translocation, the CtIP-T855A mutation also delays splenomegaly in λ-Myc mice. Collectively, our study revealed a role of CtIP-T855 phosphorylation in lymphomagenesis beyond A-EJ–mediated chromosomal translocation.

The consequences of damage to the mitochondrial genome (mtDNA) are poorly understood, although mtDNA is more susceptible to damage resulting from some genotoxicants than nuclear DNA (nucDNA), and many environmental toxicants target the mitochondria. Reports from the toxicological literature suggest that exposure to early-life mitochondrial damage could lead to deleterious consequences later in life (the “Developmental Origins of Health and Disease” paradigm), but reports from other fields often report beneficial (“mitohormetic”) responses to such damage. Here, we tested the effects of low (causing no change in lifespan) levels of ultraviolet C (UVC)-induced, irreparable mtDNA damage during early development in Caenorhabditis elegans. This exposure led to life-long reductions in mtDNA copy number and steady-state ATP levels, accompanied by increased oxygen consumption and altered metabolite profiles, suggesting inefficient mitochondrial function. Exposed nematodes were also developmentally delayed, reached smaller adult size, and were rendered more susceptible to subsequent exposure to chemical mitotoxicants. Metabolomic and genetic analysis of key signaling and metabolic pathways supported redox and mitochondrial stress-response signaling during early development as a mechanism for establishing these persistent alterations. Our results highlight the importance of early-life exposures to environmental pollutants, especially in the context of exposure to chemicals that target mitochondria., Graphical abstract Image 1, Highlights • Early life mtDNA damage led to lifelong deficits in mitochondrial function. • C. elegans developed slowly and were sensitive to chemical exposures as adults. • Redox signaling is a mechanism that establishes these persistent alterations. • Data are consistent with the Developmental Origins of Health and Disease model.

Companies have recently begun to sell a new service to patients considering in vitro fertilization: embryo selection based on polygenic scores (ESPS). These scores represent individualized predictions of health and other outcomes derived from genomewide association studies in adults to partially predict these outcomes. This article includes a discussion of many factors that lower the predictive power of polygenic scores in the context of embryo selection and quantifies these effects for a variety of clinical and nonclinical traits. Also discussed are potential unintended consequences of ESPS (including selecting for adverse traits, altering population demographics, exacerbating inequalities in society, and devaluing certain traits). Recommendations for the responsible communication about ESPS by practitioners are provided, and a call for a society-wide conversation about this technology is made. (Funded by the National Institute on Aging and others.).

Mild suppression of mitochondrial activity has beneficial effects across species. The nematode Caenorhabditis elegans is a versatile, genetically tractable model organism widely employed for aging studies, which has led to the identification of many of the known evolutionarily conserved mechanisms regulating lifespan. In C. elegans the pro-longevity effect of reducing mitochondrial function, for example by RNA interference, is only achieved if mitochondrial stress is applied during larval development. Surprisingly, a careful analysis of changes in mitochondrial functions resulting from such treatments during the developmental windows in which pro-longevity signals are programmed has never been carried out. Thus, although the powerful C. elegans genetics have led to the identification of different molecular mechanisms causally involved in mitochondrial stress control of longevity, specific functional mitochondrial biomarkers indicative or predictive of lifespan remain to be identified. To fill this gap, we systematically characterized multiple mitochondrial functional parameters at an early developmental stage in animals that are long-lived due to mild knockdown of twelve different mitochondrial proteins and correlated these parameters with animals' lifespan. We found that basal oxygen consumption rate and ATP-linked respiration positively correlate with lifespan extension and propose the testable hypothesis that the Bioenergetic Health Index can be used as a proxy to predict health-span outcomes.

Mitochondria are key targets of many environmental contaminants, because specific chemicals can interact directly with mitochondrial proteins, lipids, and ribonucleic acids. These direct interactions serve as molecular initiating events that impede adenosine triphosphate production and other critical functions that mitochondria serve within the cell (e.g., calcium and metal homeostasis, apoptosis, immune signaling, redox balance). A limited but growing number of adverse outcome pathways (AOPs) have been proposed to associate mitochondrial dysfunction with effects at organismal and population levels. These pathways involve key events such as altered membrane potential, mitochondrial fission/fusion, and mitochondrial DNA damage, among others. The present critical review and analysis reveals current progress on AOPs involving mitochondrial dysfunction, and, using a network-based computational approach, identifies the localization of mitochondrial molecular initiating events and key events within multiple existing AOPs. We also present 2 case studies, the first examining the interaction between mitochondria and immunotoxicity, and the second examining the role of early mitochondrial dysfunction in the context of behavior (i.e., locomotor activity). We discuss limitations in our current understanding of mitochondrial AOPs and highlight opportunities for clarifying their details. Advancing our knowledge of key event relationships within the AOP framework will require high-throughput datasets that permit the development and testing of chemical-agnostic AOPs, as well as high-resolution research that will enhance the mechanistic testing and validation of these key event relationships. Given the wide range of chemicals that affect mitochondria, and the centrality of energy production and signaling to ecologically important outcomes such as pathogen defense, homeostasis, growth, and reproduction, a better understanding of mitochondrial AOPs is expected to play a significant, if not central, role in environmental toxicology. Environ Toxicol Chem 2019;38:1625-1634. © 2019 SETAC.

BACKGROUND Parenteral nutrition (PN) is a complicated therapy in which having specially trained clinicians can provide benefit, but it can be difficult to provide this level of expertise to all patients requiring PN. Creation of a virtual nutrition support team (NST) model allows patients across multiple hospitals to receive care from NST clinicians using remote conferencing technology on a daily basis. This study reviewed retrospective data from before and after implementation of the virtual model to assess quality indicators. METHODS The NST was developed including a team of dietitians and pharmacists with a physician medical director. Practice guidelines were developed to provide consistent methods for ordering and monitoring patients receiving PN. Patient charts in both the preintervention and postintervention groups were reviewed for indication for PN, duration of therapy, blood glucose levels, and demographic data. RESULTS A greater proportion of patients in the postintervention period had appropriate orders (97.2%) compared with patients in the preintervention period (58.9%) (P

Axolotls (Ambystoma mexicanum) from a research colony presented with multifocal, white chalky to gray skin lesions, a diffuse whitish to blue hue to the integument, and friable gill filaments. Skin scrapings and wet mounts revealed Chilodonella, Ichthyobodo, and a trichodinid species. The average overall burden (that is, all 3 species) per axolotl ranged from 0 to 25 parasites per 40 × field (p40f; mean ± 1 SD, 2.6 ± 5.5), with a prevalence of 12%, 60%, and 48%, respectively. Concurrent with husbandry modifications, axolotls were treated with an 8-h static immersion bath that contained 0.025 mL/L 37% formaldehyde. Chilodonella organisms were no longer observed after the initial treatment, and Ichthyobodo decreased from 2.4 ± 5.6 to 0.6 ± 1.8 organisms p40f. However, the average overall burden increased 4-fold to 10.5 ± 9.8 parasites p40f, and the trichodinid organisms increased 13-fold from 0.8 ± 2.3 to 10.4 ± 9.2 organisms p40f. A second treatment consisted of an 8-h immersion bath that contained 0.05 mL/L 37% formaldehyde on 2 consecutive days. A significant change was noted in the average overall burden of 0.5 ± 1.1 parasites p40f, a greater than 5- and 21-fold decrease from pretreatment and after the initial treatment, respectively. No significant change between the first and second treatment was observed for Ichthyobodo, with 0.6 ± 1.2 organisms p40f, but this number represented a significant decrease from pretreatment. After the second treatment, the trichodinid organism was detected in only one axolotl, with a low overall burden of 0.2 ± 0.4 organisms p40f and resulting in a significant decrease in the trichodinid count to 0.01 ± 0.04 organisms p40f. Treatment with formalin (37% formaldehyde), in conjunction with husbandry improvements, was effective in significantly reducing ectoparasite burden and eliminating clinical symptoms in axolotls but did not fully eliminate all protozoa.

Significance Randomized experiments—long the gold standard in medicine—are increasingly used throughout the social sciences and professions to evaluate business products and services, government programs, education and health policies, and global aid. We find robust evidence—across 16 studies of 5,873 participants from three populations spanning nine domains—that people often approve of untested policies or treatments (A or B) being universally implemented but disapprove of randomized experiments (A/B tests) to determine which of those policies or treatments is superior. This effect persists even when there is no reason to prefer A to B and even when recipients are treated unequally and randomly in all conditions (A, B, and A/B). This experimentation aversion may be an important barrier to evidence-based practice., Randomized experiments have enormous potential to improve human welfare in many domains, including healthcare, education, finance, and public policy. However, such “A/B tests” are often criticized on ethical grounds even as similar, untested interventions are implemented without objection. We find robust evidence across 16 studies of 5,873 participants from three diverse populations spanning nine domains—from healthcare to autonomous vehicle design to poverty reduction—that people frequently rate A/B tests designed to establish the comparative effectiveness of two policies or treatments as inappropriate even when universally implementing either A or B, untested, is seen as appropriate. This “A/B effect” is as strong among those with higher educational attainment and science literacy and among relevant professionals. It persists even when there is no reason to prefer A to B and even when recipients are treated unequally and randomly in all conditions (A, B, and A/B). Several remaining explanations for the effect—a belief that consent is required to impose a policy on half of a population but not on the entire population; an aversion to controlled but not to uncontrolled experiments; and a proxy form of the illusion of knowledge (according to which randomized evaluations are unnecessary because experts already do or should know “what works”)—appear to contribute to the effect, but none dominates or fully accounts for it. We conclude that rigorously evaluating policies or treatments via pragmatic randomized trials may provoke greater objection than simply implementing those same policies or treatments untested.

Tumor buds in colorectal cancer are hypothesized to undergo a (partial) epithelial-mesenchymal transition (EMT). If so, cytokeratin (CK) and vimentin (VIM) co-expression is expected. CK+/VIM+ can also be found in some stromal cells; however, their origin remains unclear. Here, we determine the frequency of CK+/VIM+ tumor cells and characterize the CK+/VIM+ stroma in colorectal cancer. Three cell populations (CK+, VIM+, CK+/VIM+) were sorted using DepArray and fluorescence-activated cell sorting (FACS). Tumor areas were selected to include tumor center, stroma and tumor budding. Fluorescence microscopy was used to visualize co-expressing cells on whole slides. A next-generation tissue microarray (ngTMA) of matched Pan-CK-positive and -negative stroma was constructed and stained for E-cadherin, VIM, Snail1, Twist1, Zeb1 and Zeb2, COL11A1, SPARC, CD90, α-SMA, FAP and WT1. CK+/VIM+ co-expressing tumor cells were detected using all three methods. With DepArray, only tumor budding areas contained CK+/VIM+ cells. The proportion of CK+/VIM+ tumor cells was low (1.5%-22%). CK+ stroma was associated with aggressive tumor features like distant metastasis (P = .0003), lymphatic invasion (P = .0009) and tumor budding (P = .0084). CK+/VIM+ stroma was characterized by positive WT1 (P .001), ZEB2 (P .001), TWIST1 (P = .009), and FAP (P = .003). Our data suggest that CK+/VIM+ tumor cells exist, albeit in low numbers and could represent a subgroup of tumor buds in partial EMT. CK+/VIM+ stroma may be of mesothelial origin and shows features of mesenchymal cells and cancer-associated fibroblasts. These results, together with the association with metastasis point to cells in mesothelial-mesenchymal transition (MMT). This atypical stroma may be a potential target for therapy.

Controllable, precise, and stable rotational manipulation of model organisms is valuable in many biomedical, bioengineering, and biophysics applications. We present an acoustofluidic chip capable of rotating Caenorhabditis elegans (C. elegans) in both static and continuous flow in a controllable manner. Rotational manipulation was achieved by exposing C. elegans to a surface acoustic wave (SAW) field that generated a vortex distribution inside a microchannel. By selectively activating interdigital transducers, we achieved bidirectional rotation of C. elegans, namely counterclockwise and clockwise, with on-demand switching of rotation direction in a single chip. In addition to continuous rotation, we also rotated C. elegans in a step-wise fashion with a step angle as small as 4° by pulsing the signal duration of SAW from a continuous signal to a pulsed signal down to 1.5 ms. Using this device, we have clearly imaged the dopaminergic neurons of C. elegans with pdat-1:GFP expression, as well as the vulval muscles and muscle fibers of the worm with myo-3::GFP fusion protein expression in different orientations and three dimensions. These achievements are difficult to realize through conventional (i.e., non-confocal) microscopy. The SAW manipulations did not detectably affect the health of the model organisms. With its precision, controllability, and simplicity in fabrication and operation, our acoustofluidic devices will be well-suited for model organism studies.

Background Glioblastoma multiforme (GBM) is the most prevalent and aggressive malignant tumor of the central nervous system. With a median survival of only one year, GBM patients have a particularly poor prognosis, highlighting a clear need for novel therapeutic strategies to target this disease. Transcriptional cyclin-dependent kinases (CDK), which phosphorylate key residues of RNA polymerase II (RNAPII) C-terminal domain (CTD), play a major role in sustaining aberrant transcriptional programs that are key to development and maintenance of cancer cells. Material and Methods We used pharmacological inhibition and genetic ablation to study effects of CDK12/CDK13 depletion on the proliferatory and migratory capacity of GBM cells and mouse xenografts. SLAM-seq, CUT&RUN and cell cycle assays were used to study the mechanistic effects of CDK12/CDK13 depletion in GBM cells. Results CDK12/CDK13 depletion markedly reduced the proliferatory and migratory capacity of GBM cells, as well as in vivo growth. CDK12/CDK13 inhibition potentiated existing chemotherapeutic treatments. Mechanistically, inhibition of CDK12/CDK13 leads to a genome-wide abrogation of RNAPII CTD phosphorylation, which in turn disrupts transcription and cell cycle progression in GBM cells. Conclusion These results provide proof-of-concept for the potential of CDK12 and CDK13 as therapeutic targets for GBM.

Rural Eastern Cape, South Africa.To identify steps in the TB preventive care cascade from routinely collected data among TB patients at a district hospital prior to the implementation of a novel TB program.This was a retrospective study. We adapted the TB prevention cascade to measure indicators routinely collected at district hospitals for TB using a cascade framework to evaluate outcomes in the cohort of close contacts.A total of 1,722 charts of TB patients were reviewed. The majority of patients (87%) were newly diagnosed with no previous episodes of TB. A total of 1,548 (90%) patients identified at least one close contact. A total of 7,548 contacts were identified with a median of 4.9 (range 1-16) contacts per patient. Among all contacts identified, 2,913 (39%) were screened for TB. Only 15 (0.5%) started TB preventive therapy and 122 (4.4%) started TB treatment. Nearly 25% of all medical history and clinical information was left unanswered among the 1,722 TB charts reviewed.Few close contacts were screened or started on TB preventive therapy in this cohort. Primary care providers for TB care in district health facilities should be informed of best practices for screening and treating TB infection and disease.Le Cap Est rural, le Cap, Afrique du Sud.Identifier les étapes de la cascade de soins préventifs de la TB à partir des données de routine recueillies parmi des patients dans un hôpital de district avant la mise en œuvre d’un nouveau programme TB.Ceci était une étude rétrospective. Nous avons adapté la cascade de prévention de la TB pour mesurer les indicateurs recueillis en routine dans les hôpitaux de district pour la TB en utilisant un cadre en cascade afin d’évaluer les résultats dans la cohorte des contacts étroits.Un total de 1 722 dossiers de patients TB a été revu. La majorité des patients (87%) avait un diagnostic nouveau sans épisode de TB préalable. Un total de 1 548 (90%) patients ont identifié au moins un contact étroit ; 7 548 contacts ont été identifiés avec une médiane de 4,9 (fourchette 1–16) contacts par patient. Parmi tous les contacts identifiés, 2 913 (39%) ont eu une recherche de TB. Seulement 15 (0,5%) ont initié le traitement préventif et 122 (4,4%) ont mis en route le traitement de TB. Près de 25% de tous les antécédents et autres informations cliniques n’était pas remplis dans les 1 722 dossiers TB revus.Peu de contacts étroits ont été dépistés ou mis sous traitement préventif de TB dans cette cohorte. Les prestataires de soins de santé primaires pour la TB dans les structures de santé des districts doivent être informés des meilleures pratiques pour le dépistage et le traitement de la TB infection et maladie.

Mitochondrial DNA (mtDNA) copy number is a critical component of overall mitochondrial health. In this chapter, we describe methods for simultaneous isolation of mtDNA and nuclear DNA (nucDNA), and measurement of their respective copy numbers using quantitative PCR. Methods differ depending on the species and cell type of the starting material, and availability of specific PCR reagents. We also briefly describe factors that affect mtDNA copy number and discuss caveats to its use as a biomarker.

Background There are limited data on outcomes of moderate to severe coronavirus disease 2019 (COVID-19) among patients treated with remdesivir and dexamethasone in a real-world setting. We sought to compare the effectiveness of standard of care (SOC) alone versus SOC plus remdesivir and dexamethasone. Methods Two population-based nationwide cohorts of individuals hospitalized with COVID-19 during February through December 2020 were studied. Death within 30 days and need of mechanical ventilation (MV) were compared by inverse probability of treatment weighted (ITPW) logistic regression analysis and shown as odds ratio (OR) with 95% confidence interval (CI). Results The 30-days mortality rate of 1694 individuals treated with remdesivir and dexamethasone in addition to SOC was 12.6% compared to 19.7% for 1053 individuals receiving SOC alone. This corresponded to a weighted OR of 30-day mortality of 0.47 (95% CI: .38–.57) for patients treated with remdesivir and dexamethasone compared to patients receiving SOC alone. Similarly, progression to MV was reduced (OR 0.36; 95% CI: .29–.46). Conclusions Treatment of moderate to severe COVID-19 during June through December that included remdesivir and dexamethasone was associated with reduced 30-day mortality and need of MV compared to treatment in February through May., Treatment of moderate to severe coronavirus disease 2019 (COVID-19) with remdesivir and dexamethasone was associated with a lower risk of mechanical ventilation and a significant better overall 30-day survival as compared to standard care alone.

Ovarian cancer (OVCA) patients may carry genes conferring cancer risk to biological family; however, fewer than one-quarter of patients receive genetic testing. “Traceback” cascade testing —outreach to potential probands and relatives—is a possible solution. This paper outlines a funded study (U01 CA240747-01A1) seeking to determine a Traceback program’s feasibility, acceptability, effectiveness, and costs. This is a multisite prospective observational feasibility study across three integrated health systems. Informed by the Conceptual Model for Implementation Research, we will outline, implement, and evaluate the outcomes of an OVCA Traceback program. We will use standard legal research methodology to review genetic privacy statutes; engage key stakeholders in qualitative interviews to design communication strategies; employ descriptive statistics and regression analyses to evaluate the site differences in genetic testing and the OVCA Traceback testing; and assess program outcomes at the proband, family member, provider, system, and population levels. This study aims to determine a Traceback program’s feasibility and acceptability in a real-world context. It will account for the myriad factors affecting implementation, including legal issues, organizational- and individual-level barriers and facilitators, communication issues, and program costs. Project results will inform how health care providers and systems can develop effective, practical, and sustainable Traceback programs.

Polygenic indexes (PGIs) are DNA-based predictors. Their value for research in many scientific disciplines is rapidly growing. As a resource for researchers, we used a consistent methodology to construct PGIs for 47 phenotypes in 11 datasets. To maximize the PGIs’ prediction accuracies, we constructed them using genome-wide association studies—some of which are novel—from multiple data sources, including 23andMe and UK Biobank. We present a theoretical framework to help interpret analyses involving PGIs. A key insight is that a PGI can be understood as an unbiased but noisy measure of a latent variable we call the “additive SNP factor.” Regressions in which the true regressor is the additive SNP factor but the PGI is used as its proxy therefore suffer from errors-in-variables bias. We derive an estimator that corrects for the bias, illustrate the correction, and make a Python tool for implementing it publicly available.

Rationale: We aim to describe the clinical characteristics and outcomes of patients who received Tocilizumab for COVID-19 pneumonia at our institution between March 20 and October 26, 2020. Methods: In this single center, retrospective, observational study, we identified 55 adults admitted with COVID-19 pneumonia who received Tocilizumab. Demographic data, symptoms, laboratory values, treatments, and clinical outcomes were collected. Data was compared between those who received Tocilizumab and all patients admitted with COVID-19. Primary outcome was 28-day mortality. Secondary outcomes included role of concomitant steroid use and change in eosinophil counts, ferritin, AST, CRP and D-Dimer values. Results: Of the 589 patients admitted with COVID-19 pneumonia, 55 received Tocilizumab as part of their treatment course. Patient demographics of those who received Tocilizumab include a mean age of 58 years with 73% male, 51% with diabetes, and 58% with hypertension. 4/55 (7.3%) were immunocompromised. Common presenting symptoms on admission were fever (62%), cough (78%) and dyspnea (89%). 35/55 (64%) were admitted to the ICU during their hospitalization;their mean P/F ratio was 127. Tocilizumab was administered on average admission day 4 (1-19). A second dose was given to 17 (31%) of patients, with 11 given the following day. Average hospital length of stay (LOS) postadministration was 17 days. Average white blood cell (WBC) count on day of Tocilizumab administration was 11, with an absolute lymphocyte count of 0.96. Mean IL-6 on hospital admission was 48.3. Two days post Tocilizumab administration there was a peak in ferritin, percent eosinophils, and AST. Both two-and five-day post-Tocilizumab CRP levels decreased while D-Dimer increased (Table 1). All Tocilizumab patients received antibiotics. In addition, three received hydroxychloroquine, 16 Remdesivir, and 51 convalescent plasma. 31 (56%) received steroids. On Day 2, those who did not receive steroids had, on average, more than double the percent of eosinophils in their blood (3.21% vs 1.53%). This difference decreased by Day 5. In time period of interest, COVID-19 admission mortality was 63/589 (10.6%) and 40/77 (52%) for mechanically ventilated patients. For Tocilizumab recipients, 25/55 patients were mechanically ventilated and 12/25 (48%) died. Overall, 28-day mortality was 11/55 (20%), with hospital mortality up to 16/55 (29%). This was similar to our larger cohort ICU mortality of 29.3%. Conclusion: Tocilizumab recipients in our cohort had a mortality similar to overall COVID ICU mortality. It appeared to be well tolerated except for an increase in eosinophilia if with no concomitant steroid use.

What is a good workday for a software developer? What is a typical workday? We seek to answer these two questions to learn how to make good days typical. Concretely, answering these questions will help to optimize development processes and select tools that increase job satisfaction and productivity. Our work adds to a large body of research on how software developers spend their time. We report the results from 5,971 responses of professional developers at Microsoft, who reflected about what made their workdays good and typical, and self-reported about how they spent their time on various activities at work. We developed conceptual frameworks to help define and characterize developer workdays from two new perspectives: good and typical. Our analysis confirms some findings in previous work, including the fact that developers actually spend little time on development and developers’ aversion for meetings and interruptions. It also discovered new findings, such as that only 1.7 percent of survey responses mentioned emails as a reason for a bad workday, and that meetings and interruptions are only unproductive during development phases; during phases of planning, specification and release, they are common and constructive. One key finding is the importance of agency, developers’ control over their workday and whether it goes as planned or is disrupted by external factors. We present actionable recommendations for researchers and managers to prioritize process and tool improvements that make good workdays typical. For instance, in light of our finding on the importance of agency, we recommend that, where possible, managers empower developers to choose their tools and tasks.

Many Americans fail to get life-saving vaccines each year, and the availability of a vaccine for COVID-19 makes the challenge of encouraging vaccination more urgent than ever. We present a large field experiment ( N = 47,306) testing 19 nudges delivered to patients via text message and designed to boost adoption of the influenza vaccine. Our findings suggest that text messages sent prior to a primary care visit can boost vaccination rates by an average of 5%. Overall, interventions performed better when they were 1) framed as reminders to get flu shots that were already reserved for the patient and 2) congruent with the sort of communications patients expected to receive from their healthcare provider (i.e., not surprising, casual, or interactive). The best-performing intervention in our study reminded patients twice to get their flu shot at their upcoming doctor’s appointment and indicated it was reserved for them. This successful script could be used as a template for campaigns to encourage the adoption of life-saving vaccines, including against COVID-19.

We present a new method, Multi-Ancestry Meta-Analysis (MAMA), which combines genome-wide association study (GWAS) summary statistics from multiple populations to produce new summary statistics for each population, identifying novel loci that would not have been discovered in either set of GWAS summary statistics alone. In simulations, MAMA increases power with less bias and generally lower type-1 error rate than other multi-ancestry meta-analysis approaches. We apply MAMA to 23 phenotypes in East-Asian- and European-ancestry populations and find substantial gains in power. In an independent sample, novel genetic discoveries from MAMA replicate strongly.

Metformin is found in the majority of lakes and streams in the United States, leading to widespread environmental exposure. Results of the present study indicate that extended duration metformin exposure at critical developmental periods leads to decreased survival rates in zebrafish (danio rerio), an NIH approved human model. Significant abnormalities are seen with extended duration metformin exposure from 4 h post fertilization up to 5 days post fertilization, although short term metformin exposure for 24 h at 4–5 days post fertilization did not lead to any significant abnormalities. Both extended and short term duration did however have an impact on locomotor activity of zebrafish, and several genes involved in neurological and cardiovascular development were differentially expressed after exposure to metformin. The changes seen in behavior, gene expression and morphological abnormalities caused by metformin exposure should be examined further in future studies in order to assess their potential human health implications as metformin prescriptions continue to increase worldwide.

Objective Hydroxychloroquine has been proposed as a primary prophylactic agent against coronavirus disease 2019 (COVID-19). This study aimed to investigate if patients treated with hydroxychloroquine for a non-COVID-19 indication had a lower risk of verified infection with severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) compared with matched controls. Methods A cohort comprising all persons in Denmark collecting hydroxychloroquine prescriptions in 2020 and 2019 (i.e., both during and before SARS-CoV-2 was confirmed in Denmark), matched by age and sex with controls, was studied. Data were collected using the Danish national registries, which contain complete information on patient health data, prescriptions and microbiological test results. The main outcome was microbiologically verified SARS-CoV-2 infection. Results In total, 5488 hydroxychloroquine users were matched with 54,486 non-users. At baseline, the groups differed in terms of diagnoses of pulmonary disease, cardiovascular disease, renal disease, gastrointestinal/metabolic disease and dementia, as well as treatment with antirheumatic drugs. The final model was adjusted for these potential confounders. Use of hydroxychloroquine for non-COVID-19 indications was not associated with any change in confirmed SARS-CoV-2 (hazard ratio 0.90, 95% confidence interval 0.76–1.07). This result was robust in the propensity-score-matched sensitivity analysis. Conclusion This study, which is the largest to date to investigate the primary prophylactic effect of hydroxychloroquine against SARS-CoV-2, does not support any prophylactic benefit of hydroxychloroquine in the prevention of infection with SARS-CoV-2.

For many vaccine-preventable diseases like influenza, vaccination rates are lower than optimal to achieve community protection. Those at high risk for infection and serious complications are especially advised to be vaccinated to protect themselves. Using influenza as a model, we studied one method of increasing vaccine uptake: informing high-risk patients, identified by a machine learning model, about their risk status. Patients (N=39,717) were evenly randomized to (1) a control condition (exposure only to standard direct mail or patient portal vaccine promotion efforts) or to be told via direct mail, patient portal, and/or SMS that they were (2) at high risk for influenza and its complications if not vaccinated; (3) at high risk according to a review of their medical records; or (4) at high risk according to a computer algorithm analysis of their medical records. Patients in the three treatment conditions were 5.7% more likely to get vaccinated during the 112 days post-intervention (p < .001), and did so 1.4 days earlier (p < .001), on average, than those in the control group. There were no significant differences among risk messages, suggesting that patients are neither especially averse to nor uniquely appreciative of learning their records had been reviewed or that computer algorithms were involved. Similar approaches should be considered for COVID-19 vaccination campaigns.

This survey study queried employees of a health care system before COVID-19 vaccine distribution to assess their intentions to receive a vaccine and to understand their reasons for hesitancy to do so.

Caenorhabditis elegans has emerged as a major model in biomedical and environmental toxicology. Numerous papers on toxicology and pharmacology in C. elegans have been published, and this model has now been adopted by investigators in academic toxicology, pharmacology, and drug discovery labs. C. elegans has also attracted the interest of governmental regulatory agencies charged with evaluating the safety of chemicals. However, a major, fundamental aspect of toxicological science remains underdeveloped in C. elegans: xenobiotic metabolism and transport processes that are critical to understanding toxicokinetics and toxicodynamics, and extrapolating to other species. The aim of this review was to initially briefly describe the history and trajectory of the use of C. elegans in toxicological and pharmacological studies. Subsequently, physical barriers to chemical uptake and role of the worm microbiome in xenobiotic transformation were described. Then a review on what is and is not known regarding the classic Phase I, Phase II, and Phase III processes was examined. In addition, the following were discussed (1) regulation of xenobiotic metabolism; (2) review of published toxicokinetics for specific chemicals; and (3) genetic diversity of these processes in C. elegans. Finally, worm xenobiotic transport and metabolism was placed in an evolutionary context; key areas for future research highlighted; and implications for extrapolating C. elegans toxicity results to other species discussed.

Background. There has been a steady increase in the use of electronic media and instant messaging among healthcare professionals, where it has been almost universally adopted in the workplace. The use of WhatsApp and its perceived benefits in healthcare have been extensively studied; however, there are concerns regarding the potential for ethical breaches in confidentiality through shared electronic patient information. Objectives. To identify the usage characteristics and incidence of shared patient information with WhatsApp use in a team of medical doctors in an unobserved and unregulated setting. Methods. We conducted a retrospective cross-sectional content analysis of WhatsApp messages ( n =3 340) among a team of 20 doctors in a South African district hospital over 6 months. All messages found within this time period were allocated unique identifiers. The text and image messages were thematically grouped into four categories, i.e. clinical care, resource allocation, social and administrative. Messages that contained patient-identifying information were included in the analysis. Results. Of a total of 3 340 messages sent, 220 (6.6%) contained patient-identifying information. Of these, 109 (3.3%) contained non-anonymised patient information, while in 111 (3.3%) messages, the information was anonymised. The likelihood of sharing patient identifiers was proportionally much higher in shared images (odds ratio (OR) 5.1; 95% confidence interval (CI) 3.2 - 8.2; p

The neural processes that support inhibitory control in the face of stimuli with a history of reward association are not yet well understood. Yet, the ability to flexibly adapt behavior to changing reward-contingency contexts is important for daily functioning and warrants further investigation. This study aimed to characterize neural and behavioral impacts of stimuli with a history of conditioned reward association on motor inhibitory control in healthy young adults by investigating group-level effects as well as individual variation in the ability to inhibit responses to stimuli with a reward history. Participants (N = 41) first completed a reward conditioning phase, during which responses to rewarded stimuli were associated with money and responses to unrewarded stimuli were not. Rewarded and unrewarded stimuli from training were carried forward as No-Go targets in a subsequent go/no-go task to test the effect of reward history on inhibitory control. Participants underwent functional brain imaging during the go/no-go portion of the task. On average, a history of reward conditioning disrupted inhibitory control. Compared to inhibition of responses to stimuli with no reward history, trials that required inhibition of responses to previously rewarded stimuli were associated with greater activity in frontal and striatal regions, including the inferior frontal gyrus, insula, striatum, and thalamus. Activity in the insula and thalamus during false alarms and in the ventromedial prefrontal cortex during correctly withheld trials predicted behavioral performance on the task. Overall, these results suggest that reward history serves to disrupt inhibitory control and provide evidence for diverging roles of the insula and ventromedial prefrontal cortex while inhibiting responses to stimuli with a reward history.

Introduction: Loneliness and low social support can be detrimental to the health of individuals living with Alzheimer's and related dementias (ADRD) and family care partners. Restrictions on gatherings to prevent the spread of COVID-19 create an even greater risk for social isolation. Memory Cafés are a highly replicated program that provide individuals living with ADRD and care partners an opportunity to socialize in an inclusive and supportive environment without fear of judgment, pressure, or stigma. Following restrictions on in-person gatherings, virtual Memory Cafés offer regular social engagement opportunities in an online format. While the Memory Café model has been replicated globally, their effects on loneliness and perceived social support are generally unknown. Even less is known about their impact when operating in a virtual environment.Methods: Semi-structured interviews in Spanish and English were conducted with individuals living with dementia and family care partners who regularly attend Memory Cafés hosted by partners in a Texas Memory Café Network. Interviews took place online using video conferencing software, were transcribed, then analyzed for common themes using a combined inductive and deductive approach.Results: A total of 17 interviews were conducted with persons living with dementia (n = 5) and family care partners (n = 12) who attend Memory Cafés to learn about their perceived experiences of social connectedness since COVID-19. Care partners included spouses (n = 8) and adult children (n = 4). Interviews included attendees of different Memory Café models, including in-person only (n = 2), virtual only (n = 9), and those who attend both models (n = 6). Five key themes were identified: (1) Reprieve; (2) What is still possible; (3) Connectedness; (4) Inclusivity; and (5) Value added, with ten sub-themes supporting these main themes.Discussion: Findings substantiate evidence that Memory Cafés offer important benefits for families living with dementia, providing vital new insight into the potential for virtual Memory Cafés to offer similar benefits. Findings have implications beyond the context of COVID-19, where virtual models may support the social connectedness of those living in geographically marginalized and underserved areas. Virtual models may not address the needs of all families experiencing dementia due to lack of access to technology and limitations for virtual engagement with those experiencing later stage dementia.

Impacts of dust grains on spacecraft are known to produce typical impulsive signals in the voltage waveform recorded at the terminals of electric antennas. Such signals are routinely detected by the Time Domain Sampler (TDS) system of the Radio and Plasma Waves (RPW) instrument aboard Solar Orbiter. We investigate the capabilities of RPW in terms of interplanetary dust studies and present the first analysis of dust impacts recorded by this instrument. We discuss previously developed models of voltage pulses generation after a dust impact onto a spacecraft and present the relevant technical parameters for Solar Orbiter RPW as a dust detector. Then we present the statistical analysis of the dust impacts recorded by RPW/TDS from April 20th, 2020 to February 27th, 2021 between 0.5 AU and 1 AU. The study shows that the dust population studied presents a radial velocity component directed outward from the Sun, the order of magnitude of which can be roughly estimated as $v_{r, dust} \simeq 50$ km.$s^{-1}$. This is consistent with the flux of impactors being dominated by $\beta$-meteoroids. We estimate the cumulative flux of these grains at 1 AU to be roughly $F_\beta \simeq 8\times 10^{-5} $ m$^{-2}$s$^{-1}$, for particles of radius $r \gtrsim 100$ nm. The power law index $\delta$ of the cumulative mass flux of the impactors is evaluated by two differents methods (direct observations of voltage pulses and indirect effect on the impact rate dependency on the impact speed). Both methods give a result $\delta \simeq 0.3-0.4$. Solar Orbiter RPW proves to be a suitable instrument for interplanetary dust studies. These first results are promising for the continuation of the mission, in particular for the in-situ study of the dust cloud outside the ecliptic plane, which Solar Orbiter will be the first spacecraft to explore., Comment: 13 pages, 8 figures