University Of Florida

NIH Research Projects · FY 2025 · 2023-09

Ever since Lyndon Johnson declared a War on Poverty in the 1960s, developmental scientists have been assessing the effects of poverty on child development, especially language acquisition. Ample evidence shows that some of the negative effect can be explained by social factors: how parents talk to their children, parental stress, and parents’ knowledge of child development. But these factors fail to account for all, or even most of the language deficit seen in children of poverty. The goal of this project is to explore the hypothesis that poverty-related biological factors can explain previously unaccounted for variance in those deficits. Two novel mechanisms are proposed to account for additional variance in the language deficits of children of poverty: (1) Children in poverty are more at risk than other children to experience delays in the development of auditory functions, beyond raised thresholds; (2) Deficits in suprathreshold auditory functions negatively impact language acquisition for children in poverty more than for children from middle-class backgrounds because optimal social factors buffer effects for the latter group. With these possibilities in mind, the overall goal of this current project is to explore the comprehensive hypothesis that being born into poverty imposes myriad risk factors for delayed development of language skills, arising from both social and biological factors. The social factors to be studied include: (1) impoverished parental language input; (2) enhanced parental stress; and (3) poorer parental knowledge of child development. The biological factors include: (1) premature birth and (2) chronic otitis media with effusion (OME). The most innovative aspect of this work is that it will explore the hypothesis that prematurity and OME are responsible for delays in development of suprathreshold auditory processes critical to language acquisition. Children born into poverty are both more likely to experience these auditory deficits and less likely to benefit from the mitigating effects of a highly supportive environment. This project seeks: (1) to measure the contributions of each of these sources of deficit by examining auditory (spectral and temporal processing) and language (lexical, morphosyntactic, and phonological) functioning in young children born with varying SES; (2) to expand our understanding of the connections from perinatal/postnatal health conditions to development of auditory functions to language learning by including children varying in gestational age and histories of otitis media, as well. These goals will be addressed with three Specific Aims. Aim #1 will quantify the direct effects of the social and biological factors described above on language skills. Aims #2 and #3 will focus on the proposed biological factors, assessing the effects of premature birth and OME on the emergence of spectral and temporal processing (Aim #2) and assessing the contributions of spectral and temporal processing to language acquisition across the socioeconomic continuum (Aim #3). Results will enhance our understanding of the root sources of language deficit for children in poverty, as well as for all children born too soon or experiencing OME.

NIH Research Projects · FY 2025 · 2023-09

PROJECT SUMMARY Ribonuclease L (RNase L) is a key component of the mammalian innate antiviral response. For decades, RNase L was presumed to reduce viral protein synthesis by cleaving ribosomes to arrest translation. However, we and others recently demonstrated that RNase L-cleaved ribosomes are translation-competent, and that pathogenic viruses can synthesize proteins despite activating RNase L. These observations have revealed a significant gap in knowledge regarding how RNase L functions and how viruses evade it. We have demonstrated that RNase L rapidly degrades nearly all cellular mRNAs upon activation. This activity regulates three cellular processes that have expanded our understanding of RNase L and that have elucidated how pathogenic viruses evade and potentially hijack RNase L functions. First, RNase L reprograms translation to an antiviral state by degrading constitutively expressed cellular mRNAs while sparing host mRNAs encoding antiviral proteins (e.g., type I interferons), which permits antiviral protein synthesis. Importantly, the mRNAs encoded by several pathogenic viruses (e.g., dengue virus) similarly evade RNase L-mediated mRNA decay, thus permitting viral protein synthesis. This observation has elucidated how pathogenic viruses synthesize proteins despite activating RNase L. This application proposes to characterize the RNase L-mediated mRNA decay pathway and determine how host and viral mRNAs evade it. Second, RNase L activation triggers the inhibition of nuclear mRNA export. This is a critical antiviral mechanism that antagonizes influenza A virus protein synthesis, but it also downregulates the expression of host antiviral proteins (e.g., type I interferons). Importantly, pathogenic viruses (e.g., dengue virus) activate this RNase L-dependent pathway, resulting in sequestration of host antiviral mRNAs in the nucleus. This observation suggests that viruses potentially hijack this function of RNase L to limit host antiviral protein production. This application aims to determine how RNase L inhibits mRNA export, the breadth of viruses it antagonizes, how it impacts host antiviral gene expression during pathogenic viral infections. Third, RNase L regulates the assembly of cytoplasmic antiviral ribonucleoprotein complexes. Specifically, RNase L inhibits the assembly of stress granules and promotes the assembly of an alternative stress granule-like ribonucleoprotein complex termed RNase L-dependent body. RNase L-dependent bodies are the predominant antiviral granule assembled in response to SARS-CoV-2 or dengue virus infection, yet their function is completely unknown. This application aims to determine the function of antiviral stress granules and RNase L-dependent bodies and to determine how their regulation by RNase L alters the antiviral response. Understanding the mechanisms and functions of these cellular processes will advance our understanding of the OAS/RNase L pathway, innate immune antiviral gene induction, and virology. Moreover, it will promote general medicine by broadly characterizing fundamental cellular, molecular, and RNA biology that is relevant to non-infectious diseases, including autoimmune diseases, neurodegeneration, and cancer. Lastly, the proposed research will support the development of promising antiviral, immunomodulatory, and anticancer therapies based on RNase L biology.

NIH Research Projects · FY 2025 · 2023-09

PROJECT SUMMARY/ABSTRACT Hepatitis C virus (HCV) infection has markedly increased in the United States, primarily resulting from injection drug use (IDU) associated with the ongoing opioid epidemic. Furthermore, >50% of 3.2 million individuals with chronic HCV remain undiagnosed, leading to significant morbidity and mortality despite the availability of effective direct-acting antiviral therapy. Due to shared routes of transmission, HCV infection occurs in 15%-40% of persons infected with human immunodeficiency virus (HIV) and may be used as a marker of HIV exposure. Emergency departments (EDs) play major roles in screening for HCV infection and HIV infection. Several ED- based HCV screening programs have been implemented and have identified previously unrecognized HCV infections, but many challenges remain. Because targeted screening programs use methods that often fail to detect high-risk behaviors (e.g., self-reported information on prescreening questionnaires or review of patient problem lists at time of visit), they do not effectively identify persons at high risk of HCV infection (e.g., IDU). Nontargeted HCV screening strategies require less assessment of risk behaviors. However, concerns such as high costs and unnecessary tests make nontargeted screening strategies difficult to implement and sustain. Therefore, an innovative, effective, and sustainable HCV screening strategy is urgently needed. We propose to develop, implement, and evaluate a tailored, effective, and sustainable, prediction algorithm- based screening tool called Hepatitis C Emergency Department (HepC-EnD) that can be used by health care systems to identify patients at high risk of HCV infection. We will achieve these goals through three specific aims. Aim 1 will develop and validate prediction algorithms using machine learning and natural language processing to identify patients at risk of HCV infection through Florida’s all-payer electronic health records (EHRs) accessed via the OneFlorida+ Clinical Research Consortium. In Aim 2, we will design a HepC-EnD prototype that incorporates the best prediction algorithms to provide automatic notification to ED providers of patients at high risk of HCV infection. Informed by implementation science frameworks, we will enhance the functionality and usability of HepC-EnD through a workshop and qualitative interviews. In Aim 3, we will integrate HepC-EnD into the University of Florida Health EHR system to deploy and test HepC-EnD in two EDs (Gainesville and Jacksonville) and compare the performance of HepC-EnD with nontargeted screening using a difference-in- differences approach. Performance will be assessed by evaluating the usability, acceptability, effectiveness, and cost-effectiveness of the tool. Our proposed research is highly significant in its integration of a cutting-edge machine-learning–based prediction and risk stratification tool into an e-platform that will better inform clinical practice for improving HCV/HIV screening and linking patients with care. Our findings will provide timely data and adaptable strategies that are key in attaining the national and global goals of eliminating HCV infection.

NIH Research Projects · FY 2025 · 2023-09

PROJECT SUMMARY/ABSTRACT Osteoarthritis affects approximately 30 million people in the United States and is the leading cause of pain and disability among older adults. Total joint arthroplasty (TJA) is among the most common elective surgery for patients with refractory knee and hip arthritis. Although a proportion of patients achieve benefits from TJA, up to 34% of patients develop chronic postsurgical pain (CPSP) following the procedure, with approximately 33% of patients experiencing high impact or bothersome pain. While risk factors underlying CPSP following TJA have been well documented, the mechanisms of resilience that predict treatment success are poorly characterized and not routinely assessed. Building on prior research, we propose a novel conceptual model of pain resilience that defines resilient functioning as a dynamic and multidimensional process that engages numerous systems, a concept referred to as multisystem pain resilience (MSPR). In our preliminary work, we have found that individuals with a greater degree of protective resources across multiple domains (i.e., psychological, social, health) exhibit more optimal psychological, physical, and pain-related functioning when compared to those with a lower resilient phenotype. Thus, investigating a broad range of adaptive factors may provide important predictive insights into the unique combination of resources that account for resilient response trajectories following TJA. In line with the Precision Medicine Initiative, the aims of this prospective observational study are to explore mechanisms associated with treatment response in patients undergoing knee and hip arthroplasty, and to characterize associations between MSPR phenotypic profiles with pain impact and physical function trajectories. To address this goal, 300 patients ages 18+ years undergoing knee and hip arthroplasty will undergo prospective assessments of MSPR factors and outcomes at baseline and at five postsurgical time points over a 9-month period. Patients will complete measures across the following MSPR domains comprised of modifiable factors associated with pain: 1) demographic, 2) health, 3) biological, 4) behavioral, 5) psychological, and 6) sociological. Collectively, these aims have the potential to advance understanding of phenotypic mechanisms underlying postsurgical pain impact and physical function trajectories and may be a step toward the development of therapeutic modalities aimed at promoting enhanced recovery and reducing overall pain burden in patients undergoing arthroplasty.

NIH Research Projects · FY 2025 · 2023-09

The University of Florida (UF) College of Dentistry and East Carolina University (ECU) School of Dental Medicine are collaborating to establish an academic multidisciplinary practice-based research network within and between their respective dental schools to provide clinical faculty, residents, and predoctoral students with a competency-based training program for skills development, research mentorship, and participation in practice-oriented clinical research. The partnership addresses NIDCR’s opportunity for Practice-Based Research Integrating Multidisciplinary Experiences in Dental Schools (PRIMED). There is a need in dental education to promote research skill training and experiences, at both the pre- and post-doctoral levels, and to integrate these processes with interprofessional education and transdisciplinary research. This proposal is designed to foster environmental change to embrace research as a critical component of both dental education and dental practice. The proposed project outlines the Development of Opportunities for Research (DOOR): Future Academic Interdisciplinary Workforce and Collaborators for the National Dental Practice-Based Research Network (PBRN). Along with clinical research skills development and mentoring, included are two clinical research studies with common databases across the two schools, to involve dental students, residents, clinical faculty, and research-intensive faculty. The studies will be focused on diabetes detection chairside (Advancing Pre-diabetes/Periodontal Research in Oral Academic Clinical Healthcare; APPROACH) or pain conditions affecting dental treatment (Chronic Overlapping Pain Evaluation Study; COPES). This project will leverage the resources of UF as a research-intensive dental school (#5 in NIDCR funding) to promote a pipeline of dental students and residents at both UF and ECU who are engaged with clinical research. As a newer dental school, ECU (#47 in NIDCR funding) will advance its research activities and lend its resources, adding to the generalizability in this initiative given the different settings and approaches to dental education. For both institutions, the resources of the UF Clinical and Translational Science Institute will be harnessed for research training and implementation, also relying on UF’s experience with the Dental Practice Based Research Network. Additionally, the Dental Clinical Research Unit at UF will be used as a resource, as will the UF Pain Research and Intervention Center of Excellence.

NIH Research Projects · FY 2024 · 2023-09

PROJECT SUMMARY This application aims to use existing data within the All of Us Researcher Workbench to improve our ability to identify hypertension (HTN) patients at increased risk for apparent treatment resistant hypertension (aTRH) and adverse cardiovascular outcomes. Resistant hypertension describes a subset of hypertensive individuals with uncontrolled blood pressure (BP) despite the use of three or more antihypertensive medications, or BP control that requires four or more antihypertensive medications. The term aTRH is used for resistant hypertension when pseudoresistance (e.g. medication nonadherence, white coat effect) cannot be excluded. The prevalence of aTRH was recently estimated at ~17-19% among adults taking antihypertensive medications. HTN is a major risk factor for adverse cardiovascular outcomes such as acute myocardial infarction, stroke, and heart failure. Additionally, when compared to controlled hypertension patients, patients with aTRH are at increased risk for adverse cardiovascular outcomes, target organ damage, and all-cause mortality. Although there are numerous first-line antihypertensive drugs to lower blood pressure and ultimately prevent adverse cardiovascular outcomes, there is great inter-patient variability in antihypertensive drug response. It is poorly understood why patients respond differently to the same drug, why some patients develop aTRH, and why some patients experience adverse cardiovascular outcomes. Our central hypothesis is that HTN patients and subpopulations at increased risk for aTRH and adverse cardiovascular outcomes associated with HTN can be identified through clinical factors, biochemical factors, genomic factors, and patient reported data. To test our central hypothesis we will complete the following Specific Aims: 1) Characterize aTRH and adverse cardiovascular outcomes in HTN patients by health care institutions, urban versus rural areas, and geographic regions, using longitudinal electronic health record (EHR)-based data, and 2) Identify early signs of aTRH and adverse cardiovascular outcomes in HTN patients by health care institutions, urban versus rural areas, and geographic regions, using EHR-based data, genomic data, and data from surveys and wearables. To achieve these aims, we will utilize existing data from the All of Us Researcher Workbench. The All of Us Research Program is enrolling a diverse group of persons in the United States, and including multiple types of real-world data (e.g. EHR, demographic, wearables, patient surveys, genomic). We will deploy our validated HTN algorithms to determine observed rates of HTN, aTRH, and adverse cardiovascular outcomes. We will identify characteristics of aTRH and adverse cardiovascular outcomes in HTN patients. We will also use multivariable regression analyses and machine-learning models to identify predictors (EHR-based, genomic, patient reported) of aTRH and adverse cardiovascular outcomes. We will examine characteristics and predictors of aTRH and adverse cardiovascular outcomes in HTN patients by health care institutions, urban versus rural areas, and geographic region.

NIH Research Projects · FY 2025 · 2023-09

ABSTRACT Through our multidisciplinary clinical and research collaboration, we have established a rare cohort of children and adolescents who fulfilled the 2016 adult Sjögren’s disease (SjD) criteria (namely, cSjD for childhood Sjögren’s disease). Since autoimmune SjD typically affects elderly females, whether cSjD is a unique disease entity or an early manifestation of SjD is completely unknown. The overall goal is to characterize mechanisms by which mitochondrial dsRNAs (mtdsRNA) are released into the cytoplasm and activate the cytosolic dsRNA sensor, protein kinase R (PKR), for the proinflammatory signature found in cSjD CD14+ monocytes. Our preliminary data from scRNA-seq of cSjD monocytes revealed the aberrant expression of mtdsRNA processors, such as SUV3 (ATP-dependent RNA helicase, suppressor of variegation 3, SUPV3L1 gene) and PNPase (polyribonucleotide nucleotidyltransferase, PNPT1 gene), PKR, type I IFN signature (I-IFN), STAT1, and CD52. A higher percentage of cSjD monocytes were positive for particulate cytoplasmic dsRNA with downregulated SUV3 and upregulated PNPase when compared to healthy control monocytes. SUV3-deficient cell lines concurred consistently with cytoplasmic mtdsRNA accumulation, PKR activation, and ISG induction. In addition, we found mitochondrial RNA binding proteins (mtRBPs) crucial in maintaining mtdsRNA stability within the matrix and novel endogenous short duplex RNAs (e-sdRNA) bound to PKR, which may sequester PKR as a monomer to prevent its activation. Furthermore, our scRNA-seq data of PBMC clearly pinpoint CD52 downregulation detected only in cSjD monocytes compared to monocytes from other pediatric groups or adult SjD. Therefore, we propose our novel central hypothesis that defective mtdsRNA degradation by the aberrant SUV3 and PNPase expression in the mitochondria leads to cytoplasmic mtdsRNA accumulation, which activates the cytosolic dsRNA sensor, PKR, in cSjD monocytes. The three proposed aims included: Aim 1. Characterize PKR sensing of mtdsRNA in cSjD monocytes. We will determine if PKR is the key cytosolic sensor for autologous mtdsRNAs in I-IFN+CD14+cSjD Mo and if novel e-sdRNAs bound to PKR regulate its activation. Aim 2. Identify molecules involved in impaired mtdsRNA degradation and release into the cytoplasm. We will identify the impact of upregulated PNPase in cSjD Mo by hypothesizing that PNPase overexpression amplifies PKR activation through the degradation of e-sdRNAs bound to PKR. Additionally, novel mtRBPs will be tested to determine if they permit mtdsRNA to escape into the cytoplasm by altering the methylation status of mitochondrial transcripts. Aim 3. Stratify biological and clinical features of cSjD for precision medicine. We will test if the downregulation of anti- inflammatory, anti-adhesion CD52 in cSjD Mo is mediated by the mtdsRNA-PKR-STAT1 pathway to enhance extravasation through endothelial cells for tissue homing. We will also stratify transcriptomic, immunological, genetic, molecular, clinical, and laboratory features of cSjD by expanding our ongoing scRNA-seq of PBMC and paired biopsy tissues, and RNA-seq of Mo for validation. Our novel mechanistic studies on mtdsRNA, along with NGS, will lead to cSjD molecular barcodes and pertinent RNA therapy involving e-sdRNAs to dampen PKR activation in cSjD.

NIH Research Projects · FY 2025 · 2023-09

Summary/Abstract Cancer-associated cachexia is a multifactorial syndrome characterized by the involuntary loss of body and skeletal muscle mass (with or without fat loss) that reduces tolerance to cancer treatments, increases complications following surgery and is strongly predictive of reduced survival. However, there are currently no effective therapies to preserve, or reverse the loss of, muscle mass in cancer patients, highlighting a major gap in treatment. Unpublished work from our lab implicates a key role for Cellular Communication Network Factor 2 (CCN2), also known as connective tissue growth factor (CTGF), in mediating cachexia induced by pancreatic ductal adenocarcinoma (PDAC), a cancer type with high prevalence of cachexia. CTGF is a hypoxia-inducible matricellular protein produced by pancreatic cancer cells and PDAC tumors which functions locally to induce stromal remodeling, tumor growth and metastasis. In a mouse models of PDAC, we found that Ctgf and Hif1a are upregulated in tumors at time points corresponding to cachexia initiation and progression, suggesting CTGF production by hypoxic PDAC tumors could also be involved in cachexia. In preliminary studies we found that genetic or pharmacological targeting of CTGF inhibited cachexia and blocked host- and tumor cell-secretion of key circulating mediators of cachexia, despite controlling for CTGF-dependent effects on tumor growth, leading us to hypothesize that CTGF promotes PDAC cachexia, at least in part, through promoting cytokine-dependent signaling in peripheral tissues, which will be investigated in Aim 1. In addition to CTGF production within PDAC tumors, CTGF is also upregulated in skeletal muscles of cachectic patients and mice with PDAC. We therefore hypothesize that local production of CTGF within muscle tissue may also play a direct role in muscle wasting in response to PDAC, which was supported through targeting of Ctgf-shRNA to muscle tissue using AAV. Through single nucleus RNAseq we further identified Ctgf to be upregulated in both respiratory and peripheral skeletal muscles of PDAC mice in a cell type-specific manner, with Ctgf commonly upregulated within a subpopulation of mature skeletal muscle nuclei that show increased expression of atrophy-related genes. Using both in vitro and in vivo models, Aim 2 will thus further investigate the cell-autonomous role of Ctgf in mediating skeletal muscle wasting and dysfunction in response to PDAC, and the mechanisms involved. We anticipate that our findings will elucidate key tissue-specific mechanisms of muscle wasting and weakness associated with cancer, with high translational potential.

NIH Research Projects · FY 2025 · 2023-09

Project Summary Kidney transplantation is the most effective modality for treating end stage kidney disease. It provides superior quality of life and significantly improves survival over dialysis. However, the demand for kidney transplants has surpassed the supply of usable organs. Because of this deficit, it is important to improve the outcomes of first- time transplant recipients through intelligent management, thereby optimizing donor organ allocation and reducing the need for secondary transplants. In assessing the health of a renal allograft, time is of critical importance. Being able to precisely predict delayed graft dysfunction and modifying treatment strategies accordingly would be greatly impactful in decreasing chronic rejection events. Existing clinical methods, such as the Kidney Donor Profile Index, which are based solely on donor demographics and clinical data, are minimally to moderately predictive of allograft outcomes. Further, current visual, semi-quantitative transplant biopsy scoring metrics, e.g., Banff, the Maryland Aggregate Pathology Index, and Remuzzi are often not predictive of renal graft function. Digital image analytical methods that quantify chronic changes in kidney that cannot be done visually, may offer clues to long-term allograft outcome. Therefore, to address the unmet need of intelligent renal transplant management, we propose a comprehensive multimodal framework, integrating high-resolution renal transplant biopsy digital whole-slide images (WSIs), and donor and recipient clinical, demographic, and social determinants of health data. Using this framework we will combine computer vision and explainable artificial intelligence (XAI) tools to derive autonomous diagnostic and prognostic models for data-driven, long-term management of renal allografts. As part of their preliminary work, the investigator team has developed a computational tool to quantify interstitial fibrosis and tubular atrophy, a chronicity measure in renal transplant biopsies, and demonstrated that the prediction of estimated glomerular filtration rate at a later time-point after biopsy using machine learning (ML)-derived image features outperforms those based on routine visual assessment. This tool will be expanded to incorporate a variety of additional analyses including robust segmentation of renal compartments in WSIs, leveraging pathologist guided attention to train deep-learning models, state-of-the-art transformer models for multi-task learning, and XAI to increase interoperability and accessibility of ML-derived predictions to pathologists. The performance of this pipeline to predict renal allograft function in a future time-point will be compared with existing methods used in a clinical setting as well as ML- based methods used for explainable prediction of disease progression in other areas of digital pathology. The tool will be deployed on a cloud-based platform and the usability by important stakeholders, namely, transplant renal pathologists, nephrologists, and surgeons will be studied with a goal to eventually include the tool in clinical workflows. The proposed work will be an invaluable asset for clinicians to take advantage of large collections of renal transplant biopsy WSIs and inform treatment decisions towards improving renal allograft function.

NIH Research Projects · FY 2025 · 2023-09

PROJECT SUMMARY Virus-associated lymphomas cause significant morbidity and mortality in HIV-infected individuals – indeed, the oral pathogen Epstein-Barr virus (EBV) contributes to up to 90% of diffuse large B-cell lymphomas (DLBCL) and 40% of Burkitt lymphomas (BL). Although combined antiretroviral therapy (cART) and chemotherapy have improved outcomes for AIDS lymphomas, challenges remain particularly with virus-associated AIDS lymphomas, prompting efforts to better understand virus-associated factors and pathways. In particular, EBV-driven cellular genome replication which is essential to lymphoma proliferation remains underexplored. Upon infection of B cells, EBV drives host DNA replication which is essential for establishment of viral latency as well as proliferation of cancer cells. However, such viral oncoprotein-driven DNA replication is plagued with physical and functional obstacles, resulting in replication stress. Such replication stress is a barrier to cancer. And yet, how EBV-cancer cells overcome such stress at replication forks to successfully proliferate is not well understood. In addressing this knowledge gap, we combined isolation of proteins on nascent DNA (iPOND) and mass spectrometry to discover novel fork proteins. This revealed a critical role for ZC3H18 (or ZC3) as a replication dependency factor that EBV upregulates to ensure host genome replication and lymphoma cell proliferation; notably, ZC3 had not been previously linked to DNA replication. Indeed, EBV+ DLBCL from AIDS patients have elevated ZC3 expression compared to EBV- lymphomas. An intrinsically disordered protein, ZC3 has the potential to concentrate a variety of proteins at replication forks. We find a direct interaction between ZC3 and MCM7 (a core component of the replicative helicase complex), further pointing to ZC3’s influential role in proliferation of EBV transformed cells. Importantly, ZC3’s partnership with other replication dependency factors exposes EBV-lymphoma cells to synthetic lethality – such therapies exploit the property that cancer cells tolerate perturbation of a single gene but succumb to co-disruption of multiple genetic events. In this application, we will test the hypothesis that EBV modulates the DNA replication machinery, ensuring proliferation of transformed cells in the face of replication stress and enhancing the potential for susceptibility to synthetic lethality. We will perform the following aims using ex vivo models and translate our results to patient- derived EBV+ AIDS lymphomas obtained from the AIDS and Cancer Specimen Resource (ACSR). Aim 1. Investigate how novel dependency factors unmask synthetic lethal vulnerabilities in EBV- transformed cells & Aim 2. Investigate mechanisms of ZC3 upregulation, replication machinery rewiring, and contribution of replication dependency factors to EBV+ AIDS lymphomas. These studies will identify mechanisms and generate new paradigms that reveal how an opportunistic virus modulates the host replication machinery to maintain the transformed state. Our long-term goal is to identify novel druggable targets that demonstrate synthetic lethality against EBV+ lymphomas in persons with HIV/AIDS.

NIH Research Projects · FY 2025 · 2023-09

PROJECT SUMMARY/ABSTRACT Comprehensive behavioral treatments are effective at promoting weight loss and improving clinical outcomes among adults with obesity. However, a large majority of individuals with obesity never take the first step of initiating these treatments (even when barriers to cost and access are reduced), severely limiting the population impact of evidence-based weight management treatments. To address this challenge and take advantage of growing access to comprehensive weight loss treatments, our team has developed a tool designed to increase initiation of evidence-based behavioral weight loss treatments among eligible but non- treatment seeking adults (“mobilization tool”). As part of this tool, patients answer brief questions and receive automated, individually tailored feedback. This feedback targets empirically and theoretically relevant constructs for treatment initiation using a Motivational Interviewing approach that supports patient autonomy. The tool is completed by patients in the days prior to a scheduled primary care appointment and contains explicit endorsement by patients’ primary care provider (PCPs), taking advantage of PCPs’ influence while not relying on PCPs to initiate weight counseling. It is also designed to be low burden and high acceptability to PCPs to facilitate dissemination, if effective. We previously conducted a cluster randomized feasibility pilot trial to inform plans for a fully powered test of the effectiveness of the mobilization tool. The pilot showed that the tool was highly usable, informative, and enjoyable; feasibility goals were met; and a signal of an effect was observed. We are now prepared to conduct an adequately powered cluster randomized clinical trial to compare the effects of the mobilization tool and a static treatment description (comparator tool), and to examine how effects differ across key demographic factors. We will recruit and randomize PCPs (n=36) and patients with obesity (n=828) who have an upcoming appointment with enrolled PCPs. All enrolled patients will complete either the mobilization tool or the comparator tool, depending on their randomization arm, and will be informed that they have free access to comprehensive weight loss treatment. We will compare the two study arms on the proportion who initiate weight loss treatment (primary outcome), session attendance, and weight loss outcomes at 6 months (secondary outcomes). Because weight loss programs historically have had lower uptake by individuals who are men, Black/African American, younger, have lower income, and have lower educational attainment, we will recruit a diverse population to allow for comparisons of effects across gender, race, age, income, and education. If effective, the proposed mobilization tool could be disseminated within primary care practices to increase the number of adults with obesity who initiate evidence-based weight loss treatment, resulting in greater population weight loss and meaningful changes in population health.

NIH Research Projects · FY 2025 · 2023-09

ABSTRACT NCI-sponsored clinical trials represent some of the highest priority clinical trials conducted by the UF Health Cancer Center (UFHCC). These trials include participation by patients seen at our center as well as those through our academic research consortium across the entire state of Florida to deliver trial access to where patients are. The UFHCC is committed to the success of NCI-sponsored clinical trials through robust partnerships with the National Clinical Trials Network (NCTN) through the Children's Oncology Group (COG), NRG Oncology as well as the Experimental Therapeutics Clinical Trial Network (ETCTN). This commitment is demonstrated at multiple levels including, but not limited to, prioritizing investigator participation in NCI-sponsored clinical trials, facilitating rapid and efficient activation, financially subsidizing the costs of trial conduct, supporting faculty to attend meetings serving as content experts/committee members, hosting the COG Statistical and Data Center, embedding expectations for participation into salary support, and providing a mechanism to allow regional community oncology practices to access trials. My involvement and effort at each of these institutional levels has resulted in improved productivity, efficiency, and impact. These include my leadership as the inaugural UFHCC Associate Director for Clinical Research (ADCR), Director of the Experimental Therapeutics Group, and Director of the GI Oncology Program. Portfolio priority, oversight and enrollments to the NCI-sponsored clinical trials are a demonstrated institutional priority. Within the NCI-sponsored clinical trial networks, I am also active and provide impact through administrative leadership. I serve as the NRG Oncology Co-Chair of the GI Committee, Chair of the Colorectal Cancer Subcommittee, and member of the Communications Committee. I also serve as the UFHCC institutional PI to the ETCTN and Yale LAO member. I am personally involved in the NCI peer-review process by which concepts are evaluated for scientific merit, impact, and feasibility prior to being approved as trials. These reviews occur in the NCI disease-specific Task Forces, specifically, the Esophagogastric, Hepatobiliary, Pancreatic, Neuroendocrine, Colon, and Rectal-Anal Cancer NCI Task Forces as well as trial final approval as a voting member of the NCI Gastrointestinal Steering Committee. Through these positions and with additional contributions, I have actively and positively influenced the current and future landscape of the NCTN and ETCTN programs, particularly the GI Oncology trial portfolio. My career goals include continued service to the NCI-sponsored clinical trial networks through leadership, new member engagement and trial development/completion while simultaneously increasing my institutional engagement through expanded trial conduct and expanding the next generation of capable and diverse NCI clinical investigators leveraging my leadership, experience, mentorship, and sponsorship.

NIH Research Projects · FY 2025 · 2023-09

ABSTRACT Asthma affects nearly 6 million children in the United States, and on average, each child with asthma experiences at least one exacerbation per year. Pediatric asthma accounts for over 790,000 emergency department visits, 64,000 hospitalizations, and nearly $6 billion in direct healthcare costs annually. Asthma disproportionately affects minority children, who are at risk for more severe outcomes. Asthma is a heterogeneous disease with a range of etiologies, triggers, severities, and treatment responses (i.e., subtypes). Despite that well-known heterogeneity, asthma subtypes are largely confined to a simple dichotomous classification of allergic versus non-allergic, which does not account for overlapping subtypes, subtype evolution, severity, nor do they include social determinants of health (SDOH). As such, if we are to reduce the burden of asthma at both the individual and population level, we must improve asthma subtype characterization to help clinicians craft more personalized primary and emergency care. To date, however, asthma subtyping studies have been limited by small sample sizes, ignored temporal information, and/or focused on individual or a handful of sites. The proliferation of large clinical research networks (CRNs) with real-world data (RWD) from electronic health records (EHRs), combined with advancements in machine learning offer unique opportunities to improve subtyping of pediatric asthma patients. Our study team’s preliminary analysis of asthma exacerbations in the OneFlorida+ CRN using only structured data found five pediatric asthma subtypes which varied by race/ethnicity, severity, digital biomarkers, and comorbidities. Our work supports that there is further heterogeneity in pediatric asthma beyond the classically defined subtypes of allergic vs non-allergic. In this project, we will leverage the OneFlorida+ CRN’s large repository of RWD (covering nearly 20 million patients in the southeast) and a novel privacy-preserving federated machine learning-based framework to: (1) identify pediatric asthma patients, their severity, subtypes, and disease progression (i.e., progression subtypes), and (2) fine-tune those global models to local OneFlorida+ sites with site-specific data to account for between-site heterogeneity. In addition to structured EHR data, we will include spatiotemporally linked environmental data and use natural language processing to include clinical note data such as symptoms and SDOH. To guide our work and inform implementation efforts, we will establish a stakeholder advisory committee with pediatric asthma, healthcare system, and public health stakeholders, and conduct focus groups with local OneFlorida+ site clinicians to develop and test EHR prototypes that integrate subtype data. Pediatric asthma progression subtypes built using RWD from diverse populations combined with stakeholder engagement will move the field closer to precision primary and emergency care that improves outcomes. Our novel privacy-preserving federated machine learning methods address several challenges of RWD analysis and will be a generalizable framework for other CRNs to adopt, facilitating widespread dissemination of this work, and paving a path forward for progression subtype analyses of other chronic diseases.

NIH Research Projects · FY 2024 · 2023-09

ABSTRACT Total joint arthroplasty (TJA) surgeries are among the most common elective surgeries in the US and projected to increase in frequency. Recent TJA clinical practices have effectively reduced post-operative hospital length of stay, yet despite improvements in the efficiency of TJA care, the effects of TJA surgical procedures on pulmonary and respiratory muscle function are less widely appreciated. Even during surgeries as short as TJA, the administration of anesthesia, neuromuscular inactivity, and mechanical ventilation acutely deteriorate pulmonary function and trigger rapid, significant proteolysis of the primary inspiratory muscle, the diaphragm. However, conventional TJA preoperative care does not conventionally address these issues. Older adults, smokers, and those with significant pre-existing lung disease, multiple medical comorbidities, or socioeconomic disadvantage face the greatest risk for declines in post-operative pulmonary and respiratory neuromuscular function. Most patients who utilize our urban safety-net academic medical center have at least one of these risk factors, which can interfere with acute rehabilitation, increase the risk for post-operative pulmonary complications, and extend hospital length of stay. Thus, we propose a clinical study of preoperative inspiratory muscle training (IMT) among individuals with increased risk for pulmonary post-operative complications. Preoperative IMT has been shown to counteract post-operative inspiratory weakness and reduce postoperative pulmonary complications following prolonged cardiac surgeries, but its potential benefits have not been investigated in shorter surgeries such as TJA, with expected brief post-operative hospitalizations. The central hypothesis of this project is that preoperative IMT is feasible and will improve respiratory strength, hasten the early postoperative respiratory recovery, and optimize functional mobility for hospital discharge. Adults scheduled for TJA with pre-existing respiratory muscle or lung impairment will be randomized to complete either: daily IMT in advance of surgery (dIMT), a single acute IMT session immediately before surgery (aIMT), or usual surgical standard of care (SOC). Follow-up testing on the day of surgery and during the acute post-operative hospitalization will identify the feasibility of IMT (Aim 1), distinguish IMT effects on inspiratory and cough strength, (Aim 2), and evaluate patient readiness for discharge (Aim 3). This high risk, proof-of-principle proposal will provide the first controlled evidence concerning disturbances in the regulatory functions of breathing following TJA. Our plan is that data generated from this study will form the basis for future mechanistic studies of IMT to restore breathing strength and further optimize early rehabilitation following TJA.

NIH Research Projects · FY 2024 · 2023-09

Project summary/Abstract Alzheimer's disease (AD) is a common and costly neurodegenerative disease that is characterized by a long pre-clinical stage, including a prodromal stage of AD also referred to as mild cognitive impairment (MCI). Many, but not all, MCI patients progress to AD dementia at varying rates. Among MCI patients, late stage MCI patients progress to AD faster than early stage MCI patients: a faster annual cognitive decline with loss of memory. As potential disease modifying drugs are tested for their ability to delay AD dementia, it becomes critical to have tools that can better accurately predict MCI-to-AD dementia conversion. This would allow selection of cohorts most likely to decline during the study period, maximizing the ability to detect a drug/placebo difference. The proposed project will respond to PA-20-200: NIH Small Research Grant Program (Parent R03 Clinical Trial Not Allowed). In Aim 1, we will develop new deep survival models to predict MCI-to- AD dementia conversion using baseline measures, by using data from the AD Neuroimaging Initiative (ADNI) study. We will use data from the NIH funded Center for Neurodegeneration and Translational Neuroscience (CNTN) as the test data. The majority of the existing deep survival models were developed for right censored data, but MCI-to-AD dementia conversion is interval censored. When interval censored data are analyzed by using the methods developed for right censored data, the survival rates are always over-estimated that leads to the delay in AD dementia diagnosis. We will develop separate prediction models for early stage MCI and late stage MCI with biomarkers from cerebrospinal fluid (CSF), positron emission tomography (PET), magnetic resonance imaging (MRI), and clinical measures. Recently, several new biomarkers have been discovered for AD that are of interest to this study. These include plasma phosphorylated-tau181 (p-tau181), p-tau217, and the ratio of amyloid-β 42 and amyloid-β 40, and glial fibrillary acidic protein (GFAP). In progressive disorders like AD, most clinical events are very strongly correlated with the dynamics of the disease. In Aim 2, we will develop novel survival models for interval-censored data with time-varying longitudinal biomarker data. Built on our developed penalized survival model for interval censored data using baseline measures, we propose to extend that model to leverage longitudinal biomarker data to produce more accurate predictions about future conversion. Biomarkers along with clinical and demographic features were shown to improve the model performances for right censored data. We expect that the new survival models will be able to improve model prediction for interval censored data as compared to state-of-the-art models. This project will develop optimal deep survival models to predict MCI-to-AD dementia conversion for each MCI subgroup. The results of this project will provide important understanding of how each feature contributes to prediction of MCI-to-AD dementia conversion.

NIH Research Projects · FY 2025 · 2023-09

Project Summary: A large body of experimental and clinical evidence has demonstrated that dysregulation of the renin angiotensin system (RAS), resulting in elevated concentrations of Angiotensin II (Ang II), contributes to increased inflammation, oxidative stress, and development of metabolic syndrome, obesity, diabetes and its complications including DR. In addition to circulating RAS, components of RAS are also expressed in different tissues including the eye. Local RAS dysfunction contributes to tissue pathophysiology and end-organ damage in diabetes. However, the exact mechanisms by which ocular RAS contribute to retinal pathophysiology in diabetes are still not well-understood. Prorenin, a precursor of the active renin, the rate-limiting enzyme in RAS cascade, is highly elevated plasma of diabetic patients and ocular fluid of DR patients. The discovery of its receptor, pro/renin receptor (PRR), provided a mechanistic link of elevated prorenin in pathogenesis of DR. Activation of this pathway has been shown to increase Ang II production at tissue level, as well as direct activation of downstream signaling independent of Ang II action, both of which contribute to end-organ damage. In addition to function as a crucial component of RAS, PRR is an integral component of vacuolar H+-ATPase (V-ATPase), which plays central roles in the acidification of intracellular compartments and cellular pH homeostasis. PRR also acts an adaptor protein between the Wnt signaling complex and V-ATPase. Moreover, a soluble form of PRR (sPRR) is produced by protease-mediated cleavage and is elevated under various pathological conditions including DR. Increasing evidence implicates a pathological role of elevated sPRR; however, the mechanisms by which sPRR contribute to pathogenesis of these conditions are still not fully understood. We hypothesize that elevated prorenin, PRR and its soluble form (sPRR) contribute to pathogenesis of DR by multiple pathways, leading to local RAS activation, as well as signaling events independent of Ang II action. The goal of this proposal is to (1) determine the mechanisms of prorenin-induced, PRR-mediated signaling pathways in pathogenesis of DR; (2) determine whether elevated sPRR mediate prorenin-stimulated effects and activates Ang II-dependent pathways in the retina; and (3) determine the effects and mechanisms of prorenin and sPRR on V-ATPase function and associated cellular processes. Collectively, the proposed studies will determine the mechanism(s) and signaling pathways by which prorenin, pro/renin receptor, and its soluble form contribute to retinal neurovascular dysfunction in diabetes. Knowledge of the mechanisms and relationship between these pathways will drive the development of more effective therapies for diabetic retinopathy.

NIH Research Projects · FY 2025 · 2023-09

Project Summary Degraders are chemical dimerizers that recruit a target protein (TP) to an E3 Ubiquitin (Ub) ligase. In favorable cases, this results in TP poly-Ubiquitylation and subsequent destruction by the proteasome. Degraders are difficult to develop because of the requirement that the degrader promote significant protein-protein interactions between the TP and E3 Ub ligase in such a way that a TP lysine residue is placed appropriately to attack the activated Ub molecule. Here we propose to develop a new class of degraders that will recruit TP directly to the proteasome. We anticipate that this mechanism of action will result in potent TP destruction without the need for Ubiquitylation, which, in turn, will make the development of these degraders far more straightforward.

NIH Research Projects · FY 2025 · 2023-09

Project Summary Pain is a complex phenomenon that elicits somatosensory and motor reflexive responses together with marked and long-lasting changes in emotional and autonomic states. While acute pain provides protection from tissue damage, chronic or long-lasting pain, provides no protective function and is often incapacitating. Chronic pain conditions are debilitating to patients, their families, and society by reducing quality of life and creating enormous financial consequences that total more than 630 billion USD annually for the United States of America alone. Neuropathic pain is a type of chronic pain that arises from a lesion or disease affecting the somatosensory system and affects 7-8% of the general population. However, neuropathic pain is poorly responsive to analgesic drugs, including opioids, and alternative therapeutics for treatment are desperately needed. The underlying mechanisms of the development and maintenance of neuropathic pain are poorly understood. A recent wave of high-profile publications implicates the parabrachial nucleus (PBN) as a sensory hub for pain and aversion. The PBN is, a small, bilateral, pontine brain structure that has long been known to receive alarming, noxious, or threatening homeostatic information such as taste aversion, nociception, or danger cues. Promising preliminary data within the Taylor (UPitt) and Betley (UPenn) laboratories implicate glutamatergic PBN neurons expressing the neuropeptide Y (NPY) Y1 receptor (Npy1r-expressing) in the maintenance of neuropathic pain. First, application of a cool (acetone droplet) or light rub (cotton swab) stimulus to the hindpaw of a mouse following peripheral nerve injury produces significant Fos activation within Npy1r-expressing PBN neurons. Second, pharmacological inhibition of PBNNpy1r-expressing neurons via a selective agonist for the NPY Y1 Gi receptor reduces behavioral symptoms of neuropathic pain, whereas chemogenetic activation of Npy1r-expressing neurons produces conditioned place aversion. Third, application of a heat stimulus produces calcium transients in PBNNpy1r-expressing neurons assessed via in vivo fiber photometry. These observations provide the premise for my central hypothesis that the Npy1r-expressing subset of PBN neurons are necessary for neuropathic pain-like behaviors. Specific Aim 1 will utilize in vivo fiber photometry and in situ hybridization to assess the activation of PBN Npy1r-expressing neurons in both sham and neuropathic animals. Specific Aim 2 will apply in vivo chemogenetics to inhibit PBN Npy1r-expressing neurons in sham and neuropathic animals to assess their necessity for the behavioral reflexive (mechanical and cold) and affective (conditioned place preference) components of pain. Specific Aim 3 will examine both the anatomy (anatomical tracing) and functional role (inhibitory chemogenetics) of the supraspinal targets of PBNNpy1r-expressing efferent projections to uncover the specific ciruits responsible for both the reflexive and affective components of neuropathic pain.

NIH Research Projects · FY 2026 · 2023-09

ABSTRACT Zinc (Zn) is an essential trace metal to all forms of life that becomes toxic at high concentrations. Because it has both antimicrobial and anti-inflammatory properties, Zn is used as a therapeutic agent to treat a variety of infectious and non-infectious human conditions. While the efficacy of Zn as an anticaries agent is somewhat controversial, Zn salts are used in several oral healthcare products to prevent calculus formation, treat gingivitis and halitosis, and to control dental plaque accumulation. However, the consequences of rising salivary Zn levels above physiological concentrations to microbial and host-pathogen interactions are poorly understood and warrant further investigation. Recently, we discovered that S. mutans, a keystone pathogen in dental caries, is inherently more tolerant to the toxic effects of Zn than other streptococci, including commensal species associated with oral health. Using transcriptome and mutational analysis approaches, we identified a previously uncharacterized P1B-type ATPase exporter and cognate transcriptional factor, which we respectively named ZccE and ZccR, as primarily responsible for the remarkable high Zn tolerance of S. mutans. Searching public databases, we found that ZccE is unique to S. mutans providing an opportunity for the development of Zn-based antimicrobial therapies specifically tailored to eliminate S. mutans. Our working hypotheses are that the ability to overcome Zn toxicity is an important aspect of S. mutans pathophysiology, and that the identification of ZccE inhibitors can pave the way for the development of a species-specific Zn-based therapeutic modality. With the long-term goal of developing new anticaries therapies in mind, the specific goals of this conceptually, technically, and translationally innovative application are: (i) to uncover the regulatory mechanisms and pathways that mediate Zn tolerance in S. mutans; (ii) to determine the implications of increasing Zn concentrations, above physiological levels, to the composition and homeostasis of the oral microbiome; and (iii) to explore and then develop ZccE as an antimicrobial target. To accomplish these goals, the PI assembled a multidisciplinary team of investigators with complementary expertise in molecular microbiology and animal models (Abranches and Lemos), structured-based computer-aided drug design (Li), and medicinal chemistry (Huigens and Li). Completion of this study will: (i) significantly advance our understanding of the regulatory mechanisms and pathways that mediate bacterial Zn tolerance with the potential of revealing new therapeutic targets; (ii) shed light onto the implications and potential of Zn-based therapies in oral health and, more specifically, in caries control; and (iii) facilitate the rational design of new antimicrobial therapies to prevent/control the emergence of cariogenic biofilms.

NIH Research Projects · FY 2024 · 2023-08

PROJECT SUMMARY/ABSTRACT Chlamydiae are obligate intracellular bacterial pathogens that cause disease in human and animal populations Chlamydia trachomatis is the major cause of both bacterial sexually transmitted disease and infectious blindness in the world. Despite great strides over the past decade, tools for genetic study and manipulation of Chlamydia spp. remain severely limited. This proposal will develop new and powerful genetic tools including tightly negatively regulated promoter expression systems, a Tn7-based single chromosomal insertion transgene system, and application of the latter to build new, smaller shuttle plasmids for transformation. Specific Aim 1 – Development of a tightly negatively regulated inducible promoter for Chlamydia. This aim will provide one of the key missing tools in the repertoire of genetic tools for Chlamydia: a tightly negatively regulated, inducible, and titratable promoter for performing expression / overexpression studies. In bacterial physiology studies, it is important to be able to express and overexpress cloned genes in a mutant or wild type genetic background. One also must be able to tightly repress the cloned gene to avoid undesirable phenotypes associated with leaky expression of the cloned gene. The two subaims will design and test inducible promoter systems derived from two well-established Escherichia coli systems: the lac operon and the arabinose operon. Both systems are tightly repressed in the absence of inducer and rapidly upregulated when inducer is present. Our rigorous validation plan will use three reporter genes to measure promoter activity in the presence and absence of inducer. The lac operon promoters will allow us to develop a range of promoters of varying induction strength and induction ratios to provide flexibility for future genetic studies. Specific Aim 2 – Development of a chromosomal Tn7 transgene insertion system for single copy gene complementation studies in Chlamydia. Shuttle plasmids used in Chlamydia complementation studies are based on the native Chlamydia plasmid, which is present at 7.6 copies per genome. The major shortcoming of expressing a cloned gene from a multi-copy plasmid is that even low copies of the plasmid may lead to non-physiological levels of gene expression and aberrant phenotypes that complicate interpretation of the complementation results. The Tn7 transgene system provides high frequency, site- specific, single copy chromosomal insertion of any gene with the additional advantage of removing the need for antibiotic selection for transgene maintenance. We will adapt the Tn7 transgene system to Chlamydia. Our proposal includes a rigorous strategy to test the transgene system and plans to optimize the system by selection for transposase mutants that more efficiently recognize the chlamydial Tn7 attachment site. We will apply the Tn7 transgene system to build and validate new, smaller shuttle plasmids, which should improve transformation efficiencies. We will make all tools developed in this proposal available to the research community. Thus, successful completion of these aims will enhance genetic studies in Chlamydia.

NIH Research Projects · FY 2024 · 2023-08

ABSTRACT Pulmonary arterial hypertension (PH) is a progressive disease leading to pulmonary vascular remodeling, increased pulmonary arterial pressures, declining right ventricular (RV) function, right heart failure, and death. No curative therapies are currently available. In this F31 application, the PI will train in the acclaimed translational laboratory of Joe GN Garcia, MD and seek to address the unmet need for novel effective PAH therapeutic strategies by focusing on eNAMPT (extracellular nicotinamide phosphoribosyltransferase). eNAMPT is a novel damage-associated molecular pattern protein (DAMP) and PAH target identified by the Garcia lab utilizing genomic–intensive approaches. Intracellular NAMPT mainly serves as a rate-limiting enzyme in nicotinamide adenine dinucleotide (NAD) synthesis. However, secreted extracellular eNAMPT (from leukocytes, lymphocytes, endothelium, epithelium) ligates the Toll-like receptor 4 (TLR4) to potently activate this inflammatory signaling cascade. eNAMPT/TLR4 participation in innate immunity inflammatory responses is key to the severity of several serious inflammatory disorders (ARDS, lung fibrosis) including PH with elevated plasma eNAMPT levels in PH subjects correlating with RV dysfunction. Increased eNAMPT secretion is influenced by NAMPT promoter SNPs and NAMPT transcriptional regulation by anti-oxidant response elements and hypoxia response elements induced by transcription factors such as HIF-2α. We have demonstrated that the eNAMPT/TLR4 pathway is highly druggable as a humanized eNAMPT-neutralizing mAb effectively reduced the severity of preclinical PH. Several key gaps remain, however, in fundamentally understanding the role of eNAMPT in PH pathobiology. For example, it remains unclear as to whether endothelial cells (ECs) or smooth muscle cells (SMCs) are the primary target cell for eNAMPT involvement in PH. In Specific Aim #1 (SA), the PI will examine EC- and SMC-specific NAMPT promoter responses to PAH stimuli that increase NAMPT transcriptional activities and translation. SA #2 will investigate the EC- and SMC-specific intracellular mechanisms contributing to eNAMPT secretion into the circulation by PAH stimuli focusing on the role of NAMPT dimerization, inflammasome and ABC transporter activation and generation of extracellular vesicles. Finally, utilizing human and rodent tissues, SA #3 will define EC- and SMC-specific eNAMPT-induced responses by examining cytosolic Ca2+ signaling, cell proliferation/activation, cell survival, EMT activity and angiogenic activity as readouts. Together, these studies will provide key novel mechanistic insights into eNAMPT’s influence on vascular remodeling and PH pathobiology.

NIH Research Projects · FY 2025 · 2023-08

PROJECT SUMMARY / ABSTRACT Acute respiratory infection and diarrheal disease are the two leading causes of pediatric death between 1 month and 5 years of age globally. These common problems have low-cost treatments, but these treatments are most effective when administered early. This is difficult in resource-limited settings, especially at night. Based on over 5 years of formative NIH-funded implementation science research, our team has built and deployed a telemedicine and medication delivery service (TMDS) called MotoMeds to improve nighttime access to care for children. The rationale is that a TMDS generates a multiplier effect to reduce mortality and morbidity to a greater extent than the provision of in-person emergency medical services (EMS) alone. A TMDS differs from EMS in that it mobilizes resources from a centralized location and transports these resources to households. An EMS, on the other hand, identifies patients at households and transports the patients to centralized resources. EMS logistics, clinical guidelines and decision-support tools do not readily apply to a TMDS. Novel methods are needed to assure that TMDS models of healthcare delivery safely reach their potential. MotoMeds was launched in Haiti in 2019 as a pre-pilot focused on safety and logistical feasibility and configured for scale in a pilot study in 2021. We developed paper-based decision-support tools for virtual call center exams that were derived from in-person World Health Organization (WHO) guidelines. Our research exposed a critical need for electronic clinical decision support (eCDS) to assure guideline adherence at scale and a need to confidently assign triage levels despite limitations in virtual telemedicine environments. Accurate triage is essential to determine TMDS pathways of care: delivery alone (mild cases), medication delivery plus an in-person exam (moderate cases) and EMS/hospital referral (severe cases). In the R21, we will address these needs by concurrently 1) designing an alpha prototype of the eCDS tool through human centered design followed by qualitative research analysis, while 2) using existing data from our prior two studies to derive and internally validate a disease severity prediction (DSP) algorithm for integration into the eCDS tool. In the R33, we will externally validate the accuracy of the DSP and evaluate the performance of a beta eCDS prototype in a pilot stepped wedge cluster randomized trial. The performance of the beta eCDS prototype will be determined by comparing rates of guideline adherence (primary outcome measure) and logistical metrics among providers using paper (control) vs electronic (intervention) decision support. This single-site pilot will provide essential experience and metrics for a future multi-site randomized controlled trial of the eCDS integrated with the DSP.

NIH Research Projects · FY 2024 · 2023-08

PROJECT SUMMARY In this proposal, we aim to characterize the multi-enzymatic and chondroprotective functions of a bioactive biomaterial, manganese dioxide (MnO2) nanoparticles (NPs), as a therapeutic strategy to mitigate oxidative stress in osteoarthritis (OA). The motivation for this work is the critical need to address limitations for treating OA as a looming public health crisis, projected to affect 130 million people worldwide by 2050 due to an aging population. Oxidative stress, the imbalance between reactive oxygen species (ROS) generation and antioxidant function, is known to contribute to OA progression and may represent an important therapeutic target. There have been numerous studies to evaluate the use of antioxidants and small molecules as therapeutic agents, however these therapies are limited by poor bioavailability and stability within the joint. The objective of this proposal is to utilize a metal-oxide biomaterial (MnO2) to overcome limitations of retention and bioavailability and seeks to explore enzyme-mimicking functions to reduce the effects of oxidative stress. We have previously shown that MnO2 can be engineered with cartilage-targeting properties, such as size and charge, that can overcome limitations of traditional antioxidant therapies. Leveraging these properties we have seen improved retention of MnO2 NPs in healthy and OA joints. Due to the barriers for targeting cartilage, this advancement is critical in the development of a chondroprotective therapy. We hypothesize that MnO2 NPs possess enzyme mimicking properties that will reduce oxidative stress in the joint thereby alleviating pain and disease pathogenesis. Characterization of enzyme mimicking functions is critical in the use of MnO2 NPs for biomedical applications and may further classify the biomaterial as a ‘nanozyme.’ Our lab has already characterized the hydrogen peroxide scavenging properties of MnO2 NPs and we anticipate ‘nanozyme’ classification will outline catalase-like, superoxide-like, and peroxidase-like functions of MnO2. In Aim 1, we will examine how MnO2 NPs influence compartment specific H2O2 production and the downstream effects of oxidative stress. Specifically, we will characterize the antioxidant-like properties of MnO2 NPs and their impact on redox signaling, chondroprotection, and inflammatory effects. In Aim 2 we will evaluate the therapeutic efficacy of MnO2 NPs in vivo using a rodent model of post traumatic OA (PTOA) through comprehensive evaluation of NP retention in the joint, joint remodeling, and behavior. Immediate treatment following joint trauma, which leads to PTOA, is a critical opportunity for translation of a cartilage targeting therapy by leveraging cartilage that is still intact and may be responsive to mitigating oxidative stress. The proposed work is significant and innovative by revealing key mechanisms for mitigating oxidative stress and advancing the use of an enzyme-mimicking therapy that may facilitate translation of strategies to slow the progression of joint disease.

NIH Research Projects · FY 2024 · 2023-08

Abstract Dental caries is the most prevalent chronic infectious disease, with an estimated treatment cost of ~$70 billion/year. While a multifactorial disease, Streptococcus mutans is recognized as a keystone caries pathogen because of its capacity to modulate the oral biofilm in a way that promotes the establishment of a highly acidogenic (cariogenic) microbiota. The trace metals iron, manganese, and zinc (Zn) play structural, catalytic, and regulatory roles in proteins and so are essential to all forms of life. Conversely, these same metals are toxic when in excess such that the ability to maintain metal homeostasis is a critical aspect of host-pathogen interactions. The toxicity of Zn derives from its top position in the Irving-Williams series of metal stability, which allows Zn to avidly bind to non-cognate metalloproteins rendering them nonfunctional. Because it has both antimicrobial and immunomodulatory properties and relatively low toxicity to mammalian cells, Zn has been used for decades in medicine, including incorporation into oral health products for the treatment of gingivitis, halitosis, and prevention of calculus formation. Recently, it was discovered that S. mutans is intrinsically and substantially more tolerant to toxic levels of Zn than all other streptococci and that this high tolerance is mediated by a unique P-type ATPase exporter that has been named ZccE. Expression of zccE is controlled by a unique MerR-type regulator, ZccR for zccE regulator, that strongly activates zccE transcription in response to high Zn stress. Because both ZccE and ZccR are unique to S. mutans, the Lemos lab proposes that both can be targeted for the development of a Zn-based therapy tailored against S. mutans. While other projects in the Lemos lab are exploring the antimicrobial potential of ZccE, the specific aims of this application center around the ZccR regulator. The goals of this application are to determine the structure-function relationships of ZccR and explore its potential as an antimicrobial target. To accomplish these goals, the PI will (i) use crystallography and computational-based approaches to determine the structure of ZccR and identify its critical functional residues, and (ii) utilize the rat model to determine the contribution of ZccR to oral biofilm colonization and caries development alone or in combination with topical Zn treatment. Knowledge gained from this study will facilitate the structure-guided design of small molecule inhibitors for ZccR and reveal the potential of targeting mechanisms of Zn homeostasis to combat S. mutans infections, with the long-term objective of developing novel therapies for the prevention of dental caries and treatment of systemic S. mutans infections (i.e., infective endocarditis). This work will be conducted in a highly supportive and collaborative research environment with Aim 1 establishing a collaboration between the Lemos and McKenna labs. Moreover, the comprehensive training plan provided will accelerate the PI's academic and scientific growth through participation in formal course work and workshops, opportunities to learn new methodologies, mentoring students, establishing new collaborations, and improving scientific writing and presentation skills.

NIH Research Projects · FY 2026 · 2023-08

PROJECT SUMMARY Injury to the ankle syndesmosis is common in orthopaedic injuries like ankle fractures and sprains. Surgical repair of the ankle syndesmosis involves rigid fixation of the fibula to the tibia. The etiology of poor patient outcomes following syndesmotic repair, such as pain and osteoarthritis, is not well understood. The central hypothesis of this work posits that syndesmotic repair disrupts the biomechanics of the entire lower limb. Humans comprise one of only two orders in the Animal Kingdom with specialized, fully-mobile fibulae. Fibular motion facilitates shock absorption and stabilization throughout the lower limb. As the lower limb forms an interdependent, mechanical chain, fibular fixation could disrupt both the biomechanics and function of the entire lower limb from the hip to the foot. Our long-term goal is to advance diagnostic and treatment paradigms for syndesmotic injury by better understanding the biomechanical role of the mobile fibula. The objective of this work is to characterize fibular biomechanics and associated sequelae through comparative examination of subjects with healthy, mobile fibulae and surgically immobilized fibulae. We will first evaluate biomechanical differences between healthy individuals and individuals with surgically repaired ankle syndesmoses (Aim 1). We will record motion capture, force, and electromyography data during locomotion, functional, and athletic tasks. Using our experimental data, we will leverage musculoskeletal simulations to assess the effect of fibular mobility on hindfoot joint reaction forces (Aim 2). Finally, we will use explainable machine learning to predict syndesmotic injury state from biomechanical data and identify high-impact predictors (Aim 3). By combining innovative experimental and computational methods, we will improve the biomechanistic understanding of implications of fibular fixation during syndesmotic repair. Understanding what biomechanical differences and functional deficits are associated with syndesmotic repair will provide evidence for new surgical and rehabilitative protocols. Identifying which biomechanical changes are high impact predictors of syndesmotic repair will lay the groundwork to develop data-driven diagnostics and prognostics for syndesmotic injury. Through this proposal, the applicant will obtain training on a unique combination of experimental biomechanics methods (e.g., motion capture, surface and intramuscular electromyography (EMG), ultrasound imaging) and quantitative data-driven approaches (e.g., musculoskeletal simulation, machine learning). The University of Florida will provide the applicant outstanding opportunities for interdisciplinary research, exceptional mentors, and a phenomenal training environment. Further, the University’s AI Initiative provides an unparalleled opportunity to develop world-class AI expertise. These experiences will enhance the applicant’s technical and professional skills, providing the training needed for a successful career as an academic researcher.