University Of Colorado Denver

NIH Research Projects · FY 2025 · 2023-09

PROJECT SUMMARY We propose a continuation of the annual Strategic Conference of Zebrafish Investigators (SCZI), the main meeting of principal investigators using the zebrafish model. Previously, this was a biannual conference held in Asilomar, USA. In 2019, the International Zebrafish Society collaborated with the European Zebrafish Society to adopt a 3-year rotation schedule for both their larger International Zebrafish Conference (IZFC) gathering zebrafish researchers at all career stages and the SCZI so that the locations will rotate between US/Canada, Europe, and Asia-Pacific. This new, focused meeting structure aims to promote greater international collaboration by reducing the total number of international conferences, supporting the goals of both societies to increase inclusivity and to reduce the travel-associated carbon footprint. The SCZI is a prime venue for zebrafish principal investigators to discuss current research and promote coordinated advancement of research using zebrafish as a model system (as well as connections to related, emerging aquatic models). The meeting will accommodate up to 200 principal investigators. The proposed 10th SCZI meeting will take place in 2024 in Asilomar, CA; continuation of the series will follow in Europe 2025, Asia-Pacific in 2026, and so on. The zebrafish has become a powerful model organism for the genetic analysis of vertebrate development. Moreover, the zebrafish has emerged as a versatile vertebrate system to model human diseases and discover underlying mechanisms, especially when used in conjunction with other vertebrate models, and to screen for therapeutics and toxins. Advances in forward and in particular, reverse genetics, imaging techniques, transgenesis, and chemical screening are rapidly and widely disseminated throughout an active, collaborative community that has grown in coherence and connectivity through international meetings. The proposed SCZI meeting will continue to foster the critical development of resources, establishment of collaborations, and expansion of interdisciplinary interactions. Importantly, one third of past participants have been new investigators: in addition to the open access to new findings in the field and to establish collaborations, new investigators will have the opportunity to attend discussion sessions on topics such as grant writing and balancing professional development and personal growth chaired by senior investigators. Past meetings have had a solid record of inclusion of women and minority participants, and we will continue to provide inclusive meeting environment. This grant will provide financial assistance for up to 30 US-based junior laboratory heads to attend the conference. Recipients of financial support will be chosen based on the impact and originality of their abstracts as well as considering gender, race, career stage, and need.

NIH Research Projects · FY 2025 · 2023-09

PROJECT SUMMARY/ABSTRACT Gynecologic malignancies such as ovarian cancer (OvCa) are among the deadliest cancers affecting women due to therapy resistance and limited understanding of disease etiology and risk. An under-explored risk factor is Wnt ligand WNT4, which is central to ovarian organogenesis. Over 20 studies link WNT4 polymorphisms with increased risk for gynecologic pathologies; one polymorphism at a key WNT4 regulatory site (rs3820282) is associated with 10-25% increased risk for OvCa, but the mechanism(s) is unknown. Our work links WNT4 to cancer cell growth, metabolism, and therapy resistance. We find WNT4 over-expression is sufficient to mediate chemotherapy resistance in vitro, and resistance with increased metastatic outgrowth in vivo, and that WNT4 expression is strongly induced in OvCa cells surviving neoadjuvant chemotherapy. Importantly, the rs3820282 variant allele in the WNT4 regulatory site creates a binding site for nuclear receptor-class transcription factors. CRISPR knock-in of the rs3820282 variant in mice increases Wnt4 expression in gynecologic tissues. Accordingly, in a protein array study of more than 100 OvCa tumor tissues, we found that AMPK activation and downstream signaling were increased in variant allele tumors. Conversely, glucose metabolism proteins were increased in wild-type tumors and inversely correlated with AMPK signaling, suggesting WNT4 genotype underpins metabolic remodeling. These observations suggest that the rs3820282 variant activates WNT4 to drive cancer phenotypes. However, the rs3820282 variant allele frequency (VAF) is widely divergent across ethnic populations, occurring at ~0% in African populations, ~15% in Caucasians, 20-40% in Latinx populations, and 45-55% in Asian populations, paralleling high incidence of aggressive, treatment-resistance OvCa subtype clear cell carcinoma (CCC) in Asian populations. Our goal is to determine how rs3820282 mediates disparities in ovarian cancer outcomes, mechanistically define genotype-driven tumor etiology, and identify therapies to exploit dependence on WNT4. Toward this goal, we will: 1) define how the rs3820282 variant activates WNT4-dependent metabolic remodeling; 2) define rs3820282-driven tumorigenesis and therapeutic response in a model of ovarian clear cell carcinoma (CCC); 3) determine how rs3820282 genotype impacts outcomes for patients with OvCa. With a foundation of rigorous supporting data from human specimens, we will undertake highly mechanistic studies to define the contribution of this common polymorphism to a cancer disparity, tumor metabolic reprogramming, gynecologic tumorigenesis, treatment response, and patient outcomes. We will leverage cutting-edge global metabolomics, tumorigenesis modeling, and human survival studies. Our approach can define the genotype-to-phenotype link, determine how this SNP drives OvCa cancer disparities, and identify approaches to exploit the underlying biology.

NIH Research Projects · FY 2025 · 2023-09

Project Summary/Abstract The Colorado Clinical and Translational Sciences Institute’s (CCTSI) T32 Pre-Doctoral Training Program aligns and supports the goals of NCATS to ensure that a diverse pool of highly trained scientists is available in adequate numbers and in appropriate research areas to carry out the nation’s clinical and translational science research (CTSR) agenda. Our proposed Team Oriented Training across the Translational Sciences Spectrum (TOTTS) program will support trainees’ competency development and foster characteristic development for translational research scientists. We request 8 slots for diverse trainees that represent the translational spectrum (biomedical/bioengineering PhD students, population health/health outcomes PhD students, clinician doctoral students) to complete our 2-year program, with clinician doctoral students having a 1-year option. Our overall goal is to meet the needs of the translational science and research community by preparing diverse interdisciplinary doctoral trainees that are team oriented and have developed characteristics and associated skills necessary for successful CTSR careers, including Domain Expert, Rigorous Researcher, Boundary Crosser, Process Innovator, Team Player, Skilled Communicator, and Systems Thinker. Our program’s value is that Diversity Accelerates Research and Translation (DART). Integration into the clinical and translational fabric strengthens a person’s connection, self-efficacy, and identity formation, thus increasing the likelihood that T32 trainees remain in the translational workforce. Our program’s curriculum and approaches are based on our colleague’s conceptual framework for diversifying the CTSR workforce, Model for Persistence of a Diverse Clinical and Translational Research Workforce. A program objective is to apply evidence informed mentoring practices to support developing trainees from all backgrounds for career persistence and success in CTSR. Evidence informed practices to be applied include 1) signed mentoring agreements, 2) development and regular review of Individualized Career Development Plans, 3) completion of Mentoring3: Mentor, Mentee and Peer, an effective mentoring training program attended by both mentors and mentees, 4) annual mentor-mentee dyad reviews with feedback, and 5) trainee Mentoring Advisory Team reviews. Several programmatic elements are novel to promote skills development and successful transitions into careers as translational scientists: translational mentor supported immersion experiences (clinical, industry, community, lab), Leading and Teaming, Writing Accountability Groups, F-Grant Review and Mock Study Section, Communicating Research to the Public, and Café Scientifiques. Evaluative emphasis will be placed on our ability to achieve our goal of attracting and supporting the training and persistence of individuals from diverse (especially underrepresented) backgrounds who have the potential to infuse CTSR with new perspectives and approaches that are efficient and effective.

NIH Research Projects · FY 2024 · 2023-09

Project Summary/Abstract While advances in resuscitation science have improved cardiac arrest survival, we lack therapies to improve cognitive-affective outcomes in this patient population. Our lab has previously identified cognitive dysfunction in a mouse model of global cerebral ischemia (GCI) which has been attributed to hippocampal neurodegeneration and impaired hippocampal plasticity. However, no study has attempted to identify amygdala dysfunction after GCI, despite clinical evidence of emotional dysfunction, such as anxiety and Post-Traumatic Stress Disorder (PTSD). Therefore, it is important to identify the effect that GCI has on the amygdala, the emotional center of the brain. Our lab has a well-developed, translatable mouse model of GCI, the cardiac arrest/cardiopulmonary resuscitation model (CA/CPR), that has been instrumental in assessing amygdala function after GCI. I have utilized the amygdala-dependent delay-fear conditioning (DFC) paradigm to assess associative learning and memory and have performed field excitatory post-synaptic potential (fEPSP) recordings in two circuits within the amygdala, as measures of amygdala function. I have found a sexually dimorphic and circuit specific deficit in amygdala function after GCI and am working toward identifying the mechanism of this dysfunction. I have found that there is a male specific impairment in amygdala-dependent associative learning and a concomitant deficit of long-term potentiation (LTP) in the cortical input to the basolateral amygdala. I have also found no evidence that these deficits of amygdala function can be attributed to neurodegeneration within the amygdala. I have verified that there are differential mechanisms of LTP induction between the two circuits and this difference has led to the development of my hypothesis put forth in this proposal. The difference being, LTP of the cortical input to the BLA requires functional NMDA receptors and L-type calcium channels (LTCCs), whereas the intra amygdala circuit only requires functional NMDA receptors. Thus, I hypothesize that GCI induces dysfunction of LTCC's within the BLA of male mice, thereby contributing to the deficits in amygdala-dependent behavior and LTP. To assess this hypothesis, I have isolated and recorded LTCC mediated currents from BLA pyramidal neurons. This method, while powerful, has yielded no significant difference between CA/CPR and sham animals. However, a caveat of the method is that only somatic and peri somatic LTCCs can be measured. Therefore, to fully evaluate my hypothesis, I have proposed more site-specific experiments that will evaluate the contribution of LTCC's to synaptic transmission at individual distal dendritic spines. I will use two-photon calcium imaging of individual spines in the BLA while electrically inducing LTP of the cortical input to the BLA. I will then use pharmacology to identify the LTCC component of the calcium response and compare between sham and CA/CPR animals.

NIH Research Projects · FY 2025 · 2023-09

PROJECT SUMMARY The proper functioning of essentially all cells is due in part to spatially defined regions that are populated by specific proteins to perform specific activities. Neurons, due to their elongated size and morphology, must deal with large distances when navigating protein localization with regard to axons and dendrites. This problem is solved by transporting RNA molecules to specific locations within neurons. When these RNA molecules are translated, they immediately produce correctly localized proteins. The importance of this key process is evidenced by multiple neurological diseases that are associated with its misregulation. The RNA-binding protein TDP-43 is known to be involved in RNA localization, and mutations in TDP-43 are associated with Amyotrophic Lateral Sclerosis (ALS). Yet the RNAs that depend on TDP-43 for localization, how TDP-43 recognizes these RNAs and promotes their transport, and the phenotypic consequences of their mislocalization are all currently unknown. This means that an entire regulatory modality contributing to ALS pathology may be undiscovered. Here, we propose to use our combined expertises in subcellular transcriptomics, stem cell differentiation, and live cell microscopy to engage these critical problems and provide novel insights into ALS disease mechanisms that likely will result in novel treatment modalities. To do so we have developed a novel technique to monitor neuronal RNA mislocalization transcriptome-wide, allowing us to identify transcripts that need functional TDP-43 for proper transport. By combining this approach with our expertise in stem cell differentiation, we will use this technique to probe TDP-43-mediated RNA localization in functional human motor neurons. We will identify mislocalized RNAs in engineered TDP-43-null motor neurons, motor neurons expressing ALS-associated TDP-43 mutants, and motor neurons derived from ALS patients. We will use this information to derive the RNA sequence requirements that define transcripts that depend on TDP-43 for transport and test their necessity and sufficiency with reporter transcripts. We have also developed a technique for imaging the transport of single RNA and protein molecules in live Drosophila brains. We will use this approach to visualize TDP-43-mediated RNA transport in live brains and learn about its dynamics and regulation. Drosophila TDP-43 mutants display phenotypes similar to those seen in ALS patients. However, as of now data attributing cellular or organismal phenotypes to the mislocalization of specific transcripts for ALS or any other neurological disease is missing. This may be due in part to the fact that RNA localization is often overlooked and the transcripts that are mislocalized in disease states are not known. To engage this problem, we will mislocalize specific transcripts in Drosophila brains that depend on TDP-43 for transport by removing sequences that TDP-43 recognizes within the transcript. We will then ask if these flies display the locomotor and physiological defects that characterize both TDP-43 mutant flies and ALS patients. Overall, we intend to define new connections between misregulated molecular mechanisms and ALS patient phenotypes, thus opening up new avenues of research and therapeutic development.

NIH Research Projects · FY 2025 · 2023-09

Abstract With an estimated annual cost of $12.5 billion in the United States, epilepsy affects approximately 3.4 million Americans and carries a lifetime risk of around 3%. The most common form of epilepsy is focal in nature, meaning seizures arise from a restricted part of the brain. It remains a significant challenge to predict who will respond well to treatment despite modern technology and research, largely due to the heterogeneity of focal epilepsy. Existing statistical methods to analyze seizure data do not appropriately address the within- and between-individual variation in epileptic seizures over time. We propose to leverage our access to the Human Epilepsy Project (HEP1 and HEP2), an observational study of 450 patients with focal epilepsy that tracked seizures longitudinally, and our statistical and clinical expertise to develop novel dynamic prediction models for seizure frequency over time. The daily seizure data from HEP show (1) subgroups of individuals with different seizure trajectories and (2) clumping of seizures, in which a patient is more likely to experience subsequent seizures following a seizure episode. Dynamic prediction models have been used successfully in other clinical areas besides epilepsy, but they do not allow for subgroups of trajectories. Similarly, clumping of events has been handled with the Hawkes process in association models where a homogenous population is assumed. Instead, we seek to predict occurrence of events and understand covariate effects on processes where subgroups of trajectories exist. We hypothesize that accounting for subgroups of individual trajectories and clumping of seizures will provide more accurate and precise prediction of seizure outcomes. We plan to develop novel models for prediction of seizure events through the following aims: (i) Develop a Bayesian nonparametric models for dynamic personalized prediction of seizures over time. We will use the HEP1 dataset to predict longitudinal seizure count and occurrence that allows for subgroups of trajectories and will evaluate the methods using HEP2 data, (ii) Develop a novel Dirichlet Process Mixture Hawkes process model for personalized prediction of recurrent event data that will allow for subgroups and clumping of events; and will compare to existing approaches to handle clumping, and (iii) Develop an R package and a shiny application to implement and illustrate the novel methods. The expected outcomes of this work are both clinically and statistically significant: a) a shiny application tool to obtain tailored predictions of longitudinal seizure trajectory based on seizure history, treatment and other clinically relevant covariates that will help patients and clinicians identify optimal treatment earlier in the personalized course of the patient’s disease; and b) the new methods will be relevant to other epilepsy types and other conditions, for example in modeling relapses in multiple sclerosis.

NIH Research Projects · FY 2025 · 2023-09

PROJECT SUMMARY/ABSTRACT The goal of this project is to define vulnerabilities that can be leveraged to comprehensively eradicate MDS stem cells (MDS-SCs). This objective is based on the concept that MDS-SCs lie at the root of disease and are not effectively targeted by current therapies. A significant challenge in achieving this goal is the heterogeneity of the MDS-SC population, as recent studies have identified distinct subsets of MDS-SC with varying responsiveness to therapy. Thus, the premise of our work is that understanding and targeting the unique properties of heterogenous MDS-SCs is critical to improve outcomes in MDS. The foundation of our project is built upon laboratory studies coupled with an active clinical trial (NCT03564873) evaluating the clinical impact of targeting protein synthesis with the global translation inhibitor omacetaxine (oma) in MDS-SC from individuals with high- risk MDS. While the results from our trial using oma in combination with the hypomethylating agent azacytidine (aza) are indeed promising and represent a substantial improvement over standard of care (aza alone), instances of disease progression in patients suggest oma may not eradicate all MDS-SC subsets. These outcomes, alongside toxicities associated with global protein translation inhibition have motivated us to better characterize the unique biological and functional properties of heterogenous MDS-SC subsets. With the goal of developing next-generation approaches that more effectively target MDS-SC while minimizing dose-limiting toxicities to patients, here we will address two primary questions: 1) what is the overall efficacy of protein synthesis inhibition amongst varying MDS-SC subtypes, 2) what are the optimal therapeutic strategies to eradicate MDS-SC based on their sensitivity to protein synthesis inhibition? Hence, we will leverage comprehensive CITE-seq based analyses of serial pre- and post-treatment specimens from our oma/aza clinical trial alongside PDX-based functional studies to identify the link(s) between features such as protein synthesis activity, MDS-SC genotype and clonal behavior with oma response at single cell resolution. We will define the mechanisms governing reliance on protein synthesis in MDS-SC, focusing on pharmacological and molecular genetic-based targeting of key pathways linked with oma sensitivity. Lastly, we will develop improved approaches to comprehensively target heterogenous MDS-SC populations, with focus on next-generation strategies that more selectively target aberrant metabolic features of MDS-SC uncovered by our ongoing CITE-seq characterization of patient specimens. Taken together, the proposed studies will use advanced functional models and single-cell resolution analyses to identify key vulnerabilities of MDS-SC that can be leveraged to design rational therapeutic approaches to improve outcomes for MDS patients.

NIH Research Projects · FY 2025 · 2023-09

PROJECT SUMMARY. Down syndrome (DS), the genetic condition caused by trisomy 21 (T21), is the most prevalent chromosomal abnormality and a leading cause of intellectual and developmental disability. T21 affects the development and/or function of nearly every organ system, predisposing individuals with DS to many co-occurring conditions such as Alzheimer’s disease, congenital heart defects, and autoimmune disorders, among others. Although it is accepted that T21 causes genome-wide dysregulation of gene expression programs, little is known about how T21 affects gene expression across different tissues and organs, and how these effects contribute to the etiology of the co-occurring conditions of DS. Therefore, there is a clear need to understand the complex cause-effect relationships between T21, altered gene expression, and organ development and pathophysiology. Several mouse models of DS recapitulate key phenotypes of DS and have been used to study gene expression dysregulation in DS. Additionally, induced pluripotent stem cell (iPSC) systems have been employed to study the effects of T21 in human cell types. Therefore, in clear response to the Funding Opportunity Announcement PAR- 22-247, we propose here to generate a Trisomy 21 Model Atlas of tissue-specific murine and human transcriptomes, matched to detailed metrics of organ development and architecture. The use of mouse models will provide insights into tissue- and developmental stage-specific transcriptome changes and pathophysiology, while human cell types differentiated from iPSCs will dissect cell-intrinsic features of T21 and validate findings from the mouse models. Our Specific Aims are: 1. Generate an atlas of gene expression programs affected by trisomy 21 across model systems. Supported by strong preliminary data, we propose here to complete an atlas of transcriptome changes in ten organs across three developmental stages in mice and matched data from human cell types derived from iPSCs. 2. Define the impact of trisomy 21 on organ development and pathophysiology at the cellular level. To investigate the link between dysregulated gene expression and tissue dysfunction, we will assess organ development and pathophysiology through histopathological profiling, multiplexed immunofluorescence imaging, single-cell RNA-sequencing, and characterization of human organoids derived from iPSCs. 3. Curate, organize, and share all data for open access through the INCLUDE Data Hub. We will ensure that all data is made publicly available in the INCLUDE Data Hub by completing careful file curation and organization, developing a metadata schema, and creating a code repository. This will ensure that the atlas becomes a lasting resource that can be used beyond the scope of this grant. Altogether, this resource will advance our understanding of the effects of T21 on organ development and pathology, while enabling findings cross-validated in mouse models and human cell types.

NIH Research Projects · FY 2025 · 2023-09

PROJECT SUMMARY/ABSTRACT: The long-term objective of this proposal is to understand formation of the midface skeleton, both during normal development, and in human genetic disease conditions. The midface consists of the structures around the nose, the eye, and the upper jaw. Human genetic disorders, like frontonasal dysplasia, affect these midface structures. Specifically, the ALX transcription factor encoding genes have been implicated in multiple types of frontonasal dysplasia. Uniquely, we model these diseases in zebrafish by using genetic mutants. We propose that the ALX genes function to specify an identity code which patterns the vertebrate midface. Our model draws from examples like the DLX and HOX codes, which specify identity in the dorsoventral and anteroposterior axes of the craniofacial skeleton, respectively. We propose three specific aims to test our innovative “alx-bullseye code” hypothesis, that nested alx gene expression directly regulates frontonasal skeletal identity. In Aim 1 we define the skeletal structures arising from the alx-bullseye code in wild types using in situ hybridization, live cell tracking, and lineage tracing. Aim 2 will determine whether alx gene combinations function to specify frontonasal identity. To identify these functions, we will compare wild types to alx mutants examining gene expression, skeletal cell differentiation, and misexpression phenotypes. Aim 3 will uncover how alx genes molecularly control cellular identity. We will examine an in vivo Alx direct transcriptional reporter, ChIP-seq to identify Alx protein occupancy across the genome, and Hi-C to reveal enhancer-promoter contacts mediated by Alx. Significance of this proposal is high, the genes that we propose function in the alx bullseye code have direct human orthologs that are associated with midface dysmorphologies. This proposal is innovative, as there has not yet been a patterning code proposed for the vertebrate midface. Success of this proposal will enhance our understanding of midface craniofacial development and human disease. By using the strengths of the zebrafish system to elucidate the molecular, cellular, and genetic mechanisms underlying frontonasal dysplasia, we are fulfilling the mission of the NIDCR to improve human health through research.

NIH Research Projects · FY 2025 · 2023-09

Clinician scientists fulfil a unique role by integrating discovery science with clinical practice and therapeutic intervention. However, their numbers are in decline, creating the need for flexible training and research opportunities to ensure their future. The Colorado Clinical and Translational Sciences Institute’s (CCTSI) T32 Post-Doctoral Program will address the need for highly qualified and competent translational scientists by providing a responsive and evidence-informed program, a stimulating environment with well-equipped facilities, a community of practice for translational scientists. We request support for 4 post-doctoral trainee slots of two years each. Our program is novel in that it expands the translational spectrum by recognizing the importance of shared, naturally occurring models of disease among animals and humans to push discovery, which we coin T0.5. Our program will involve both doctorally prepared clinicians (physicians (MD), pharmacists (PharmD), physical therapists (DPT), nurses (DPN) and others) at the University of Colorado who will complete the Master of Science in Clinical Science (MSCS) degree and veterinarians completing a PhD from Colorado State University, a partnering university. Trainees of our interprofessional program will be highly qualified and thoroughly prepared to be effective team members and translational scientists that fuel discovery that is translated to communities for improved health and lives of its citizens. Development of translational scientists that characterize Domain Expert, Rigorous Researcher, Boundary Crosser, Process Innovator, Team Player, Skilled Communicator, and Systems Thinker will be emphasized through completion of a degree program (MSCS or PhD), meaningful immersion experiences for translational science and translational research, effective mentorship and peer support, and alleviating “pull factors” by creating a community of practice through our Translational Scientist seminars. Pull factors are demands that compete or conflict with trainee engagement, integration, and performance that ultimately diminish persistence. Clinical and translational science and research requires a shift from an individual-based approach to a teamwork model. Team member competency development will be kick-started by early participation in our Teaming and Leading program. The One Health framework will be used to highlight the need for teaming and permit application of all translational scientist competencies recognizing that achievement of optimal health is reliant on the interconnection among people, animals, and their shared environment across eco-health system levels (local, regional, national and global). A culminating activity will be trainee led interdisciplinary teams participating in a One Health Hackathon focused on antimicrobial resistance. Long term-evaluation and continuous quality improvement will be used to guide programmatic efforts and respond to changing demands. Evaluative efforts will focus on the program’s ability to develop a successful group of persistent team oriented translational scientists.

NIH Research Projects · FY 2025 · 2023-09

Instruction: Funded by NIH since 2008, the Colorado Clinical and Translational Sciences Institute (CCTSI) at the University of Colorado Denver (CU Denver) has transformed the clinical and translational research and training enterprise in the Colorado region with the vision of accelerating the translation of discoveries into improved patient care and public health for all. The CCTSI, headquartered at the CU Anschutz Medical Campus, is a partnership of CU Denver, CU Boulder, Colorado State University (CSU), 5 hospital systems and 20 community organizations, which has advanced translational science by educating a successful workforce, creating a collaborative environment that supports local and multi-site research, engaging Colorado communities through its unique community engagement program, and greatly enhancing our informatics and data science capabilities. Despite the successes, there remain many challenges to overcoming inefficiencies and roadblocks in clinical translational science (CTS) processes locally and nationally. This application maps our collaborative path forward to meet these challenges. In the next grant cycle, we will enhance our efforts to improve efficiency and coordination with Partners and across the CTSA Consortium, reinforce our extensive community partnerships, develop innovative informatics and technology solutions to advance CTS, and develop a highly skilled translational workforce for the future. This UM1 will be tightly coordinated with our other 6 CTSA grant applications, to accomplish the following 6 Overall Strategic Goals: Goal 1: Advance CTS by developing, demonstrating, and disseminating innovative programs to improve the efficiency and impact of translation across the entire T0.5 to T4 spectrum. Goal 2: Promote collaboration, team and data science, and partnerships to accelerate CTR locally, regionally and nationally. Goal 3: Partner locally, regionally and nationally with institutions, stakeholders and communities to develop innovative research programs that will address the nation’s most pressing health issues. Goal 4: Further develop operational efficiencies to enhance the quality, safety, efficiency, effectiveness and informativeness of clinical research. Goal 5: Promote a nimble research environment that can rapidly respond to urgent public health needs. Goal 6: Develop and disseminate CTS training programs that educate and sustain a resilient clinical research professional workforce. Progress will be monitored and improved by our Evaluation Core and Continuous Quality Improvement program and we will make mid-course corrections as needed to achieve these goals and ultimately improve the health of our state and the nation.

NIH Research Projects · FY 2025 · 2023-09

PROJECT SUMMARY/ABSTRACT Nicotine vaping among adolescents has surged in prevalence over the past decade. Vaping poses clear health risks, is highly addictive, and leads to the use of combustible cigarettes. Our multi-disciplinary team, which includes expertise in youth participatory approaches, substance use prevention, nicotine use and cessation, and statistics, has developed an innovative school-based approach that promotes positive health behavior while also addressing academic standards, and thus incorporates both the academic mission of schools and youth health. YES-CAN! (Youth Engaged Strategies to Change Adolescent Norms) integrates the following evidence-based strategies: youth-adult collaboration; youth-developed narrative videos to convey health messages; peer leaders as change agents; and sustained implementation to change the normative environment. It rests on evidence that adolescents are more likely to change attitudes, norms, and behaviors when exposed to messages developed and delivered by peer role models rather than teachers, parents, or other adults. For this pilot/feasibility study, two middle/high school communities will receive the intervention. In each school community, we will implement a credit-earning high school class in which a trained teacher will deliver the one-year classroom-based program to 25-30 high school students, who will produce 6-8 short videos intended to increase refusal skills; promote stress management and positive coping; change social norms; prevent vaping initiation; and promote vaping cessation among current users. Videos will use a narrative approach and integrate known determinants of vaping. High school students will collaborate with the teacher and researchers to develop discussion guides and skills-building activities based on best practices for substance use prevention. In 6-8 eight sessions, high school students will deliver their videos to all students in the associated middle school. A text messaging component will reinforce and boost the effectiveness of the classroom sessions. Our overarching hypothesis is that this intensive approach that involves both middle and high school students in the same community will result in a new normative environment and a reduction in youth vaping. Aims for this pilot/feasibility study are to: 1) Determine the feasibility and acceptability of implementing the YES-CAN! program; and 2) Determine the feasibility and acceptability of the research protocols that will be used in a future efficacy trial. Primary efficacy outcomes include vaping ever use, past month use, daily use, susceptibility, behavioral intentions, and vaping-related knowledge, attitudes, perceived norms, resistance skills, and self-efficacy. Additionally, for high school students who participate in the intervention development and delivery, outcomes include positive youth development. Outcomes will be measured over a 13 month period.

NIH Research Projects · FY 2025 · 2023-09

To maintain organismal energy balance, energy molecules extracted from the diet or liberated from stored forms must be distributed appropriately throughout the body. By integrating and distributing signals to and from disparate tissues and organs, the brain plays a major role as a command center in organismal energy balance. The brain is also a hungry organ, consuming a disproportionate amount of energy relative to its size. Higher-level cognitive functions like learning and forming memories burn even more energy. Energy imbalance, such as a chronic high-calorie diet, perturbs cognitive functions like learning and memory, but the underlying mechanism is not clear. Metabolic syndromes like obesity are also associated with neurodevelopmental and neurodegenerative disorders. Most studies of organismal energy balance focus on how the brain uses a few known pathways to mediate inter-organ communication, but it is not known how cognitive functions specifically signal the brain’s demand for fuel and mobilize energy from stores in other parts of the body. The proposed studies test an entirely new model in which virus-like particles synthesized during learning/memory activity in brain neurons travel to fat storage tissues and induce mobilization of stored energy. Arc (activity-regulated cytoskeleton-associated protein) was known for decades to be induced by learning/memory activity in neurons, where Arc oligomers promote synaptic activity and plasticity. Arc proteins evolved from a retrovirus and retained the ability to assemble into virus-like capsids that spread from cell to cell. A ground-breaking hypothesis to be tested here proposes that Arc capsids travel from the brain to fat storage cells, where they signal brain activity and trigger release of energy into circulation. Levels of circulating energy feed back onto Arc expression via metabolic control of N6-methyladenosine (m6A) modification of Arc mRNA. Together, these coupled processes are proposed to comprise a homeostatic circuit that integrates the brain’s need for fuel and maintains organismal energy balance. The experimental system addresses the basic features of this circuit from the behavioral to the molecular level, including a conserved requirement for Arc in associative learning and cognitive dysfunction when excess dietary calories overwhelm the system. The planned research will determine properties of Arc required for communication with fat storage cells and how it alters organismal metabolism to supply energy to the brain. Other experiments will identify the key components of diet that alter m6A modification and virus-like Arc assembly and test custom diets designed to ameliorate cognitive dysfunction. This project will establish the mechanistic details of a previously unknown brain–adipose signaling axis and a homeostatic circuit where uncoupling leads to neurodevelopmental and neurodegenerative disease.

NIH Research Projects · FY 2025 · 2023-09

Project Summary Overview: This K24 mid-career investigator award in patient-oriented research and mentorship proposes to expand the research and mentorship activities of a highly qualified candidate, Dr. Lisa Abuogi, focused on improving HIV and health outcomes for people living with HIV in high burden, low resource settings. Candidate: Dr. Abuogi is a physician-scientist trained in pediatrics, global health, and HIV and an Associate Professor at the University of Colorado Denver (UCD). She is also a Senior Investigator in the UCD Center for Global Health and Medical Director of the Children's Hospital HIV Prevention Program. Dr. Abuogi conducts patient-oriented research to address disparities in HIV health outcomes, including retention in care and viral suppression, among pregnant women, adolescents and children living with HIV. She has a sustained track record of extramural funding and demonstrated dedication to successful mentorship of new investigators. Mentoring Plan: Dr. Abuogi has a demonstrated commitment to highly successful mentorship of a diverse group of mentees. She has the ability to recruit multidisciplinary mentees from a wide range of research collaborations and networks, a structured approach to ensure high quality mentorship, and a commitment to build the next generation of investigators committed to research aimed addressing knowledge gaps and at improving HIV outcomes. Research Plan: Dr. Abuogi's proposed research activities address NIH's Office of AIDS Research Priority Research agenda in Cross Cutting research that includes implementation and behavioral sciences, health disparities, and training and capacity building. Leveraging her ongoing NIH-funded research evaluating behavioral interventions to improve HIV care outcomes for both pregnant/postpartum and on 1, viral women and adolescents young adults living with HIV (AYA), Dr. Abuogi proposes innovative new patient-oriented research focused the sustainable impact f interventions aimed a improving engagement in care and viral suppression. In Aim Dr. Abuogi will examine the effect of short-term peer navigation on long-term sustained re-engagement and suppression in AYA on antiretroviral treatment after navigation is discontinued. In Aim 2, o t determine the impact of behavioral interventions on care transitions in these populations. Finally, in Aim 3, she will explore the perceptions of patients and LHW on how the interventions support treatment success as well as unintended consequences of peer-led interventions to inform iterative adaptations of these interventions. proposed evidence-based provide The newly research described in this application will expand our understanding of the sustainable impact of interventions aimed at improving treatment outcomes for vulnerable populations with HIV and research opportunities for new mentees.

NIH Research Projects · FY 2025 · 2023-09

Project Summary ADAM17 loss of function (LOF) is the cause of a pediatric enteropathy termed Neonatal Inflammatory Skin and Bowel Disease 1 (NISBD1), which is associated with intestinal inflammation and diarrhea. While little is known about NISBD1, two patient biopsies revealed abnormal intestinal crypt/villus morphology. The crypt/villus structure is the fundamental repeating intestinal unit lined by an epithelium that contains crypt restricted intestinal stem cells (ISCs). ISCs give rise to differentiated cells and are integral to maintaining epithelial function. ISCs are supported by specialized epithelial cells and underlying mesenchymal cells that provide ISC niche signals, including EGF-like ligands. ADAM17 is a ubiquitously expressed protease that cleaves and sheds proteins from the cell membrane. Its substrates include EGF-like ligands, suggesting a role for ADAM17 in the ISC niche. Recent work has revealed that ErbB3 ligand neuregulin 1 (NRG1) promotes human ISC maturation and differentiation. Human fetal intestinal tissue sections reveal mesenchyme restricted expression of NRG1 and epithelium restricted expression of its receptor ErbB3. This suggests a pattern of ADAM17 mediated mesenchyme to epithelium signaling crosstalk. I will test the hypothesis that ADAM17 mediates cellular crosstalk, regulating human intestinal epithelial development. First, I am establishing a matched isogenic set of NISBD1 patient iPSC lines with one CRISPR corrected to be ADAM17 proficient. I will differentiate both lines into human intestinal organoids containing organized epithelium and mesenchyme. Then, I will interrogate the ADAM17 LOF phenotype using cellular, transcriptomic, and functional analyses. Next, I will investigate ADAM17 mediated cellular crosstalk in a targeted approach by studying NRG1-ErbB3 signaling. To identify additional candidate ADAM17 substrates, I will take an unbiased mass spectrometry and proteomics approach to determine all human intestinal ADAM17 substrates, i.e. the sheddome. This analysis will establish datasets for future targeted investigation of ADAM17 substrates in the human ISC niche. In summary, I will perform an innovative patient specific disease modeling study using iPSC-derived organoids to characterize NISBD1 and study the role of ADAM17 in the developing human intestine.

NIH Research Projects · FY 2025 · 2023-09

PROJECT SUMMARY Trauma is the leading cause of death worldwide for people under 45 years old, and hemorrhagic shock remains the primary cause of early death after trauma. Later trauma deaths are frequently attributable to endotheliopathy of trauma (EOT), a systemic response to activated endothelial cells characterized by impaired blood flow, barrier integrity, and coagulation. Clinically, EOT manifests as a pro-inflammatory state of microcirculation leak and tissue edema, contributing to acute respiratory distress syndrome (ARDS), multi- organ dysfunction, and, eventually, death. Fibrinogen replacement restores the endothelial glycocalyx in vitro and in mice via stabilization of syndecan-1, a glycocalyx-based proteoglycan. Fibrinogen stabilization of syndecan-1 then mitigates EOT and restores microcirculation barrier integrity. However, it is unclear if fibrinogen-based restoration of endothelial integrity translates to improved clinical outcomes in humans. Further, current transfusion protocols in trauma provide fibrinogen too little or too late. This is a problem because trauma patients who develop EOT are twice as likely to die than those who do not. Therefore, restoring endothelial barrier integrity is essential to mitigating late morbidity and mortality in trauma. Accordingly, there is a critical need to determine the effect of early fibrinogen replacement on endothelial and organ dysfunction in critically ill trauma patients. Our preliminary data indicate improved patient-centered outcomes when fibrinogen is replaced within 6 hours of hospital arrival. However, we do not know whether endothelial restoration was the primary mechanism. To address this gap, we will test our overarching hypothesis that preserving endothelial function with early fibrinogen replacement will prevent ARDS and multi- organ dysfunction after trauma. We will test this hypothesis with the following specific aims: 1) determine the association between early fibrinogen replacement and multi-organ dysfunction; 2) determine the effect of early fibrinogen replacement on endothelial function; and 3) determine the cumulative effect of endotheliopathy on supplemental oxygen-free days. To achieve these aims, the candidate, David Douin, MD, will leverage his background in clinical research and the existing research infrastructure within the emergency medicine, trauma surgery, and anesthesiology departments. As an anesthesiologist and surgical intensivist, Dr. Douin is uniquely positioned to accomplish the proposed K23 research and career development aims. His long-term goal is to become an expert in novel interventions to prevent and treat multi-organ dysfunction and improve outcomes in critically ill trauma patients. Dr. Douin has assembled a multidisciplinary team of mentors with extensive clinical and translational research experience and topical expertise in traumatic injury, critical care, clinical trials, endothelial function, and lung injury to ensure his success in achieving the stated specific aims and career goals. This proposal will allow Dr. Douin to transition to an independent physician-scientist and prepare him for future NIH R61/R33 funding and, eventually, NIH UG3/UH3 or R01 funding.

NIH Research Projects · FY 2025 · 2023-09

PROJECT SUMMARY - R01 (Veress) Inhalation of sulfur mustard (SM) at high doses causes acute cardiopulmonary failure from a hypercoagulable state within the lungs, presenting as acute airway obstruction by fibrin airway casts. Acute and early systemic hypercoagulability also occurs, resulting in macro- and microvascular thrombosis within at least the pulmonary vessels, and likely other organs as well. However, most people who are exposed to SM inhale a lower dose of SM, resulting in minimal acute symptoms and acute fatalities. Nevertheless, their SM inhalation exposures results in the development of significant pulmonary morbidities, including delayed, long-term (often progressive) cardiovascular sequelae months to years after the acute exposure event. These late-onset morbidities from SM inhalation include chronic lung diseases, and chronic progressive cardiovascular diseases, such as pulmonary hypertension (PH), arterial hypertension (HTN) and cardiac dysfunction/failure. These disorders decrease quality of life for survivors, have no cure, and their pathogenesis are poorly understood. Conversely to SM, high dose chlorine (Cl2) inhalation is only fatal within a few hours after exposure, due to severe epithelial and airway edema, severe acute nervous system dysfunction, abnormal calcium storage/release, and acute vasoconstrictive pulmonary hypertension (PH). No acute fibrin casts form in the airways with Cl2, and intriguingly, recovery after Cl2 exposure does not result in any long-term cardiovascular morbidities. Mechanisms that protect from development of chronic cardiopulmonary diseases after Cl2 gas is of high interest. We developed, characterized and validated two relevant recovery models of SM and Cl2 inhalation in Sprague Dawley rats (LD50- 28d), both of which mimic the human recovery syndrome after these injuries. Preliminary data show that rats exposed to low dose SM inhalation develop not only late pulmonary fibrosis, but also significant progressive (worsening over time) PH, RV dysfunction, cardiac failure, and increased systemic arterial resistance late (>14- 21 days) after exposure – as measured by rat echocardiography and hemodynamics techniques. Additionally, we found that after acute recovery from exposure to Cl2 inhalation, a complete cardiovascular recovery occurs by 29 days (or earlier). Proteomics pathway analysis and histopathologic studies from 29 days after SM inhalation indicate significant ongoing endothelial cell pathway dysfunction, cardiomyocyte/myocyte pathway dysfunction, and continued coagulation abnormalities past the acute recovery phase. We hypothesize, that continued endothelial cell dysfunction during recovery following SM but not Cl2 inhalation will trigger persistent pro- coagulant and pro-remodeling pathways within the lungs, heart and systemic vasculature, and that this will result in the development of chronic thrombosis and myofibrillar hypertrophy, leading to late-onset chronic progressive pulmonary arterial dysfunction (PH), systemic vascular dysfunction (HTN), and cardiac ventricular dysfunction and failure. This proposal will develop hits for future therapeutic development against chronic cardiovascular sequelae of SM inhalation.

NIH Research Projects · FY 2025 · 2023-09

PROJECT SUMMARY The human gut microbiome harbors microbes with the capacity to cause infection or drive pathogenic inflammation. Immune status often determines risk for microbiome-associated disease, which is typically attributed to immune impacts on microbial community composition. But pathogenic or commensal lifestyles can also be dynamically regulated within individual microbes, and there is far less understood about immune impacts on microbial inherent pathogenic potential. Candida albicans is a morphologically and transcriptionally dynamic commensal fungus that can cause life-threatening infections and exacerbate pathogenic inflammation. The ability for C. albicans to cause disease depends on its phenotypic state. Of particular importance is the formation of hyphae, which are elongated cells specialized for adherence and invasion, and promote disease in both infection and inflammatory settings. Immune status is crucial for determining risk for C. albicans-associated disease and both immune deficiencies and active inflammation are linked to C. albicans pathogenesis. However, the role of immune environment on in vivo C. albicans pathogenic potential is not well understood. Here, I will investigate the impact of two human relevant immune environments on C. albicans pathogenic potential. Project 1 will focus on IgA regulation of C. albicans biology. Anti-C. albicans IgA antibodies are found in the gut of most people, and I previously found that C. albicans hyphae and associated effectors are heavily targeted by IgA during colonization. In mouse models, IgA targeting is associated with reduced hyphae and reduced capacity to exacerbate colitis. Here, I will interrogate mechanisms by which IgA regulates C. albicans biology in vivo using a mouse colonization model that permits investigation of immediate regulatory impacts of IgA targeting on C. albicans biology. Using this model, I will interrogate IgA impacts on C. albicans morphology and gene expression, which will include single cell transcriptional profiling to investigate gene expression in individual IgA-targeted cells. The goal of Project 2 is to define the impacts of inflammation on C. albicans pathogenic potential. Evidence from human IBD studies and mouse models of colitis suggest that this fungus exploits inflammation to bloom and perpetuate disease. Here, I will use a mouse model of intestinal colitis to define inflammation-dependent impacts on C. albicans morphology and transcriptional profile, with the goal of defining C. albicans pathways responsible for disease exacerbation. This proposal will reveal fundamental mechanisms by which immune environment regulates C. albicans biology and advance our understanding of how C. albicans becomes pathogenic in certain people. Broadly, these efforts will provide a foundation for our long-term goal of identifying targeted therapeutic strategies to prevent commensal C. albicans reservoirs from causing disease.

NIH Research Projects · FY 2024 · 2023-09

SUMMARY Giardia lamblia is a single-cell eukaryote that infects hundreds of millions of people every year. Because Giardia has many molecular pathways that are simplified compared to other eukaryotes, it has potential as a nontraditional model system for studying the diversity and evolution of key biological processes. We recently serendipitously discovered that the 2A ‘self-cleaving’ peptide sequences work very poorly in Giardia, surprising because 2A peptides are thought to work universally in eukaryotes. Found in picornaviruses like foot-and-mouth disease virus and poliovirus, 2A peptides are an essential part of the viral life cycle because they enable two polypeptides to be produced from one open reading frame. Although often referred to as ‘self- cleaving,’ 2A peptides operate by causing the ribosome to skip a peptide bond. The mechanism of this is unknown but must involve specific interactions between the 2A nascent peptide chain and the exit tunnel of the ribosome. Thus, our discovery that 2A peptides work poorly in Giardia points at fundamental differences in its ribosomes compared to other eukaryotes and can be exploited to understand the mechanism of 2A action. Examination of our recently solved structure of the Giardia 80S ribosome reveals a compelling difference in the structure of ribosome protein uL4 in the exit channel: Giardia lacks a specific loop in uL4. We hypothesize that this loop is important for the peptide bond-skipping mechanism of 2A peptides, and its absences can partially explain why 2A peptides operate poorly in Giardia. Here, we will test this hypothesis and in so doing (1) define the mechanism by which 2A peptides induce bond skipping and (2) determine why it fails in Giardia. We will combine genetic, biochemical, and structural approaches in two aims. In the first aim, we will determine the extent to which 2A sequence variants can function in Giardia, with the goal of finding novel efficient and functional sequences that will serve as powerful tools for Giardia researchers. In the second aim, we will directly test the functional role of the uL4 loop and solve the structure of a T2A-ribosome complex by cryo-EM with the goal of describing the mechanism of peptide bond skipping in eukaryotes. Overall, this work will provide critical knowledge about the function of the Giardia ribosome, the fundamental workings of the eukaryotic translational machinery, and the mechanism of 2A peptide function. Our discoveries will facilitate the development of Giardia as a model organism and help lay the foundation for new anti-viral therapeutics that block 2A peptide activity.

NIH Research Projects · FY 2025 · 2023-09

PROJECT SUMMARY/ABSTRACT: Type 1 diabetes (T1D) is an increasing public health burden for which current therapies are focused on insulin replacement. Future T1D treatments will try to preserve endogenous pancreatic beta cell function in new onset T1D and replace beta cells via transplantation in long standing diabetes. Classically, T1D is viewed as an autoimmune disease but beta cell loss is also driven by genetic risk factors and metabolic stress. Much has been done to try to delay T1D progression from preclinical multiple autoantibody positivity to clinical disease, but progression is highly heterogenous. C-peptide preservation can improve diabetes control and decrease the risk for diabetes complications. There are a wide array of C-peptide measurements to examine endogenous beta cell function and stress and many genes have been associated with C-peptide preservation in T1D. Less is known about the influence of genetics on the preservation of endogenous beta cell function during the partial remission period (PRM) or “honeymoon” after diagnosis. These factors are also likely important for patients with chronic pancreatitis (CP) who have risk for islet loss but for whom genetic risk and metabolic stressors remain largely unexplored. Patients with CP have severe pain and progressive endocrine insufficiency due to persistent inflammation and hepatic insulin resistance. For patients with severe CP, total pancreatectomy with islet auto- transplantation (TPIAT) is an attempt to preserve a patient’s endogenous beta cell function while removing the source of their pain. Younger age and higher beta cell mass during transplantation are predictors for functional graft survival. Less is known about the role of beta cell genetics in these patients, which I will evaluate in this proposal. I am an emerging researcher with experience in basic science beta cells studies and clinical research defining heterogeneity in T1D. My goal in this career development award is to hone clinical research skills by studying the impact of genetics and markers of beta cell function in patients with newly diagnosed T1D and in a model of cell therapy (CP patients who have undergone TPIAT) with these aims: 1: Analyze if baseline markers of beta cell function and stress can predict PRM C-peptide preservation. 2: Evaluate if known pre-specified T1D SNPs are associated with PRM C-peptide preservation 3: Evaluate if pre-specified SNPs associated with C-peptide preservation predict insulin use and C-peptide in CP patients who have undergone TPIAT As part of this K23 award proposal, I outline educational, training, and scientific goals that will support my pathway to independence. I have assembled a diverse and broad mentorship/collaborator team to support this endeavor. This career development award will support my career goal to elucidate heterogeneity in beta cell (dys-)function in diabetes and to develop novel cell therapy interventions.

NIH Research Projects · FY 2025 · 2023-09

PROJECT SUMMARY/ABSTRACT Our overarching goal is to recruit and follow a new cohort of pregnant women to implement longitudinal data collection using the ECHO Cohort Protocol, and to lead ECHO-wide analyses that advance understanding of when and how exposures during preconception, prenatal, and postnatal/lactational periods influence a broad range of child health outcomes. We will lead solution-oriented science to target adverse environmental exposures that may be prevented through policy, programs, and practices, and to identify positive influences in the preconception, prenatal, and postnatal periods that may be promoted to mitigate health risks associated with adverse early-life exposures. We will leverage our local investigator and staff expertise to enroll pregnant women and their children from a large Federally Qualified Health Center in Denver, Colorado. In Aim 1, we propose to utilize the breadth of Core data elements in the ECHO Cohort Protocol to answer solution-oriented research questions regarding the link between early-life exposures and childhood health outcomes, and to identify modifiable prenatal and postnatal factors that may minimize risk. In Aim 2, we propose to implement novel and scalable Specialized measures to determine biological pathways linking parent and family lifestyle factors and the physical/chemical environment of early life with child obesity and neurodevelopment, and to identify susceptible periods for these exposures in early life. In Aim 3, we describe our plans to recruit and follow a cohort of over 800 pregnant women with and their children, and to additionally enroll over 300 participants with a high probability of subsequent pregnancy into a preconception study. In Aim 4, we propose research questions that utilize data elements collected during the preconception period, to identify parental preconception exposures that adversely affect child health outcomes, and to describe molecular markers that may mediate these effects. Our team of expert investigators is well- positioned to contribute to the development of the ECHO specialized exposure and outcome protocols, and to the design and implementation the preconception pilot study. Our experienced staff and MPI team will ensure successful recruitment and implementation of the ECHO Cohort Protocol to provide a robust sample size and high-quality data. We will lead ECHO-wide analyses to address pressing questions in the early-life origins of child health and disease, and to identify potential targets for prevention of adverse environmental exposures and for promotion of protective factors in early life.

NIH Research Projects · FY 2025 · 2023-09

PROJECT SUMMARY/ABSTRACT Adolescent increases in psychopathology across multiple dimensions have a significant detrimental impact on morbidity, mortality and well-being during this period of life, and set the stage for adult physical and mental health difficulties. Prevalence rates of internalizing psychopathology, which is higher on average in females, and externalizing psychopathology, higher in males, diverge during adolescence, and this trajectory continues on subsequently over the lifespan, implicating sex-differentiated mechanisms. Consistent with this, a robust literature supports individual differences in pubertal timing (e.g., onset) and pubertal tempo (e.g., rate of change) as risk-factors for increased psychopathology. However, reliance on single sex-specific indicators, retrospective reports, and cross-sectional data to measure this dynamic period of maturation have made identification of mechanisms driving puberty and psychopathology links difficult. The current study aims to capitalize on measured genomics approaches integrated with longitudinal data in both sexes to inform the genomic signal of puberty across multiple physical and hormonal indicators, and examine genetic covariation between puberty and psychopathology across the lifespan. The first aim will leverage the novel method of genomic structural equation modeling (genomic SEM; training aim 1) to combine summary statistics from published genome-wise association studies (GWAS) of pubertal timing (i.e., age of menarche, relative age of voice break, relative age of first facial hair), pubertal growth spurt, pubertal maturation (i.e., Tanner staging), testosterone, estradiol, and sex hormone binding globulin (SHBG) to identify latent pubertal genomic factors both specific to and unified across sex. Polygenic scores (PGS; training aim 2) derived from the multivariate pubertal genomic factors will be validated by out-of-sample prediction of longitudinally measured pubertal timing and tempo characterized by multiple pubertal markers in the Adolescent Brain Cognitive Development (ABCD) Study. The second aim is to investigate measured genetic covariance of multivariate pubertal genomic signal with lifespan sex-differentiated psychopathology (training aim 3). This will be achieved through (a) examining correlations between the pubertal genomic factor model and previously established factor models of the genetic architecture of adult psychiatric traits using genomic SEM, and estimating pubertal PGS prediction of sex-specific lifetime psychiatric diagnoses in the UKBiobank; and (b) probing sex-specific and sex-unified pubertal PGS effects on longitudinally modeled adolescent symptoms of psychopathology in the ABCD Study, both directly and in conjunction with longitudinally measured pubertal timing and tempo. This research will yield a comprehensive model of measured genomic signal of puberty across multiple related phenotypes in both sexes, and provide improved tools for parsing genetic and non-genetic sources of covariation to disentangle multilayered mechanisms underlying pubertal risk for psychopathology.

NIH Research Projects · FY 2024 · 2023-09

PROJECT SUMMARY/ABSTRACT Skeletal muscle insulin resistance with type 2 diabetes (T2D) is, in part, caused by adipose inflammation and endocrine signaling. Intermuscular adipose tissue (IMAT) is a particularly problematic adipose depot as it resides under the muscle fascia and between myofiber bundles. Thus, it is uniquely situated for paracrine-mediated signaling towards muscle. Total IMAT content and its pro-inflammatory transcriptome relate to insulin resistance in humans, with the obese IMAT secretome milieu inhibiting myotube insulin sensitivity in vitro. However, it is unknown if the pro-inflammatory IMAT secretome is dynamic, contributing to muscle insulin resistance with obesity and T2D. There is a critical need to address this gap in knowledge in order to provide the framework for interventions to minimize IMAT-induced muscle metabolic dysfunction. The overall objective of this project is to quantify the IMAT secretome across BMI and insulin sensitivity and evaluate the importance of mitogen-activated protein kinase (MAPK) signaling in IMAT-induced muscle insulin resistance. We will utilize IMAT from the vastus lateralis of individuals with obesity or obesity and prediabetes/T2D in comparison to lean controls through a currently funded R01 (under the sponsor Dr. Bryan Bergman) to achieve the following specific aims. Aim I: Evaluate the extent to which intermuscular adipose tissue pro-inflammatory secretion differs across BMI and insulin sensitivity in humans. Based on our preliminary evidence that IMAT with obesity has a more potent pro-inflammatory secretome compared to other adipose depots, our hypothesis is that IMAT secretome scales to BMI and insulin resistance. A high-throughput quantitative proteomic assay will measure inflammatory proteins in IMAT conditioned media. Aim II: Elucidate the importance of MAPK signaling in intermuscular adipose tissue-induced skeletal muscle insulin resistance. Our preliminary evidence demonstrates that obese IMAT paracrine milieu upregulates c-Jun N-terminal Kinase (JNK1) and p38 activity, and downregulates insulin sensitivity, in human myotubes. Our hypothesis is that inhibition of MAPK signaling will help to alleviate IMAT- induced muscle insulin resistance. This will be tested by evaluating IMAT conditioned media-induced insulin resistance with either JNK1 or p38 inhibition through either viral siRNA knockdown or pharmacological inhibition in human myotubes. Our expected contribution is significant because IMAT is positioned as a strong candidate to explain the pathology of muscle insulin resistance that is central to T2D. This training plan includes sponsorship by Dr. Bergman at the University of Colorado Anschutz Medical Campus (CUAMC). The proposed research will enhance the skillset of the postdoctoral fellow in both clinical and in vitro methods. The postdoctoral fellow will complete additional training in endocrinology, obesity metabolism, and biostatistics through the Nutrition Obesity Research Center and Clinical and Translational Sciences Institute at CUAMC. This research, sponsor, and institution will collectively position me as an independent translational scientist investigating the role of skeletal muscle insulin resistance in T2D risk and progression.

NIH Research Projects · FY 2026 · 2023-09

Abstract/Summary Rheumatoid arthritis (RA), a common autoimmune rheumatic condition, has no cure and even with novel treatments, is associated with significant irreversible joint damage, physical disability, and numerous comorbidities. The appearance of serum anti-citrullinated protein antibodies (ACPAs) indicates RA-related autoimmunity and defines the start of the preclinical period of RA, which is the ideal time to identify relevant disease risk biomarkers and approaches for disease prevention. The etiology of RA has a genetic component, which may interact with environment in the development of disease. RA is characterized by excessive chronic inflammation, suggesting a failure in the ability to control/resolve inflammation. Mechanisms for both initiating and resolving inflammation are important for physiological homeostasis. Lipid mediators, products of the metabolism of omega-3 and omega-6 polyunsaturated fatty acids, are involved in both initiation and resolution of inflammation. We have shown that an elevated level of an individual omega-6 lipid mediator (5-HETE) increased risk of progression from RA-related autoimmunity to inflammatory arthritis. However, as lipid mediators share common pathways and enzymes, we propose that individual lipid mediators do not act in isolation and therefore should be analyzed in combination as a profile. We propose to conduct a study in three novel at-risk cohorts: the Targeting Immune Responses for Prevention of Rheumatoid Arthritis (TIP-RA) cohort of 81 ACPA+ individuals, the Studies of the Etiologies of RA (SERA) cohort of 79 ACPA+ individuals, and the StopRA cohort of 144 ACPA+ individuals in the preclinical period of RA that have been followed over time for the development of RA. We will create lipid mediator profiles (a composite score of combinations of highly correlated lipid mediators) indicating the ability to resolve inflammation by performing principal components analysis of the lipid mediators and look at the trajectories of these profiles over time. Aim 1 will determine if the association of these profiles with progression from ACPA+ to RA differs by genetic susceptibility to RA. We will also explore whether the association is mediated by cytokine profiles or trajectories. Aim 2 will explore the underlying mechanism by examining whether the lipid mediator profiles are associated with DNA methylation differences or trajectories. Aim 3 will examine inflammation resolution in the lung by identifying sputum lipid mediator profiles and cytokines associated with progression from RA-related autoimmunity to RA. Results from this work will provide the foundation for designing prevention studies by elucidating which combinations of lipid mediators play a role in inflammation resolution in preclinical RA. The inability to resolve inflammation can lead to chronic inflammation; a common pathogenic element of RA. Elucidating mechanisms as well as the site (ie lung) of inflammation resolution during the preclinical period of RA significantly contributes to the understanding of pathogenesis and development of innovative interventions for RA.

NIH Research Projects · FY 2025 · 2023-09

Project Summary During early brain development, GABA is the main excitatory neurotransmitter due to a reverse chloride gradient mediated via the chloride co-transporter genes NKCC1 and KCC2. Signaling via the nicotinic receptor, a7nAChR, is a key driver of the GABA excitatory/inhibitory [E/I] shift necessary for normative neurodevelopment. Dysfunctional maturation of GABAergic neurotransmission and E/I balance in the brain is implicated in the pathogenesis of several neurodevelopmental disorders (NDDs), including autism spectrum disorder (ASDs). Recently, our studies have identified a functional polymorphism in the Neuregulin 1 gene promoter, rs6994992, which is associated with elevated brain transcription of a novel NRG1 isoform, (NRG1-IV) and more critically lower levels of α7nAChR in the human brain. Signaling via the α7nAChR is a critical driver of the GABA excitatory/inhibitory (E/I) shift, mediated via the chloride transporters, NKCC1 and KCC2 and is also a known genetic risk factor for several NDDs, especially ASD, in the form of 15q13.3 microdeletion syndrome. In rodents, α7nAChR knockdown prevents the switch, resulting in aberrant maturation of GABAergic hyperpolarization and NRG1 is a key regulator of α7nAChRs; the specific role of NRG1-IV is unknown. In human studies, rs6994992 is associated with human cognition, sensory processing and anxiety behaviors, and data from our lab suggest attenuated sensory processing in babies carrying the T allele, and an interaction with maternal gestational dietary choline supplementation. Choline is a α7nAChR agonist, and a novel prenatal nutrient supplementation strategy shown to improve childhood behaviors and early brain development. In summary, the objective of this multidisciplinary proposal is the developmental characterization of a novel biological pathway linking NRG1, α7nAChR and NKCC1/KCC2 with regulation of E/I balance maturation, using a novel transgenic mouse (NRG1- IVtgNSE-tTA) genetically modified to express human NRG1-IV in brain and cutting-edge experimental approaches. We will test two main hypotheses.1). NRG1-IV overexpression contributes to atypical maturation of neocortical GABAergic signaling and E/I cortical balance, mediated via reduced α7nAChR and developmental expression of the chloride co-transporters NKCC1 and KCC2 and 2). Prenatal stimulation of α7nAChR, via dietary choline supplementation, will correct development of the switch, improve adult E/I cortical imbalance and neurobehavioral outcomes relevant to several NDDs, mechanistically via a7nAChR. This research will improve our knowledge of basic mechanisms of typical and atypical development related to a key signaling pathways involved in several NDDs, especially ASD, and identify new treatment/intervention approaches for people with developmental disabilities.