• PhD nursing programs: Where are we headed?

  Fain, James A; Sullivan-Bolyai, Susan L; Bova, Carol A (2024-08-29)

  Thank you for your timely editorial, The PhD in Nursing - Questions about a Credential at a Crossroads. We agree that the future of PhD education in nursing is in peril and needs rapid solutions. Low enrollment and threats of PhD program closures are on the rise. As three former Directors of PhD Programs, we feel it is essential that we do not walk away from PhD education. Here are some of our thoughts.
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  Preventing diabetes: What overweight and obese adults with prediabetes in the United States report about their providers' communication and attempted weight loss

  Demosthenes, Emmanuella J; Freedman, Jason; Hernandez, Camila; Shennette, Lisa; Frisard, Christine; Lemon, Stephenie C; Gerber, Ben S; Amante, Daniel J (2024-08-11)

  Objective: To investigate what overweight or obese adults with prediabetes in the United States report being told by providers about 1) having prediabetes, 2) diabetes risk, and 3) losing weight and the associations of these communications with attempted weight loss. Methods: Data from 2015 to 2018 National Health and Nutrition Examination Surveys (NHANES) for adults with a body mass index in the overweight or obesity ranges and HbA1c in the prediabetes range were examined (n = 2085). Patient reported data on what providers told them about having prediabetes, being at risk for diabetes, and losing weight were compared with attempted weight loss. Results: Most participants (66.4%) reported never being told they had prediabetes nor being at risk for diabetes, 13.0% reported being told they had prediabetes, 10.6% at risk for diabetes, and 8.0% both messages. 18.3% of participants reported being told to lose weight. Participants who reported being told they had prediabetes and at increased diabetes risk were more likely to report attempted weight loss (adjusted odds ratio (AOR) 1.8, 95% confidence interval (CI) 1.1-3.2). Reporting that they were told to lose weight was not significantly associated with an increase in reported weight loss attempts. Conclusions: In this cohort of individuals with overweight/obesity and prediabetic HbA1c values, low rates communications with providers about prediabetes and diabetes risk were reported. When both were discussed, patients reported greater attempted weight loss. These findings draw attention to the potential impact that provider communications about prediabetes and diabetes risk may have on lifestyle behavior change.
• Racial Disparities and Trends in Anticoagulant Use among Ambulatory Care Patients with Atrial Fibrillation and Atrial Flutter in the United States from 2007-2019

  Kan, Vincent (2024-08-08)

  Introduction Atrial fibrillation (AF) is the most common sustained cardiac arrhythmia, significantly increasing the risk of stroke. The introduction of direct oral anticoagulants (DOACs) since 2010 has transformed anticoagulation therapy, offering an alternative to warfarin with improved safety profiles. Despite the increased adoption of DOACs, disparities in their use among different racial and ethnic groups in the United States remain understudied. Methods This study utilized a repeated cross-sectional design, analyzing data from the National Ambulatory Medical Care Survey (NAMCS) from 2007 to 2019. The study population included adults diagnosed with AF or atrial flutter (AFL). We analyzed the temporal trends of DOAC and warfarin use from 2007 to 2019. We examined the prevalence of DOAC versus warfarin use and assessed associations between race/ethnicity, patient characteristics, and DOAC utilization from 2011 to 2019. Multivariable modified Poisson regression models were used to calculate adjusted prevalence ratios (aPR) for the associations. Results From 2011 to 2019, NAMCS recorded 3,224 visits involving AF or AFL, representing a weighted estimate of 103.6 million visits. DOAC use increased significantly, with apixaban becoming the predominant anticoagulant by 2016. Non-Hispanic Black patients were less likely to use DOACs compared to non-Hispanic White patients over time (aPR 0.75; 95% CI, 0.63-0.90). Patients with Medicaid insurance were also less likely to use DOACs (aPR 0.14; 95% CI: 0.04-0.46). Conclusion Despite the shift from warfarin to DOACs for AF and AFL treatment, significant racial and socioeconomic disparities persist. Non-Hispanic Black patients and those with Medicaid insurance are less likely to use DOACs. These findings highlight the need for targeted strategies to ensure equitable access to advanced anticoagulant therapies.
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  Identification of WNK1 as a Therapeutic Target to Suppress IgH/MYC Expression in Multiple Myeloma

  Ye, Tianyi (2024-08-08)

  Multiple myeloma (MM) remains an incurable hematological malignancy demanding innovative therapeutic strategies. Targeting MYC, the notorious yet traditionally undruggable oncogene, presents an appealing avenue. This thesis aims to identify and characterize novel regulators of MYC expression as therapeutic targets in MM. Using a genome-scale CRISPR/Cas9 screen, we identify the WNK lysine deficient protein kinase 1 (WNK1) as a regulator of MYC expression in MM cells. Genetic and pharmacological inhibition of WNK1 reduces MYC expression and, further, disrupts the MYC-dependent transcriptional program. Mechanistically, WNK1 inhibition attenuates the activity of the immunoglobulin heavy chain (IgH) enhancer, thus reducing MYC transcription when this locus is translocated near the MYC locus. Furthermore, we show in MM cells possessing distinct translocation profiles that WNK1 inhibition also downregulates other oncogenes frequently translocated near the IgH locus, including CCND1, FGFR3, and NSD2, broadening its potential therapeutic implications. WNK1 inhibition profoundly impacts MM cell behaviors, leading to growth inhibition, cell cycle arrest, senescence, and apoptosis. Importantly, the WNK inhibitor WNK463 inhibits MM growth in primary patient samples as well as xenograft mouse models, and exhibits synergistic effects with various anti-MM compounds. Collectively, this study uncovers WNK1 as a promising therapeutic target in MM, and suggests the utility of IgH translocations as useful biomarkers.
• Computationally Detecting Viral Infection and Characterizing Host-Virus Dynamics in scRNA-seq Datasets

  Cao, Yuming (2024-07-26)

  Viruses pose significant threats to human health, with their impacts varying by type. Advances in single-cell RNA sequencing (scRNA-seq) have enhanced our understanding of viruses and host responses by mapping human and viral transcripts within individual cells. However, ambient RNA contamination complicates the accurate identification of viral infections in scRNA-seq datasets. To address this, we introduced scVirusFinder, a method that uses a zero-inflated negative binomial model followed by a support vector machine classifier to identify virus-infected cells. This approach improves the detection of true viral infections in scRNA-seq datasets of virus infected cells. We applied this method to scRNA-seq data from nasal washes of healthy donors and those with acute influenza during the 2017-18 season. We identified seventeen cell populations, including a novel epithelial cell population with high MHC class II gene expression in infected individuals. Influenza virus infections were found in most cell populations, primarily in epithelial cells and major immune cells such as macrophages and neutrophils. Using viral reads from the scRNA-seq data, we discovered that each donor harbored a unique influenza variant with distinct non-synonymous mutations. Additionally, we observed interferon production and response in infected samples, with type III interferon particularly produced in infected ciliated epithelial cells. This study highlights the challenge of identifying infected cells from scRNA-seq datasets and provides a robust solution applicable to clinical samples, enhancing our understanding of viral infections and paving the way for therapeutic discoveries.
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  To treat or not to treat: a comparative effectiveness analysis of oral anticoagulant outcomes among U.S. nursing home residents with atrial fibrillation

  Chen, Qiaoxi; Baek, Jonggyu; Goldberg, Robert J.; Tjia, Jennifer; Lapane, Kate L; Alcusky, Matthew J (2024-07-19)

  Background: Nursing home residents with atrial fibrillation are at high risk for ischemic stroke, but most are not treated with anticoagulants. This study compared the effectiveness and safety between oral anticoagulant (OAC) users and non-users. Methods: We conducted a new-user retrospective cohort study by using Minimum Data Set 3.0 assessments linked with Medicare claims. The participants were Medicare fee-for-service beneficiaries with atrial fibrillation residing in US nursing homes between 2011 and 2016, aged ≥ 65 years. The primary outcomes were occurrence of an ischemic stroke or systemic embolism (effectiveness), occurrence of intracranial or extracranial bleeding (safety) and net clinical outcome (effectiveness or safety outcomes). Secondary outcomes included total mortality and a net clinical and mortality outcome. Cox proportional hazards and Fine and Grey models estimated multivariable adjusted hazard ratios (aHRs) and sub-distribution hazard ratios (sHRs). Results: Outcome rates were low (effectiveness: OAC: 0.86; non-users: 1.73; safety: OAC: 2.26; non-users: 1.75 (per 100 person-years)). OAC use was associated with a lower rate of the effectiveness outcome (sHR: 0.69; 95% Confidence Interval (CI): 0.61-0.77), higher rates of the safety (sHR: 1.70; 95% CI: 1.58-1.84) and net clinical outcomes (sHR: 1.20; 95% CI: 1.13-1.28) lower rate of all-cause mortality outcome (sHR: 0.60; 95% CI: 0.59-0.61), and lower rate of the net clinical and mortality outcome (sHR: 0.60; 95% CI: 0.59-0.61). Warfarin users, but not DOAC users, had a higher rate of the net clinical outcome versus OAC non-users. Conclusions: Our results support the benefits of treatment with OACs to prevent ischemic strokes and increase longevity, while highlighting the need to weigh apparent benefits against elevated risk for bleeding. Results were consistent with net favorability of DOACs versus warfarin.
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  Conquering hypertension in Vietnam: 12- month follow up results from a cluster-randomised controlled trial

  Nguyen, Hoa L; Ha, Duc A; Tran, Oanh T; Phan, Van H; Nguyen, Cuc T; Nguyen, Giang H; Nguyen, Thang T; Le, Thanh T; Goldberg, Robert J.; Wang, Bo; et al. (2024-07-01)

  Background: Approximately 20% of adults in Vietnam have hypertension, and management of this chronic condition remains challenging. This study aimed to assess the effectiveness of a multicomponent intervention in reducing blood pressure (BP) in adults with uncontrolled hypertension. Methods: This cluster randomised controlled trial was conducted in sixteen communities (8 intervention and 8 comparison) in a rural setting in Vietnam (2017-2022). Consenting adults (aged ≥ 18 years) with uncontrolled hypertension were enrolled. Sixteen communities were equally randomised to an intervention or comparison group stratified by district. Consenting adults were assigned to study groups by community. The comparison arm received training sessions about hypertension prevention and management for health workers, and patient education materials. The intervention arm received training sessions about hypertension prevention and management for health workers, and patient education materials and three enhancement components including a storytelling intervention, home BP self-monitoring, and expanded community health worker services. The primary outcome was the difference in changes in patient's levels of systolic BP between the study groups over a 12-month follow-up period. Patients and outcome assessors were masked. Findings: A total of 671 patients (340: intervention, 331: comparison) were enrolled in the trial. The mean age was 66 years and 45% were men. At the 12-month follow-up, the mean systolic BP declined by 18.4 mmHg in the intervention group and 3.7 mmHg in the comparison group (differential decline of 14.7 mmHg [95% CI: 11.8-17.6]). The intervention group also achieved better BP control and medication adherence than the comparison group. There were no serious adverse events related to study participation. Interpretation: The results of this trial demonstrate that a multicomponent intervention can effectively reduce elevated BP in individuals with uncontrolled hypertension in Vietnam. Trial registration: This trial was registered at ClinicalTrials.gov, NCT03590691. Funding: National Heart, Lung, and Blood Institute.
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  Factors Influencing Margin Clearance and the Number of Stages of Mohs Micrographic Surgery in Basal Cell Carcinoma: A Retrospective Chart Review

  Daniel, Vijaya T; Azzolino, Vincent; Abraham, Maria; Leonard, Nicholas; Blankenship, Kaitlin; Lal, Karan; Flahive, Julie; Brown, Regina; Tkachenko, Elizabeth; Teymour, Shereen; et al. (2024-06-28)

  How patient and tumor factors influence clearance margins and the number of Mohs Micrographic Surgery (MMS) stages when treating basal cell carcinoma (BCC) remains widely uncharacterized. It is important to elucidate these relationships, as surgical outcomes may be compared nationally between colleagues. Our objective is to evaluate the relationships between defect size and patient demographics, as well as between BCC subtypes and the number of MMS stages. Our second objective is to compare practice patterns and characteristics of patients requiring MMS at academic centers and private practices. A retrospective chart review was performed using data collected at academic centers (2015-2018) and private practices (2011-2018) of BCC patients older than 18 years old who underwent MMS. In total, 7651 patients with BCC requiring MMS were identified. Academic center adjusted analyses demonstrated clearance margins 0.1 mm higher for every year's increase in age (p < 0.0001) and 0.25 increase in MMS stages for high-risk BCC (p < 0.0001). Private practice adjusted analyses demonstrated clearance margins 0.04 mm higher for every year's increase in age (p < 0.0001). Clearance margins correlate with older age, and additional MMS stages correlate with high-risk BCC, suggesting the role patient and tumor factors may play in predicting tumor clearance and MMS stages.
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  Ethical Frameworks in Periviable Decision-Making: Patient Perspectives and Provider Patterns

  Delaney, Kathryn (2024-06-25)

  Background: Periviability counseling involves engaging in difficult ethical decisions. The ethical frameworks used by patients and their healthcare providers to discuss this topic have not been previously studied. Objectives: This study assessed the ethical frameworks used by patients and by providers during periviability counseling and subsequent decision making. Study Design: This mixed methods study included patients between gestation ages 21 weeks 0 days through 24 weeks 6 days who required periviability counseling, and the providers performing this counseling. Counseling sessions between providers and patients were recorded, as were semi-structured follow-up interviews with patients. These recordings were transcribed and analyzed using thematic analysis. Demographic surveys were given to providers, and patient demographic, obstetric, and delivery information was abstracted from medical records. Results: A total of 11 patients and 11 providers participated in either a recorded counseling session, a follow-up interview, or both. Qualitative analysis revealed the following themes: 1) decisions regarding periviable intervention are the patient’s to make, 2) desire to avoid pain or suffering of the fetus/newborn, especially as it related to CPR, 3) doing what is best for the family, including partners and other children at home, 4) “doing everything” as a good in itself, and 5) periviable complications disrupt expectations about parenthood or pregnancy which then need to be reimagined. These themes corresponded to four distinct ethical frameworks: principlism, care ethics, virtue ethics, and narrative ethics. All of the patients, and approximately three-quarters of providers used at least one of these ethical frameworks; most used a combination of frameworks. Conclusions: Patients and providers engaging in decision making surrounding periviable intervention use commonly accepted ethical frameworks to discuss and decide how to proceed with these pregnancies.
• Analyzing the Role of MS4A Receptors in Mouse Olfaction

  Jiang, Hao-Ching (2024-06-17)

  How the brain interprets sensory information to generate appropriate behaviors remains a fundamental, unanswered question in neurobiology. In mammals, the olfactory system detects external chemicals through a vast array of odorant receptors expressed by peripheral olfactory sensory neurons (OSNs). These OSNs connect to brain circuits responsible for creating olfactory percepts and triggering odor-driven behaviors. The olfactory system has evolved distinct subsystems to initiate various behaviors, each containing anatomically and molecularly distinct OSNs for different functions. The olfactory epithelium (OE) is the largest and best-studied subsystem, crucial for odor discrimination and learning. Smaller olfactory subsystems detect ethologically relevant odorants, necessary for innate behaviors like foraging, mating, and predator avoidance. Unlike conventional OSNs in the OE, receptors in smaller subsystems are largely uncategorized, leaving their roles in odor-driven behaviors poorly understood. I focused on the function of the MS4A gene family, which is expressed in a specific olfactory subsystem linked to mouse innate behaviors. I demonstrated that multiple genes in this family act as chemoreceptors in vivo. Additionally, I identified one family member, MS4A1, expressed in a subset of previously uncharacterized OSNs in the mouse nose. MS4A1 functions as a non-canonical chemoreceptor, required for evoking innate avoidance responses to certain aversive chemicals. Beyond OSNs, the MS4A gene family is also expressed in various immune cells. Notably, I found that MS4A1/CD20, a tumor marker on B cells, acts as a potential chemoreceptor. This discovery could enhance understanding of MS4A proteins' roles in tissues outside OSNs and provide potential therapeutic targets for human diseases.
• Calcineurin coordinates cell cycle progression with adaptation to environmental stress

  Flynn, Mackenzie J (2024-06-14)

  Sudden exposure to environmental stress threatens the viability of single-celled microbes and cells within complex tissues. In order to survive, cells must sense environmental changes and coordinate a transient cell cycle arrest with the appropriate adaptive response. Cells have several stress-responsive pathways that promote adaptation to distinct stressors, but how these pathways interact with one another is poorly understood. Here, we examined the response to calcium chloride stress, which activates the phosphatase calcineurin and the MAPK Hog1 in Saccharomyces cerevisiae. We discovered that calcineurin extends Hog1 activation, which causes prolonged downregulation of cell cycle-regulated genes and delays progression through multiple cell cycle phases. At the G1/S transition, crosstalk between calcineurin and Hog1 dramatically increases the duration of calcium-induced arrest. I found that Hog1 triggers arrest independent of calcineurin by decreasing G1 cyclin transcription and calcineurin maintains this arrest by extending Hog1-dependent activation of the G1 CDK inhibitor Cip1. These results suggest that stress-response pathway interactions tailor cell cycle arrest with adaptation to environmental stress. The immediate response to stress is well-characterized, but how cells maintain viability in challenging environments after recovering from a stress-induced arrest is unknown. I investigated the response to prolonged growth in calcium stress and found that calcineurin maintains fitness by promoting cell division and suppressing death. I determined that calcineurin helps cells proliferate and survive prolonged calcium exposure by two mechanisms, which differentially require a downstream transcription factor. Together, these findings highlight the importance of stress-response pathways during both acute and chronic environmental stress.
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  Location, Location, Location: The Impact of One's Neighborhood on Outcomes Following Proctectomy for Rectal Cancer

  Onyiego, Alexandra (2024-06-13)

  Purpose: The area deprivation index (ADI) is a comprehensive assessment of the social determinants of health. The higher the composite ADI score, the more underserved the area. This study examined the impact of a patient’s ADI score on surgical, pathologic, and survival outcomes following proctectomy for rectal cancer. Methods: Data from a single tertiary care medical center’s targeted National Surgical Quality Improvement Program (NSQIP) were used to identify all patients undergoing proctectomy for rectal cancer between January 2014 – December 2022. A state ADI (ranking 1-10) was assigned to each patient using their residential address. Patients were categorized into Low (1-4), Moderate (5-6), High (7-8), and Very High (9-10) ADI groups. Results: Two-hundred and four patients who underwent proctectomy for rectal cancer were included. There were no significant between group differences in patient’s post-operative outcomes including 30-day morbidity and hospital readmissions. As well as pathologic outcomes, including completeness of mesorectal excision and rate of positive margins. A Cox multivariable adjusted regression model showed those in the Very High group had a higher risk of dying at 5 years than the Low ADI group. Patients in the Very High and High ADI groups were significantly more likely to be discharged somewhere other than home after their surgery than those in the Low ADI group, (13% vs. 0%). The median time from diagnosis to initiating treatment or surgery showed those in the Very High and High ADI groups took longer to obtain treatment. Conclusion: A patient’s ADI has an impact on their overall survival following proctectomy for rectal cancer and influences their time to receiving treatment and their postoperative disposition. Future studies with larger patient cohorts are needed to more clearly define the role of ADI in predicting patient outcomes following proctectomy for rectal cancer.
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  Preventing acute neurotoxicity of CNS therapeutic oligonucleotides with the addition of Ca and Mg in the formulation [preprint]

  Miller, Rachael; Paquette, Joseph; Barker, Alexandra; Sapp, Ellen; McHugh, Nicholas; Bramato, Brianna; Yamada, Nozomi; Alterman, Julia F; Echeverria, Dimas; Yamada, Ken; et al. (2024-06-08)

  Oligonucleotide therapeutics (ASOs and siRNAs) have been explored for modulation of gene expression in the central nervous system (CNS), with several drugs approved and many in clinical evaluation. Administration of highly concentrated oligonucleotides to the CNS can induce acute neurotoxicity. We demonstrate that delivery of concentrated oligonucleotides to the CSF in awake mice induces acute toxicity, observable within seconds of injection. Electroencephalography (EEG) and electromyography (EMG) in awake mice demonstrated seizures. Using ion chromatography, we show that siRNAs can tightly bind Ca2+ and Mg2+ up to molar equivalents of the phosphodiester (PO)/phosphorothioate (PS) bonds independently of the structure or phosphorothioate content. Optimization of the formulation by adding high concentrations (above biological levels) of divalent cations (Ca2+ alone, Mg2+ alone, or Ca2+ and Mg2+) prevents seizures with no impact on the distribution or efficacy of the oligonucleotide. The data here establishes the importance of adding Ca2+ and Mg2+ to the formulation for the safety of CNS administration of therapeutic oligonucleotides.
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  Bigtools: a high-performance BigWig and BigBed library in Rust

  Huey, Jack D; Abdennur, Nezar (2024-06-03)

  Motivation: The BigWig and BigBed file formats were originally designed for the visualization of next-generation sequencing data through a genome browser. Due to their versatility, these formats have long since become ubiquitous for the storage of processed sequencing data and regularly serve as the basis for downstream data analysis. As the number and size of sequencing experiments continues to accelerate, there is an increasing demand to efficiently generate and query BigWig and BigBed files in a scalable and robust manner, and to efficiently integrate these functionalities into data analysis environments and third-party applications. Results: Here, we present Bigtools, a feature-complete, high-performance, and integrable software library for generating and querying both BigWig and BigBed files. Bigtools is written in the Rust programming language and includes a flexible suite of command line tools as well as bindings to Python. Availability and implementation: Bigtools is cross-platform and released under the MIT license. It is distributed on Crates.io, Bioconda, and the Python Package Index, and the source code is available at https://github.com/jackh726/bigtools.
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  Kinetics of Pre-mRNA 3’ End Cleavage

  Torres Ulloa, Leslie (2024-05-31)

  3’ end cleavage and polyadenylation are required steps in pre-mRNA maturation. The rate at which 3’ end cleavage occurs can determine the temporal availability of mRNA for subsequent function throughout the cell and is likely tightly regulated. While there are numerous high-throughput methods for global profiling of RNA maturation rates, the study of pre-mRNA 3’ end cleavage kinetics has remained limited to low-throughput approaches, and the temporal regulation of polyadenylation site choice that determines the composition of the 3’ UTRs of mRNAs remains poorly understood. This research project seeks to address this gap by introducing a novel genome-wide, site-specific methodology for estimating rates of pre-mRNA 3’ end cleavage, using metabolic labeling of nascent RNA, high-throughput sequencing, and mathematical modeling. Using in-silico simulations of nascent RNA-seq data, we show that our approach can accurately and precisely estimate cleavage half-lives for both constitutive and alternative sites. In Drosophila melanogaster S2 cells, we find that cleavage rates are fast but highly variable across sites, with alternative events being slowest. This variability in rates is underpinned by distinctive sequence elements, where an A-rich region upstream of the cleavage site, a U-rich element downstream of the cleavage site, and a higher density of polyadenylation signals, lead to faster cleavage reactions. Assessment of Polymerase II dynamics around cleavage sites reveals that cleavage rates are associated with the localization of RNA Polymerase II at the end of a gene and faster cleavage leads to quicker degradation of downstream read-through RNA. This approach for estimating pre-mRNA 3’ end cleavage kinetics opens new possibilities in the study of co-transcriptional regulation of mRNA expression and transcription termination across cellular states.
• Burnout Among Young Adults With Type 1 Diabetes

  Perez, Danielle; Sullivan-Bolyai, Susan L; Bova, Carol A; Fain, James A (2024-05-30)

  Purpose: The purpose of this qualitative descriptive study was to describe the experience of diabetes burnout in young adults with type 1 diabetes (T1DM). In addition, aims included participant perspectives of risk and protective factors associated with burnout and ways to balance everyday life with diabetes self-management (DSM). Methods: Young adults with T1DM (N = 11) were recruited through social media platforms and modified snowball sampling and interviewed. Informational redundancy was achieved. Qualitative thematic coding and analysis were conducted within and across transcripts. Results: Diabetes burnout was described as the willingness to put diabetes and DSM on the "back burner" and let things slide due to exhaustion, frustration, apathy, and the desire to be like everyone else for a while. Risk and protective factors were identified along with strategies to achieve balance of DSM in everyday life. Conclusions: This study identified a clear definition of diabetes burnout and acknowledges this concept as distinct and separate from other psychosocial conditions. Health care providers can utilize this information to identify individuals at risk for diabetes burnout and offer more effective support to lessen the overall burden associated with T1DM.
• Variation in Depth of Sedation Targeted and Achieved among Mechanically Ventilated Patients and Associated Outcomes

  Rucci, Justin M (2024-05-30)

  Introduction: Sedative agents are commonly administered to patients receiving mechanical ventilation (MV). Practice guidelines recommend provision of light sedation within validated scoring systems (e.g., Richmond Agitation Sedation Scale [RASS]), but recognize some circumstances require deeper sedation. The real-world approaches to depth of sedation, and the impact of hospital sedation practices on patient outcomes, remain uncharacterized. Methods: We used the US based eICU collaborative research database to identify adult patients who received MV > 24 hours, who did not have a diagnosis that may require sedatives for indications other than facilitating MV, and who had recorded RASS goals and scores. We used mixed effects regression models to determine factors associated with initial RASS goals and rates of RASS score-goal concordance. We organized hospitals into quartiles of risk-adjusted RASS score-goal concordance, and used g-computation to evaluate differences in ventilator free days (VFD) at hospital day 28. Results: We identified a study sample of 1,650 adult patients (at 21 hospitals) who met inclusion/exclusion criteria. Hospital-level risk-adjusted initial RASS goals ranged from -1.4 to 0.2, and hospital-level risk-adjusted RASS score-goal concordance ranged from 27% to 64%. Patients admitted to hospitals in the highest quartile of score-goal concordance (quartile 4) were generally targeted for deeper sedation (median RASS goal -1.31) than patients admitted to hospitals in the lowest quartile (quartile 1) (median RASS goal -0.58). Compared to patients admitted to quartile 1 hospitals, patients at quartile 4 hospitals experienced fewer VFDs (adjusted incidence risk difference -2.4, 95% CI -4.26 to -0.36). Conclusion: US hospitals prescribe RASS goals in line with guideline recommendations for light sedation, but there is wide variation in achieving these RASS goals. Hospitals with higher RASS score-goal concordance typically prescribed deeper RASS goals, and patients admitted to these hospitals experienced fewer VFD.
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  Dual engagement of the nucleosomal acidic patches is essential for deposition of histone H2A.Z by SWR1C

  Baier, Alexander S; Gioacchini, Nathan; Eek, Priit; Leith, Erik M; Tan, Song; Peterson, Craig L (2024-05-29)

  The yeast SWR1C chromatin remodeling enzyme catalyzes the ATP-dependent exchange of nucleosomal histone H2A for the histone variant H2A.Z, a key variant involved in a multitude of nuclear functions. How the 14-subunit SWR1C engages the nucleosomal substrate remains largely unknown. Studies on the ISWI, CHD1, and SWI/SNF families of chromatin remodeling enzymes have demonstrated key roles for the nucleosomal acidic patch for remodeling activity, however a role for this nucleosomal epitope in nucleosome editing by SWR1C has not been tested. Here, we employ a variety of biochemical assays to demonstrate an essential role for the acidic patch in the H2A.Z exchange reaction. Utilizing asymmetrically assembled nucleosomes, we demonstrate that the acidic patches on each face of the nucleosome are required for SWR1C-mediated dimer exchange, suggesting SWR1C engages the nucleosome in a 'pincer-like' conformation, engaging both patches simultaneously. Loss of a single acidic patch results in loss of high affinity nucleosome binding and nucleosomal stimulation of ATPase activity. We identify a conserved arginine-rich motif within the Swc5 subunit that binds the acidic patch and is key for dimer exchange activity. In addition, our cryoEM structure of a Swc5-nucleosome complex suggests that promoter proximal, histone H2B ubiquitylation may regulate H2A.Z deposition. Together these findings provide new insights into how SWR1C engages its nucleosomal substrate to promote efficient H2A.Z deposition.
• Synthesis of Chemically Modified Nucleic Acids to Characterize and Inhibit APOBEC3 Enzymes

  Hedger, Adam K (2024-05-28)

  APOBEC3 proteins (A3s) are enzymes that catalyze the deamination of cytidine (C) to uridine (U) in single-stranded DNA (ssDNA) substrates, playing a key role in innate antiviral immunity. However, incomplete A3 restriction of viruses is shown to cause drug resistance, and erroneous deamination of host DNA is strongly implicated in many cancers and therapy resistance. Small molecule drug discovery efforts have so far largely failed in finding potent and selective inhibitors of A3 enzymes. Recent structural advances have informed the design of “substrate mimicking” competitive oligonucleotide inhibitors, with potencies in the low-µM to -nM range. Nevertheless, further medicinal chemistry efforts are needed to improve the cellular stability and potency of these compounds. With that goal, in this thesis I report the first co-crystal structure of an active A3 enzyme bound to an oligonucleotide inhibitor at high-resolution, capturing the transition-state, and further informing inhibitor design. Next, I develop new and efficient synthesis routes to novel cytidine analogs for later incorporation into oligonucleotide inhibitors, including 2′-fluoroarabino- and 2′-difluoro-1,3-diazepinone nucleosides. Finally, I carry out a detailed structure-activity relationship of chemically modified oligonucleotide inhibitors against A3 enzymes. The resulting chemically optimized inhibitors from this work are the most potent reported to date, show greatly enhanced nuclease stability, and display inhibition in a cellular inhibition assay. In summary, my thesis work combines structural biology with nucleic acid chemistry to further A3 inhibitor design on three fronts, ultimately achieving the goal of generating potent and chemically stabilized inhibitors for use in cellular assays and beyond.
• Single-cell genomics and regulatory networks for 388 human brains

  Emani, Prashant S; Liu, Jason J; Clarke, Declan; Jensen, Matthew; Warrell, Jonathan; Gupta, Chirag; Meng, Ran; Lee, Che Yu; Xu, Siwei; Dursun, Cagatay; et al. (2024-05-24)

  Single-cell genomics is a powerful tool for studying heterogeneous tissues such as the brain. Yet little is understood about how genetic variants influence cell-level gene expression. Addressing this, we uniformly processed single-nuclei, multiomics datasets into a resource comprising >2.8 million nuclei from the prefrontal cortex across 388 individuals. For 28 cell types, we assessed population-level variation in expression and chromatin across gene families and drug targets. We identified >550,000 cell type-specific regulatory elements and >1.4 million single-cell expression quantitative trait loci, which we used to build cell-type regulatory and cell-to-cell communication networks. These networks manifest cellular changes in aging and neuropsychiatric disorders. We further constructed an integrative model accurately imputing single-cell expression and simulating perturbations; the model prioritized ~250 disease-risk genes and drug targets with associated cell types.

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