A mechanism for oxidative damage repair at gene regulatory elements.

Oxidative genome damage is an unavoidable consequence of cellular metabolism. It arises at gene regulatory elements by epigenetic demethylation during transcriptional activation1,2. Here we show that promoters are protected from oxidative damage via a process mediated by the nuclear mitotic apparatus protein NuMA (also known as NUMA1). NuMA exhibits genomic occupancy approximately 100 bp around transcription start sites. It binds the initiating form of RNA polymerase II, pause-release factors and single-strand break repair (SSBR) components such as TDP1. The binding is increased on chromatin following oxidative damage, and TDP1 enrichment at damaged chromatin is facilitated by NuMA. Depletion of NuMA increases oxidative damage at promoters. NuMA promotes transcription by limiting the polyADP-ribosylation of RNA polymerase II, increasing its availability and release from pausing at promoters. Metabolic labelling of nascent RNA identifies genes that depend on NuMA for transcription including immediate-early response genes. Complementation of NuMA-deficient cells with a mutant that mediates binding to SSBR, or a mitotic separation-of-function mutant, restores SSBR defects. These findings underscore the importance of oxidative DNA damage repair at gene regulatory elements and describe a process that fulfils this function.

Evaluating Transition Readiness: Is Self-Report Valid as the Gold Standard?

OBJECTIVE: Determination of transitional readiness is an essential component of delivery of transition services to adolescents and young adults with chronic diseases, including those with inflammatory bowel disease (IBD). To date, this has been performed using the transition checklist (TC). However, clinical experience suggests the validity of the checklist may not be ideal. We sought to evaluate the validity of the TC among adolescents and young adults with IBD (AYA IBD). METHODS: We evaluated the validity of the TC compared with a skills-based practicum (TKSP) in a cross-sectional study using a real-world clinical cohort of AYA IBD. Extent of concordance between TC and TKSP was assessed using confusion matrices and calculation of Matthews correlation coefficients. RESULTS: Concordance between self-reported TC answers and TKSP performance varied based on skill/knowledge base assessed, ranging from 81.3% for mastery of knowing one's medical condition to 39.2% for mastery of knowing one's medication schedule and 29.3% for mastery of knowing how to refill medications. Matthews correlation coefficients were closer to random prediction than perfect correlation for all skills/knowledge tested. CONCLUSIONS: Self-reported TC answers lack sufficient concordance with TKSP performance. Our findings suggest that a TKSP should be performed to assess for transition readiness and to identify individual AYA IBD patient needs during the transition process.

The role of the inferior parietal lobule in writer's cramp.

Humans have a distinguishing ability for fine motor control that is subserved by a highly evolved cortico-motor neuronal network. The acquisition of a particular motor skill involves a long series of practice movements, trial and error, adjustment and refinement. At the cortical level, this acquisition begins in the parieto-temporal sensory regions and is subsequently consolidated and stratified in the premotor-motor cortex. Task-specific dystonia can be viewed as a corruption or loss of motor control confined to a single motor skill. Using a multimodal experimental approach combining neuroimaging and non-invasive brain stimulation, we explored interactions between the principal nodes of the fine motor control network in patients with writer's cramp and healthy matched controls. Patients and healthy volunteers underwent clinical assessment, diffusion-weighted MRI for tractography, and functional MRI during a finger tapping task. Activation maps from the task-functional MRI scans were used for target selection and neuro-navigation of the transcranial magnetic stimulation. Single- and double-pulse TMS evaluation included measurement of the input-output recruitment curve, cortical silent period, and amplitude of the motor evoked potentials conditioned by cortico-cortical interactions between premotor ventral (PMv)-motor cortex (M1), anterior inferior parietal lobule (aIPL)-M1, and dorsal inferior parietal lobule (dIPL)-M1 before and after inducing a long term depression-like plastic change to dIPL node with continuous theta-burst transcranial magnetic stimulation in a randomized, sham-controlled design. Baseline dIPL-M1 and aIPL-M1 cortico-cortical interactions were facilitatory and inhibitory, respectively, in healthy volunteers, whereas the interactions were converse and significantly different in writer's cramp. Baseline PMv-M1 interactions were inhibitory and similar between the groups. The dIPL-PMv resting state functional connectivity was increased in patients compared to controls, but no differences in structural connectivity between the nodes were observed. Cortical silent period was significantly prolonged in writer's cramp. Making a long term depression-like plastic change to dIPL node transformed the aIPL-M1 interaction to inhibitory (similar to healthy volunteers) and cancelled the PMv-M1 inhibition only in the writer's cramp group. These findings suggest that the parietal multimodal sensory association region could have an aberrant downstream influence on the fine motor control network in writer's cramp, which could be artificially restored to its normal function.

Harnessing the Electronic Health Record to Distribute Transition Services to Adolescents With Inflammatory Bowel Disease.

OBJECTIVES: The aim of the study was to evaluate feasibility and utility of an electronic health record (EHR) activity to assess transitional readiness, deliver services to meet individual needs, and to track patient progress. METHODS: We developed a Transition EHR activity (TEA) to track patients through a standardized process where transition readiness is annually assessed and services distributed based on need. The process assesses transition skills starting at age 12 years and sets goals through shared decision-making, delivers resources according to need, reviews patients' personal medical histories, and documents healthcare transfer to adult gastroenterology. We piloted TEA among patients with inflammatory bowel disease (IBD) ages ≥12 years. Distribution to patients was measured and tolerability assessed via patient self-report evaluations. RESULTS: Since launch, TEA has been distributed to all eligible patients (N = 53) with a median age of 16 (14,18) years (median [IQR]), 62% male, 58% white, 26% Hispanic at our weekly dedicated IBD clinic. All have performed the transition skills' self-assessment and practicum, and set transition goals with their healthcare provider. Of these individuals, 41 (77%) participated in survey feedback. On a utility rating scale of 0 (not helpful at all) to 10 (very helpful), patients reported median (IQR) utility scores of 8 (7,10) for the transition readiness assessment, 9 (7,10) for transition resources provided, and 9 (7,10) for the medical history summary. Most (91%) would recommend TEA to other patients. CONCLUSIONS: TEA standardized delivery of resources among pediatric IBD patients and was well received and friendly to clinical workflow.

Modeling pupil responses to rapid sequential events.

Pupil size is an easily accessible, noninvasive online indicator of various perceptual and cognitive processes. Pupil measurements have the potential to reveal continuous processing dynamics throughout an experimental trial, including anticipatory responses. However, the relatively sluggish (~2 s) response dynamics of pupil dilation make it challenging to connect changes in pupil size to events occurring close together in time. Researchers have used models to link changes in pupil size to specific trial events, but such methods have not been systematically evaluated. Here we developed and evaluated a general linear model (GLM) pipeline that estimates pupillary responses to multiple rapid events within an experimental trial. We evaluated the modeling approach using a sample dataset in which multiple sequential stimuli were presented within 2-s trials. We found: (1) Model fits improved when the pupil impulse response function (PuRF) was fit for each observer. PuRFs varied substantially across individuals but were consistent for each individual. (2) Model fits also improved when pupil responses were not assumed to occur simultaneously with their associated trial events, but could have non-zero latencies. For example, pupil responses could anticipate predictable trial events. (3) Parameter recovery confirmed the validity of the fitting procedures, and we quantified the reliability of the parameter estimates for our sample dataset. (4) A cognitive task manipulation modulated pupil response amplitude. We provide our pupil analysis pipeline as open-source software (Pupil Response Estimation Toolbox: PRET) to facilitate the estimation of pupil responses and the evaluation of the estimates in other datasets.

Classification of Functional Movement Disorders with Resting-State Functional Magnetic Resonance Imaging.

Introduction: Functional movement disorder (FMD) is a type of functional neurological disorder characterized by abnormal movements that patients do not perceive as self-generated. Prior imaging studies show a complex pattern of altered activity, linking regions of the brain involved in emotional responses, motor control, and agency. This study aimed to better characterize these relationships by building a classifier using a support vector machine to accurately distinguish between 61 FMD patients and 59 healthy controls using features derived from resting-state functional magnetic resonance imaging. Materials and Methods: First, we selected 66 seed regions based on prior related studies, then we calculated the full correlation matrix between them before performing recursive feature elimination to winnow the feature set to the most predictive features and building the classifier. Results: We identified 29 features of interest that were highly predictive of the FMD condition, classifying patients and controls with 80% accuracy. Several key features included regions in the right sensorimotor cortex, left dorsolateral prefrontal cortex, left cerebellum, and left posterior insula. Conclusions: The features selected by the model highlight the importance of the interconnected relationship between areas associated with emotion, reward, and sensorimotor integration, potentially mediating communication between regions associated with motor function, attention, and executive function. Exploratory machine learning was able to identify this distinctive abnormal pattern, suggesting that alterations in functional linkages between these regions may be a consistent feature of the condition in many FMD patients. Clinical-Trials.gov ID: NCT00500994 Impact statement Our research presents novel results that further elucidate the pathophysiology of functional movement disorder (FMD) with a machine learning model that classifies FMD and healthy controls correctly 80% of the time. Herein, we demonstrate how known differences in resting-state functional magnetic resonance imaging connectivity in FMD patients can be leveraged to better understand the complex pattern of neural changes in these patients. Knowing that there are measurable predictable differences in brain activity in patients with FMD may help both clinicians and patients conceptualize and better understand the illness at the point of diagnosis and during treatment. Our methods demonstrate how an effective combination of machine learning and qualitative approaches to analyzing functional brain connectivity can enhance our understanding of abnormal patterns of brain activity in FMD patients.

Quantitative epistasis analysis and pathway inference from genetic interaction data.

Inferring regulatory and metabolic network models from quantitative genetic interaction data remains a major challenge in systems biology. Here, we present a novel quantitative model for interpreting epistasis within pathways responding to an external signal. The model provides the basis of an experimental method to determine the architecture of such pathways, and establishes a new set of rules to infer the order of genes within them. The method also allows the extraction of quantitative parameters enabling a new level of information to be added to genetic network models. It is applicable to any system where the impact of combinatorial loss-of-function mutations can be quantified with sufficient accuracy. We test the method by conducting a systematic analysis of a thoroughly characterized eukaryotic gene network, the galactose utilization pathway in Saccharomyces cerevisiae. For this purpose, we quantify the effects of single and double gene deletions on two phenotypic traits, fitness and reporter gene expression. We show that applying our method to fitness traits reveals the order of metabolic enzymes and the effects of accumulating metabolic intermediates. Conversely, the analysis of expression traits reveals the order of transcriptional regulatory genes, secondary regulatory signals and their relative strength. Strikingly, when the analyses of the two traits are combined, the method correctly infers ~80% of the known relationships without any false positives.

Corticolimbic Modulation via Intermittent Theta Burst Stimulation as a Novel Treatment for Functional Movement Disorder: A Proof-of-Concept Study.

Neuroimaging studies suggest that corticolimbic dysfunctions, including increased amygdala reactivity to emotional stimuli and heightened fronto-amygdala coupling, play a central role in the pathophysiology of functional movement disorders (FMD). Transcranial magnetic stimulation (TMS) has the potential to probe and modulate brain networks implicated in neuropsychiatric disorders, including FMD. Therefore, the objective of this proof-of-concept study was to investigate the safety, tolerability and preliminary efficacy of fronto-amygdala neuromodulation via targeted left prefrontal intermittent theta burst stimulation (iTBS) on brain and behavioral manifestations of FMD. Six subjects with a clinically defined diagnosis of FMD received three open-label iTBS sessions per day for two consecutive study visits. Safety and tolerability were assessed throughout the trial. Amygdala reactivity to emotionally valenced stimuli presented during an fMRI task and fronto-amygdala connectivity at rest were evaluated at baseline and after each stimulation visit, together with subjective levels of arousal and valence in response to affective stimuli. The FMD symptom severity was assessed at baseline, during treatment and 24 h after the last iTBS session. Multiple doses of iTBS were well-tolerated by all participants. Intermittent TBS significantly decreased fronto-amygdala connectivity and influenced amygdala reactivity to emotional stimuli. These neurocircuitry changes were associated to a marked reduction in FMD symptom severity. Corticolimbic modulation via iTBS represents a promising treatment for FMD that warrants additional research.

Assessment of Prevalence of Adolescent Patient Portal Account Access by Guardians.

Importance: Patient portals can be configured to allow confidential communication for adolescents' sensitive health care information. Guardian access of adolescent patient portal accounts could compromise adolescents' confidentiality. Objective: To estimate the prevalence of guardian access to adolescent patient portals at 3 academic children's hospitals. Design, Setting, and Participants: A cross-sectional study to estimate the prevalence of guardian access to adolescent patient portal accounts was conducted at 3 academic children's hospitals. Adolescent patients (aged 13-18 years) with access to their patient portal account with at least 1 outbound message from their portal during the study period were included. A rule-based natural language processing algorithm was used to analyze all portal messages from June 1, 2014, to February 28, 2020, and identify any message sent by guardians. The sensitivity and specificity of the algorithm at each institution was estimated through manual review of a stratified subsample of patient accounts. The overall proportion of accounts with guardian access was estimated after correcting for the sensitivity and specificity of the natural language processing algorithm. Exposures: Use of patient portal. Main Outcome and Measures: Percentage of adolescent portal accounts indicating guardian access. Results: A total of 3429 eligible adolescent accounts containing 25 642 messages across 3 institutions were analyzed. A total of 1797 adolescents (52%) were female and mean (SD) age was 15.6 (1.6) years. The percentage of adolescent portal accounts with apparent guardian access ranged from 52% to 57% across the 3 institutions. After correcting for the sensitivity and specificity of the algorithm based on manual review of 200 accounts per institution, an estimated 64% (95% CI, 59%-69%) to 76% (95% CI, 73%-88%) of accounts with outbound messages were accessed by guardians across the 3 institutions. Conclusions and Relevance: In this study, more than half of adolescent accounts with outbound messages were estimated to have been accessed by guardians at least once. These findings have implications for health systems intending to rely on separate adolescent accounts to protect adolescent confidentiality.

In vivo assessment of neurodegeneration in Spinocerebellar Ataxia type 7.

Spinocerebellar Ataxia type 7 (SCA7) is a neurodegenerative disease characterized by progressive cerebellar ataxia and retinal degeneration. Increasing loss of visual function complicates the use of clinical scales to track the progression of motor symptoms, hampering our ability to develop accurate biomarkers of disease progression, and thus test the efficacy of potential treatments. We aimed to identify imaging measures of neurodegeneration, which may more accurately reflect SCA7 severity and progression. While common structural MRI techniques have been previously used for this purpose, they can be biased by neurodegeneration-driven increases in extracellular CSF-like water. In a cross-sectional study, we analyzed diffusion tensor imaging (DTI) data collected from a cohort of 13 SCA7 patients and 14 healthy volunteers using: 1) a diffusion tensor-based image registration technique, and 2) a dual-compartment DTI model to control for the potential increase in extracellular CSF-like water. These methodologies allowed us to assess both volumetric and microstructural abnormalities in both white and gray matter brain-wide in SCA7 patients for the first time. To measure tissue volume, we performed diffusion tensor-based morphometry (DTBM) using the tensor-based registration. To assess tissue microstructure, we computed the parenchymal mean diffusivity (pMD) and parenchymal fractional anisotropy (pFA) using the dual compartment model. This model also enabled us to estimate the parenchymal volume fraction (pVF), a measure of parenchymal tissue volume within a given voxel. While DTBM and pVF revealed tissue loss primarily in the brainstem, cerebellum, thalamus, and major motor white matter tracts in patients (p < 0.05, FWE corrected; Hedge's g > 1), pMD and pFA detected microstructural abnormalities in virtually all tissues brain-wide (p < 0.05, FWE corrected; Hedge's g > 1). The Scale for the Assessment and Rating of Ataxia trended towards correlation with cerebellar pVF (r = -0.66, p = 0.104, FDR corrected) and global white matter pFA (r = -0.64, p = 0.104, FDR corrected). These results advance our understanding of neurodegeneration in living SCA7 patients by providing the first voxel-wise characterization of white matter volume loss and gray matter microstructural abnormalities. Moving forward, this comprehensive approach could be applied to characterize the full spatiotemporal pattern of neurodegeneration in SCA7, and potentially develop an accurate imaging biomarker of disease progression.

Increase in Pediatric Respiratory Visits Associated with Santa Ana Wind-Driven Wildfire Smoke and PM2.5 Levels in San Diego County.

Rationale: There is significant evidence of increased healthcare utilization from cardiopulmonary causes in adults from exposure to wildfire smoke, but evidence in pediatric age groups is limited.Objectives: To quantify and examine the healthcare utilization effects of the December 2017 Lilac Fire in San Diego County among pediatric patients at the Rady Children's Hospital (RCH) emergency department and urgent care (UC) clinics.Methods: Using data from 2011 to 2017, including data on daily particulate matter <2.5 µm (PM2.5) in an inverse-distance interpolation model and RCH electronic medical records, we retrospectively analyzed pediatric respiratory visits at the RCH emergency department and UC clinics during the Santa Ana wind (SAW)-driven Lilac Fire from December 7 to 16, 2017. An interrupted time series study design was applied as our primary analysis to compare the observed pediatric respiratory visits from December 7 to 16, 2017 to what would have occurred in a counterfactual situation, namely, if the Lilac Fire had not occurred. A complementary descriptive spatial analysis was also used to evaluate the geographic distribution of respiratory visits in relationship to satellite imaging of the Lilac Fire and the associated wind pattern.Results: The Lilac Fire was associated with 16.0 (95% confidence interval [CI], 11.2-20.9) excess respiratory visits per day at the RCH emergency department across all pediatric age groups. Children aged 0 to 5 years had the highest absolute excess respiratory visits per day with 7.3 (95% CI, 3.0-11.7), whereas those aged 6 to 12 years had the highest relative increase in visits, with 3.4 (95% CI, 2.3-4.6). RCH UC clinics had similar results. The top five ZIP codes in San Diego County with the highest standard deviations of age-adjusted respiratory visits were all located generally downwind of the fire perimeter, as expected for the SAW pattern.Conclusions: We have demonstrated an increase in pediatric respiratory visits during the SAW-driven Lilac Fire in San Diego County in a patterned geographic distribution that is attributable to an increase in PM2.5 exposure. Younger children were particularly affected. Climate change is expected to result in more frequent and extensive wildfires in the region and will require greater preparedness and adaptation efforts to protect vulnerable populations, such as young children.

The m6A-methylase complex recruits TREX and regulates mRNA export.

N6-methyladenosine (m6A) is the most abundant internal modification of eukaryotic mRNA. This modification has previously been shown to alter the export kinetics for mRNAs though the molecular details surrounding this phenomenon remain poorly understood. Recruitment of the TREX mRNA export complex to mRNA is driven by transcription, 5' capping and pre-mRNA splicing. Here we identify a fourth mechanism in human cells driving the association of TREX with mRNA involving the m6A methylase complex. We show that the m6A complex recruits TREX to m6A modified mRNAs and this process is essential for their efficient export. TREX also stimulates recruitment of the m6A reader protein YTHDC1 to the mRNA and the m6A complex influences the interaction of TREX with YTHDC1. Together our studies reveal a key role for TREX in the export of m6A modified mRNAs.

UCHL3 Regulates Topoisomerase-Induced Chromosomal Break Repair by Controlling TDP1 Proteostasis.

Genomic damage can feature DNA-protein crosslinks whereby their acute accumulation is utilized to treat cancer and progressive accumulation causes neurodegeneration. This is typified by tyrosyl DNA phosphodiesterase 1 (TDP1), which repairs topoisomerase-mediated chromosomal breaks. Although TDP1 levels vary in multiple clinical settings, the mechanism underpinning this variation is unknown. We reveal that TDP1 is controlled by ubiquitylation and identify UCHL3 as the deubiquitylase that controls TDP1 proteostasis. Depletion of UCHL3 increases TDP1 ubiquitylation and turnover rate and sensitizes cells to TOP1 poisons. Overexpression of UCHL3, but not a catalytically inactive mutant, suppresses TDP1 ubiquitylation and turnover rate. TDP1 overexpression in the topoisomerase therapy-resistant rhabdomyosarcoma is driven by UCHL3 overexpression. In contrast, UCHL3 is downregulated in spinocerebellar ataxia with axonal neuropathy (SCAN1), causing elevated levels of TDP1 ubiquitylation and faster turnover rate. These data establish UCHL3 as a regulator of TDP1 proteostasis and, consequently, a fine-tuner of protein-linked DNA break repair.

Variability in Delivered Dose from Pressurized Metered-Dose Inhaler Formulations Due to a Delay Between Shake and Fire.

BACKGROUND: Pressurized metered-dose inhalers (pMDIs) should be shaken before use to prevent creaming or sedimentation of the drugs in solution; however, data published on this topic are limited, and it is rarely specified how soon after shaking the device should be actuated. Delays between shaking and firing the pMDI have previously been shown to cause significant inhomogeneity in delivered dose. We studied the effect of various shake-fire delays on the drug delivered from five commercially available pMDIs commonly prescribed for asthma and chronic obstructive pulmonary disease to assess the potential variability in delivered dose. METHODS: The pMDI formulations tested were the Flovent HFA, Ventolin Evohaler, Airomir Inhaler, and Symbicort (suspension pMDIs), and the QVAR 100 Inhaler (solution pMDI). Each pMDI was shaken for 5 seconds before attachment to a dosage unit sampling apparatus collection tube and filter, and it was actuated once with shake-fire delays of 0, 5, 10, 20, 30, 40, 50, and 60 seconds. Analysis of the eluates from the collection tubes and filters was performed by using high-performance liquid chromatography. Three of each pMDI were tested twice with each time delay. RESULTS: All of the suspension pMDIs produced variable amounts of drug over the shake-fire delays tested. A comparison of the delivered doses after the 0- and 60-second delays showed that the drug delivered increased for the Flovent HFA (320%), Ventolin Evohaler (346%), and Airomir Inhaler (230%) pMDIs; decreased for the Symbicort budesonide (75%) and formoterol fumarate (76%) pMDI; and remained consistent for the QVAR 100 Inhaler pMDI. CONCLUSIONS: The amount of drug delivered can vary widely over different shake-fire delays with suspension pMDIs. Therefore, guidance should be given to users/caregivers on the timing of firing after shaking their device, particularly with pediatrics, who may take time to become receptive to accepting their medication after pMDI shaking and before dose administration.