University Of Colorado Denver

NIH Research Projects · FY 2025 · 2025-09

Project Summary An illicit drug enterprise is currently duping United States citizens by marketing synthetic cannabinoids to them as a safe alternative to cannabis. Professionals and students are also drawn to these compounds to cheat conventional drug tests. Rather than a safe alternative, most synthetic cannabinoids are more dangerous than cannabis itself. Indeed, the primary psychoactive component of cannabis, a chemical called delta-9- tetrahydrocannabinol, only partially activates the primary cannabinoid receptor in the brain. In contrast, the most commonly abused class of synthetic cannabinoids fully activate this receptor. While we already know that this increased cannabinoid receptor activation can produce more substantial drug effects, the neurobehavioral consequences of repeated exposures remain unclear. This statement is particularly true across different stages of lifespan development and between sexes. Synthetic cannabinoids are one of the most commonly abused drugs amongst adolescents and cannabinoid abuse rates are at all-time highs across all age ranges, including senior citizens. Elucidating the neurobehavioral consequences of chronic synthetic cannabinoid exposure is necessary to validate and raise public awareness of the dangers posed by this harmful new class of drugs. Our preliminary data demonstrate that chronic exposure to synthetic cannabinoids renders them less effective at evoking dopamine release in adult rats. This finding is concerning because we have shown that dopamine- release events are necessary for adaptive behavior and resiliency. Because little is known regarding the effects of chronic cannabinoid abuse across the developmental lifespan, we will first test whether this form of neurochemical tolerance develops in both female and male early adolescent, adult, and aged rats. It is also essential to consider whether chronic cannabinoid exposure interferes with fundamental behaviors that allow us to thrive in society. Thus, we will use an innovative behavioral economics framework to assess whether the value female and male adult rats place on the pursuit of reward and the avoidance of harm changes following chronic synthetic cannabinoid exposure. We recently used this behavioral economics approach to show that dopamine value signals represent price and causally modify the price rats will pay for both reward and avoidance. Finally, we will investigate the interaction between chronic cannabinoid abuse and exercise. We will test whether a history of exercise protects against the development of neurochemical tolerance in female and male adult rats. Existing data reveal exercise creates a state of heightened dopamine release and that this neurobiological adaptation may protect against adverse events. The collection of these data will advance our understanding of the neurobiology of cannabinoid abuse in a manner that can be applied to improve individual and public health.

NIH Research Projects · FY 2025 · 2025-09

PROJECT SUMMARY Understanding the metabolism of human red blood cells (RBCs) is important for improving medicine surrounding blood transfusion and inborn errors of metabolism. This proposal aims to use computational genome-scale metabolic models (GEMs) to advance translational applications for blood transfusions and inborn errors of metabolism. By population, RBCs are the most common cell in the human body, numbering approximately 25 trillion RBCs per person. The main role of the RBC is to carry oxygen using hemoglobin and, with approximately 260 million units of hemoglobin per cell, RBCs are highly evolved to carry oxygen. Mature RBCs lack mitochondria and organelles so they cannot synthesize new proteins. Hence, they have evolved elaborate metabolic mechanisms to regulate their physiology and adjust to environmental stressors. One such stressor is RBC storage in the blood bank, a crucial part of meeting patients' transfusion needs. Another RBC stressor is inborn errors of metabolism, such as G6PDH deficiency. Studying RBC metabolism sheds light on these medical treatments and conditions. The first specific aim is to expand the current state-of-the-art human RBC GEM with data from our lab's most recent proteomics study of ultra-pure human RBCs. This will enable computational models of the human red blood cell to reflect RBC metabolism more accurately. The second specific aim is to use the update or current RBC genome-scale metabolic model to identify metabolic profiles linked to RBC failure (hemolysis) in storage. To this end, we will use the Recipient Epidemiology and Donor Evaluation Study (REDS) operated by the NIH NHLBI to enhance knowledge of RBC storage and transfusion medicine.

NIH Research Projects · FY 2025 · 2025-09

Project Summary Hundreds of taste buds (TBs) are located in the epithelial trenches of the circumvallate taste papilla (CVP) at the posterior midline of the tongue. Each TB contains taste receptor cells (TRCs) that transduce taste information to the brain and are continually replaced from stem/progenitor cells outside of TBs. Cancer patients undergoing chemo- or radiotherapy often experience dysgeusia, or taste dysfunction, likely due to perturbation to TRC renewal. Patients experiencing dysgeusia have increased risk of depression, malnutrition, and poor treatment outcomes. Therefore, understanding the mechanisms regulating maintenance of taste epithelium and TRC renewal will inform development of treatments to prevent or restore taste loss in these patients. CVP homeostasis occurs through proliferation and differentiation of progenitor cells that generate both TRCs and the surrounding non-taste epithelium. Our lab has recently identified new, long-term SOX9+ progenitors that, as a population, generate TRCs and non-taste epithelium (NTE); however, the potential of individual SOX9+ progenitors is unexplored. SOX9+ progenitors are located distant from taste buds, in the epithelial junction linking CVP trenches to the ducts of the underlying von Ebner's minor salivary glands (VEG). Each CVP contains a dozen or more junctions containing SOX9+ cells, suggesting that multiple independent progenitor reservoirs contribute to taste epithelium. Additionally, in contrast to the highly proliferative CVP, the junction contains few proliferative cells, indicating SOX9+ cells are slow cycling. These findings lead to my hypothesis that individual SOX9+ progenitors from multiple junctions produce progeny that move into local CVP trench epithelium, become highly proliferative, and differentiate into TRCs and NTE. Thus, in Aim 1, I will use sparse SOX9 lineage trace, whole-tissue clearing, and EdU birth dating, to determine the lineage and proliferative potential of individual progenitors from multiple junctions. Our previous work has focused on SOX9+ progenitor contribution in adult homeostasis, but if and when these taste stem cells are established embryonically is unknown. Embryonic immunostaining reveals SOX9+ cells are present during initial formation of the CVP and have overlapping expression with Ptch, the Shh signaling receptor. Shh is necessary for anterior tongue taste bud development, and proper CVP trench invagination in the posterior tongue. These findings lead to my second hypothesis that embryonic Shh signaling is required to establish adult SOX9+ taste progenitors. In Aim 2, I will use embryonic lineage tracing and genetic deletion of Shh signaling in SOX9+ cells to determine when SOX9+ cells are established, begin contributing to TBs, and if Shh signaling is required in these processes. Overall, this work may lead to approaches to leverage concentrated pools of taste stem cells to restore taste function in patients.

NIH Research Projects · FY 2026 · 2025-09

PROJECT SUMMARY/ABSTRACT Fetal growth restriction (FGR) impacts 10-20% of pregnancies worldwide and increases the offspring’s risk for later development of obesity and type 2 diabetes due to incompletely understood mechanisms. The focus of this proposal is the nexus of intestinal development and gut microbiome establishment. Gut microbial composition represents an important and modifiable factor that contributes to postnatal intestinal function and systemic health and has been understudied in FGR. Our preliminary studies demonstrate impaired glucose tolerance in adult FGR males after high fat diet challenge and gut microbial dysbiosis into adulthood. The purpose of this work is to test the reciprocal relationship between the intestine and microbiome in FGR, and how a microbiome-directed nutritional intervention can impact that relationship. The central hypothesis is that impaired intestinal and gut microbial development in FGR increases risk for adverse outcomes which can be mitigated by postnatal dietary interventions. Aim 1 will use our established mouse model of FGR to test the function of intestinal stem, goblet, Paneth, and enteroendocrine cells. Organoid formation and single cell RNA sequencing of ileal and colonic cells will further characterize intestinal development differences between control and FGR animals. Aim 2 examines whether FGR animals can sustain transplanted fecal microbiome from control animals, testing for intrinsic defects of microbiome establishment. Aim 3 tests the ability of early supplementation with human milk oligosaccharide 2’- fucosyllactose (2’FL) to protect against adverse metabolic outcomes in adult FGR mice. Control and FGR offspring will be challenged with a high fat diet with or without 2’FL supplementation. At age 24 weeks we will assess glucose homeostasis, body composition, energy balance, and microbiome composition. The interdisciplinary approach is innovative as it examines a potential mechanistic role for the gut microbiome in adverse metabolic outcomes after FGR. The proposal is significant as it addresses a critical and prevalent clinical problem with high translation potential. The proposed research is translational to human health as 2’FL has demonstrated safety in infants and known associations with growth and intestinal health. Complementary to the proposed research plan, a five-year mentored career development training plan has been devised, incorporating didactic learning in statistical methods and organoids in addition to hands-on training in bioinformatic analyses, intestinal pathophysiology, intestinal organoids, fecal microbiome transplant, and evaluation of glucose metabolism. The candidate requires mentorship by a multidisciplinary team and has assembled a group with expertise in fetal growth restriction, intestinal pathophysiology, microbiome, host- intestine interactions, and glucose homeostasis. The candidate’s long-term career goal is to become an independent investigator studying early childhood dietary interventions to improve lifelong after FGR.

NIH Research Projects · FY 2025 · 2025-09

PROJECT SUMMARY Cardiovascular disease (CVD) is the leading cause of death in U.S. women. Hypertensive disorders of pregnancy greatly increase risk for developing CVD later in life for both mothers and children. Hypertensive disorders of pregnancy affect 16% of pregnancies, and are more prevalent among pregnant people of color, highlighting the need for a health-equitable approach to understanding etiology and prevention/treatment. The human microbiome is a compelling target for etiologic investigations of hypertensive disorders of pregnancy because, unlike the human genome, it can be modified. Cross-sectional studies, case-control studies, and murine models suggest oral, gut, and vaginal microbes can cause cardiovascular diseases, including hypertensive disorders in pregnancy, through their production metabolites. However, previous human studies have not integrated and jointly assessed the oral, gut, and vaginal microbiomes in pregnancy; have not concomitantly examined the role of microbe-dependent targeted and untargeted metabolites; have been limited to a single time point in pregnancy; and have been in populations that lack racial and ethnic diversity. To address these gaps, we will conduct an ancillary study to an ongoing NIH-funded cohort to investigate the associations of maternal microbiomes and metabolomes with hypertensive disorders of pregnancy and maternal child CVD risk factors. Key objectives are to: (1) characterize the distinct body-site specific microbiomes (oral, gut, vaginal) and metabolomes of pregnant persons in a racially and ethnically diverse population, and (2) determine the relative importance of different microbiome niches, microbiome features within these niches, and critical windows for possible intervention to reduce hypertensive disorders of pregnancy and future CVD risk for mothers and their offspring. In particular, we propose to perform whole genome shotgun metagenomics, targeted quantification of short chain fatty acids, trimethylamine N-oxide, and nitrite and nitrate, along with untargeted metabolomics for discovery. We will jointly investigate the microbiomes and metabolomes measured during the 1st/2nd and 3rd trimesters in pregnancy. Drs. Mueller (contact MPI) and Moore (MPI) will carry out this research with an outstanding group of interdisciplinary co-investigators in the collaborative and eminent environments of the University of Colorado Anschutz Medical Campus. Dr. Mueller and Moore’s co-investigators have complementary expertise in statistics (Zhao), -omics analyses (Perng), computational biology and informatics (Olm), body composition and cardiometabolic diseases (Tilves), and community-based dissemination (Rinehart). With the support of their research team, Drs. Mueller and Moore are well positioned to complete the proposed activities. The findings have great potential to: (a) identify early predictors of hypertensive disorders in pregnancy that can be targeted for prevention, (b) reveal novel mechanisms of these hypertensive disorders, (c) offer new disease prevention strategies and therapeutic possibilities, and (d) inform use of the microbiome-metabolome nexus for precision nutrition/medicine.

NIH Research Projects · FY 2025 · 2025-09

PROJECT SUMMARY Degenerative diseases that cause retinal neuronal cell death often result in permanent vision loss. This is because retinal neurons, like rod and cone photoreceptors, do not regenerate. The most promising potential therapeutic strategies for restoring lost vision include artificially stimulating endogenous neuronal regeneration or programming human stem cells into transplantable retinal tissue. However, realizing such strategies is hindered by our limited understanding of the developmental mechanisms used to build the retina. During retinal development, proliferative multipotent progenitor cells choose between seven major fate outcomes. This choice process, called fate specification, is determined by a combination of a cell’s potential (i.e., competence) and instructive factors that select between competing fate choices. Retinal fate specification is a dynamic, probabilistic process that is controlled by the intersection of intrinsic gene regulatory networks and environmental cell-cell signaling mechanisms. The Notch signaling pathway impacts competence and fate choice decisions in the retina. However, its mechanisms are poorly understood due to a lack of genetic tools that can dynamically manipulate signaling in specific subpopulations of competent cells over time. To overcome this barrier, we identified enhancer sequences for the key transcription factor Otx2 that drive discrete spatial and temporal activity patterns in the mouse retina. Using these narrowly tailored enhancer tools, our initial findings show that Notch signaling plays multiple discrete fate choice roles throughout development. Our objective is to finely dissect how Notch signaling functions to understand the probabilistic nature of retinal cell fate specification. In Aim 1 of this proposal, we will investigate how Notch signaling regulates multiple different fate decisions throughout retinal development. We will activate Notch signaling at discrete stages of retinal development and use single cell RNA sequencing and histological approaches to determine how cells change competence and fate choices over time. These data will be used to determine whether Notch signaling delays decision making or acts by specifically instructing fate choices in competent cells at different stages of development. In Aim 2, we will use developmental and genetic techniques to explore how Notch signaling exposure (dosage and duration) differentially impacts competence and retinal cell fate decisions. Leveraging our unique genetic tools, this project will reveal how the multifaceted Notch signaling pathway impacts competence and dynamic fate choice probabilities in the developing retina. This knowledge is essential for creating regenerative and cell-based therapies to replace lost neurons, which may restore vision in millions of people suffering from retinal degenerative diseases.

NIH Research Projects · FY 2025 · 2025-09

Abstract Aging is a natural process resulting in the decline of tissue functionality. In the skin, aging results from degenerative changes in dermal and epidermal compartments. However, the molecular mechanisms behind skin aging are not fully understood. One of the poorly explored characteristics of the aged skin is the reduced number of anchoring fibrils, which connect the dermis to the epidermis, and the reduced level of their main protein component, type VII collagen (COL7A1), which is critical for the stability of the extracellular matrix. Interestingly, the skin of patients with recessive dystrophic epidermolysis bullosa (RDEB), a severe skin fragility condition caused by biallelic mutations in the gene COL7A1, shows clinical similarities with the skin of the elderly. This suggests that COL7A1 deficiency promotes premature aging in RDEB patients. However, it is not clear whether the pro-inflammatory background of chronic non-healing wounds is the only cause of premature skin aging in RDEB patients or functional COL7A1 plays an important, currently unknown role in protecting the skin from aging. My preliminary proteomic analysis of primary patient derived fibroblasts (COL7A1-/- PPF(s)), as well as patient specific organoid derived fibroblasts (iF(s)) generated from primary fibroblasts transitioned through induced pluripotency that were either uncorrected (COL7A1-/-) or genetically corrected using CRISPR/Cas9 (COL7A1+/-), revealed COL7A1 dependent perturbations in insulin-like binding protein 2 (IGFBP2) with COL7A1- /- PPF(s) and COL7A1-/- iF(s) showing marked deficiencies in its expression relative to genetically corrected iF(s). Therefore, I hypothesize that COL7A1 has a moonlighting role regulating fibroblast secretory profiles, and, in its deficiency, leads to accelerated cellular senescence and aberrant perturbations in fibroblast functionality, specifically through the IGF axis. Inhibition of IGFBP2 has been shown to lead to an overexpression of p21, p16, and p19, all powerful promotors of senescence and a potential mechanism by which premature acceleration senescence is induced in COL7A1 deficiency. During my fellowship, I will dissect the role of COL7A1 in senescence of skin cells. In aim 1, I will further analyze COL7A1 dependent perturbations in the proteome and secretome of COL7A1 deficient and COL7A1 corrected lines that are generated from induced pluripotent stem cells. Reprogramming into iPSCs erases aging associated marks in cells that arose from pro-inflammatory and fibrotic background and allows us to generate and characterize COL7A1 deficient and corrected cell lines without the influence of external factors that can mask the role of COL7A1 in skin aging. In aim 2, I will assess other age-associated marks in COL7A1 deficient and corrected cell lines to validate the connection between IGF signaling and COL7A1 functionality. If successful, this proposal will result in our better understanding of the role of COL7A1 in protecting the skin from aging and in developing novel therapies for RDEB and anti-aging interventions.

NIH Research Projects · FY 2025 · 2025-09

ABSTRACT Bias in pulse oximetry accuracy is associated with significant delays in care and unrecognized eligibility for therapeutics among patients with darker skin pigmentation. However, very few pulse oximetry studies have incorporated direct measures of skin pigmentation, and none have incorporated objective measurements. This is a problem because the bias of modern pulse oximetry devices is believed to be related to skin pigmentation. Most studies on pulse oximetry bias are retrospective and rely on self-reported race and ethnicity. We know pulse oximetry bias contributes to worse outcomes among non-White hospitalized patients, disproportionately impacting Black and Hispanic patients. However, significant variance in skin pigmentation exists within racial and ethnic groups. Therefore, race and ethnicity are poor surrogates for skin pigmentation. Further, race and ethnicity are social constructs that do not address the root cause of pulse oximeter bias: skin pigmentation. In 2024, the US Food and Drug Administration (FDA) recommended directly measuring skin pigmentation when evaluating pulse oximeters. Accordingly, there is a critical need to prospectively determine the effects of directly measured skin pigmentation on pulse oximeter performance. To address this gap, we will test our overarching hypothesis that darker skin pigmentation will be associated with increased pulse oximetry bias. Pulse oximeter performance, as defined by the correlation between a non-invasive/continuous pulse oximeter (SpO2) and an invasive/intermittent laboratory measure of arterial oxygen saturation (SaO2), is sub-optimal among patients with darker skin pigmentation. Landmark studies demonstrated that non-White patients experience a greater incidence of hypoxemia (SaO2<88%) compared to White patients when SpO2 values are normal (i.e., unrecognized hypoxemia). Unrecognized hypoxemia increases the risk of cardiac arrest, organ dysfunction, and is associated with an up to 3-fold increase in mortality among hospitalized patients. Skin pigmentation may affect the accuracy of SpO2 measurements by introducing error from systematic bias (e.g., consistently higher SpO2 values at a given SaO2). However, retrospective studies can neither directly measure skin pigmentation nor collect arterial blood gases for paired SpO2-SaO2 comparisons. We will leverage data from our ongoing, federally funded SAVE-O2 AI multicenter clinical trial (NCT06374225), comparing a closed- loop autonomous oxygen titration device versus usual care in 300 acutely ill adult patients at three US tertiary care hospitals. We are directly measuring skin pigmentation in all patients via two validated scales plus an objective spectrophotometer. We will use data from the SAVE-O2 AI trial to determine the association between skin pigmentation and pulse oximetry bias (Aim 1). We will then create a prediction tool incorporating skin pigmentation to establish personalized SpO2 targets for hospitalized patients (Aim 2). These specific aims will generate preliminary data to support a clinical trial that will address our long-term goal of validating personalized SpO2 targets based on skin pigmentation to mitigate the risks of unrecognized hypoxemia.

NIH Research Projects · FY 2025 · 2025-09

PROJECT SUMMARY The lungs are exposed to billions of potentially harmful particles daily. Robust defense is crucial, but protection must cause minimal physiologic disruption. Accordingly, a thin and easily transported layer of mucus lines the airways in health. In asthma, mucus aggregates on airway surfaces and obstructs airflow, but this is not effectively treated with existing therapies. We seek to close this gap by building a better mechanistic understanding of how mucus function and dysfunction are mediated by the chief macromolecules in airway mucus--MUC5AC and MUC5B. Their importance is well-recognized through studies in humans and animal models. MUC5B is required for mucociliary clearance, host defense, and survival in both species. Muc5ac is dispensable in healthy mice, but in human asthma and in mouse models, it causes airway obstruction. Both mucins also have host defense and pathogenic roles in non-respiratory tissues. Accordingly, despite clear significance, there is a need to find treatments that prevent mucus dysfunction while also preserving defense. To investigate this, we are focusing on the defining characteristic of mucins – heavy O-glycosylation – and how MUC5AC and MUC5B glycosylation is carried out in the Golgi apparatus. Mucin glycosylation terminates when sugars such as fucose (Fuc) are attached. This terminal fucosylation process in mucins is mediated by the enzyme fucosyltransferase 2 (FUT2), which catalyzes transfer of Fuc to galactose (Gal) acceptors forming Fuc(α1-2)Gal bonds. FUT2 absence is protective in people with asthma and in mouse models where its deficiency protects against mucus plugging and obstruction. Fucosylation increases mucus viscoelasticity, transport, and aggregation, and it is associated with excessive mucus viscoelasticity, and asthma severity. Accordingly, a mechanistic understanding of how FUT2 mediates mucin fucosylation will reveal novel strategies that could be used to target mucus dysfunction in asthma. We hypothesize that trans Golgi- localized FUT2-dependent mucin fucosylation mediates allergic airway mucus dysfunction. We will test this in three Specific Aims that 1) test the hypothesis that mucin α1,2-fucosylation requires FUT2 localization and activity within trans Golgi compartments; 2) test the hypothesis that conserved catalytic sites and non-catalytic regions in FUT2 are required for mucin α1,2-fucosylation; and 3) test the hypothesis that Golgi specific FUT2- dependent α1,2-fucosylation mechanisms mediate mucociliary dysfunction and mucus obstruction. Our focused investigations could reveal ways to target the important, but currently untreatable, problem of mucus hypersecretion while also helping resolve unanswered basic science questions.

NIH Research Projects · FY 2025 · 2025-09

PROJECT SUMMARY/ABSTRACT This project seeks to efficiently and effectively obtain input regarding important immunization issues through nationally representative surveys of healthcare professionals (HCPs) and the public. This initiative includes several innovative elements: (1) the ability to survey a diverse range of HCPs who play key roles in vaccine administration and policy implementation, (2) the integration of nationally representative public voices to inform recommendations, and (3) a novel feedback mechanism that provides participants with accurate vaccine information to address knowledge gaps. The proposed study team – the Vaccine Policy Collaborative Initiative (VPCI) – has extensive experience in immunization-related survey research among both HCPs and the public. We bring deep and broad knowledge and experience with immunization policy and delivery, including current and past ACIP membership, with several of our investigators currently serving on one or more ACIP Work Groups. We will collaborate closely with CDC technical advisors to identify critical survey topics, and additionally elicit input from national groups of advisors—an Expert Advisory Committee and Community Advisory Committee—as well as from a Public Advisory Committee. The team will conduct a minimum of four surveys per year among HCPs and/or the public with the ability to provide preliminary data to CDC within two weeks of survey launch. Leveraging WebMD's Medscape subscriber network, we will survey HCPs, including primary care physicians, obstetricians, subspecialists, pharmacists, and advanced practice providers, to assess their knowledge, attitudes, and barriers related to vaccination. Simultaneously, the Understanding America Study (UAS) will allow us to gather real-time insights from the public, including parents, pregnant women, persons living in rural areas, and adults at high risk of severe disease, to understand their vaccine perceptions, hesitancies, and access challenges. Findings will be rapidly disseminated to policymakers, professional societies, and public health organizations, with results presented to CDC, ACIP, and relevant stakeholders within weeks of survey completion. Based on our prior work, we expect to produce an average of five peer-reviewed manuscripts per year, each within 6-12 months from final data collection. This project will advance knowledge and understanding of vaccine policy-related issues and directly impact important policy decisions regarding vaccination of the US population.

NIH Research Projects · FY 2025 · 2025-09

PROJECT SUMMARY / ABSTRACT The study of physiology is at the core of modern biomedical research, which more than ever relies on integrating fundamental concepts of whole animal and organ physiology with sophisticated molecular and cellular approaches to investigate important questions related to human health and disease. Understanding mechanisms underlying the function of various systems in the body that contribute to both normal and pathological physiology is the fundamental prerequisite for all applied research in biomedical sciences allowing scientists to make important and translationally relevant discoveries. The Integrated Physiology PhD Program (IPHY) on the University of Colorado Anschutz Medical Campus (CU AMC) is a multidisciplinary PhD training program that prepares students for careers in biomedical research. Students in Integrated Physiology have opportunities to explore how molecules, cells, organ systems, and organisms regulate complex physiological functions through the integration of molecular, cellular, and physiological mechanisms. Our goal for this T32 program is to provide interdisciplinary training in areas focused on (1) Metabolism, Nutrition, and Obesity; (2) Cardiovascular, Pulmonary, Renal, and GI Physiology; and (3) Reproductive Physiology in an inclusive environment to support a diverse group of IPHY students. Our students and faculty are united by common interests in the understanding of the physiology of human health and disease; this program is designed to train students to become the next leaders contributing to innovative and groundbreaking scientific discoveries. We will accomplish our goal through the following objectives: (1) Improve the national need for interdisciplinary systems biology biomedical research focused on human health and disease; (2) Fill a campus-wide need to advance training in biomedical integrated physiology and translational science at the University of Colorado Anschutz Medical Campus; (3) Provide student-oriented well-balanced training and career development opportunities; (4) Develop a diverse and inclusive training environment focused on career development. To accomplish these goals, we will establish a program named the Physiology of Human Health and Disease (PHHD) Training Program that will build upon the success of the current IPHY PhD program. The PHHD T32 Training Plan will emphasize five specific areas that will allow trainees to develop professional skills, including project management, leadership, teamwork, and communication, including: i). Knowledge skills; ii). Innovative Technical Research skills; iii). Communication skills; iv). Professional skills; and v). Training in Diversity and Inclusion. The overall vision is to support predoctoral students in an inclusive and diverse environment that facilitates opportunities for a future generation of physiologists to obtain training in integrative and translationally relevant research focused on human health and disease, to foster creative, independent thinking combined with strong communication skills, and to provide career development, professionalism, and leadership opportunities.

NIH Research Projects · FY 2025 · 2025-09

Abstract The folding of many proteins occurs co-translationally and involves the orchestration of numerous cellular factors, including a large set of ribosome-associated proteins and chaperones. In the co-translational folding process, the folding is intimately coupled to translation. If the ribosome elongates too fast or stalls for too long at the wrong place and time, folding kinetics can be severely perturbed, leading to protein misfolding and/or aggregation. In extreme cases, this can lead to diseases. A better understanding of protein co-translational folding kinetics in the native context of translation is therefore critical to human health. The major challenge in the field is the lack of spatiotemporal resolution needed to track and quantify co-translational folding in a living intracellular environment. To address this challenge, we have developed a novel co-Translational Folding Tracking (coTFT) technology that enables us to directly visualize co-translational folding with single mRNA precision and sub- second resolution in living cells. With coTFT, we will focus on investigating co-translational folding in living cells. Specifically, we will quantify the kinetics of co-translational folding occurring in diverse intracellular environments, such as in cytosol and on the surfaces of organelles, to identify any location heterogeneity in co-translational folding as well as heterogeneity in mRNAs in each location. Then we will further investigate an ongoing question in the protein co-translational folding field – how alterations in translation elongation rate affects folding. Moreover, we will develop a novel method that specifically labels and isolates the short-lived co-translational folding complexes with high spatiotemporal resolution, which is challenging to achieve with current technologies. We will apply this method to investigate the interactome that is highly specific for co-translational folding in diverse subcellular environments. For discovered novel interactors, we will study their impact on co-translational folding using our established coTFT technology. The discoveries from this study will help us further understand co- translational folding in a native intracellular environment. In addition, our developed technologies are highly adaptable. In the long run, we expect to adapt our technologies to study a group of disease related transmembrane proteins and discover general as well as target-specific regulation mechanisms of co- translational folding. The discovered novel regulation mechanisms have the potential to lead to new therapeutics that have never been explored before for protein misfolding-related diseases.

NIH Research Projects · FY 2025 · 2025-09

SUMMARY To optimize behavior, animals must balance responding to important stimuli with an ability to ignore less critical stimuli. Behavioral thresholds provide a mechanism for this: animals respond to stimuli that exceed a particular threshold and ignore stimuli below the threshold. Palmitoyltransferases (or PATs, of which there are 24 in mammals) catalyze a reversible post-translational lipid modification and are emerging as key molecular regulators that tune behavioral responses. That they are important for human health is exemplified by their association with multiple disorders impacting behavior: schizophrenia, Alzheimer’s disease, and Huntington’s disease to name a few. Despite their ubiquity and importance, major questions regarding PAT function in vivo remain unanswered. Using larval zebrafish, we recently showed that the PAT enzyme Hip14 regulates behavioral thresholds for acoustic stimuli and the ability to tune thresholds through plasticity mechanisms such as habituation. This provides a tractable system to address how Hip14 specifically, and PATs more broadly, function in vivo to regulate behavior and ideally positions us to address the following open questions: How does PAT canonical enzymatic function contribute to behavioral plasticity? How PATs contribute to behavior has not been systematically examined in vertebrates. We will use high-throughput assays to rapidly identify which PATs are crucial for regulating behavior and behavioral plasticity in vivo using the larval zebrafish. In parallel, we will examine which palmitoylation substrates act downstream by generating point mutations in key palmitoylated residues and examining behavior. How is PAT substrate specificity regulated? Most PATs are expressed in the brain, but each has a unique expression pattern. Moreover, PATs exhibit substrate specificity, but the underlying mechanisms are not known. We will investigate the extent to which PAT protein sequence versus localization contribute to substrate specificity using structure-function and tissue-specific overexpression experiments. In parallel, we will use BioID to probe how localization and domain structure influence binding partners. Do PATs have enzymatic-independent functions in vivo? The best understood molecular function for PATs, including Hip14, is catalyzing the post-translational attachment of fatty acids to target proteins. However, non-enzymatic functions for Hip14 and other PATs have been identified in vitro. For example, Hip14 can function as a Mg2+ channel, and our pilot data indicate that Hip14 can regulate behavior even when its catalytic (palmitoyltransferase) domain is mutated. We will explore non-enzymatic functions for Hip14 and interrogate whether other PATs function in vivo as cation channels to regulate behavior. To answer these questions, we use zebrafish behavior as a readout. Successful completion of these projects will uncover basic mechanisms through which palmitoyltransferases function in vivo and provide new insights into how these key enzymes regulate behavior.

NIH Research Projects · FY 2025 · 2025-09

Abstract This proposal seeks support for a conference series dedicated to improving the outcomes of older adults by addressing the complexities of enhancing patient mobility within skilled nursing facilities (SNFs). Mobility, a component of physical function, describes the ability to move and has a significant impact on older adult’s health, independence, and quality of life. Despite evidence indicating that inactive older adults can benefit from engaging in mobility and movement at any intensity, those receiving care in SNFs have higher average sedentary time compared to individuals in other care facilities. Thus, mobility interventions are currently most needed in SNFs and are most likely to demonstrate value. Aligned with the unique theme of prioritizing the mobility of older adults, our aims are: 1) to develop and advance a collaborative research, policy, and care delivery improvement agenda to explore and develop innovative approaches for enhancing patient mobility and outcomes in SNFs 2) to develop and support workgroups during the five-year conference period to identify and enhance clinical adoption of priority mobility initiatives in SNFs, and 3) to generate and disseminate knowledge and approaches for adoption to enhance patient function, movement, mobility, and outcomes through national, regional, and local networks. The innovative conference series will consist of in-person events and quarterly structured and facilitated virtual workgroup meetings. This combination will maintain momentum for implementing strategies to promote mobility in SNFs and will facilitate ongoing progress and collaboration. The first conference will explore barriers and challenges to promoting mobility in SNFs to set the stage to identify innovative solutions that effectively address these barriers. This conference will set the landscape for collaboration and discussion regarding: 1) national policy, 2) clinical & research challenges/opportunities, 3) innovations, 4) operational considerations, and 5) implementation strategies. The second conference focuses on identifying promising implementation strategies, and the focus of the third conference shifts to implementation, dissemination, and actionable strategies. We will employ rigorous evaluation methods for each conference and the conference series overall. We will also develop research priorities and produce a white paper to comprehensively document the challenges and solutions highlighted during the conference. The innovation of this initiative lies in the intersection of all this expertise and resources assembled to address the complexities of enhancing patient mobility within SNFs. Through the conference series and engagement of representative stakeholders, we will align incentives to drive innovation for practice-based adoption. These novel advancements can improve patient experiences and outcomes for this vulnerable population. Furthermore, the strategies developed hold promise for broader application in other healthcare settings, underscoring the scalability of our interventions beyond SNFs.

NIH Research Projects · FY 2025 · 2025-08

PROJECT SUMMARY This is an exploratory developmental grant aimed at the design, development, and implementation of a future national, interdisciplinary, collaborative, team science research network that will advance the study of the exposome in autoimmune diseases (Exposome in Autoimmune Diseases Collaborating Teams, EXACT). We propose to plan a study of the exposome during the pre-clinical period of rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE). We assert that it is important to measure the exposome prior to the development of disease to ensure that what we are measuring precedes the disease and is not a consequence of the disease (which might happen if the exposome were measured in diseased individuals). This will allow us to draw inference about the temporality and potential causality of exposures on the risk of RA and SLE. This can be accomplished using prospective cohorts of at-risk individuals that are followed for the development of clinical disease (ie classifiable RA and SLE) with which we have a long history of working. Investigators at the University of Colorado Anschutz Medical Campus (Norris, Holers and Abraham) will work with investigators at the Oklahoma Medical Research Foundation (James and Munroe) to conduct a landscape analysis of relevant existing 1) resources, populations, databases, registries, repositories and consortia, 2) data on relevant exposures, 3) exposome laboratories, and 4) exposome analytic techniques. This will provide a background that we will use to plan activities for a new project to study the exposome in RA and SLE. As part of the planning process, we will hold a summit of exposome experts and investigators with cohorts/populations of individuals at-risk for RA and SLE, to develop and expand existing research collaborations to conduct a prospective study in the exposome of these diseases. We will use our existing novel cohorts of at-risk individuals who have been followed to the development of classifiable RA and SLE to explore feasibility of harmonization of questionnaire and biomarker data and conduct secondary data analysis to support the hypothesis and research strategy of the future EXACT Network. We will engage stakeholders such as scientific or professional associations and patients and patient-advocates in the areas of RA and SLE for better translation to inform the EXACT Network research agenda. The overall goal of this proposal is to plan research strategies and develop partnerships, infrastructure, and capabilities needed to address the major goals of a future collaborative EXACT network and to develop a research framework and strategies to support coordination among studies, collaborative research projects, and sites. Specific Aim 1. To conduct a multipronged approach to determine the needs and currently available resources to prospectively and effectively measure the exposome in RA and SLE (via a landscape analysis). Specific Aim 2. To design a prospective study to examine the role of the exposome in the etiology of RA and SLE, with an interdisciplinary research team, rigorous exposome measurements and novel analytic approaches.

NIH Research Projects · FY 2025 · 2025-08

PROJECT SUMMARY/ABSTRACT The goal of the proposed K23 Career Development Award is to provide the PI with the mentorship, knowledge, and skillset to develop into a patient-oriented independent investigator studying scalable, accessible behavioral and mind-body interventions for youth with migraine and other pain conditions. The mentoring team will provide the PI with training in randomized clinical trials, implementation science, and mind-body interventions. The training is integrated in a research proposal and planned clinical trial evaluating a remotely-delivered mindfulness-based intervention (MBI) group program adapted for adolescents with migraine. Migraine is the second leading cause of disability worldwide. Negative affectivity (depression, anxiety) is associated with worse pain-related disability and is particularly prevalent in the adolescent developmental period. Existing pharmacological and behavioral interventions for adolescent migraine do not target or change negative affectivity, are difficult to access, and for pharmacological intervention, are often no more effective than placebo. Scalable and accessible approaches that target negative affectivity may yield more efficacious treatment and reach more patients in need. MBIs hold promise to consider these needs. However, minimal research has studied MBIs for adolescents with migraine and none have considered patient-centered adaptations or designing for dissemination principles. The PI’s preparatory work for this K23 focused on Phase I (Design) of the ORBIT Model for Developing Behavioral Treatments for Chronic Diseases framework. In this phase, the PI adapted and refined a virtual group-based MBI to meet the unique needs of adolescents with migraine in partnership with interest holders (patients, parents, providers). Now for ORBIT Phase II (Preliminary Testing), the PI proposes a pilot feasibility randomized trial to test the feasibility and acceptability of the adapted intervention, BREATHE- Headache (BREATHE-HA) and Enhanced Standard-of-Care (attention-matched virtual, group headache education program) in 72 adolescents with frequent migraine (Aim 1). Parallel implementation science work will assess potential facilitators and barriers to implementation of virtual group-based interventions (like BREATHE- HA) in tertiary care settings (Aim 2) to bridge the translational science-to-practice gap. If this pilot study meets pre-determined feasibility and acceptability targets, it will provide a strong platform for the PI to undertake an adequately powered, hypothesis-testing pragmatic clinic-based efficacy trial in a future R01. Together, this work will provide a critical foundation for the PI to advance efficacy of and access to evidence-based behavioral and mind-body interventions for youth with migraine and chronic pain.

NIH Research Projects · FY 2025 · 2025-08

PROJECT SUMMARY. Novel strategies are urgently needed to enhance the efficacy of exercise in improving physical function of older people with HIV (PWH). Our team has identified impairments in measures of physical function across several cohorts of middle-aged and older PWH. Exercise is the standard of care for prevention and treatment of impaired physical function in people without HIV, and we previously demonstrated that exercise also improves physical function in older PWH; however, the effectiveness of responses varied. The gut microbiome plays a pivotal role in the regulation of gut health, and exercise has been associated with changes in the gut microbiome and improvements in physical function measures in people without HIV, yet little is known in regards to the gut microbial features associated with an exercise-gut microbiome-physical function axis in older PWH. HIV-1- associated disruption of the gastrointestinal tract is characterized by perturbations in gut microbiota (dysbiosis), epithelial barrier breakdown and microbial translocation which, in turn, has been linked to systemic inflammation and the development of multiple age-associated co-morbidities. In our prior studies of older adults with and without HIV, we established that relationships between the gut microbiome and relevant outcomes (systemic inflammation, physical function) differ based on HIV serostatus. Moreover, systemic levels of intestinal fatty acid binding protein (IFABP), a biomarker indicative of gut epithelial barrier damage, were reduced in older PWH following an exercise intervention. In preliminary studies, we found that in older PWH, higher levels of myostatin, a muscle biomarker associated with the negative regulation of skeletal muscle growth, significantly associated with higher systemic levels of IFABP, and myostatin levels were reduced following a supervised exercise intervention. Similar observations were not observed in people without HIV. Taken together, our observations suggest a unique exercise-gut microbiome-physical function axis also exists in older PWH. The central hypothesis of this proposal is that a supervised exercise intervention will lead to changes in the gut microbiome of older PWH with associated improvements in gut epithelial barrier integrity and reduced microbial translocation, ultimately resulting in improved muscle function and physical function outcomes. These ‘health-positive’ changes in the microbiome will be observed to a lesser extent (if at all) in participants with limited improvements in physical function. We propose to determine the impact of a 16 week supervised exercise intervention on the structure and function of fecal microbial communities in older PWH with viral suppression (Aim 1), and to identify microbial and metabolomic features associated with epithelial barrier damage, microbial translocation and muscle health (Aim 2), and with physical function outcomes (Aim 3). Identification of specific gut microbiome features will provide the groundwork for the effective design of future therapeutic trials that enhance the efficacy of exercise in improving physical function of older PWH.

NIH Research Projects · FY 2025 · 2025-08

Project Summary Despite advances in the treatment of rheumatoid arthritis (RA) with biologic therapies, a substantial number of patients do not have adequate disease control and specifically fail commonly used treatments. This suggests an unmet need for new and personalized therapies for RA. Current studies have found that tissue macrophages in the synovium of patients with RA can interact with complement factors, a complex innate immune system that plays critical roles in rheumatic diseases by promoting inflammation and disrupting tissue homeostasis. Specifically, our recent work has revealed that the MERTK+ tissue macrophages are expanded in lymphocyte- low synovial RA tissues and display multiple complement factors, including the receptors C5AR1 and C3AR1. In particular, we observed that a new MERTK+ macrophage subtype we refer to as the MERTK+HBEGF+ tissue macrophage is marked by high C5AR1, TNF, and CXCL2 and CXCL3 expression. Through experimental validation, we further found that the addition of an inhibitor of C5aR1 suppressed the inflammatory and interferon response pathways while upregulated tissue protective phagocytic programs. Thus, we hypothesize that modulating specific complement pathways in the MERTK+ tissue macrophages could improve homeostatic synovial macrophage function in this apparently dysregulated subtype, and further relieve deleterious pro- inflammatory cytokine and other mediator signals propagated by neighboring synovial fibroblasts. Herein, we propose to determine the complement component signatures and responses for the MERTK+HBEGF+ tissue macrophages (Aim 1), define how complement pathway crosstalk impacts synovial macrophages and fibroblasts (Aim 2), and evaluate how macrophage phagocytic function is enhance by complement pathway modulation (Aim 3). Together with our established synovial RA tissue cohort, developed single-cell omics integrative methods, and ex vivo and in vivo experimental assay and macrophage-fibroblast co-cultured system in place, these aims will provide novel insights into a new complement pathway-modulated macrophage target population that upon rewiring may restore tissue homeostasis and suppress tissue inflammation (Significance). Our proposed work will also generate new computational-experimental integrative methods and single-cell spatial transcriptomic approaches that are urgently needed for translational research toward personalized treatments of RA and other autoimmune diseases that are revealed through emerging single-cell technologies (Innovation).

NIH Research Projects · FY 2025 · 2025-08

PROJECT SUMMARY Loneliness and is a significant health issue for older adults and is a major risk factor for morbidity mortality, including for older adults with Alzheimer's disease and related dementias (ADRD). Loneliness interventions are a promising strategy to improve health outcomes in older adults with ADRD. As care t he frontline entry to health care for most older adults in the US, primary is an ideal setting to intervene on loneliness.Unfortunately, there is a lack of loneliness interventions tailored to older adults with cognitive impairment in the primary care setting. This proposal aims to address this gap by adapting an evidence-based creative storytelling intervention for older adults with cognitive impairment, called TimeSlips, for the primary care setting. aims older 0), (NIH Time2Connect, support has coursework, human-centered Mullen experience and funded Guided by the NIH Stage Model for Behavioral Intervention Development, the study are: 1) Assess potential psychosocial mechanisms of loneliness in community-dwelling adults with a high prevalence of mild cognitive impairment (MCI) or mild ADRD (NIH Stage 2) Adapt the TimeSlips intervention for older adults with MCI or mild ADRD in primary care Stage 1A), and 3) Conduct a pilot randomized trial of the adapted i ntervention, called for older adults with MCI or mild ADRD (NIH Stage 1B). This proposal will the research and career development of the candidate, Dr. Rebecca Mullen . Dr. Mullen developed a detailed career development plan consisting of mentorship, didactic and hands-on experience t o expand her knowledge in mixed methods research, design for intervention adaptation and development, and clinical trials. Dr. has assembled multidisciplinary team of mentors with extensive clinica l research and content expertise to ensure her success in achieving the stated research aims career goals. This K23 proposal wil l allow Dr. Mullen to transition to an independently primary care physician-scientist a with expertise in loneliness intervention to improve the health outcomes for older adults.

NIH Research Projects · FY 2025 · 2025-08

Summary Abstract Chronic pain is highly prevalent (38%) condition in older adults and impacts quality of life with increased depression, anxiety, existential distress, and social isolation, and associated with an increased risk in overall morbidity and mortality. Existing pharmacologic therapies for older adults have limited efficacy and significant side effects. Psychedelic Assisted Therapy (PAT) has demonstrated preliminary safety and efficacy for improving mental health (e.g., depression, existential distress), addiction, and chronic pain in non-elderly adults. However, published research to date has not included older adults who have been severely under- represented in studies to date. This lack of safety and efficacy data in older adults is a critical public health need. To respond to this research gap, we propose the INSPIRE Network (INnovation in Science of Psychedelics with Inclusive Research in Elders), a geographically diverse, transdisciplinary consortium comprised of sites with established psychedelic research programs and expertise in geriatrics, chronic pain, psychiatry, palliative care, and multi-site clinical trials. Initial research will focus on psilocybin, due to its favorable safety profile and evidence for preliminary efficacy in related psychiatric conditions (e.g. depression, anxiety, existential distress), and on lysergic acid diethylamide (LSD), due to early evidence suggesting safety and efficacy in treating pain, as well as psychiatric and existential distress in advanced cancer pain syndromes. In the UG3 phase, we will evaluate the pharmacokinetic (PK) and pharmacodynamic (PD) characteristics of these two drugs in healthy older adults, with results informing selection of drug(s) and doses for the UH3 phase. In the UH3 phase, we will conduct two randomized trials of PAT in two mechanistically distinct pain conditions: (i) a predominantly nociplastic condition, chronic back pain, testing psilocybin combined with an emerging evidence-based therapy for nociplastic pain—Pain Reprocessing Therapy, and (ii) a pain condition with a substantial nociceptive element, cancer-induced bone pain, testing LSD combined with an existential psychotherapy. The overarching hypothesis for the UH3 phase studies is that participants randomized to intervention arm (moderate dose followed by high dose PAT) will have improved pain outcomes compared to the control arm (placebo followed by moderate dose PAT) at all study time points (4,8,12 weeks) with a primary outcome of pain interference at 8 weeks. In addition, general measures of mood, sleep, and wellbeing will be improved in the intervention arm compared to the control arm.

NIH Research Projects · FY 2025 · 2025-08

PROJECT SUMMARY/ABSTRACT Patients with chronic lung disease and comorbid cognitive impairment without dementia represent a large, high-risk group with poor quality of life. Patients with chronic lung disease experience premature aging, frequent hospital admissions, a high burden of symptoms, and poor quality of life. Patients with chronic lung disease are more likely to have comorbid cognitive impairment without dementia than the general population, and these patients have even worse quality of life. Serious illness communication is an iterative, complex process that includes conversations between patients and clinicians in which patients explore their values and priorities in order to ensure patients receive goal-concordant care.1 Thus serious illness communication with this high-risk group is of paramount importance. Prior studies of serious illness communication among patients with chronic lung disease have not characterized patients’ cognitive function or have excluded patients with cognitive impairment. Understanding how serious illness communication differs among patients with chronic lung disease with cognitive impairment without dementia compared to those with normal cognition is an essential first step in learning how to best communicate with this high-risk population. The overall goal of the proposed research is to determine how serious illness communication differs between patients with chronic lung disease with comorbid cognitive impairment without dementia compared to those with normal cognition. This will be accomplished in the following specific aims: 1) Determine differences in self-report of a serious illness conversation, completion of a medical durable power of attorney for healthcare, and completion of a living will among those who have lung disease and cognitive impairment without dementia compared to those with normal cognition, and 2) Determine patterns of communication which are associated with patient engagement in serious illness communication that occurred with patients who screened positive for cognitive impairment. For Aim 1, data from the Health and Retirement Study, a nationally representative study of approximately 20,000 adults >50 years, will be used. For Aim 2, a Conversation Analysis of serious illness communication between a patient and a nurse and/or social worker as a part of the intervention in a clinical trial will be performed to identify strategies leading to patient engagement.

NIH Research Projects · FY 2025 · 2025-08

Project Summary/Abstract Fibrosis is defined as excess deposition of extracellular matrix (ECM), resulting in tissue scarring and organ dysfunction. In the heart, fibrotic remodeling in the context of chronic comorbidities such as hypertension and metabolic disease is associated with increased passive myocardial stiffness and the development of diastolic dysfunction (DD), a contributor to the pathogenesis of a multitude of cardiac disorders, including heart failure with preserved ejection fraction (HFpEF). The adult heart contains resident cardiac fibroblasts (CFs), which, in response to stress, undergo a cell state transition to become activated fibroblasts, sometimes referred to as myofibroblasts. Activated CFs are characterized by transcriptional reprogramming that results in enhanced production and secretion of fibrotic ECM proteins. Despite the well-recognized roles of CFs in fibrotic remodeling of the heart, there are no targeted therapies to prevent or reverse the phenotypic conversion of these cells into an activated state. Epigenetic regulatory proteins, including those that ‘erase’ (histone deacetylases [HDACs]) and ‘read’ (bromodomain-containing protein 4 [BRD4]) acetyl-histones, have clearly been shown to control of CF activation. Acetyl-histone ‘writers’ (histone acetyltransferases [HATs]) have also been suggested to regulate ECM production by CFs, but much of the prior work with HATs in the context of cardiac fibrosis was performed with pharmacological tool compounds with suboptimal selectivity and potency profiles. Following up on a recent phenotypic high-throughput screening campaign, our unpublished data reveal that highly optimized, drug-like inhibitors of the p300 and CREB-binding protein (CBP) HATs profoundly and dose-dependently suppress agonist-induced CF activation. Inhibition of p300/CBP catalytic activity with A-485, or neutralization of the acetyl- histone binding bromodomain in these HATs using PF-CBP1, blocked transforming growth factor- (TGF-)- mediated activation of cultured murine CFs through a SMAD-independent, non-canonical mechanism. Additionally, inhibition of p300/CBP catalytic activity blocked CF activation and cardiac fibrosis in a mouse model. Remarkably, A-485 and PF-CBP1 are also able to dramatically reverse constitutive activation of failing human CFs, highlighting the translational potential of our findings. Mechanistically, we provide evidence to suggest that p300/CBP inhibition squelches formation of activating acetyl-histone marks on gene regulatory elements and results in transcriptional rewiring in CFs, at least in part, through inhibition of the forkhead box protein M1 (FOXM1) transcription factor. Three specific aims are designed to extend this new field of cardiac research and test the overall hypothesis that inhibition of p300/CBP prevents and reverses the epigenomic and transcriptional reprogramming that culminates in CF activation and pathological fibrosis of the heart.

NIH Research Projects · FY 2025 · 2025-08

PROJECT SUMMARY Binaural hearing allows localization of sounds and confers advantages in complex environments, workplaces, classrooms, etc., where competing speech, noise, and reverberation abound. Unfortunately, an increasing population, spanning infancy through elderly and of diverse etiology, experiences difficulty in such environments despite having normal audiometric thresholds. Such difficulties are a hallmark of auditory processing disorders, or listening difficulties (LDs), which refers to difficulties in processing sound in the central auditory system as demonstrated by poor performance, often specifically in binaural hearing tasks. LDs can emerge from noise exposure, temporary hearing loss, aging, autism, and neurodegenerative diseases. The consequences of LDs can be severe; in children, LDs impact speech and language learning and academic performance and in adults, quality of life, job performance, fitness for duty, etc. Regardless of etiology, a limitation in LDs is that clinical diagnosis is based on a cluster of symptoms, many of which overlap with other disorders such as attention deficit disorder, learning disabilities and language deficits. An additional barrier to diagnosis and development of future treatments to alleviate LDs is that neither the neural mechanisms nor the loci in the auditory system that produce these LDs are yet fully understood. We hypothesize that many LDs are attributable to reductions in peripheral temporal coding that in turn impact binaural processing at the brainstem level. We think binaural hearing is essentially ‘a canary in the coal mine’ for LDs that aren’t associated with audiologically-diagnosed hearing loss. Here we test the hypothesis that two hearing deficits, namely cochlear synaptopathy and extended high frequency hearing loss (EHFL), can cause LDs. The LDs resulting from these deficits are often called “hidden hearing loss” because the hearing impairment is typically hidden from standard audiological assessments. To better identify and treat subjects with such LDs requires understanding the mechanisms that lead to and therefore predict LDs. In this proposal, we hypothesize that noise-induced cochlear synaptopathy causes reduced monaural peripheral temporal coding leading to impaired binaural brainstem function and deficient binaural hearing abilities. The underlying contributions of synaptopathy to LDs are unclear, and there may be confounding/coexisting factors occurring following noise exposure. One suspected factor is loss of hair cells in extended high frequency regions of the cochlea, resulting in EHFL. Therefore, we also study the joint impact of synaptopathy and EHFL. Finally, based on preliminary data we explore the hypothesis that synaptopathy leads to activity-dependent demyelination, due to persistently reduced neural activity, in the brainstem circuits that support initial binaural processing. Because myelin is essential for the precise encoding of temporal information, we hypothesize that even subtle changes in myelin in these circuits can impair binaural processing and thus monaural temporal and binaural hearing ability.

NIH Research Projects · FY 2025 · 2025-08

PROJECT SUMMARY: Shigella flexneri is a gram-negative bacterial pathogen that causes diarrhea in humans. Essential for the disease process is the ability of the bacterium to invade epithelial cells lining the colon and to spread between the cells of the epithelium. In order to spread, S. flexneri requires actin-based motility to move to the cell periphery, it deforms the plasma membrane into a protrusion enabling it to push into neighboring cells, where the protrusion is actively engulfed and resolves into a double membrane vacuole that the bacterium escapes. Although the biological steps required for spread are well-established, the molecular mechanisms that enable them to occur are poorly understood. Our preliminary data establish that the short-chain fatty-acid butyrate promotes S. flexneri spread in a manner that is dependent on the actin-binding protein synaptopodin. Synaptopodin enhances the intercellular spread of S. flexneri and Listeria monocytogenes, indicating it may be generally required among bacterial pathogens that spread between cells. We show synaptopodin localizes to S. flexneri as they initiate protrusions, but it is largely excluded from the protrusion structure as it elongates. Synaptopodin recruitment to S. flexneri is dependent on the bacterial proteins IcsA and Spa15. These data highlight that synaptopodin appearance around bacteria is not solely attributable to its actin binding properties and show its spatial location around bacteria is regulated during protrusion formation. Yet, the manner of synaptopodin regulation and the requirement of synaptopodin during protrusion formation is unclear. We anticipate that synaptopodin forms a protein complex that promotes protrusion initiation, and to our knowledge, this would be the first such complex described for intracellular pathogens that spread. We hypothesize that S. flexneri infection alters synaptopodin function in a manner that enables the generation of forces to deform the plasma membrane and initiate protrusion formation. We aim to test this model with the following aims. AIM 1: To define the functional requirement of synaptopodin during S. flexneri infection. AIM 2: To determine how S. flexneri infection alters synaptopodin function. Several important human pathogens require intercellular spread to cause disease, the proposed approaches are likely to define a new mechanistic step that enables these pathogens to cause disease in humans by defining components of a synaptopodin-dependent protrusion initiation complex, its mechanisms of regulation, and its functional contribution to protrusion formation. We anticipate these studies may also resolve long-standing gaps in our understanding as to how pathogens that do not polymerize actin in the protrusion can generate force at the membrane to form protrusion structures. Moreover, synaptopodin is a poorly characterized protein that has not been described to have a role in infection; the investigations proposed here will provide fundamental mechanistic insights into the role of synaptopodin in cells and its requirement for bacterial pathogenesis.

NIH Research Projects · FY 2025 · 2025-08

PROJECT SUMMARY/ABSTRACT This K08 proposal presents a comprehensive five-year research and career development plan to address critical questions pertaining to impaired placentation associated with fetal growth restriction (FGR). As a pediatric and fetal surgeon, this research project, along with its accompanying didactic efforts will equip me with unique cross- disciplinary skills to facilitate my transition to independence as a physician-scientist in placental biology. The catalyst for this proposal is based on the rigor of previous research and our preliminary data in the field. FGR is a significant problem that carries a high rate of perinatal mortality and a high frequency of lifelong poor health. Pathology of fetal growth restricted placentas demonstrates reduced extravillous trophoblast invasion and poor feto-placental angiogenesis. In addition, microRNAs, which are short non-coding RNAs, have been implicated in the pathogenesis of FGR. They can be exported from one cell to communicate with and induce effect on other cells via small transporters known as extracellular vesicles (EVs) and are critical for normal placentation. Based on these data, our central hypothesis is that anti-angiogenic miRNAs within human extravillous derived EVs inhibit extravillous trophoblast invasion and feto-placental endothelial angiogenesis through paracrine signaling, resulting in FGR. In Aim 1, we will model the metabolic stress associated with FGR by culturing extravillous trophoblasts in a nutrient restricted media. The miRNA cargo within extravillous trophoblast-released EVs will then be sequenced to determine the effect nutrient-restriction has on anti-angiogenic miRNA expression. In Aim 2, nutrient-restricted EVs derived from extravillous trophoblasts will be incubated with extravillous trophoblasts and microvascular endothelial cells (MVECs) to determine if their cargo impairs trophoblast invasion and placental feto-placental angiogenesis, respectively. Lastly, in Aim 3, we will utilize a blastocyst trophectoderm transduction model to upregulate anti-angiogenic miRNAs only within the placenta. We have previously identified miRNAs that are upregulated in mouse FGR placental tissue and will determine their effects on implantation and fetal growth in this novel in vivo model of pregnancy. The proposed experiments will impact multiple clinical disciplines including obstetrics, neonatology, and fetal care by providing mechanistic data relevant to the role of miRNAs in placental pathologic changes evident in FGR. This foundational work will provide an excellent vehicle for my maturation into an independent investigator. Experiments will be performed in an environment with an established history of successful mentorship of junior faculty to independence. With the support of this application, I will 1) advance my technical skills (RNA- sequencing, invasion assays, angiogenic assays, and blastocyst transduction) and 2) learn advanced molecular principles and biostatistics and improve my mentoring and writing skills.