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Intestinal absorption is an important contributor to systemic cholesterol homeostasis. Niemann-Pick C1 Like 1 (NPC1L1) assists in the initial step of dietary cholesterol uptake, but how cholesterol moves downstream of NPC1L1 is unknown. We show that Aster-B and Aster-C are critical for nonvesicular cholesterol movement in enterocytes. Loss of NPC1L1 diminishes accessible plasma membrane (PM) cholesterol and abolishes Aster recruitment to the intestinal brush border. Enterocytes lacking Asters accumulate PM cholesterol and show endoplasmic reticulum cholesterol depletion. Aster-deficient mice have impaired cholesterol absorption and are protected against diet-induced hypercholesterolemia. Finally, the Aster pathway can be targeted with a small-molecule inhibitor to manipulate cholesterol uptake. These findings identify the Aster pathway as a physiologically important and pharmacologically tractable node in dietary lipid absorption.

Introduction: Urinary tract infections (UTI) are a common reason for an emergency department (ED) visit. The majority of these patients are discharged directly home without a hospital admission.  After discharge, emergency physicians have traditionally managed the care of the patient if a change is warranted (as a result of urine culture results). However, in recent years clinical pharmacists in the ED have largely incorporated this task into their standard practice. In our study, we aimed to 1) describe our unique process in having a pharmacist-led, urinary culture follow-up, and 2) compare it to our previous, more traditional process.Methods: In our retrospective study, we evaluated the impact of a pharmacist-led, urinary culture follow-up program after discharge from the ED. We included patients prior to and after the implementation of our new protocol to compare the differences. The primary outcome was time to intervention after urine culture result was released. Secondary outcomes included rate of documentation of intervention, appropriate interventions made, and repeat ED visits within 30 days.Results: We included a total of 265 unique urine cultures from 264 patients in the study: 129 cultures were from the period prior to implementation of the protocol, and 136 were from the post-implementation period. There were no significant differences between pre- and post-implementation groups for the primary outcome. Appropriate therapeutic intervention based on positive urine culture results was 16.3% in the pre-implementation group vs 14.7% in the post-implementation group (P=0.72). Secondary outcomes of time to intervention, documentation rates, and readmissions were similar between both groups.Conclusion: Implementation of a pharmacist-led, urinary culture follow-up program after discharge from the ED led to similar outcomes as a physician-run program.  An ED pharmacist can successfully run a urinary culture follow-up program in an ED without physician involvement.

Purpose: Dyslexia is characterized by its diverse causes and heterogeneous manifestations. Chinese children with dyslexia exhibit orthographic, phonological, and semantic deficits across character and radical levels when writing. However, whether character dictation can be used to distinguish children with dyslexia from their typically developing peers remains unexplored. Method: A dataset of written characters from 1,015 Chinese children with and without dyslexia from Grades 2-6 was used to train multiple machine models with different learning algorithms. Results: The multi-level multidimensional model reached a predictive accuracy of 78.0%, with stroke, grade, lexicality, and character configuration manifesting as the most predictive features. The accuracy of the model improved to 80.0% when only these features were included. Conclusion: These results not only provide evidence for the multidimensional causes of Chinese dyslexia, but also highlight the utility of machine learning in distinguishing children with dyslexia from their peers via Chinese dictation, which elucidates a promising area of future research.

Background: Face-to-face class suspensions during the coronavirus disease (COVID-19) pandemic in 2019 increased telepractice in speech and language therapy. However, little is known about speech and language therapists' (SLTs) perceived effectiveness of telepractice and its antecedents. Aims: To examine the use of telepractice and the factors affecting its perceived effectiveness in Hong Kong mainstream schools during COVID-19. Methods & Procedures: Seventy-two school-based Hong Kong SLTs completed a 110-item online survey with six structural components: (1) concerns, (2) adoption, (3) student selection criteria, (4) perceived effectiveness, (5) continuous professional development and (6) attitudes/beliefs. Outcomes & Results: Over 90% of respondents adopted telepractice during the pandemic. Confirmatory factor analysis identified reliable constructs from their component measures. These participants reported great telepractice difficulties (especially in therapy preparation and managing students' attention and/or communication). Mixed-response analysis revealed that psychosocial factors (i.e., students' engagement and SLTs' attitudes towards telepractice) but not professional practice skill factors (i.e., student client selection criteria and SLTs' years of experience in school settings) contributed to SLTs' self-perceived effectiveness of telepractice. Conclusions & Implications: Our findings suggest that psychosocial factors play a more important role than professional practice skill factors in the self-perceived effectiveness of telepractice.

Regulation of hepatocyte proliferation and liver morphology is of critical importance to tissue and whole-body homeostasis. However, the molecular mechanisms that underlie this complex process are incompletely understood. Here, we describe a role for the ubiquitin ligase BRCA1-associated protein (BRAP) in regulation of hepatocyte morphology and turnover via regulation of MST2, a protein kinase in the Hippo pathway. The Hippo pathway has been implicated in the control of liver morphology, inflammation, and fibrosis. We demonstrate here that liver-specific ablation of Brap in mice results in gross and cellular morphological alterations of the liver. Brap-deficient livers exhibit increased hepatocyte proliferation, cell death, and inflammation. We show that loss of BRAP protein alters Hippo pathway signaling, causing a reduction in phosphorylation of YAP and increased expression of YAP target genes, including those regulating cell growth and interactions with the extracellular environment. Finally, increased Hippo signaling in Brap knockout mice alters the pattern of liver lipid accumulation in dietary models of obesity. These studies identify a role for BRAP as a modulator of the hepatic Hippo pathway with relevance to human liver disease.

Neurons of the peripheral nervous system (PNS) are tasked with diverse roles, from encoding touch, pain, and itch to interoceptive control of inflammation and organ physiology. Thus, technologies that allow precise control of peripheral nerve activity have the potential to regulate a wide range of biological processes. Noninvasive modulation of neuronal activity is an important translational application of focused ultrasound (FUS). Recent studies have identified effective strategies to modulate brain circuits; however, reliable parameters to control the activity of the PNS are lacking. To develop robust noninvasive technologies for peripheral nerve modulation, we employed targeted FUS stimulation and electrophysiology in mouse ex vivo skin-saphenous nerve preparations to record the activity of individual mechanosensory neurons. Parameter space exploration showed that stimulating neuronal receptive fields with high-intensity, millisecond FUS pulses reliably and repeatedly evoked one-to-one action potentials in all peripheral neurons recorded. Interestingly, when neurons were classified based on neurophysiological properties, we identified a discrete range of FUS parameters capable of exciting all neuronal classes, including myelinated A fibers and unmyelinated C fibers. Peripheral neurons were excited by FUS stimulation targeted to either cutaneous receptive fields or peripheral nerves, a key finding that increases the therapeutic range of FUS-based peripheral neuromodulation. FUS elicited action potentials with millisecond latencies compared with electrical stimulation, suggesting ion channel–mediated mechanisms. Indeed, FUS thresholds were elevated in neurons lacking the mechanically gated channel PIEZO2. Together, these results demonstrate that transcutaneous FUS drives peripheral nerve activity by engaging intrinsic mechanotransduction mechanisms in neurons [B. U. Hoffman, PhD thesis, (2019)].

A compelling demonstration of implicit learning is the human ability to unconsciously detect and internalize statistical patterns of complex environmental input. This ability, called statistical learning, has been investigated in people with dyslexia using various tasks in different orthographies. However, conclusions regarding impaired or intact statistical learning in dyslexia remain mixed. This study conducted a systematic literature search of published and unpublished studies that compared statistical learning between people with and without dyslexia using different learning paradigms in different orthographies. We identified 49 papers consisting of 59 empirical studies, representing the data from 1,259 participants with dyslexia and 1,459 typically developing controls. The results showed that, on average, individuals with dyslexia performed worse in statistical learning than age-matched controls, regardless of the learning paradigm or orthography (average weighted effect size d = 0.47, 95% confidence interval [0.36, 0.59], p < 0.001). Meta-regression analyses further revealed that the heterogeneity of effect sizes between studies was significantly explained by one reader characteristic (i.e., verbal IQ) but no task characteristics (i.e., task paradigm, task modality, and stimulus type). These findings suggest domain-general statistical learning weakness in dyslexia across languages, and support the need for a new theoretical model of statistical learning and reading, that is, the SLR model, which elucidates how reader and task characteristics are regulated by a multicomponent memory system when establishing statistically optimal representations for deep learning and reading.

We recently showed that NOTUM, a liver-secreted Wnt inhibitor, can acutely promote browning of white adipose. We now report studies of chronic overexpression of NOTUM in liver indicating that it protects against diet-induced obesity and improves glucose homeostasis in mice. Adeno-associated virus (AAV) vectors were used to overexpress GFP or mouse Notum in the livers of male C57BL/6J mice and the mice were fed an obesifying diet. After 14 weeks of high fat, high sucrose diet feeding, the AAV-Notum mice exhibited decreased obesity and improved glucose tolerance compared to the AAV-GFP mice. Gene expression and immunoblotting analysis of the inguinal fat and brown fat revealed increased expression of beige/brown adipocyte markers in the AAV-Notum group, suggesting enhanced thermogenic capacity by NOTUM. A β3 adrenergic receptor agonist-stimulated lipolysis test suggested increased lipolysis capacity by NOTUM. The levels of collagen and C-C motif chemokine ligand 2 (CCL2) in the epididymal white adipose tissue of the AAV-Notum mice were significantly reduced, suggesting decreased fibrosis and inflammation, respectively. RNA sequencing analysis of inguinal white adipose of 4-week chow diet-fed mice revealed a highly significant enrichment of extracellular matrix (ECM) functional cluster among the down-regulated genes in the AAV-Notum group, suggesting a potential mechanism contributing to improved glucose homeostasis. Our in vitro studies demonstrated that recombinant human NOTUM protein blocked the inhibitory effects of WNT3A on brown adipocyte differentiation. Furthermore, NOTUM attenuated WNT3A's effects on upregulation of TGF-β signaling and its downstream targets. Overall, our data suggest that NOTUM modulates adipose tissue function by promoting thermogenic capacity and inhibiting fibrosis through inhibition of Wnt signaling.

Introduction: During the coronavirus disease 2019 (COVID-19) pandemic, a reduction in emergency department (ED) visits was seen nationally according to the US Centers for Disease Control and Prevention. However, no data currently exists for the impact of ED transfers to a higher level of care during this same time period. The primary objective of the study was to determine whether the COVID-19 pandemic affected the rate of non-COVID-19 transfers from a rural community ED.Methods: We completed a retrospective chart review of all ED patients who presented to Kingman Regional Medical Center in Kingman, Arizona, from March 1–June 31, 2019 and March 1–June 31, 2020. To ensure changes were not due to seasonal trends, we examined transfer rates from the same four-month period in 2019 and 2020. Patients were included in the study if they were transferred to an outside facility for a higher level of care not related to COVID-19.Results: Between the time periods studied there was a 25.33% (P = 0.001) reduction in total ED volume and a 21.44% (P = 0.009) reduction in ED transfers to a higher level of care. No statistical difference was noted in ED transfer volume following adjustment for decreased ED volumes. Transfers for gastroenterology (45%; P = 0.021), neurosurgery (29.2%; P = 0.029), neurology (76.3%; P < 0.001), trauma (37.5%; P = 0.039), urology (41.8%; P = 0.012), and surgery (56.3%; P = 0.028) all experienced a decrease in transfer rates during the time period studied. When gender was considered, males exhibited an increased rate of transfers to psychiatric facilities (P = 0.018).Conclusion: Significant reductions in both ED volume and transfers have coincided with the emergence of the COVID-19 pandemic. Further research is needed to determine how the current pandemic has affected patient care.

Soft microfluidic systems that capture, store, and perform biomarker analysis of microliter volumes of sweat, in situ, as it emerges from the surface of the skin, represent an emerging class of wearable technology with powerful capabilities that complement those of traditional biophysical sensing devices. Recent work establishes applications in the real-time characterization of sweat dynamics and sweat chemistry in the context of sports performance and healthcare diagnostics. This paper presents a collection of advances in biochemical sensors and microfluidic designs that support multimodal operation in the monitoring of physiological signatures directly correlated to physical and mental stresses. These wireless, battery-free, skin-interfaced devices combine lateral flow immunoassays for cortisol, fluorometric assays for glucose and ascorbic acid (vitamin C), and digital tracking of skin galvanic responses. Systematic benchtop evaluations and field studies on human subjects highlight the key features of this platform for the continuous, noninvasive monitoring of biochemical and biophysical correlates of the stress state.

Liver X receptors limit cellular lipid uptake by stimulating the transcription of Inducible Degrader of the LDL Receptor (IDOL), an E3 ubiquitin ligase that targets lipoprotein receptors for degradation. The function of IDOL in systemic metabolism is incompletely understood. Here we show that loss of IDOL in mice protects against the development of diet-induced obesity and metabolic dysfunction by altering food intake and thermogenesis. Unexpectedly, analysis of tissue-specific knockout mice revealed that IDOL affects energy balance, not through its actions in peripheral metabolic tissues (liver, adipose, endothelium, intestine, skeletal muscle), but by controlling lipoprotein receptor abundance in neurons. Single-cell RNA sequencing of the hypothalamus demonstrated that IDOL deletion altered gene expression linked to control of metabolism. Finally, we identify VLDLR rather than LDLR as the primary mediator of IDOL effects on energy balance. These studies identify a role for the neuronal IDOL-VLDLR pathway in metabolic homeostasis and diet-induced obesity.

Immune cells are vital constituents of the adipose microenvironment that influence both local and systemic lipid metabolism. Mice lacking IL10 have enhanced thermogenesis, but the roles of specific cell types in the metabolic response to IL10 remain to be defined. We demonstrate here that selective loss of IL10 receptor α in adipocytes recapitulates the beneficial effects of global IL10 deletion, and that local crosstalk between IL10-producing immune cells and adipocytes is a determinant of thermogenesis and systemic energy balance. Single Nuclei Adipocyte RNA-sequencing (SNAP-seq) of subcutaneous adipose tissue defined a metabolically-active mature adipocyte subtype characterized by robust expression of genes involved in thermogenesis whose transcriptome was selectively responsive to IL10Rα deletion. Furthermore, single-cell transcriptomic analysis of adipose stromal populations identified lymphocytes as a key source of IL10 production in response to thermogenic stimuli. These findings implicate adaptive immune cell-adipocyte communication in the maintenance of adipose subtype identity and function.

The mechanisms underlying sterol transport in mammalian cells are poorly understood. In particular, how cholesterol internalized from HDL is made available to the cell for storage or modification is unknown. Here, we describe three ER-resident proteins (Aster-A, -B, -C) that bind cholesterol and facilitate its removal from the plasma membrane. The crystal structure of the central domain of Aster-A broadly resembles the sterol-binding fold of mammalian StARD proteins, but sequence differences in the Aster pocket result in a distinct mode of ligand binding. The Aster N-terminal GRAM domain binds phosphatidylserine and mediates Aster recruitment to plasma membrane-ER contact sites in response to cholesterol accumulation in the plasma membrane. Mice lacking Aster-B are deficient in adrenal cholesterol ester storage and steroidogenesis because of an inability to transport cholesterol from SR-BI to the ER. These findings identify a nonvesicular pathway for plasma membrane to ER sterol trafficking in mammals.

In the United States, the lives of undocumented people have become increasingly precarious due to increased surveillance, enforcement, criminalization, and detention. In this context, deferred action, a form of prosecutorial discretion in which the government declines to pursue removal and provides temporary work authorization, has become a source of both hope and vulnerability. Based on fieldwork, interviews, and legal analysis, we delineate the forms of partial inclusion experienced by deferred action recipients and explore the position from which they can make claims on the US state. Our analysis advances citizenship theory by detailing the relationship between the discretionary state and its transitory, noncitizen subjects, as well as how this relationship is complicated by resistance from youth activists and their allies. The liminal legality afforded by deferred action provides partial but insecure relief from the precarity experienced by the undocumented.

BackgroundRecognizing the factors associated with falling in Parkinson's disease (PD) would improve identification of at-risk individuals.ObjectiveTo examine frequency of falling and baseline characteristics associated with falling in PD using the National Institute of Neurological Disorders and Stroke (NINDS) Exploratory Trials in PD Long-term Study-1 (NET-PD LS-1) dataset.MethodsThe LS-1 database included 1741 early treated PD subjects (median 4year follow-up). Baseline characteristics were tested for a univariate association with post-baseline falling during the trial. Significant variables were included in a multivariable logistic regression model. A separate analysis using a negative binomial model investigated baseline factors on fall rate.Results728 subjects (42%) fell during the trial, including at baseline. A baseline history of falls was the factor most associated with post-baseline falling. Men had lower odds of post-baseline falling compared to women, but for men, the probability of a post-baseline fall increased with age such that after age 70, men and women had similar odds of falling. Other baseline factors associated with a post-baseline fall and increased fall rate included the Unified PD Rating Scale (UPDRS) Activities of Daily Living (ADL) score, total functional capacity (TFC), baseline ambulatory capacity score and dopamine agonist monotherapy.ConclusionFalls are common in early treated PD. The biggest risk factor for falls in PD remains a history of falling. Measures of functional ability (UPDRS ADL, TFC) and ambulatory capacity are novel clinical risk factors needing further study. A significant age by sex interaction may help to explain why age has been an inconsistent risk factor for falls in PD.

C/EBPε is a critical transcriptional factor for granulocyte differentiation and function. Individuals with germline mutations of C/EBPε fail to develop normal granulocytes and suffer from repeated infections. In order to gain a global view of the transcriptional machinery regulated by C/EBPε, we performed whole-genome ChIP-Seq using mouse bone marrow cells. To complement the C/EBPε DNA binding analyses, RNA-Sequencing was done in parallel using sorted mature and immature granulocytes from WT and C/EBPε KO bone marrow. This approach led to the identification of several direct targets of C/EBPε, which are potential effectors of its role in granulocytic differentiation and function. Interestingly, Trem1, a gene critical to granulocyte function, was identified as a direct C/EBPε target gene. Trem1 expression overlaps very closely with expression signature of C/EBPε during hematopoietic development. Luciferase reporter and EMSA assays revealed that C/EBPε binds to the regulatory elements of Trem1 and regulates its expression during granulocytic differentiation. In addition, we provide evidence that inflammatory stimuli (LPS) can also control the expression of Trem1 independent of C/EBPε. Overall, this study provides comprehensive profiling of the transcriptional network controlled by C/EBPε during granulopoiesis and identifies Trem1 as one of its downstream effectors involved in eliciting an immune response.

A highly orchestrated gene expression program establishes the properties that define mature adipocytes, but the contribution of posttranscriptional factors to the adipocyte phenotype is poorly understood. Here we have shown that the RNA-binding protein PSPC1, a component of the paraspeckle complex, promotes adipogenesis in vitro and is important for mature adipocyte function in vivo. Cross-linking and immunoprecipitation followed by RNA sequencing revealed that PSPC1 binds to intronic and 3'-untranslated regions of a number of adipocyte RNAs, including the RNA encoding the transcriptional regulator EBF1. Purification of the paraspeckle complex from adipocytes further showed that PSPC1 associates with the RNA export factor DDX3X in a differentiation-dependent manner. Remarkably, PSPC1 relocates from the nucleus to the cytoplasm during differentiation, coinciding with enhanced export of adipogenic RNAs. Mice lacking PSPC1 in fat displayed reduced lipid storage and adipose tissue mass and were resistant to diet-induced obesity and insulin resistance due to a compensatory increase in energy expenditure. These findings highlight a role for PSPC1-dependent RNA maturation in the posttranscriptional control of adipose development and function.

Although much progress has been made in identifying the mechanisms that trigger endothelial activation and inflammatory cell recruitment during atherosclerosis, less is known about the intrinsic pathways that counteract these events. Here we identified NOTCH1 as an antagonist of endothelial cell (EC) activation. NOTCH1 was constitutively expressed by adult arterial endothelium, but levels were significantly reduced by high-fat diet. Furthermore, treatment of human aortic ECs (HAECs) with inflammatory lipids (oxidized 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphocholine [Ox-PAPC]) and proinflammatory cytokines (TNF and IL1β) decreased Notch1 expression and signaling in vitro through a mechanism that requires STAT3 activation. Reduction of NOTCH1 in HAECs by siRNA, in the absence of inflammatory lipids or cytokines, increased inflammatory molecules and binding of monocytes. Conversely, some of the effects mediated by Ox-PAPC were reversed by increased NOTCH1 signaling, suggesting a link between lipid-mediated inflammation and Notch1. Interestingly, reduction of NOTCH1 by Ox-PAPC in HAECs was associated with a genetic variant previously correlated to high-density lipoprotein in a human genome-wide association study. Finally, endothelial Notch1 heterozygous mice showed higher diet-induced atherosclerosis. Based on these findings, we propose that reduction of endothelial NOTCH1 is a predisposing factor in the onset of vascular inflammation and initiation of atherosclerosis.

Abstract Developing central synapses exhibit robust plasticity and undergo experience-dependent remodeling. Evidently, synapses in sensory systems such as auditory brainstem circuits mature rapidly to achieve high-fidelity neurotransmission for sound localization. This depends on a developmental switch in AMPAR composition from slow-gating GluA1-dominant to fast-gating GluA4-dominant, but the mechanisms underlying this switch remain unknown. We hypothesize that patterned stimuli mimicking spontaneous/sound evoked activity in the early postnatal stage drives this gating switch. We examined activity-dependent changes in evoked and miniature excitatory postsynaptic currents (eEPSCs and mEPSCs) at the calyx of Held synapse by breaking through the postsynaptic membrane at different time points following 2 min of theta burst stimulation (TBS) to afferents in mouse brainstem slices. We found the decay time course of eEPSCs accelerated, but this change was not apparent until > 30 min after TBS. Histogram analyses of the decay time constants of mEPSCs for naive and tetanized synapses revealed two populations centered around τfast ≈ 0.4 and 0.8 ms, but the relative weight of the τ0.4 population over the τ0.8 population increased significantly only in tetanized synapses. Such changes are blocked by NMDAR or mGluR1/5 antagonists or inhibitors of CaMKII, PKC and protein synthesis, and more importantly precluded in GluA4−/− synapses, suggesting GluA4 is the substrate underlying the acceleration. Our results demonstrate a novel form of plasticity working through NMDAR and mGluR activation to trigger a gating switch of AMPARs with a temporally delayed onset of expression, ultimately enhancing the development of high-fidelity synaptic transmission.

RationaleThe E3 ubiquitin ligase inducible degrader of the low-density lipoprotein receptor (IDOL) triggers lysosomal degradation of the low-density lipoprotein receptor. The tissue-specific effects of the IDOL pathway on plasma cholesterol and atherosclerosis have not been examined.ObjectiveGiven that the liver is the primary determinant of plasma cholesterol levels, we sought to examine the consequence of effect of chronic liver-specific expression of a dominant-active form of IDOL in mice.Methods and resultsWe expressed a degradation-resistant, dominant-active form of IDOL (super IDOL [sIDOL]) in C57Bl/6J mice from the liver-specific albumin promoter (L-sIDOL transgenics). L-sIDOL mice were fed a Western diet for 20 or 30 weeks and then analyzed for plasma lipid levels and atherosclerotic lesion formation. L-sIDOL mice showed dramatic reductions in hepatic low-density lipoprotein receptor protein and increased plasma low-density lipoprotein cholesterol levels on both chow and Western diets. Moreover, L-sIDOL mice developed marked atherosclerotic lesions when fed a Western diet. Lesion formation in L-sIDOL mice was more robust than in apolipoprotein E*3 Leiden mice and did not require the addition of cholate to the diet. Western diet-fed L-sIDOL mice had elevated expression of liver X receptor target genes and proinflammatory genes in their aortas.ConclusionsLiver-specific expression of dominant-active IDOL is associated with hypercholesterolemia and a marked elevation in atherosclerotic lesions. Our results show that increased activity of the IDOL pathway in the liver can override other low-density lipoprotein receptor regulatory pathways leading to cardiovascular disease. L-sIDOL mice are a robust, dominantly inherited, diet-inducible model for the study of atherosclerosis.