Indiana University Indianapolis

NIH Research Projects · FY 2026 · 2026-02

Project Summary/Abstract Vascular contributions to cognitive impairment and dementia (VCID) are a significant yet underexplored factor in dementia pathology, frequently coexisting with Alzheimer’s disease (AD). While pericytes, astrocytes, and microglia have been studied for their roles in cerebrovascular damage, platelets—typically found beyond the blood-brain barrier—may also contribute to vascular dysfunction. Platelets drive endothelial injury through pro- inflammatory signaling, extracellular matrix degradation, and thrombotic activity, yet their role in cerebrovascular decline remains unclear. Given their accessibility and known involvement in vascular disease, targeting platelets may provide a novel therapeutic avenue for VCID, an area currently lacking effective treatments. This proposal aims to investigate platelet-mediated mechanisms of vascular injury in VCID and determine whether inhibiting platelet surface receptor function can mitigate disease progression. We hypothesize that platelets adopt a hyperactive, pro-inflammatory phenotype that exacerbates cerebrovascular dysfunction and that inhibiting platelet glycoprotein VI (GPVI) signaling will alleviate these effects. To test this, we will assess platelet activation and vascular interactions over time in two VCID mouse models: diet-induced hyperhomocysteinemia (HHcy) and amyloidogenic Tg2576 mice. Using flow cytometry, multiphoton imaging, and histological analyses, we will characterize platelet activation states, surface receptor expression, and their infiltration into cerebrovascular structures. Additionally, we will determine whether GPVI depletion reduces platelet-driven vascular damage and inflammation, thereby improving cognitive and physiological outcomes. The successful completion of these aims will provide crucial insights into the overlooked contributions of platelets to VCID pathology and establish their potential as therapeutic targets. Furthermore, this project will enhance my technical expertise in neurovascular imaging, histology, RNAseq, and mouse modeling of neurovascular disease while contributing to my professional development as an independent researcher. With the resources available at Indiana University – Indianapolis, including the Alzheimer’s Disease Research Center and mentorship from Dr. Wilcock, this proposal will advance both scientific knowledge and my career trajectory in dementia-related research.

NIH Research Projects · FY 2026 · 2026-02

PROJECT SUMMARY This study aims to understand kidney stone formation by examining interactions between calcium oxalate (CaOx) crystals, bacteria, and host factors, with certain bacterial genes playing crucial roles. Kidney stones are increasingly common, especially among children, and current treatments have remained unchanged since the 1960s, focusing on preventing crystal formation rather than addressing the mechanisms that turn crystals into complex stones. Research has identified bacteria like Enterobacteriaceae in kidney stones, with experiments showing E. coli increases CaOx deposits in mice. The urinary microbiome differs in stone formers compared to healthy individuals, with some bacteria promoting stone formation and others offering protection. Genetic sequencing found two biofilm-related genes consistently present in stone-associated bacteria, suggesting potential targets for new treatments. Host immune proteins also contribute to stone formation, with stones containing layers of crystals and organic material. Preliminary data show certain proteins are more active when both E. coli and CaOx are present, forming layered stones. The hypothesis is that interactions between CaOx crystals, immune proteins, and bacteria facilitate stone formation. Aim 1 involves measuring how stone matrix proteins and bacteria interact to promote stone formation, while Aim 2 focuses on studying biofilm-related genes in stone-associated bacteria, using genomics and transcriptomics to identify and validate these genes. This research could lead to innovative treatments targeting specific bacterial genes and mechanisms to prevent and treat kidney stones.

NIH Research Projects · FY 2026 · 2026-02

Nationwide, over one million adults suffer from Exercise-Induced Laryngeal Obstruction (EILO) – also termed paradoxical vocal fold motion. EILO is a debilitating disorder characterized by inappropriate episodic adduction of the vocal folds and/or supraglottic structures during exercise, resulting in shortness of breath and/or loss of consciousness. Progress in the management of EILO is limited due to a lack of understanding of its pathophysiology. A significant gap is the lack of real-time pathophysiological data that captures dynamic upper airway changes during episodic EILO events in field settings. The current standard-of-care for EILO diagnosis requires continuous laryngoscopy during exercise; however, it is suboptimal, and unpredictable, with a 44% missed rate of EILO events. Hence, there is an urgent need for examination of EILO pathophysiology in real-time field-based settings. However, there are many unknowns about the causes and underlying pathophysiology of EILO that hinders development of effective treatments. There is consensus that the incidence of EILO peaks at higher ventilation levels during exercise; however, data supporting a pulmonary basis for EILO are lacking. Smaller upper airway size and associated changes in upper airway aerodynamics have been suggested as contributing factors, but their functional and clinical relevance for EILO remains unknown. To address these needs, this proposal leverages current advances in mechanical engineering, machine learning, exercise and respiratory physiology, as well as computational fluid dynamic modeling and medicine to provide quantitative predictive measures of EILO in realtime in field settings using a novel wireless mechano-acoustic device developed in our lab and delineate the biophysiological signatures of pulmonary abnormalities, size, and upper airway aerodynamics contributing to EILO in young adults. Our long-term goals are to determine the pathophysiology of EILO to enhance clinical assessment and establish efficacious treatments. Therefore, the specific aims of this application are to: 1) identify real-time physiological changes associated with EILO during exercise in field settings using an innovative, optimized wearable mechano-acoustic sensor developed in our lab, 2) identify the role of abnormal exercise ventilation and hypocapnia in laryngeal closure during symptomatic breathing in young adults with EILO, and 3) identify unique biophysiological factors of size and upper airway aerodynamics contributing to EILO among exercisers with and without EILO using magnetic resonance imaging and computation fluid dynamic modeling. A total of 120 young adults (60 with and 60 without EILO), 18-26 years of age, will be recruited to this study. The proposed research is significant in filling a crucial gap in our knowledge concerning the role of real-time pathophysiology from field-based monitoring in EILO and in yielding new insights into the pulmonary and biophysiological signatures of EILO. The fundamental knowledge from this research will have a significant clinical impact, benefiting both children and adults, and reducing the burden of illness caused by EILO.

NIH Research Projects · FY 2026 · 2026-01

Project Summary/Abstract: This NRSA proposal, tailored to Ms. Hibbard, provides high-quality predoctoral research training and career development centered upon her future goals. The sponsor’s excellent mentoring record, collaborations with leading biomedical researchers, and the outstanding environment at the IUSM and Indiana Center for Musculoskeletal Health (ICMH) will foster the successful completion of this project. Additionally, participation in the Preparing Future Faculty and Professionals program for ethics and grant writing courses, manuscript preparation, departmental seminars and journal clubs, as well as national meetings will enhance Ms. Hibbard’s career development towards becoming a well-rounded, independent investigator. Patients with chronic kidney disease (CKD) develop renal fibrosis, which is a common pathological manifestation of virtually all etiologies of CKD, and one of the major causes of end-stage renal failure. Currently, there are no direct therapies for this manifestation. My preliminary data demonstrate that fibroblast activation protein (FAP) is highly expressed in CKD kidney, and our initial RNAseq datasets support that FAP is associated with altered myofibroblast motility and matrix composition. Thus, our models of the interactions between fibrosis and the effects of FAP on this patient phenotype remain incompletely understood. The primary goal of the present application is to test new hypotheses regarding drivers of CKD fibrosis, including FAP as well as its direct targeting in pre-clinical studies. Although my initial results show increased Fap mRNA and protein in the kidney of mice with CKD, the effects of targeting FAP to reduce renal fibrosis, are unstudied. Thus, the central hypothesis is: FAP increases renal fibrosis onset and progression in CKD through enhancing matrix secretion and cell migration, and CD5/LNP-FAPCAR will target FAP+ activated fibroblasts to reduce pathologic CKD outcomes. In Aim 1, the role of Fap in progression of CKD fibrosis will be tested using FAP-KO mice, and in isolated cell culture studies. Aim 2 will test the translational, pre-clinical rescue of renal fibrosis during CKD using a novel targeted anti-fibrotic therapy and scRNAseq. By performing these studies, Ms. Hibbard will gain new research skills in utilizing state-of-the-art translational mouse models, bioinformatic skills, and CKD treatments. Collectively, this proposal will also provide excellent research, ethics, and written and oral presentation training to Ms. Hibbard, as well as test important disease mechanisms that result in kidney fibrosis, and its potential resolution.

NIH Research Projects · FY 2025 · 2026-01

Type 2 diabetes (T2D) has reached epidemic levels in the United States and, alarmingly, cases of early-onset T2D are steadily rising, with nearly 305,000 new cases diagnosed among 18 to 44 year-olds in the United States each year.1,2 Early-onset T2D (diagnosed before age 40) progresses more rapidly than older-onset T2D3 and is associated with increased risk of developing comorbid cardiovascular and kidney diseases and life-altering complications.2-5 Emerging adults (ages 18 to 29) with family histories of T2D have an especially high risk of developing early-onset T2D and associated complications.2,6 National Diabetes Prevention Programs (NDPPs) are the gold standard for T2D prevention,7 and emerging adults compared to older adults have lower participation and retention in NDPPs.8,9 The underlying reasons for emerging adults’ low NDPP participation, including this population’s T2D prevention-related health beliefs, have been ill-explored.8,9 High perceived threat and low perceived benefits related to the belief that T2D is inevitable paired with high perceived barriers and limited cues to action related to limited healthcare usage may have a powerful influence on emerging adults’ NDPP participation.10 Understanding the T2D prevention-related health beliefs of emerging adults with family histories of T2D can help identify opportunities to intervene to increase NDPP participation. The purpose of this study is to describe T2D prevention- and NDPP-related health beliefs of emerging adults with family histories of T2D, focusing on the perceived threat of developing T2D, perceived benefits and barriers of T2D prevention behaviors, and cues to action. A sample of 20 to 30 emerging adults will be recruited using three methods: recruitment from Indiana University Indianapolis and Bloomington and local community organizations, social media recruitment, and online recruitment from an Indiana-based research registry. Participants will be eligible if they are 18 to 29 years old, have at least one biological parent or sibling diagnosed with T2D, have a body mass index greater than or equal to 25, and have not participated in an NDPP. Data will be collected with a demographic survey and a semi-structured interview about T2D prevention- and NDPP-related health beliefs. Interviews will be transcribed and analyzed using qualitative descriptive methods as described by Sandelowski.11 Findings will elucidate how health beliefs influence T2D prevention behaviors of emerging adults with family histories of T2D including their NDPP participation and inform intervention development and NDPP adaptation to increase NDPP participation in this population to lower their risk of early-onset T2D. This grant will also support training of the principal investigator in conducting research with emerging adult populations, community-based participatory and intervention research, and postdoctoral fellowship preparation.

NIH Research Projects · FY 2025 · 2025-09

ABSTRACT ICU admission is a major life crisis that can result in severe distress for the patient's family, both during and after the hospitalization. This includes psychological distress such as anxiety and posttraumatic stress and also spiritual distress, including crises of meaning, purpose, or religious faith. Facing life and death decisions as the surrogate decision maker or experiencing the patient's death further increases distress. Prior interventions to support ICU surrogate decision makers have affected end of life care such as ICU length of stay for patients who die during hospitalization, but few have impacted surrogate well-being. We propose that for an ICU intervention to improve surrogate well-being, it must provide intensive spiritual and psychological support. Healthcare chaplains are highly trained professionals who provide spiritual and emotional support to patients and family members of all faiths and those with no faith affiliation in the healthcare setting. Chaplains are trained to listen deeply to an individual's religious and cultural experiences, providing opportunities to improve the care of marginalized groups who have faced discrimination in healthcare. Connecting religious or spiritual concerns and medical decisions also is a chaplain's skill. Although chaplains are part of the healthcare team in many US ICU's, research about the outcomes associated with chaplains' spiritual care is just emerging, and nearly all available randomized trials examined the effects of chaplains on patient outcomes. To address the need for interventions that reduce surrogate distress, our team has developed the Spiritual Care Assessment and Intervention (SCAI) Framework, an intervention for ICU surrogates that is rigorous, reproducible and consistent with spiritual care standards of practice. Our prior, single site randomized trial of the SCAI framework in the ICU demonstrated improved spiritual and psychological well-being for ICU surrogates in the intervention group, but the study population lacked religious and ethnic diversity. We now propose an NIH Stage 3 multi-center, randomized, attention controlled clinical trial of the SCAI Framework for surrogates to evaluate efficacy in a sample with geographic, religious, racial and ethnic diversity. The study will take place in 6 geographically diverse sites that are also diverse in terms of religion, race and ethnicity. The intervention will be delivered by skilled and highly trained chaplains. Specific aims are: to demonstrate the effects of chaplain- delivered spiritual care on psychological and spiritual outcomes including anxiety, distress, spiritual well-being, posttraumatic stress symptoms and complicated grief; to demonstrate the impact of chaplain-delivered spiritual care on medical decision making; and to examine the effects of spiritual care among historically underserved racial and religious subgroups. Results will guide hospital leaders, policy makers and the healthcare team regarding chaplains' role in improving surrogates' psychological and spiritual health and the quality of decisions for critically ill patients.

NIH Research Projects · FY 2025 · 2025-09

The goal of this NIA R25 application is to create an immersive research education program that will attract undergraduate science majors, ultimately increasing early-stage investigators pursuing research careers in Alzheimer's Disease (AD) and Related Dementias (AD/ADRD). The rationale for this program is based on the NIA-stated need to expand the skilled AD/ADRD research workforce by providing exposure to individuals early in their careers. One of the most daunting barriers preventing expansion in AD/ADRD research is the low number individuals obtaining STEM degrees, which is attributed to a large number of individuals leaving STEM majors. Thus, retention in STEM majors should be the key focus in expanding the AD/ADRD research workforce. Data shows that participation in undergraduate research programs have profound effects on the graduation rates of STEM majors. Additionally, research programs that aim to achieve a welcoming research environment increase the rates of students entering biomedical research. Thus, the summer research education program that we are proposing, “Investigating Neurosciences, a Summer Program to Ignite Research and Education (INSPIRE),” is intentionally designed to attract early-stage undergraduates, providing them with unique training to increase their success in obtaining STEM degrees, motivate their passion for research, and prepare them with the skills and confidence to pursue AD/ADRD research careers. Furthermore, we will create a safe and supportive research training environment through deliberate efforts to achieve excellence within our program staff and mentors through training activities and in our curriculum by intentionally recruiting high-quality speakers, educators, and discussants at multiple career levels. These goals will be met through achieving the following aims: 1) Create an immersive program that will provide an authentic, open-ended AD/ADRD summer research experience for early stage undergraduates, 2) Provide undergraduates with academic skills and AD/ADRD-neuroscience specific learning opportunities, and 3) Expose undergraduates to pathways for multiple AD/ADRD research careers. A large component of INSPIRE is to reach undergraduates who are unlikely to have exposure to transformative undergraduate research opportunities or are unaware of the benefits of such programs. Thus, the program will have extensive outreach, advertising and recruiting efforts at local and regional higher education institutions with neuroscience programs that lack extensive research opportunities.

NIH Research Projects · FY 2025 · 2025-09

PROJECT ABSTRACT This application describes my research on a plasma cell (PC) survival pathway in Multiple Myeloma (MM) to be performed within the context of a 5-year mentored career development plan. My ultimate goal is to become an independent physician-scientist in the area of MM as a laboratory-based Hematology/Oncology academic and to develop potential therapeutics for use in trials. Under the guidance of my primary mentors, Dr. David Allman and Dr. Alfred Garfall, at the University of Pennsylvania, I have developed a structured training plan consisting of intensive laboratory research, didactics, and oversight by an experienced faculty advisory committee. The proposed research will center on mechanisms of cell survival in MM mediated by the unfolded protein response (UPR) pathway by focusing on a key negative regulator of the pathway, BiP, and its potential as a therapeutic target. MM is an incurable plasma cell cancer that despite treatments is universally fatal. One key reason for lack of durable therapies is a limited understanding of survival pathways, such as the UPR. Professional protein secreting cells such as PCs are uniquely dependent on the UPR for survival. PCs produce 2-10,000 antibodies per second, and this extreme protein load actively engages the UPR to adapt to the stresses of high output protein production. If stress is unable to be resolved, the UPR transitions from pro-survival to pro-death. It is known that this transition occurs through gene transcription programs, but the specific programs and their time courses are unclear. I require further training in bioinformatic analysis of transcriptional datasets to tease apart this transition. The activation of these gene programs by the UPR is controlled by the negative regulator BiP. Its removal leads to UPR activation, making it both a good target to study the pro-survival to pro-death transition but also as a therapeutic target to force prolonged UPR activation and death. By using a novel BiP-targeting agent and novel MM model systems, this proposal will characterize changes in gene expression during the UPR- mediated pro-survival to pro-death transition and determine what death pathways are responsible for this transition (Aim1). It will also test the efficacy of novel BiP-targeting approaches in untreated and relapsed MM in a patient derived xenograft mouse model (Aim2). These findings will provide insight into the biology of MM survival mechanisms, provide the rationale for BiP-targeting as a therapeutic approach and provide data for larger grant applications. In undertaking the proposed studies and training plan, I will develop the skills and expertise necessary to establish an independent career in translational research.

NIH Research Projects · FY 2025 · 2025-09

The NIH-wide Social Determinants of Health (SDOH) Research Coordinating Committee defines SDOH as “the conditions in which people are born, grow, learn, work, play, live, and age, and the wider set of structural factors shaping the conditions of everyday life.” The interactions between SDOH and individual, family, and community level factors require researchers to address systems and structures to improve health for Americans. Nurse scientists and scientists in aligned professions have limited opportunities for training to conduct SDOH research. To address this gap, our team from the IU Schools of Nursing and Medicine (IUSON and IUSM) propose to develop and implement the Midwest Leadership in Education, Application, and Dissemination of Research on the Social Determinants of Health (LEADRS) Program.   We will engage three cohorts of 20 scientists (PhD students, post-doctoral trainees, and early/mid-career faculty) over 9 months each, with 8 seats reserved for Institutional Scholars and 12 for Regional Scholars. Institutional Scholars: the eight scholars participating via this track will include four IUSON PhD students, postdoctoral fellows, or faculty in their first three years of appointment and four doctoral students, postdoctoral fellows, or faculty in their first three years of appointment from aligned health schools at Indiana University. Regional Scholars: all twelve scholars participating via this track will be nurse scientists; eight will recruited through our partnership with the Midwest Nursing Research Society (MNRS), allowing for competitive recruitment of nurse scientists from the Society’s 13-state catchment area, and four will be recruited in partnership with two Midwest Schools of Nursing. Regional Scholars will be a) faculty at the assistant or associate level, or b) scientists who conduct or direct research in non-academic settings (e.g. major health organizations, public health departments, non-profits).  The Education Pillar will consist of the Midwest SDOH Research Intensive, a four-day workshop in SDOH principles and methods. As part of the Application Pillar, scholars will apply learned concepts to their research, and to research needs of community partners. As part of the Dissemination Pillar, scholars will present locally (home institution) and submit presentations to regional, national, and international conferences.  The overall goal of the Midwest LEADRS Program is to prepare nurse scientists and scientists from aligned health professions to conduct and evaluate research that addresses the SDOH to improve health. Its aims are: Aim 1: Educate nurse scientists and scientists in the aligned health professions on key SDOH concepts, frameworks, and research methods to address SDOH and improve health. Aim 2: Support the development of new research and refinement of existing research to address critical gaps in SDOH. Aim 3: Evaluate, refine, and disseminate key resources from the program, making them easily accessible for nurse scientists and the scientific community.

NIH Research Projects · FY 2025 · 2025-09

Health-related social needs (HRSNs), sometimes called social determinants, are drivers of morbidity, mortality, healthcare utilization, and costs. Per the Centers for Medicare & Medicaid Services (CMS) HRSN data will be used in quality reporting metrics, in referrals, in adjusting payments, for public reporting, and in value-based care. To better inform service delivery and policy, it is crucial to understand whether screening tools result in valid measurement of HRSNs. Problematically, current HRSN screening tools may be inaccurate and have unknown performance among older adults. Our long-term goal is to enable effective care delivery and optimal policy decisions that leverage information on the social needs that drive health. The objective of this proposal is to advance the science of HRSN measurement and assess the implications of current HRSN screening approaches for Medicare policy. Our central hypothesis is that current HRSN screening tools disadvantage older adults covered by Medicare and will undermine efforts to create effective patient care and population health. Drawing on strong preliminary data and extensive domain expertise, our multidisciplinary team proposes the following specific aims. 1) Compare the validity of health-related social needs screening tools for older adults will establish the sensitivity, specificity, and positive predictive values of 4 screening tools (CMS, Epic, PRAPARE, Health Leads) against reference (i.e. “gold”) standard measures in a sample of 4,000 Medicare beneficiaries. We hypothesize that HRSN screening tool performance among older adults will be worse than the accepted performance thresholds for clinical utility. 2) Compare the performance of health-related social needs screening tools in predicting healthcare quality and costs for older adults will test the performance of the 4 HRSN screening tools and reference standard measures in predicting subsequent quality and costs outcomes. We hypothesize that the reference standard measures will explain more of the variance in costs and quality outcomes than the HRSN screening tools. 3) Evaluate the content validity of health-related social need screening for older adults will use key informant interviews with older adults, their caregivers/family members, and their clinicians, we will assess the appropriateness and relevance of different HRSN screening tools. This project will advance practice from a “one-size fits all” screening policy to a scientifically justifiable approach that leverages the most effective screening tools and incorporates HRSNs most relevant to older adults. This project is significant as: 1) CMS policy prioritizes HRSN screening as a means to better population health; 2) HRSNs are critical to the well-being of older adults; and 3) unvalidated HRSN screening tools could lead to undesirable downstream effects on future operation and policy decisions. This proposal is innovative because it rigorously evaluates HRSN screening tool validity against reference standard measures and considers on the appropriateness of HRSN screening tools.

NIH Research Projects · FY 2025 · 2025-09

Children with chronic kidney disease (CKD) are plagued by debilitating cardiovascular and mineral bone disease (CKD-MBD) with cardiovascular mortality rates that are 1000 times greater and fracture rates that are 2 to 3-fold higher than in healthy children. Our previously published work demonstrated population-level differences in CKD-MBD including differences in cardiovascular mortality and differences in the bone biomarkers 25-hydroxyvitamin D and parathyroid hormone (PTH). There is limited understanding of nonclinical correlates like the impact of socioeconomic status on long- and short-term CKD-MBD outcomes. Given the reliance of this life-limiting comorbidity on dietary intake and health behaviors, there is a critical need to define intervenable social-environmental characteristics that impact CKD-MBD outcomes. As such, the objective of this application is to integrate clinical data and the social determinants of health to map the social-environmental exposome and define its relationship to CKD, particularly vitamin D status. The central hypothesis of this research is that adverse social-environmental characteristics increase the risk of CKD comorbidities like vitamin D deficiency. Aim 1 will develop a facile, secure SDOHKIDney dataset that integrates data from electronic health records and geocoded census-level data on the social determinants of health. Aim 2 will use SDOHKIDney to define social-environmental risk factors for vitamin D deficiency. This work will expand upon my current K23 which seeks to understand the genetic contributions to population-level differences in CKD-MBD and set the foundation for investigations of gene by environment interactions and the impact of early CKD interventions on vitamin D on CKD outcomes like progression and hyperparathyroidism.

NIH Research Projects · FY 2025 · 2025-09

Medicare is the dominant payer for the >800,000 US patients with end-stage kidney disease (ESKD). These patients have poor outcomes and high medical spending, and there is wide variation in the quality and value of care across communities. Medicare is increasingly using novel payment models to contain medical care spending and improve care quality, including for ESKD. Starting in 2022, nephrology practices could choose voluntarily to participate in one of Medicare’s 4 Kidney Care Choices payment models; participants would bear financial risk for the medical costs and quality of care for their attributed patients with ESKD or chronic kidney disease stage 4 or 5. As of 2024, 123 provider groups were participating in a Kidney Care Choices model, representing >2,000 practices and ~2,700 nephrologists who care for an estimated >115,000 US patients with ESKD (>15%). Evidence on these models’ precursor, the Comprehensive ESRD Care Initiative (CEC)—including key evidence prepared by the proposed study team—showed that smaller (vs. larger) practices were less likely to participate in CEC, and the mortality and medical spending outcomes of CEC-participating practices’ attributed patients tended to be improved most in high socioeconomic-status communities. Because Kidney Care Choices includes several novel provisions (e.g., emphasizing ESKD prevention and transplant access), it is unclear i) how nephrology practices are deciding whether to participate in Kidney Care Choices and the roles of physician, practice, community, and policy-level factors in these decisions, and ii) how participating practices are reforming care models to reduce medical spending and improve quality. The proposed R03’s research objective is to examine practices’ participation decisions and practice reforms under Kidney Care Choices through a rigorous qualitative study. Leveraging PI Wilk’s strong training and applied practice in qualitative methods under his K01 Award (DK128384), we will conduct ~24 in-depth interviews with nephrology practice leaders to learn their perspectives on these topics. Aim 1 will examine how multi-level factors affect provider decisions about participating (or not) in Kidney Care Choices through in-depth interviews with nephrologists and practice managers, including model participants and non-participants. Aim 2 will explore how providers are approaching changing their care practices to improve financial performance and quality under Kidney Care Choices through in-depth interviews with model-participating nephrologists and practice managers. Thematic analyses of our rich interview data will yield valuable evidence on nephrology practices’ decisions to participate (or not) in Kidney Care Choices and how they are innovating in their care practices under one of Medicare’s largest payment reforms in specialty care. Our findings will guide future R01-level studies evaluating the health and health care impacts of novel payment reforms and practice leaders’ strategies for implementing care practice reforms intended to achieve system-wide goals of reducing health care spending and improving care quality for patients with chronic illness.

NIH Research Projects · FY 2025 · 2025-09

Project Summary Alcohol-associated liver disease (ALD) is a complex disease and represents a spectrum of histopathological changes in the liver. Accumulating evidence suggests that multiple types of immune cells are involved in the pathogenesis of alcohol-associated hepatitis (AH), particularly neutrophils. However, the mechanisms underlying neutrophils in mediating AH pathogenesis are not well understood. Our exploratory experiments had led to an important and clinical observation on the inter-patient variability and two distinct patterns of hepatic neutrophil infiltration. Our data also indicated that higher level of hepatic parenchymal neutrophils was associated with AH disease severity. The key question we would like to address in this application is how neutrophils drives disease severity and mediates liver injury in ALD. Mechanistically, neutrophils, as an innate inflammatory response, mediates tissue injury by producing inflammatory mediators and reactive oxygen species (ROS). ROS production in neutrophils is regulated by the multi-meric transmembrane enzyme complex, nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX). Among them, the NCF1 gene, which encodes the phagocytic oxidase (phox) unit p47phox, is predominantly expressed in neutrophils and plays an important role in controlling ROS production in neutrophils. Our data suggested that miR-223, the most abundant neutrophilic miRNA, may be a downstream target of NCF1 and its induction ameliorates alcohol-induced liver injury. In specific aim#1, we will determine the molecular mechanism on the inhibition of neutrophilic miR-223 by NCF1. Our data and planned experiments in specific aim#1 will expand our current knowledge on the signaling pathway in the neutrophils, neutrophilic Ncf1-induced oxidative stress, and miR- 223 expression. The next important question to address is how the inhibition of miR-223 by NCF1-induced oxidative stress leads to hepatocyte injury. In specific aim #2, we will determine the downstream crosstalk between neutrophil-induced ROS generation and hepatocytes via extracellular vesicles (EVs) leading to alcohol-induced liver injury. Taken together, we have developed animal and cellular models to mechanistically examine the roles of neutrophils in ALD pathogenesis. Understanding the mechanism linking neutrophils to ALD pathogenesis is of importance; this will pave a way for the discovery of potential targeted therapy for patients with ALD.

NIH Research Projects · FY 2025 · 2025-09

Alcohol use disorder (AUD) is a complex condition influenced by both genetic and environmental factors, significantly affecting millions worldwide. This project aims to elucidate the genetic and regulatory mechanisms underlying AUD in several brain regions previously implicated in alcohol consumption and addiction, namely, the caudate nucleus, extended amygdala, anterio-lateral hypothalamus, and middle temporal gyrus. Utilizing advanced single-cell multi-omics techniques, we will identify cell-type-specific gene expression and chromatin accessibility differences in these brain regions between individuals with and without AUD. Specific Aim 1 will involve performing differential expression and chromatin accessibility analyses using single-nucleus RNA-seq and ATAC-seq data. We will identify cis-regulatory mechanisms driving these differences by integrating expression quantitative trait loci (eQTL) and genome-wide association studies (GWAS) data. Additionally, we will infer trans-regulatory mechanisms using advanced bioinformatics tools to identify transcription factors and regulatory proteins involved in AUD. Specific Aim 2 will focus on identifying genes likely to significantly contribute to development of AUD, using Mendelian randomization and elucidating their functions through high-throughput perturbation experiments and real-world validation. We will perform eQTL analysis to identify variants associated with gene expression in specific cell types, followed by two-sample Mendelian randomization to determine causal relationships. High-throughput Perturb-seq will be used to elucidate the downstream functions of these causal genes, and we will leverage drug-target perturbation databases to identify potential therapeutic targets for AUD. The fellowship training plan includes developing expertise in cutting-edge genomics and bioinformatics techniques, gaining hands-on experience with experimental methods such as Perturb-seq, and enhancing scientific communication skills. Training will take place in Dr. Yunlong Liu's lab at Indiana University School of Medicine, providing access to state-of-the-art facilities and a collaborative research environment. This comprehensive training will prepare me for a successful career as an independent researcher in genomics and neurobiology, contributing to the development of targeted treatments for AUD and advancing public health.

NIH Research Projects · FY 2025 · 2025-09

PROJECT SUMMARY/ABSTRACT Dysfunctional cellular processes contribute to the pathogenesis of many human diseases. Genes and their functional products are central to the molecular mechanisms of all cellular processes. As such, determining the regulation and molecular mechanisms of uncharacterized genes may reveal mechanisms controlling cellular processes that could be exploited to improve disease. Long noncoding RNA genes (lncRNAs) are a type of regulatory noncoding RNA that contain 200 or more nucleotides and don’t encode proteins. They are increasingly recognized as critical cell and context specific regulators of cellular functions. There are potentially tens of thousands of lncRNA genes; however, the function and molecular mechanism of most are unknown. Determining these may reveal mechanisms regulating cellular processes that could lead to more specific ways of targeting cellular defects that contribute to human disease. LncRNAs are not supposed to code for proteins, but recent systematic analyses found an appreciable proportion contain unrecognized small open reading frames (sORFs) that are translated into functional microproteins of less than 100 amino acids. Some microproteins regulate key cellular processes; however, the vast majority of putative microproteins encoded by lncRNAs remain functionally and mechanistically unvalidated. Furthermore, some microproteins encoded by lncRNA genes exhibit functions distinct from those of their parent lncRNAs. These “bifunctional” genes (i.e., possess coding and noncoding functional elements) are poorly understood and pose intriguing questions about their regulation and cellular mechanisms. Our long-term goal is to understand how bifunctional genes are regulated and how they control fundamental cellular processes. Towards this goal, we have identified a bifunctional gene currently annotated as a lncRNA that is mainly expressed in T cells and cortical neurons. Its microprotein-dependent effects play a crucial role in protecting against pathogenic T cell responses, while both its microprotein-dependent and independent effects regulate cytokine responses of T cells. However, its transcriptional and translational regulation and the molecular mechanisms of its microprotein and lncRNA molecule are undefined. In addition, its regulation, function, and molecular mechanism in neurons are unknown. Here, we propose using this bifunctional gene as a model and leveraging our expertise in lncRNA and microprotein cellular and molecular biology to address the following questions: 1) What are the transcriptional and translational mechanisms regulating its RNA and microprotein expression? 2) What are the molecular mechanisms of its RNA molecule and microprotein? 3) How do different cellular contexts influence the regulation of expression and the molecular mechanisms of its microprotein and RNA products? By addressing these questions, we will establish a template for characterizing bifunctional genes thereby enhancing understanding of this growing group of genes and their contributions to human disease.

NIH Research Projects · FY 2025 · 2025-09

Despite population-level improvement in cardiovascular disease (CVD) incidence and mortality, CVD still disproportionately burdens Black adults. Recognizing that elevated stress exposure among these groups may contribute to CVD risk, the American Heart Association recently called for the development of interventions for blood pressure (BP) and cardiovascular health (CVH) that reduce stressors and enhance one’s stress response. The latest research on stress indicates Black adults report greater stress exposure and are a logical focus for advancing stress science. Thus, I will work with Black patients receiving care from federally-qualified health centers to iteratively design and pilot test the operable feasibility of a stress management intervention among Black adults with elevated blood pressure. Changes in stress and ambulatory blood pressure will be examined as well. Due to the design process, the resulting intervention will address patient preferences and thus may be an engaging and sustainable strategy in an externally valid setting for the promotion of cardiovascular health in a high-risk subpopulation. This has the potential to complement existing interventions, as cardiovascular health is not solely affected by traditional CVH risk factors (e.g., Life’s Essential 8). The stress management intervention will be informed through both primary data collection (Aims 1-2) as well as existing literature on supraindividual factors that protect against stress and its cardiovascular health consequences. In a user-centered design approach, the intervention will be iteratively developed and prototyped with Black adults with stress. In Aim 3, I propose to feasibility test, in a Stage 1b pilot, the stress management intervention among Black adults with stress and elevated blood pressure. Measures will be at baseline and posttest and assess feasibility; stress and blood pressure will also be measured. A successful pilot would support a large-scale stress-management intervention across safety-net primary care sites with the objectives of cardiovascular disease prevention and reduction of CVH disparities. These aims provide experiential learning in CVH, user-centered design, and primary data collection and behavioral intervention trial methods. By the end of this award period, I will be an independent scientist in behavioral intervention research with a focus on stress and CVH.

NIH Research Projects · FY 2025 · 2025-09

PROJECT SUMMARY/ABSTRACT Duchenne muscular dystrophy (DMD) is a devastating X-linked skeletal and cardiac myopathy resulting from a defect in the gene coding for dystrophin. DMD myopathy leads to loss of ambulation, respiratory failure, cardiomyopathy (CM), and premature death. Since the discovery of the DMD gene nearly three decades ago, the field has been dominated by the expectation of a “cure”; this has not yet transpired. Gene therapies have been approved but the cardiac effects remain unknown. Supportive care advances have decreased death from respiratory failure but simultaneously unmasked the lethal and fully penetrant CM phenotype, which is now the leading cause of death in DMD. We have documented that DMD CM progresses from normal imaging to diastolic dysfunction with subtle strain abnormalities to manifest systolic dysfunction; these changes correspond pathologically with myocardial inflammation and progressive increase in myocardial fibrosis and fibrofatty infiltration. Guideline-directed medical therapy (GDMT) has become standard of care for patients with DMD, but individual response to these therapies is suboptimal and often only serves to minimally delay the inevitable progression to heart failure and early death. Sodium/glucose cotransporter-2 inhibitors (SGLT2i) have resulted in mortality benefits in adult CM patients. The mechanism of this beneficial effect is unknown. Several factors specific to DMD suggest that SGLT2i could have potential for DMD CM – 1) improved cardiac energetics and metabolism (specifically, switch from carbohydrate towards fatty acids, ketone bodies, and amino acids metabolism); 2) improved mitochondrial function and reduced oxidative stress; 3) anti-inflammatory; 4) myocardial fibrosis inhibition; 5) reduction in sarcoplasmic calcium leak; 6) improved sympathetic overdrive. Our prior work, funded by the NIH/NHLBI and the FDA, has created the DMD Cardiovascular Care Consortium (DMDCCC), a multi-center consortium of 9 institutions following over 1350 DMD patients. The primary goal of the DMDCCC is to discover surrogate outcome measures of CM for use in clinical trials and to leverage the DMDCCC infrastructure for future clinical trials for DMD CM. The central hypothesis of this proposal is that SGLT2 inhibition is safe and well-tolerated in DMD patients with early CM. Aim 1 will define the SGLT2i pharmacokinetic (PK) profile in pediatric DMD patients. Aim 2 will determine the safety and tolerability of SGLT2i in younger DMD patients with early CM. Aim 3 will evaluate the clinical efficacy and blood biomarker alterations as a result of SGLT2i therapy in DMD, with a goal of informing future, larger efficacy studies. This study performs a critical evaluation of the pharmacokinetics, safety, and tolerability of SGLT2i in DMD. The potential of SGLT2i’s in DMD is immense, but a more complete evaluation must be performed before equipoise is lost.

NIH Research Projects · FY 2025 · 2025-09

Not Applicable

NIH Research Projects · FY 2025 · 2025-09

Project Summary / Abstract Alzheimer’s Disease (AD) is a devastating neurodegenerative disease characterized behaviorally by memory loss typically later in life and is ultimately fatal. Early detection of potential AD biomarkers in asymptomatic patients will promote disease prevention strategies. Migraine disorder is a disabling chronic disease condition that affects over 15% of the general population. Clinical reports show that AD is prevalent in individuals with a history of migraine, and that the incidence of AD is significantly higher in those that suffer from migraine at a younger age. There is a lot of overlap in the CNS neuroinflammatory pathways involved in the pathogenesis of both migraine disorders and AD, however, a mechanistic and behavioral understanding of the molecular contributors to migraine-induced AD remains unknown. A major barrier to progress is that the AD field has not reverse translated this increased AD incidence in migraine patients to preclinical animal models. Here, we show that transgenic AD mice exposed to chronic migraine via a series of injections with the migraine-causing drug, Nitroglycerin, don’t learn as well as non-migraine AD mice during a cognitive task. Additionally, we show increased amyloid accumulation in the brains of AD mice treated with Nitroglycerin. The central hypothesis of this proposal is that chronic migraine accelerates the development of AD pathology and associated cognitive decline in preclinical AD mouse models. In Aim 1, we will determine how chronic migraine affects multiple domains of cognition in a preclinical mouse model of AD. We will utilize an identical test that is currently used in the clinic to diagnose AD. In Aim 2, we will work with the IUSM Biomarkers Core to run spatial transcriptomics and proteomics in order to determine whether migraine progresses the activation of particular gene and protein expression patterns known to be altered in AD transgenic mice. We will measure key pathological features of both AD and migraine including amyloid plaques, multiple forms of tau, and CNS inflammatory markers in hippocampus, an area of the brain known for its role in spatial learning and memory processes. In both Aims, we will assess for potential sex differences. Indeed, female patients with a history of migraine have a much higher risk of developing AD compared to age-matched males. This area of research is highly novel to the AD field in that we are the first to reverse translate the increased human AD incidence in chronic migraine sufferers to preclinical animal models and the first to identify overlapping transcriptome and proteome targets associated with both diseases. Based on the spatial-omics data collected here, we will have the necessary rationale in our future R01 application to determine whether targeting various proteome biomarkers restores cellular function in various regions of the brain including hippocampus, frontal cortex, and in areas of the brain that greatly contribute to the onset of migraine and AD pathology including the locus coeruleus. A large percentage of the human population suffer from migraine disorders and will greatly benefit from this area of work that will allow for us to identify early biomarkers of AD in chronic migraine sufferers.

NIH Research Projects · FY 2025 · 2025-09

PROJECT ABSTRACT Diabetes is a key driver of serious non-AIDS events in people with HIV (PWH). Depression potentially contributes to new-onset diabetes in PWH. Collectively, our preliminary data indicate that successful depression treatment with internet cognitive behavioral therapy for depression (iCBT-D) could reduce diabetes risk in PWH by improving gut integrity, reducing monocyte activation, and preventing worsening insulin resistance. In addition, our data suggest that examining the metabolome may elucidate new mechanistic and potentially therapeutic pathways linking successful depression treatment to improved insulin resistance. However, these promising findings are preliminary and require confirmation in a sufficiently powered RCT. Our multidisciplinary group of investigators propose the following Specific Aims: Aim 1: To determine the effects of iCBT-D on insulin resistance in depressed PWH on ART. We hypothesize that iCBT-D, compared to active control, will prevent worsening insulin resistance as measured by HOMA-IR at 24 weeks. In secondary analyses, we will also compare changes in HbA1c and glycated albumin as endpoints more proximal to diabetes diagnosis. Aim 2: To determine the effects of iCBT-D on monocyte activation in depressed PWH on ART. We hypothesize that iCBT-D, compared to active control, will lead to reduced circulating sCD14, sCD163, and CD14+CD16+ intermediate monocyte levels at 24 weeks. In secondary analyses, we will examine the effects of iCBT-D on markers of gut barrier integrity (REG3α) and microbial translocation (16S rDNA and β-D-glucan). Aim 3: To determine the effects of iCBT-D on metabolomic profiles in depressed PWH on ART. We hypothesize that iCBT-D, compared to active control, will result in changes in the circulating metabolome at 24 weeks. We will then perform formal pathway analysis based on differential metabolite profiles to identify novel mechanistic biologic mechanisms to explain the treatment group - HOMA-IR relationship. We will test these hypotheses by conducting a 48-week RCT comparing iCBT-D with an active control comparator group (depression education, depressive symptom monitoring, and current care for depression in the HIV clinics) in 150 depressed PWH on ART. The primary analyses will be performed at the 24-week timepoint with similar secondary analyses performed at the 48-week timepoint to assess duration of response. Given the high prevalence of depression in PWH of up to 40%, depression treatment could have significant and widespread benefits by preventing diabetes in addition to its mental health importance. Thus, this application meets the stated objectives of PAS-24-163, “Priority HIV/AIDS Research within the Mission of NIDDK.”

NIH Research Projects · FY 2025 · 2025-09

Abstract The advent of fast and affordable sequencing technologies has facilitated detailed descriptive annotation of many mammalian genomes. In contrast, functional annotation connecting genetic variants to phenotypes is severely lacking, due to an inability to conduct current methods at scale. Removing this bottleneck will facilitate rapid identification and characterization of key regulators of physiological and disease processes, which is necessary for the development of targeted therapeutics. Methodologies based on high-throughput functional genomics offer a rapid, affordable, and systematic way to characterize gene functions, with CRISPR-based pooled forward genetic screens being one such method, which has been used to great effect in cultured cells. However, due to multiple barriers, the in vivo application of forward genetic screens has thus far been extremely limited, hindering our ability to study tissues that cannot be adequately modeled in vitro, such as the heart and brain. In this work, we assess the obstacles hindering extensive adoption of forward genetic screens in vivo and propose innovative techniques to overcome these challenges. We will utilize the heart for proof-of- concept studies that will examine regulation of Nppa, a gene encoding the cardiovascular disease biomarker atrial natriuretic peptide (ANP). Our methodologies can be adapted to many tissues and will provide significant value to diverse research initiatives. Furthermore, we propose a groundbreaking in vivo forward genetic screen for regulators of cellular proliferation – a critical biological process relevant to a wide variety of fields such as cancer biology, development, and tissue regeneration. By merging several cutting-edge technologies, our strategy will enable efficient identification of regulators of cardiomyocyte proliferation. This study, which is currently not possible using conventional screening methods, will serve as a blueprint for employing systematic functional genomics in tissues that are difficult to model in vitro or to deliver molecular components to in vivo. In sum, successful execution of the strategies described in this proposal will considerably reduce the level of expertise and resources that are required to take advantage of the unbiased systematic nature of in vivo forward genetic screens, leading to the improvements in functional annotation of the genome that are necessary to inform the design of therapeutics.

NIH Research Projects · FY 2025 · 2025-09

Project Summary Chronic and excessive alcohol consumption is known to cause multiple diseases including alcohol- related liver disease and liver cancer. It is estimated that approximately 25% of cirrhosis deaths and ~20% of hepatocellular carcinoma (HCC)-related deaths in the world are associated with alcohol. However, the pathogenesis of the alcohol-related HCC remains incompletely understood. Tumor suppressor genes are known to have protective functions against cancer development. But very little is known which tumor suppressor genes are crucial in the alcohol-related HCC. In this application, we will use both candidate and systemic approaches to identify and characterize key tumor suppressor genes and their roles in the alcohol-related HCC development. It is expected that the findings from this innovative research will provide insights into risk assessment, HCC prevention, and therapeutic development.

NIH Research Projects · FY 2025 · 2025-08

PROJECT ABSTRACT Post-tuberculosis lung disease (PTLD) is a major cause of chronic disease globally. Data are lacking, however, on the prevalence of PTLD and targets for its prevention in youth (ages 15-24), who are vulnerable to care engagement challenges and social determinants that may increase risk for PTLD. Tuberculosis (TB) has tremendous health burdens for youth and is a leading cause of death in this age group. Our long-term goal is to implement youth-focused TB services that target the drivers of PTLD, optimize care engagement, and promote long-term lung health and well-being. Our objective is to establish the prevalence of PTLD in youth, identify modifiable patient-level and environmental risk factors, and assess the effect of PTLD on youth quality of life. Our central hypothesis is that PTLD is prevalent in youth and has associated modifiable risk factors and quality of life impacts that can be targeted with tailored interventions and novel care models. The project will leverage the Tuberculosis Sentinel Research Network (TB-SRN) of the International epidemiology Databases to Evaluate AIDS (IeDEA). TB-SRN is enrolling 2,600 people aged ≥15 with pulmonary TB, with or without HIV, in 11 countries, and following them from TB treatment initiation to 12 months post-treatment. Assessments span clinical, adherence, lung health, mental health, substance use and social determinant domains. We will enhance existing procedures at 7 Youth TB-SRN sites in 6 African countries. Activities at Youth TB-SRN sites will ensure robust analyses of PTLD in youth, through: 1) enhanced data collection on lung health, air pollution, quality of life and well-being; 2) additional enrollment of youth with and without TB, to target 600 total; and 3) youth-friendly supports for study retention. In Aim 1, we will identify PTLD prevalence and risk factors among youth. Persistent pulmonary function impairment at 12-months post-TB treatment (and at 24-months in secondary analyses) will be tested for association with clinical, care engagement, and substance use factors. Pulmonary function will be compared in youth with and without TB. In Aim 2, we will assess air pollution as a risk factor for PTLD in youth. We will add actionable air pollution data to TB-SRN procedures. In 180 youth with TB, 72-hour personal fine particulate matter (PM2.5) exposure will be measured at month 2 of TB treatment, at end of treatment, and 6- and 12-months post-TB treatment. Among 180 youth without TB, PM2.5 exposure will be measured at two timepoints, to enable analyses of TB and air pollution as drivers of lung health. In Aim 3, we will evaluate the impact of PTLD on youth quality of life. Data on health-related quality of life and mental and social well-being will inform the development of care models and interventions to mitigate and manage PTLD. We evaluate persistent pulmonary function impairment for association with health-related quality of life (by EQ-5D-5L) at 12-months post-treatment. Quality of life will be compared in youth with and without TB. Study findings will address a critical knowledge gap about PTLD prevalence in youth and identify key targets for interventions to prevent or mitigate PTLD, as well as foci for strategies to alleviate the detrimental impacts of PTLD on youth quality of life and well-being.

NIH Research Projects · FY 2025 · 2025-08

PROJECT SUMMARY Pulmonary arterial hypertension (PAH) is a rare disease with poor long-term survival and no targeted therapies available. Prior studies have established that pulmonary artery endothelial cell (PAEC) injury and dysfunction, including abnormal proliferation, angiogenesis and apoptosis resistance, plays a critical role in the pathogenesis of disease. My long-term goal is to identify new molecules that may become therapeutic targets to treat and improve the lives of patients with PAH. RASA3, a GTPase activating protein, is a negative regulator of Rap1 and R-Ras, GTPases that regulate angiogenesis, endothelial cell adhesion and turnover, processes highly relevant to PAH. Previous in vitro studies have shown that RASA3 leads to dysregulation in pathways and processes highly relevant to pulmonary hypertension. I have found that RASA3 expression is decreased in four large cohorts of individuals with pulmonary hypertension (PH), and correlates with increased mortality in a cohort of individuals with sickle cell disease-associated PH. I have identified a single nucleotide polymorphism (SNP) within the RASA3 gene, rs9525228, that correlates with increased PH severity within four PH cohorts and is located at a predicted estrogen receptor-α and glucocorticoid binding site. We have also developed a conditional endothelial cell RASA3 knockout mouse for in vivo studies. The specific objective of the proposed research is to investigate the role of RASA3 in maintaining PAEC health in vitro and in vivo within the context of pulmonary vascular disease. I hypothesize that RASA3 is a critical regulator of homeostatic responses in PAEC. This hypothesis will be tested by investigating the following Specific Aims: AIM 1. Identify how RASA3-mediated regulation of BMPR2 impacts pulmonary vascular dysfunction in PAEC. My working hypothesis is that loss of RASA3 leads to dysregulated PAEC homeostasis through hyperactivation of Rap1 signaling altering cytoskeletal dynamics and resulting in BMPR2 downregulation. AIM 2. Determine the effects of RASA3 SNP rs9525228 on a PH-relevant phenotype in PAEC. My working hypothesis is that rs9525228 leads to decreased RASA3 expression. AIM 3. Determine the endothelial-specific role of RASA3 expression in the development of experimental pulmonary hypertension. My working hypothesis is that RASA3 mediates pulmonary vascular health at a proliferative, angiogenic and cell survival level. Upon completion of the proposed studies, I anticipate that my findings will delineate how loss of RASA3 expression leads to PAEC dysfunction and pulmonary vascular remodeling and establish RASA3 as a therapeutic target in PAH.

NIH Research Projects · FY 2025 · 2025-08

PROJECT ABSTRACT Early biomarker detection is crucial for identifying disease onset and evaluating therapy effectiveness, especially in autoimmune diseases like type 1 diabetes (T1D). Detecting T1D before symptoms arise can greatly improve prevention strategies and enable personalized treatments. However, current biomarkers are insufficient to predict each stage of T1D development1. The need for early detection biomarkers remains unmet, as current standards rely on autoantibody conversion, which occurs late in the disease process. Identifying markers that detect disease before autoantibody development could help preserve β-cell mass and delay progression. Autoantibody generation relies on interactions between B cells and activated T cells3. High- risk individuals can be distinguished from later-stage patients using pre-symptomatic biomarkers, which may uncover disease pathways and enable earlier interventions4. T cell activation, critical in T1D, results in the release of soluble lymphocyte activation gene-3 (sLAG3), a marker of immune system activation. Activated, self-reactive T cells damage β cells, triggering stress responses, including the integrated stress response (ISR), which produces extracellular vesicles (EVs) containing β-cell stress markers such as insulin mRNA. Early studies show increasing sLAG3 levels in at-risk individuals, suggesting early immune activation. We recently developed an assay system using plasma and peripheral blood leukocytes (PBLs) to detect immune system changes before autoantibody formation. By employing tetramer technology, we can detect autoreactive T cells at the single-cell level, assessing sLAG3 levels and insulin mRNA. Our objectives are to evaluate if sLAG3, tetramer-positive T cells, and insulin mRNA can predict T1D onset and to incorporate these biomarkers into a multiplex nanoplasmonic assay for simultaneous detection.