University Of Colorado Denver

NIH Research Projects · FY 2025 · 2025-09

Project Summary Technological advances like genome sequencing, RNA interference (RNAi), and CRISPR have transformed functional genomics by speeding up genetic discoveries and enhancing our understanding of diseases and personalized medicine. Dr. Molishree Joshi, the PI of the proposal and the Associate Director of the Functional Genomics Shared Resource (FGSR) at the University of Colorado Cancer Center (UCCC), has significantly contributed to this field by providing essential resources to academic labs in Colorado. UCCC, one of 71 NCI- Designated Comprehensive Cancer Centers, is funded by the NCI-P30 under Dr. Richard Schulick and supports four interdisciplinary research programs and twelve shared resources, including FGSR. Initially, FGSR functioned as a repository for small hairpin RNA (shRNA) from The RNA Consortium (TRC) library. Significant advancements began when Dr. Molishree Joshi became Manager and Scientific Consultant in 2013. As the sole full-time employee, she expanded FGSR's capabilities by strategically securing local funding and proactively integrating complementary technologies and instrumentation for automation. Today, FGSR supports three major platforms: shRNA, Open Reading Frame (ORF), and CRISPR, providing pre-made reagents and customized solutions. The addition of services to create custom CRISPR libraries, assistance with CRISPR screens, and gene editing in complex cancer cell lines has led to groundbreaking discoveries. Owing to Dr. Joshi’s relentless efforts, FGSR is one of the most utilized core facilities in Colorado; it has provided services to >360 academic labs across four campuses, contributed to >250 peer-reviewed publications, facilitated over 85 NCI-funded grants, and helped initiate several clinical trials. Dr. Joshi goes above and beyond, and she continues to innovate. The ongoing noteworthy developments include 1) Copy-number analysis and mutation detection in cancer cells and measurement of CAR-T cell dynamics in immunotherapy patients using digital PCR, 2) Robust protocol development for heterozygous gene editing in cancer cells and iPSCs, 3) Protocol optimization for single-cell and spatial CRISPR screens, 4) CRISPR editing in 3D cell cultures, 5) Repurposing of CRISPR-Cas9 technology to target tumors with genomic amplification and 6) Applying for the NIH S10 Shared Instrumentation Grant to procure CellCelector Flex for precise cell isolation. Dr. Joshi also contributes to education by giving seminars and teaching multiple graduate-level courses focused on genome editing and precision medicine on the Anschutz campus. Thus, Dr. Joshi’s scientific and administrative acumen have been the key to the evolution and success of FGSR. Dr. Joshi's outstanding contributions have garnered her the prestigious Outstanding Scientist Award from ABRF and a promotion to Associate Director of FGSR in 2023. UCCC is committed to championing both FGSR and Dr. Joshi’s career. The R50 award will be a game-changer, empowering Dr. Joshi to drive groundbreaking cancer research and ensuring a stable and impactful future for her work.

NIH Research Projects · FY 2025 · 2025-09

Project Summary/Abstract The technical and biologic expertise offered by Research Specialists provides a foundation of support that expedites the growth of research programs, shared resources, and cancer centers. Especially important in complex fields, such as the growing use of immunotherapeutics in personalized treatment of cancer, a comprehensive understanding of biologic responses requires cutting-edge technology and expertise that would be cost prohibitive to develop in individual laboratories. Monitoring immune responses in patients treated with immunotherapies is vital to understanding the mechanisms involved in these treatments and for the rational design of combinatorial clinical trials. This proposal demonstrates the need for support of the Research Specialist in the continued development of critical services provided by the Human Immune Monitoring Shared Resource (HIMSR), a University of Colorado Cancer Center (UCCC) Shared Resource at the University of Colorado Anschutz Medical Campus. The Research Specialist collaborates with UCCC members to support cancer-related investigator-initiated clinical trials with a full-service approach, from sample processing, to optimization and implementation of high parameter correlative assays for liquid and tissue biopsies, to primary data analyses. Continued support of the Research Specialist with this NCI R50 funding mechanism will allow time for the implementation of new technologies for spatial proteomic and transcriptomic imaging for use in the study of tumor microenvironments, development of novel computational approaches and improvements to existing computational tools for primary image analyses, and continued advancement of existing imaging technology with customized multiplex panels. Dr. Jordan's expertise and training in immunology and human clinical studies is a vital component of the Unit Director's Cancer Center Support Grant, many research programs of UCCC members, and to the completion of projects under the Head and Neck SPORE (NCI P50). As evidenced by rapidly expanding publications and funding opportunities that stem from its advanced techniques, the HIMSR has become an integral part of many research programs on campus and will be a vital component of the future growth of immunotherapies at CUAMC.

NIH Research Projects · FY 2025 · 2025-09

PROJECT ABSTRACT Pain and problematic alcohol use have substantial public health consequences and co-occur at high rates. Pain-related alcohol use is common and concerning; although alcohol use may provide short-term relief from pain, it may also worsen pain and alcohol use consequences over time. Understanding the factors that underlie whether physical pain leads to alcohol can inform intervention efforts that address both pain and problematic alcohol use. This project will help understand (1) potential pathways from pain to alcohol use and (2) factors that impact the pain-alcohol relationship over time. Specifically, this project combines laboratory-based pain induction and ecological momentary assessment (EMA – i.e., daily text-message surveys) to investigate two hypotheses of pain-related alcohol use. First, we hypothesize that pain increases negative affect (e.g., anxiety, anger), which then leads to alcohol use (i.e., a mediation effect). Second, we hypothesize that pain-related alcohol use is most prevalent among people with low capacity to modulate their experience of pain (i.e., pain modulatory capacity) and high impulsivity (i.e., moderation effects). To test these hypotheses, we will recruit 225 community adults (aged 25-80). At a baseline laboratory appointment, we will assess individual differences (e.g., negative emotionality, impulsivity), systemic inflammation (via circulating cytokines), and pain modulatory capacity during laboratory pain induction. After baseline, participants will complete a 30-day EMA protocol consisting of five daily assessments via smartphone app. Each smartphone survey will assess substance use, pain, craving, and negative affect. Leveraging these data, we will first test whether the pain-alcohol relationship is mediated by negative affect in daily life (Aim 1). We will also test whether the pain-alcohol relationship is stronger among people with lower pain modulatory capacity and higher impulsivity (Aim 2). Finally, we propose that pain-related alcohol use cannot be fully understood without considering biological age. Age is related to increased pain and chronic pain conditions; decreased negative affect, impulsivity, and pain modulatory capacity; and increased sensitivity to alcohol. Age is also associated with greater inflammation, and the two have similar associations with key facets of pain and alcohol use. It is unclear whether inflammation and aging are independent risk factors for pain and pain-related alcohol use. Thus, we will examine whether effects tested in Aims 1 and 2 vary as a function of age, and whether age and inflammation independently affect pain- related alcohol use (Aim 3). Findings across all aims will determine whether modifiable risk factors are promising intervention targets and whether the relevance of these factors varies as a function of age or inflammation.

NIH Research Projects · FY 2025 · 2025-09

Bronchopulmonary dysplasia (BPD), the chronic lung disease following preterm birth, is the most frequent complication of prematurity. Infants who develop BPD are at high risk for death and have substantial life-long morbidity. Despite decades of research, the biomechanical mechanisms that cause sustained lung development abnormalities after preterm birth and optimal strategies for the prevention and treatment of BPD remain poorly understood. Recent evidence suggests that the chronic respiratory failure of BPD may be due to disproportionate growth of lung compartments. Dysanapsis is the concept of disproportionate airway and distal lung growth. Prior work in experimental BPD has shown that intra-amniotic endotoxin causes dysanaptic growth of the airways and parenchyma in infant rats. It is unknown whether dysanaptic growth affects the trajectory of lung function over a lifetime. Given that vascular growth occurs in parallel with alveolarization during lung development, disruption of airway-airspace growth likely implies disrupted growth of the vasculature. This proposal will test whether there is persistent dysanaptic growth of airways, distal parenchyma, and vasculature in experimental BPD, and whether these structural changes contribute to persistent impaired biomechanics, pulmonary hypertension, and increased susceptibility to postnatal insults. First, we will determine whether intra-amniotic endotoxin causes dysanaptic lung growth that persists throughout postnatal lung development (Aim 1). Then, we will investigate whether impaired vascularization occurs in parallel with dysanaptic growth of the airways and parenchyma (Aim 2). Finally, we will test whether dysanaptic growth after intra- amniotic endotoxin increases the risk for worsened lung function trajectory after acute postnatal inflammation (Aim 3). In addition to expanding the understanding of interactions between airways, alveolar, and vascular growth and the long-term functional implications of dysanaptic growth, this K38 proposal would allow me to build scientific expertise early in my career. The K38 StARRTS program would provide the support and mentorship to transition from a resident physician in the R38 StARR program to a strong candidate for a future K08. My career development plan includes time to 1) advance my understanding and use of preclinical models to study lung structural development, 2) learn skills to design and perform experiments assessing vascular structure and development, 3) to perform robust and non-biased data analysis, and 4) develop strong written and oral communication skills.

NIH Research Projects · FY 2025 · 2025-09

Abstract Chronic Obstructive Pulmonary Disease (COPD) is a major cause of morbidity and mortality in the United States. We propose to integrate existing omics data (genetic, transcriptomic, epigenetic, proteomic, metabolomics, and spirometrics) from multiple TOPMed cohorts, medical imaging data (CT), wearable data (Fitbit and Apple), social determinants of health (SDOH), and clinical research data (text and tabular) to develop EMAI-COPD, a novel and open-source ethics-aware multimodal AI (MAI) model to enable a variety of downstream applications in preventive, diagnostic and therapeutic interventions in COPD. These include medical applications, such as disease progression and prediction modeling, disease subtyping and pathway analysis, and early exacerbation detection. We envision EMAI-COPD as both a blueprint and a starting model to be adopted and adapted to generate a family EMAI-X models for other diseases (i.e., “X”). Our team is well positioned to take on this task. The team constitutes of 1) AI experts with extensive research experience in introducing novel AI models as well as agile model development lifecycle for large- scale models, 2) domain experts with decades of research and clinical experience in COPD interventions, and 3) medical ethics experts that could help us form the ethics framework required to develop EMAI- COPD as an intrinsically ethics-aware tool, including awareness of bias, fairness, privacy, accountability and transparency. Along with a wide set of stakeholders, which have worked with the team in the past, this team will adopt an agile approach to co-design and enhance EMAI-COPD to meet all ethical and performance requirements of the model.

NIH Research Projects · FY 2025 · 2025-09

Despite launching in 2015 as an evidence-based and policy-supported cancer screening strategy, lung cancer screening using low-dose computed tomography has suffered a broad range of implementation challenges that have constrained its beneficial impact on individual and population health outcomes. Lung cancer screening constitutes one of the most significant missed opportunities to reduce cancer mortality. While the literature has identified numerous implementation challenges that explain the starkly suboptimal uptake and constrained impact, few comprehensive solutions have been developed and tested to address this devastating translational gap between the potential generated by the National Lung Screening Trial (NLST) and the ongoing failure to achieve more than a small fraction of the possible mortality-reducing impact. The Kentucky LEADS Collaborative™ developed the QUILS™ System to facilitate implementation of lung cancer screening by packaging implementation strategies and evaluating the QUILS™ System in 10 lung cancer screening programs across Kentucky. Not only did the lung cancer screening programs demonstrate significant quality improvements, but Kentucky also achieved some of the highest rates of lung cancer screening implementation in the country. Additional preparatory research has helped refine the original QUILS™ System, and the proposed research seeks to evaluate the impact of the QUILS™ System 2.0 on the scale-up and sustainment of high-quality lung cancer screening in a sample of 60 lung cancer screening programs. The overarching aims are to facilitate high-quality lung cancer screening and to understand the factors that impact the scale-up and sustainment of high-quality lung cancer screening. In the UG3 Phase, the investigative team will (1) engage and prepare 60 lung cancer screening programs, (2) co-refine the proposed research methods, and (3) co-refine the QUILS™ System to prepare for the UH3 Phase trial. In a three-group pragmatic randomized comparative effectiveness trial (UH3 Phase), Aim 1 will compare implementation strategy components of the QUILS™ System on QUILS™ Index scores (a measure of program quality) and reach (as measured by the QUILS™ Index Reach Domain). Guided by PRISM and its RE-AIM outcomes, Aim 2 applies mixed methods to evaluate (a) moderators (i.e., contextual factors) and (b) mechanisms of each arm’s implementation strategies’ effects on reach, adoption, implementation (including adaptations), and maintenance/sustainment among 300 LCSP clinicians and staff. Aim 3 involves a thorough economic evaluation of the cost-effectiveness of the three different combinations of QUILS™ implementation strategy components. Aim 4 co-develops a QUILS™ Implementation, Tailoring, and Sustainment Guidebook to consolidate learnings and prepare for future dissemination. This innovative and comprehensive quality system offers the potential to accelerate implementation of lung cancer screening and simultaneously improve outcomes and public health.

NIH Research Projects · FY 2025 · 2025-09

Inflammatory pain due to symptomatic osteoarthritis (OA) is one of the leading indications for opioid prescriptions prior to definitive treatment with surgery. The endocannabinoid (eCB) system modulates pain by regulating synaptic transmission and immune reaction/ inflammation peripherally and centrally, as well as interacting with ion channels and receptors. The rapid rise in cannabis legalization has allowed patients to self-medicate resulting in altered states of eCB synthesis and degradation, and receptor activation prior to seeking medical care. The clinical consequences of an altered eCB system due to exogenous cannabinoids are unknown. We propose a comprehensive, systematic and highly integrated multidisciplinary basic and clinical science approach to characterize the a) human eCB system response to chronic pain associated with osteoarthritis, b) acute pain directly post-surgery, as well as c) pain resolution or pain chronification in TKA patients 6-months post-surgery and d) to mechanistically study changes in eCB signaling in preclinical animal models. Our goal is to determine the role of the eCB system in the maintenance and resolution of inflammatory pain using a prospective observational clinical study in a cohort of cannabis users and non-users undergoing TKA to validate existing animal models. We will test the overarching hypothesis that a functional eCB system is necessary for inflammatory pain resolution. Aim 1 will determine the effect of cannabis use and chronic inflammatory pain on the eCB system by comparing cytokine and eCB/lipid signatures associated with pain in patients with chronic OA to healthy subjects without pain to validate animal pain models. Exogenous cannabinoid use will be assessed with self-report and verified by quantitative LC-MS/MS assays. A reverse translational approach will examine identified changes in the patients to mechanistically study effects on eCBs on pain modulation and synaptic plasticity in preclinical models of inflammation and surgical incision. In Aim 2 we will determine effects of acute surgical pain (TKA) on the eCB system in cannabis-users compared to non-users and assess mechanisms in preclinical pain models. Effects of cannabis use on eCB signaling and synaptic plasticity in will be examined in two major pain-processing regions such as dorsal horn (DH) of the spinal cord and periaqueductal gray (PAG). Aim 3 will follow patients for post-surgical pain outcomes and opioid/cannabis use with a validated, innovative dried blood spot-based home monitoring strategy as an objective outcome measure. Pre-clinical studies will assess changes in eCBs/lipids in a TKA/post-incision rodent pain model in the absence and presence of cannabis. The results of these studies will inform whether adaptations in the eCB system induced by cannabis use contribute to hyperalgesia and regression to chronic pain in patients undergoing TKA procedures. We will be able to validate and mechanistically qualify preclinical animal models of persistent inflammation and post-incision pain. Physicians will be better informed on effects of cannabis use on patient outcomes.

NIH Research Projects · FY 2025 · 2025-09

PROJECT SUMMARY The prevalence of stress-related disorders has surged globally. Although females are twice as likely as males to develop a stress-related disorder, they remain markedly underrepresented in preclinical research investigating stress resistance factors and their associated mechanistic pathways. Exercise is an important resistance factor with a broad range of stress-buffering effects that are not only highly conserved across species but develop more rapidly in females than males. Preclinical models demonstrate that voluntary wheel running (VWR) prevents the typical depressive- and anxiety-like outcomes of inescapable stress (IS) in rats, and importantly, females attain stress resistance more rapidly (3 weeks of VWR) than males (6 weeks of VWR). Exercise affords behavioral stress resistance at these timepoints by constraining the serotonergic (5HT) response in the dorsal raphe nucleus (DRN), however the mechanisms underlying this constraint are unknown. My preliminary data reveal a robust inhibitory pathway from the nucleus accumbens (NAc) to the DRN that is positioned to inhibit the activity of DRN 5HT neurons during stress and is primed by prior exercise. This proposal explores whether the NAc is critical for the protective stress effects of exercise, at least in part, through GABAergic projections to DRN 5HT neurons. My preliminary data reveal that 6 weeks of VWR in males potentiates IS-induced activity of DRN-projecting NAc neurons. Notably, the NAc displays differences in neural morphology and enhanced stimulus-evoked dopamine females, which could account for the accelerated acquisition of exercise-induced stress resistance in female rats. Aim 1 will examine whether prior VWR engages the NAc-DRN pathway during IS in both sexes, and whether this process occurs earlier in females compared to males, using viral tracing and fluorescent in situ hybridization. Given the temporal differences of stress resistance between VWR males and females, I hypothesize that prior exercise will increase IS-induced cfos in DRN-projecting GABAergic NAc neurons in males after 6 weeks and in females (but not males) after 3 weeks. Aim 2A will examine whether chemogenetic inhibition of the NAc-DRN pathway during IS will prevent exercise-induced resistance to the behavioral outcomes of IS, such as social avoidance and exaggerated fear. Aim 2B will utilize in vivo fiber photometry to examine whether chemogenetic inhibition of the NAc-DRN pathway during IS restores the DRN 5HT response to IS, despite sufficient exercise experience. Preliminary data suggest that males recruit multiple mechanisms for the stress-protective effects of exercise, such as IS- induced activity from prefrontal cortical neurons. In contrast, my preliminary data suggest that females rely on the NAc-DRN pathway as a single, distinct stress resistance mechanism. Therefore, I hypothesize that NAc- DRN pathway inhibition during IS will eliminate the protective effects of VWR on IS-induced 2A) behavioral outcomes and 2B) DRN 5HT activity in female, but not male rats. Collectively, this proposal aims to investigate the underlying mechanisms of exercise-induced stress resistance and the sex differences therein.

NIH Research Projects · FY 2025 · 2025-09

ABSTRACT Type 2 diabetes mellitus (T2D) is a common chronic disease associated with morbid and costly complications. New medication classes – glucagon-like peptide-1 receptor agonists (GLP1RA) and sodium-glucose cotransporter-2 inhibitors (SGLT2i) – have proven cardiovascular, renal, and glycemic benefits in randomized clinical trials (RCTs) in individuals with T2D. RCTs provide high quality evidence of efficacy yet leave several evidence gaps relevant to routine clinical care. First, while a well-conducted RCT has strong internal validity, the applicability of the results in real-world care – or external validity – can be uncertain, particularly for populations underrepresented in the RCT. Unfortunately, minoritized race and ethnicity groups and rural individuals bear excess burden of T2D and T2D complications but are underrepresented in recent RCTs of GLP1RA and SGLT2i. Second, most Phase 3 RCTs for GLP1RA and SGLT2i utilize a placebo control. As a result, there are little to no head-to-head data comparing efficacy and harms of GLP1RA and SGLT2i for cardiovascular and kidney endpoints – potentially valuable data for routine diabetes care. Finally, RCTs are designed to estimate an average treatment effect in the trial population but are typically underpowered to identify heterogeneous treatment effects or subgroups in which one treatment or another has particular benefit or risk of harm. Moreover, prediction models to guide T2D treatment selection may not generalize well across populations – similar to the problem of RCT external validity. The overall goal of this proposal is to bring together real-world data from diverse T2D populations from three health systems and data from pivotal RCTs of GLP1RA and SGLT2i to address each of these evidence gaps using biostatistical tools known as transportability methods. Transportability methods use weighting to balance participant characteristics between a clinical research study population (e.g., an RCT) and a target population (e.g., individuals with T2D from a population underrepresented in an RCT). This allows inference of what the study’s results would have been had the target population participated in the study. Importantly, these methods avoid several critical threats to validity of conventional comparative effectiveness research methods. In Aim 1, transportability methods will be used to extrapolate the efficacy and harm of GLP1RA and SGLT2i from recent landmark RCTs to representative real-world samples of T2D patients and to populations underrepresented in RCTs. In Aim 2, extensions of transportability methods will be used to estimate head-to-head effects of GLP1RA and SGLT2i for glycemic, cardiovascular, and kidney outcomes. Aim 3 will focus on developing prediction models to guide individualized selection of GLP1RA versus SGLT2i and utilize transportability methods to improve prediction model performance across diverse populations. In completing the Aims, the study will create a roadmap that describes best practices for designing and executing transportability analyses and will provide insights into how RCT and real-world data can be integrated to generate timely evidence relevant to all individuals with T2D.

NIH Research Projects · FY 2025 · 2025-09

PROJECT ABSTRACT Nontyphoidal Salmonella are responsible for about 180 million cases of diarrhea and 300,000 deaths a year globally. Very young and very old people, cancer patients, immunocompromised individuals coinfected with HIV, and chronic granulomatous disease (CGD) patients bearing inactivating mutations in the phagocyte NADPH oxidase are some of the populations that are at high risk from suffering disseminated nontyphoidal Salmonella infections. The high susceptibility of CGD patients to disseminated nontyphoidal salmonellosis attest to the role reactive oxygen species produced in the respiratory burst of macrophages play in resistance to this facultative enteropathogen. We still have profound gaps in knowledge regarding the mechanisms by which reactive oxygen species mediate antimicrobial activity. Reactive oxygen species directly damage a variety of biomolecules. We have made the unprecedented discovey that reactive oxygen species produced by host phagocytic cells exert potent anti-Salmonella activity by inducing expression of the Gifsy-1 prophage terminase. The Gifsy-1 terminase gains unprecedented tRNase activity upon oxidation of cysteine residues forming a structural zinc finger. The oxidation of Gifsy-1 terminase results in the redox- dependent cleavage of the anticodon loop of tRNALeu. This is a very unexpected finding if we consider that the canonical functions of phage terminases are to cleave phage DNA and load the resulting single genomes into preformed viral capsids. Cumulatively, our data support the hypothesis that the oxidation of redox active cysteine residues in the zinc finger of the Gifsy-1 terminase stimulates the formation of an oligomer with tRNase activity, thus sensitizing Salmonella to the phagocyte NADPH oxidase. Aim 1 will characterize the mechanism by which gpA senses oxidative stress. Aim 2 will identify the mechanism by which oxidized gpA gains tRNase activity. Aim 3 will reveal the determinants responsible for the selective cleavage of tRNALeu by oxidized gpA. The knowledge generated using a combination of synergistic biophysics, biochemistry, bioinformatics, bacteriology, as well as bacterial and mouse genetics will illuminate key aspects of Salmonella pathogenesis, while providing a new framework for understanding the strategies used by bacteriophages to subvert their bacterial hosts during the mammalian immune response. Terminases are highly conserved in evolutionarily distant phages and herpesviruses. Hence, the knowledge gained in our investigations may be applicable to phylogenetically diverse prokaryotic and human viruses.

NIH Research Projects · FY 2025 · 2025-09

SUMMARY/ABSTRACT Nursing homes play a crucial role in caring for individuals facing serious illness, including many individuals living with Alzheimer's Disease and related dementias and multimorbidity. Nursing home residents commonly experience burdensome symptoms, reduced quality of life, unpredictable disease trajectory and low-quality end-of-life care. Despite the widely recognized need to enhance palliative care for this vulnerable population, nursing home residents, including those with Alzheimer's Disease and cognitive impairment rarely receive such care. This is partly due to difficulties identifying residents with unmet palliative care needs. The primary objective of the proposed study is to address the national priority of improving palliative care for nursing home residents by developing an efficient, innovative, and resident-centered method to identify those with unmet palliative care needs, communicate these needs to providers, and stratify residents to the appropriate level of palliative care (primary palliative care, specialty palliative care) with focus on enhancing applicability for residents with Alzheimer's Disease and related dementias. Specifically, the candidate seeks to (1) evaluate the content and face validity and iteratively refine a palliative care screening tool; (2) develop contextual understanding, implementation strategies, and training materials for the effective deployment of a palliative care screening tool, customized to the specific needs of the nursing home setting; and (3) pilot test the screening tool and training in real-world clinical settings and evaluate the psychometric properties of the palliative care screening tool. To achieve these aims, the project will involve a Delphi study with an expert panel and cognitive interviews with nursing home staff. Co-design workshops will be used to develop contextually relevant and effective training materials and implementation strategies for deploying the screening tool in nursing homes, ultimately enhancing successful integration and impact. The pilot study will enlist nursing home staff to implement the tool, scoring the tool quarterly alongside routinely collected assessment data. The specific training objectives that will provide the knowledge and skills to complete the research aims are to gain expertise in: (1) community-engaged research (including Delphi methods), (2) implementation context and strategies, and (3) conducting research with patients who have dementia and nursing home clinical trials. The research environment at the University of Colorado features a number of high-quality, well- established resources that will facilitate Dr. Cole's development. This proposal will provide the foundation for future clinical trials that test the efficacy, effectiveness and implementation of palliative care screening interventions to enhance care and outcomes for older adults with serious illness, such as Alzheimer's Disease and Related Dementias, living in nursing homes.

NIH Research Projects · FY 2025 · 2025-09

PROJECT SUMMARY / ABSTRACT Individuals with Down syndrome (DS) experience health disparities that escalate with age. When pressed, clinicians echo the documented literature: clinicians lack confidence in providing quality care to individuals with intellectual and developmental disabilities (IDD) like DS. Dr. Sanders is an Assistant Professor, board certified in Neurology with Special Qualifications in Child Neurology and Neurodevelopmental Disabilities. Her preliminary work revealed that resident physicians in adult settings lacked confidence caring for patients with IDD due to lack of skills in communicating and engaging with adults with IDD and their supporters (caregivers). There is a communication tool for patients with autism that has been developed to help with this, but the tool has not been implemented widely or tailored for individuals with DS. To address this gap, Dr. Sanders will adapt a patient-and- supporter-initiated communication tool to meet the specific needs of adults with DS, their supporters, and clinicians. This proposed study, rooted in the RE-AIM framework for implementation science, will pursue three aims, to (1) analyze communication quality in adult primary care encounters using the unique qualitative method of Conversation Analysis; (2) adapt a patient-and-supporter-driven communication tool that meets the unique needs of adults with DS who have intellectual disability and come to clinic with a supporter; and (3) conduct a pragmatic pilot acceptability and feasibility trial of the adapted tool with adult clinicians, adults with DS, and their supporters. To achieve these aims, Dr. Sanders will pursue training in targeted research methods: Conversation Analysis, user centered design, and implementation science, including pragmatic trial design and analysis. Her multidisciplinary mentorship and advisory team brings expertise in neurodevelopmental disabilities, Conversation Analysis, communication in IDD, qualitative and mixed methods, physician behavior, user- centered design, pragmatic trial design, implementation science, and biostatistics. The University of Colorado Anschutz Medical Campus provides a unique research environment to conduct research on communication with adults with DS, supported by collaborative partnerships between the Adult and Child Center for Health Outcomes Research and Delivery Science, the Linda Crnic Institute for DS Research, and the large network of clinics that serve adults with DS. Successful completion of the research aims paired with the career development plan will provide preliminary data and allow for the refinement of data collection procedures to inform an R01 application to conduct a multi-site hybrid effectiveness implementation trial to evaluate the tool's ability to improve health care for individuals with DS.

NIH Research Projects · FY 2026 · 2025-09

The Vision of the Mountain States ASCEND Advancement in Research, Supportive Networking, and Career Development Program is to develop future academic leaders in kidney, urology, and hematology (KUH). This new application maps our path forward to create an innovative, comprehensive training program stressing translational multi-institutional science, alliances between MDs and PhDs, and incorporation of the newest scientific platforms and didactic training. ASCEND will capitalize on the interdisciplinary research communities at the University of Colorado (CU) and the University of Utah (UU) to carry out this mission. CU and UU are the premier research institutions in the Mountain West, a large geographic area currently lacking U2C/TL1 programs. ASCEND will be led by an interdisciplinary group with basic, translational, and clinical expertise: Dr. Michel Chonchol (CU Nephrology, Contact PI), Dr. Anna Malykhina (CU Urology), and Dr. Anna Beaudin (UU Hematology). To achieve our mission of recruiting, developing, and retaining KUH investigators we will apply the following: 1) Create a continuous pathway for forming strategic partnerships to achieve continuity of support for KUH trainees throughout all stages of career development; 2) Expand meaningful research opportunities to support identity formation as a KUH investigator and provide motivation to pursue and persist in KUH careers; 3) Foster effective mentorship and peer support to maintain integration into the KUH culture; 4) Advocate for supportive environments to diminish “pull” factors drawing people away from KUH careers; and 5) Support program evaluation, with a focus on longitudinal outcomes to guide and revise training efforts. We propose the following aims: Aim 1 will develop a rigorous, integrated, mentored cross-disciplinary training program (TL1 Training Core). Trainees will participate in structured, comprehensive, and specialized mentored research training that is personalized and based on the needs of the learner. Aim 2 will develop a structured professional and individualized development program that provides support and opportunities for professional career development to enhance skill development, career satisfaction, and planning (Professional Development Core). We will promote trainee engagement in their career development. All mentors will undergo comprehensive and structured mentorship training to equip them with the tools to be effective. Aim 3 will create the ASCEND Network to provide trainees with access to purposeful networking, successful role models, and peer trainees, and integration into the local and national KUH community (Network Core). ASCEND will create an inspiring online and in-person research community in which KUH trainees can exchange ideas, innovations, and scientific expertise with peers, mentors, and alumni. Ultimately, the ASCEND mission is to support, connect, and train the next generation of KUH researchers. By creating a climate for team-oriented science and integrated networks of support, we will create the training program and environment necessary for trainees to progress successfully and efficiently as KUH investigators. .

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

Acute lung injury (ALI) and its clinical manifestation, acute respiratory distress syndrome (ARDS), carry significant morbidity and mortality. However, there are intrinsic innate protective mechanisms. Harnessing those poorly understood mechanisms could enable us to exploit them therapeutically. Alveolar type II (ATII) cells are key players in acute lung injury. Alveolar macrophages (AM) are critical to the pathogenesis of ALI, where local microenvironmental cues shape their inflammatory or anti-inflammatory properties, which can be fluid and amenable to manipulation. How metabolic intermediates (such as lactate, the end product of glycolysis), released into the microenvironment, can affect macrophage phenotypes is unknown, especially in the setting of ALI. A key innate protective mechanism in ALI involves enhanced glycolysis in ATII cells. Our published work shows that macrophages co-cultured with ATII cells display a blunted response to LPS stimulation, which is dependent on lactate produced by the alveolar epithelial cells. Furthermore, locally intratracheally (i.t.) applied lactate can attenuate ALI in mouse models. AMs take up lactate via the monocarboxylase transporter (MCT) 1. Lastly, our preliminary data suggest lactate can induce an increase in histone lactylation. Histone lactylation is a recently described histone modification that renders promoters of genes accessible to transcription. The overall goal of this proposal is to delineate how lactate produced by alveolar epithelial type II cells is protective in ALI by shifting the AM towards an anti-inflammatory phenotype. We hypothesize that in the intra-alveolar compartment, lactate released by ATII cells is necessary and sufficient to induce the generation of anti-inflammatory cytokines by AM in ALI via epimetabolic regulation of transcription: we will test if uptake of ATII-cell-derived lactate by AMs via MCT1 induces an anti-inflammatory macrophage phenotype (Aim 1). We will test if ATII derived lactate specifically targets AMs and if, in AMs, lactate funnels into the Krebs cycle (Aim 2). Lastly, we will study whether lactate induces the expression of anti-inflammatory cytokines by increasing the accessibility of transcription factors through histone lactylation (Aim 3). Acid aspiration, i.t. LPS and Staph. aureus pneumonia will be used as murine models of ALI that directly target the intra-alveolar compartment. We will use a comprehensive genetic approach utilizing tissue specific knock out mice targeting macrophages and AT II cells. We will utilize primary human AMs for translational studies. Flow cytometry, ELISA assays, RNA single-cell sequencing, and qPCR will be used to characterize AMs. Functional studies with metabolomic tracer experiments with 13C-labeled lactate will be performed to characterize the metabolic flux. We will study H3 histone lactylation modifications and unbiased transcription factor analysis experiments. This proposal is novel by expanding the scope of understanding of the role of epimetabolic crosstalk between ATII cells and AMs in ALI. It involves state-of-the-art in vitro and in vivo experimental approaches and incorporates a therapeutic approach with potential for translation.

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

PROJECT SUMMARY (ABSTRACT): Enterovirus D68 (EV-D68) is a non-polio enterovirus that primarily causes respiratory illness, particularly in children. While many individuals infected with EV-D68 may have mild symptoms, others have more severe presentations including severe asthma-like respiratory illness requiring intensive care and acute flaccid myelitis (AFM) that can lead to permanent polio-like paralysis. Large outbreaks of EV-D68 and increases in AFM cases have occurred in cyclic patterns since 2014. However, no effective treatments or vaccines against EV-D68 are available. Challenges to the development of such tools include gaps in knowledge about the immune response to respiratory viruses such as EV-D68. Passive transfer of EV-D68 antibodies provides protection against EV-D68 infection and severe disease in animal models; however, the role of humoral immunity in humans acutely exposed to or infected with EV-D68 is unknown. Understanding the role of the human humoral immune response in the development of different disease presentations of EV- D68 infection is essential to inform the development of effective therapeutic and preventative tools. In this study, the candidate proposes to investigate clinical factors and the role of humoral immunity as a predictor of EV-D68 infection and disease. Through the analysis of previously and prospectively collected de- identified clinical data and biospecimens from cohorts of individuals with acute EV-D68 infection or exposure, the candidate will address the research aims of: 1) describing the clinical spectrum and identifying clinical factors predictive of different EV-D68 disease presentations and 2) characterizing the humoral immune response to EV-D68 by evaluating the dynamics of the humoral immune response and identifying immunologic predictive factors of disease presentations of EV-D68 infection in humans. This study will improve the understanding of the role the humoral immune response to EV-D68 exposure and infection has to development of disease which will inform immune correlates of protection to guide development of potential monoclonal antibody treatments and next-generation vaccine candidates. The candidate is committed to a career in research in emerging infectious diseases such as EV-D68 in which she has both clinical and research experience with children with EV-D68 infection. The objective of this career development award is for the candidate to enhance skills in patient-oriented clinical research to become a successful independent investigator. Through didactic coursework in pursuit of a PhD, experiential training, and seminars/workshops she will build on her knowledge and skillset in areas of team science, clinical research, biostatistical analysis, and clinical outcomes. The guidance of her multidisciplinary mentorship team and advisors who have expertise in EV-D68 and AFM, immunology, and clinical research, will ensure the candidate successfully achieves her goals and aims in the proposed study.

NIH Research Projects · FY 2025 · 2025-09

Project Abstract The Department of Homeland Security considers numerous chemical threat agents a concern for human health, specifically those that are acute pulmonary toxicants. In acute lung injury, inflammation is critical thus we propose that inflammation is a common mechanism of lung injury caused by chemical threat agents due to mast cell activation. We and others have shown mast cell activation to be critical in response to a wide range of xenobiotics including nitrogen mustard, ozone, diesel exhaust, insecticides/herbicides, cigarette smoke, heavy metals and nanoparticles as examples. Mast cells are a logical cell type to study in pulmonary injury from chemical threat agents due to 1) their location at interfaces with the external environment (e.g., lung); 2) their roles as sensors for initiating both innate and adaptive immune responses; and 3) their immediate response to danger signals through degranulation and release of preformed mediators. We have demonstrated that mast cell activation is a major contributor to the pulmonary toxicity and inflammation observed following nitrogen mustard (NM), chloropicrin and formaldehyde exposure. Currently there are few shared mechanisms which have been identified between these chemical threat agents, thus identification of common pathways would be beneficial for future therapeutic targets and biomarkers of exposure. Here, we propose two specific aims to provide confirmatory data on the role of mast cells in pulmonary injury resulting from exposure to chloropicrin (fumigant/pesticide) and formaldehyde (industrial chemical). Our overall hypothesis is that activation of mast cells is a common initiating step in recruitment and propagation of immune responses in the lung across several classes of chemical threat agents. In aim 1, we will Confirm the in vivo contribution of mast cells to pulmonary injury and inflammation resulting from chemical threat agents in Cpa3-Cre mast cell deficient mice. In aim 2 we will Determine role of mast cells in lung injury using a novel human lung-on-a-chip model where we have incorporated mast cells. Collectively, our goal is to establish activation of mast cells as a common mechanism across several chemical classes which are linked with pulmonary toxicity. Confirmation of these findings will allow identification and development of novel therapeutic targets for prevention and/or treatment of the effects of these potential chemical warfare agents through targeting of mast cells.

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

Project Summary The monocytic leukemia zinc-finger protein-related factor MORF is a transcriptional coactivator and a catalytic subunit of the histone acetyltransferase MORF (named after its catalytic subunit) complex. MORF regulates expression of hematologic, cardiac and developmental genes and plays a pivotal role in the development of the hematopoietic system and the formation of blood cells. Aberrant acetyltransferase activity of MORF is linked to aggressive forms of leukemia, blood diseases and cardiovascular abnormalities. The MORF driven epigenetic mechanisms in the normal and disease states remain poorly understood, and characterization of these mechanisms may pave the way for the development of novel strategies to diagnose, prevent or treat these diseases. Our preliminary data indicate a multivalent engagement of MORF with chromatin through its histone and DNA binding domains that regulate enzymatic activity and genomic localization of the complex. The molecular basis underlying novel functions of MORF and biological consequences are unknown and will be determined in the proposed studies. The results generated in this work will offer a comprehensive understanding of the molecular mechanisms by which the MORF subunit is bound to specific genomic sites and its enzymatic activity is regulated, provide atomic-resolution insight into MORF signaling pathways that may constitute new targets for therapeutic interventions, aid in our understanding of the etiology of hematological and cardiovascular diseases associated with aberrant MORF activity, and enhance our knowledge of the fundamental principles underlying the ‘epigenetic code’ writing and recognition.

NIH Research Projects · FY 2025 · 2025-09

Abstract Gene pathway analysis is a critical tool for understanding how groups of genes within biological pathways collectively contribute to complex human traits and diseases. Widely applied in both bulk and single-cell RNA- seq (scRNA-seq) studies, these methods have provided valuable insights into molecular processes and their effects on health. Despite the development of hundreds of pathway analysis tools, recent studies have exposed significant challenges that undermine the reliability of these approaches. For instance, substantial discrepancies between gene pathway scoring methods lead to inconsistencies in pathway network construction and cell type clustering, with false-positive rates in GSEA methods reaching as high as 40%. These issues create uncer- tainty for researchers in selecting appropriate tools for real-world applications, and attempts to establish a “gold standard” have proven computationally intensive and context-dependent. This project proposes a transformative framework to address these challenges. First, we introduce a latent- factor-based top-down model for unsupervised pathway interaction analysis, designed to efficiently capture population-specific variations while remaining robust to differences in pathway activity scoring algorithms. Sec- ond, we propose a set of multivariable latent factor regression models to identify pathway-phenotype associa- tions. These models enhance statistical power by leveraging individual gene expression data and accounting for pathway correlations, reducing false discoveries. Finally, we will apply these methods to extensive bulk- and single-cell RNA-seq datasets to validate existing findings, generate new insights, and tackle complex biological questions that have previously been out of reach.

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

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 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 Abstract Pulmonary arterial hypertension (PAH) is a progressive and incurable disease with high morbidity and mortality. Although PAH is significantly more prevalent in women compared to men, female PAH patients exhibit better right ventricular (RV) function and survival. However, the mechanisms underlying superior female adaptation to RV pressure overload in PAH remain unknown. This proposal seeks to identify the role of RV macrophages in mediating sexual dimorphisms in RV function in PAH. This investigation is motivated by our compelling recently published data demonstrating that macrophages play a role in RV maladaptive remodeling in experimental pulmonary hypertension (PH) through effects the extracellular matrix (ECM), and that removal of macrophages in PH results in decreased collagens and other ECM proteins, improving RV contractility and RV-pulmonary arterial (RV-PA) coupling. Furthermore, through single cell RNA sequencing experiments of cells from the murine RVs, we discovered profound sexual dimorphism in gene expression in resident cardiac macrophages from male and female mice that include upregulation of several genes involved in ECM organization and collagen binding in males. We have previously shown that endogenous and exogenous 17beta-estradiol (E2) improves RV function in ovariectomized females and in males with PH, that E2 decreases collagen accumulation in the RV, and that E2 down-regulates the pro-inflammatory and pro- fibrotic transcriptome of bone marrow-derived macrophages. We now seek to understand whether the sexual dimorphisms we identified in RV adaptation and cardiac macrophage function are explained in part by transcriptional effects of sex hormones on the pro-fibrotic programming of RV macrophages. We hypothesize that female sex hormones regulate expression of RV macrophage genes associated with fibroblast activation and ECM stiffness, resulting in sexual dimorphisms in RV contractility, stiffness, and diastolic dysfunction in adaptation to RV pressure overload. Specifically, we hypothesize that female hormones downregulate pro-fibrotic programming in resident RV macrophages, resulting in less fibroblast activation and ECM stiffening. We will utilize the experimental PH models of mouse pulmonary artery banding and rat Sugen hypoxia, and examine differences in resident RV macrophage gene expression, RV contractility and stiffness, and the RV ECM between healthy and diseased male and female animals. We will utilize a menopausal animal model and exogenous estradiol hormone replacement therapy to examine how female and male sex hormones regulate RV macrophage programming.