Ohio State University

NSF Awards · FY 2024 · 2024-07

Age at death estimates are critical for the study of demography and health in past populations and for correct identifications in forensic cases. However, the accuracy and reliability of age estimation methods based on the human skeleton have limitations resulting from biases in the age, ancestry and sex composition of the collections originally used to develop them. This project advances research on an alternative skeletal age-estimation method based on the analysis of chemical changes that occur in human DNA as we age and can be preserved in the skeleton. To date, applications of such epigenetic methods have been primarily limited to blood and cheek DNA sources. This study advances epigenetic methods that use DNA extractions obtained from bone. Workshops to learn the theory and application of this new method are open to students, faculty and other researchers. Engagement and outreach opportunities are offered through Ohio State University’s Anthropology Outreach Program. These opportunities introduce participants to epigenetic methods as applied to studies of human variation, and aging and health in the past. A high-quality short film about the anthropology of aging and the epigenetic clock is made freely accessible to other researchers and the public. This study develops a new method for biological age estimation from the human skeleton, using a genome-wide age-associated DNA methylation approach tailored to damaged/degraded DNA. The project aims to: (1) develop and validate a cost-efficient, robust-to-degradation, genome-scale method for methylation typing, (2) develop a high-quality predictive model of age from methylation signals in bone, and (3) characterize the potential impact of lifestyle factors on any discrepancies between chronological and biological (DNA methylation) ages. Parts 1 and 2 of this study are based on an integrative genomic-osteological analysis of 100 individuals with documented age, sex, and postmortem exposure to various taphonomic conditions from the University of Tennessee, Knoxville, Donated Skeletal Collection. The project integrates epigenomic, osteological, and lifestyle data, broadening the applications of this method. The study analyzes the relationship between biological and chronological age in the context of osteological signs of aging and stress, differentiating the biological and developmental roots of osteological traits used to estimate age in past populations and individuals. This project is jointly supported by the NSF Biological Anthropology program and the National Institute of Justice, Office of Investigative and Forensic Sciences This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

NIH Research Projects · FY 2025 · 2024-07

PROJECT SUMMARY/ABSTRACT Multiple sclerosis (MS) is a chronic immune-mediated inflammatory disorder of the central nervous system afflicting around one million people in the United States. The disease often presents with a relapsing-remitting phase followed by gradual conversion to secondary progressive MS (SPMS) characterized by irreversible accrual of disability and unresponsiveness to current disease-modifying therapies for MS. Age carries the highest risk for developing SPMS, yet the underlying cellular, molecular, and genetic processes contributing to biological aging are understudied in MS. Recent progress in aging research have led to the emergence of geroscience as an interdisciplinary approach that seeks to reduce the incidence and severity of chronic age- related diseases by understanding aging physiology and developing clinical interventions targeting aging mechanisms. However, no studies to date have investigated the role of biological aging in MS using established markers of cellular senescence (p16INK4a) and epigenetic aging (age-associated DNA methylation patterns, or epigenetic clocks). The Trans-NIH Geroscience Interest Group recently emphasized a need to engage researchers and clinicians across multidisciplinary specialties to translate geroscience discoveries into effective clinical practices. Through the completion of this career development award, Dr. Zhang will become one of few clinician scientists in the country whose research combines principles of geroscience with neurology. This proposed study will investigate the associations of biological aging with clinical outcomes (aim 1), neuroimaging findings (aim 2), and immune dysregulation (aim 3) in people with MS. The study will consist of a cross-sectional analysis of baseline associations of biological aging with MS outcomes and a 2-year longitudinal analysis of the rate of biological aging with changes in MS outcomes. Dr. Zhang has assembled a multidisciplinary mentorship team to achieve his career development goals of establishing a geroscience framework to study MS by integrating clinical, neuroimaging, and molecular mechanistic studies. The successful completion of this project will enable future studies to 1) use biological aging as a prognostic biomarker in MS to employ early aggressive treatment in individuals with accelerated biological aging and 2) conduct trials of anti-aging therapies in combination with MS disease-modifying therapies to mitigate or even prevent MS progression.

NIH Research Projects · FY 2025 · 2024-07

PROJECT SUMMARY Candidate: Lynn Fussner, MD, is an early stage investigator in the Division of Pulmonary, Critical Care, and Sleep Medicine at The Ohio State University who is dedicated to improving care of patients with antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV). Dr. Fussner’s patient-oriented clinical and translational research in AAV has included study of patient outcomes and single biomarkers for predicting relapses. With this proposal, Dr. Fussner’s goals are to expand her skillset to include advanced statistical modeling and bioinformatics coupled with understanding of immunological methods to investigate novel mechanisms underlying the pathogenesis of AAV, and leverage markers and modifiers of those mechanisms for personalized management of these rare diseases. Career Development Plan: To achieve these goals, Dr. Fussner and her primary mentor, Dr. Kymberly Gowdy, MS, PhD, have outlined a comprehensive career development plan incorporating coursework and experiential learning in immunology, biostatistics, bioinformatics, and strategies for study design in rare diseases. Central to this plan is strong mentorship from Dr. Gowdy (expert in translational innate immunity in lung and vascular diseases), Ohio State co-mentors, Dr. Elliott Crouser (pulmonary/critical care physician-scientist, monocytes as biomarkers) and Dr. Guy Brock (biostatistics/bioinformatics), and international leaders in vasculitis, Drs. Ulrich Specks (pulmonary physician- scientist at Mayo Clinic) and Peter Merkel (rheumatology physician-scientist at Penn, Director of the Vasculitis Clinical Research Consortium (VCRC)). Environment: Ohio State’s research environment couples a large patient population with well-funded multidisciplinary clinical, translational, and basic research infrastructure. This includes the Center for Clinical and Translational Science and the Davis Heart and Lung Research Institute. Research: Dr. Fussner will investigate the role of alpha-1 antitrypsin (A1AT) and monocytes as drivers and markers of AAV. A1AT is the primary inhibitor of proteinase 3, which is the predominant target of ANCAs in the western hemisphere. Patients with A1AT deficiency alleles have increased risk of AAV, and our preliminary data indicate that A1AT genotype impacts the clinical phenotype of AAV. Dr. Fussner’s specific aims are: Aim 1: Determine whether A1AT levels, alone or in combination with ANCA levels and monocyte inflammatory markers, predict disease activity and clinical phenotype of AAV. Aim 2: Determine whether ANCA- induced monocyte signatures predict clinical features of AAV and evaluate the impact of exogenous A1AT. Dr. Fussner will utilize clinical data and samples from two cohorts: (i) patients with AAV with and without A1AT deficiency alleles from the VCRC, the world’s largest vasculitis biorepository, and (ii) a clinical trial cohort of patients with relapsing AAV. Patient total and functional A1AT levels, and ANCA-induced monocyte responses will be analyzed for prediction of disease activity and specific manifestations of AAV. These studies will enhance understanding of AAV pathogenesis and identify new biomarkers and safer treatment strategies in AAV.

NIH Research Projects · FY 2025 · 2024-07

Summary Visceral leishmaniasis (VL), caused by Leishmania donovani complex spp. causes between 20,000- 40,000 deaths a year. L. infantum is the cause of VL in the Mediterranean basin and imported to both South and North America. L. infantum is zoonotic with canid reservoirs. Leishmania spp. are transmitted primarily between mammalian hosts by female Lutzomyia or Phlebotomus sand flies. It has been established that both sand fly and host factors modulate local dermal immunity when and where parasites first encounter skin, guiding immunity locally and systemically and impacting infection outcomes. Relatively little is known about the skin immune environment during progressive VL, how dermal immune changes alter host infectiousness or responses to immunomodulatory sand fly salivary components. Skin parasite burden in dogs with L. infantum infection correlates with transmission efficiency, more so than parasitemia, though dogs with late-stage disease were less infectious than those with mild-moderate disease. We have shown that asymptomatic VL clinical status was associated in dogs, as in humans, with productive Th1 type responses, while symptomatic infection correlated with presence of exhausted CD4+ and CD8+ T cells, significant expression of Programmed Death 1 (PD-1) and its ligand, PD-L1 on both B cells and macrophages and loss of macrophage parasite clearance. These findings collectively lead us to hypothesize that immune cell responses in subclinical or clinically infected hosts’ skin, whether from dogs or humans, dictate host infectiousness. We will address this hypothesis through three specific aims in this proposed work, 1) Identify unique dermal immune environments in subclinical vs. clinical hosts that alter infectiousness to naïve sand flies, 2) Evaluate how the functions of sand fly salivary proteins are impacted by skin changes during progressive VL leading to new understanding of vector saliva-host interactions and how they contribute to infectiousness to sand flies and 3) Evaluate how alteration of the dermal immune environment through dermal immunotherapy alters bite-site inflammation and transmission. The work proposed here is therefore significant, as it provides infection control relevant evidence regarding the dermal microenvironment and how it alters infectivity during progressive L infantum infection. The studies proposed here are significantly innovative as they quantitatively on a spatial level assess how parasite burden and different dermal inflammatory states alter infectiousness to sand flies. Using purified salivary antigens, skin biopsy, histopathologic and transcriptome analysis and a key unique natural infection cohort, we will determine how progressive VL alters the landscape in which vector salivary proteins operate with consequences for understanding both host infectiousness and the epidemiology of VL. When completed, findings from these proposed studies will both underscore how progressive dermal inflammation impacts reservoir host infectiousness and provide critical data to further assess appropriate interventions to prevent canine-sand fly-human transmission.

NIH Research Projects · FY 2025 · 2024-07

The Ohio State University (OSU) R38 StARR Program will leverage the large, collaborative and multidisciplinary research environment of Ohio State with the support of the College of Medicine (COM), Office of Graduate Medical Education (GME), and participating departments to provide 24 months of research training in immune-mediated disease, infectious disease and/or immunotherapeutics for select Resident- Investigators from the Departments of Medicine, Neurology, Pathology, Surgery, and Plastic Surgery. The R38 program will be embedded within the OSU COM Office of Physician Scientist Education and Training (PSET) that will facilitate Resident-Investigator interactions, vertical peer mentoring and networking with a full spectrum of physician scientists ranging from undergraduate trainees, MSTP students to early career physician scientist faculty. The R38 program will benefit from an experienced leadership team engaged in national efforts to fortify the physician scientist and surgeon scientist pipelines. The recruitment strategy, curricular design, ample NIH-funded faculty mentor pool and resources build upon OSU strengths while leveraging opportunities for trainee networking with the successful Physician Scientist Training Program (PSTP) in the Department of Internal Medicine (DOIM) and Department of Surgery’s Research Training Program (RTP). The R38 program will spearhead the recruitment of exceptional residents to engage in research training and entice and prepare them through tailored education, career development and mentorship to pursue physician scientist careers. The R38 program will offer two tracks: Track 1 for residents without advanced research experience (categorical, undifferentiated “Late Bloomer” residents) and Track 2 for categorical residents with advanced scientific backgrounds (including MD, MD/MS, MD/MPH, MD PhDs). Track 1 Resident-Investigators will pursue mentored research and earn a COM Masters of Medical Science (MMS) degree that has a well established core curriculum (research design, biostatistics, research ethics and grant writing) and tracks for basic/translational, clinical and health services research. Track 2 Resident-Investigators will have individualized curricular plans including required Responsible Conduct of Research (RCR) and Research Rigor coursework and selected optional coursework to enhance knowledge, skills or abilities in immunology, host defense, immunotherapeutics, biomedical informatics, clinical trial research, pharmacogenomics, computational biology, health services research. Resident-Investigators will participate in a monthly OSU-StARR physician scientist career development seminar series consisting of faculty led discussions on career topics and select workshops to enhance skills in science communication (oral and written), mentoring, utilization of search tools, databases, etc. The OSU-StARR training opportunities for less experienced as well as more experienced Resident-Investigators will create a rich environment and community for physician scientist career development that is centralized in the COM.

NIH Research Projects · FY 2025 · 2024-07

Triple-negative breast cancer (TNBC) disproportionately affects African American (AA) women, with mortality rates 65% higher than Caucasian (CA) women. The mechanisms underlying the heightened aggressiveness and metastasis in AA TNBC remain elusive. This study investigates the immune suppressive tumor microenvironment (TME) as a driving factor in AA TNBC progression. Specifically, it delves into the novel role of the S100A7 and its interplay with intrinsic IFNγ signaling, elucidating their impact on TNBC aggressiveness and metastasis in AA women. Our recent findings reveal elevated S100A7 expression in AA TNBC patient samples and cell lines relative to CA counterparts. Moreover, higher S100A7 expression correlates with increased tumor burden in various pre-clinical models, including AA TNBC patient-derived xenografts (PDX). We also noted that S100A7 knockout (KO) mouse models (generated in our lab) exhibit reduced tumor burden, while treatment with a novel S100A7-neutralizing antibody (nAb) shows promising efficacy in inhibiting TNBC growth and metastasis. Mechanistically, S100A7 is demonstrated to enhance cPLA2/PGE2/IFNGR1 signaling in AA TNBC cells, modulating intrinsic IFNγ signaling. This process generates an immune suppressive TME by upregulating PD-L1 and downregulating Fas on tumor cells. Additionally, AA TNBC tumor tissues manifest heightened immunosuppression, characterized by increased PD-L1 expression and infiltration of FoxP3+ Treg cells. Our proposed research aims to meticulously uncover how S100A7 orchestrates IFNγ responsive genes (PD-L1 and Fas) to generate an immunosuppressive TME in AA TNBC. This investigation leverages diverse AA TNBC cell lines, humanized PDX models, and genetically engineered mouse models (GEMMs) overexpressing mS100a7, mS100a7 KO, and the S100A7 nAb. The study's overarching hypothesis posits that S100A7 contributes to AA TNBC aggressiveness by fostering an immunosuppressive and immune evasive TME via regulating intrinsic IFNγ signaling, resulting in PD-L1 upregulation and Fas downregulation. The research strategy encompasses three key aims: Aim 1 will elucidate how S100A7 signaling regulates IFNγ responsive genes in AA TNBC using AA TNBC cells and mS100a7 GEMMs. Aim 2 will determine the novel role of S100A7 in regulating macrophage plasticity, T cell function, and Fas-mediated immune evasion in AA TNBC. Aim 3 will evaluate the therapeutic efficacy of S100A7 nAb in combination with chemo or immunotherapy using AA TNBC PDOs and PDX mouse models. This aim will also determine the prognostic significance of S100A7 and its downstream signaling molecules in AA TNBC. In addition, this aim will establish the association of S100A7 and various immune cells in TNBC racial disparity. The insights gained are poised to identify novel S100A7-mediated downstream signaling pathways and determine the clinical relevance of S100A7 in AA TNBC. This study holds the immense potential to inform the design of innovative therapeutic strategies tailored for AA TNBC, thereby contributing to improved outcomes in this high-risk population.

NSF Awards · FY 2024 · 2024-07

Understanding fundamentally quantum phenomena is of high interest because those phenomena are important in applications that span from quantum information processing to magnetic resonance imaging. This project will explore electronic noise at the quantum scale, what environmental factors control it, and then how to control it by design. In the larger landscape of quantum technology research, this information will enable the design of effective quantum bits for information processing and biomedical sensing applications. Beyond science, this effort will build curricula focused on quantum information science and engineering (QISE) for wide distribution and organize a local QISE-focused workshop on understanding noise at the quantum level. The technical goal of this project is to enable fast, efficient profiling of magnetic noise at the atomic level. Noise at this scale is a critical challenge in the current state of quantum computers and sensors because noise causes decoherence which destroys a qubit’s utility. To deal with said noise, technologists need to understand it: what frequency is the noise, how loud is is, and what causes it. This project, comprising an interdisciplinary chemistry and physics team from University of Colorado Boulder and Colorado State University, will explore a new technique for the efficient characterization of noise and benchmark the new method with both solid-state qubits (like the NV center in diamond) and metal-containing molecules through spectroscopic and theoretical methods. These studies will extend the understanding of magnetic noise to an atomistic level beyond the border of what it currently known. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

NIH Research Projects · FY 2025 · 2024-07

Project Summary Sudden cardiac death and arrhythmias account for ~15-20% of all deaths worldwide. Voltage gated sodium channels (Nav) in heart are major regulators of myocyte excitability and cardiac function. The Nav channel current (INa) is a large amplitude and short duration inward current that is regulated by rapid channel activation and immediate inactivation. However, a small late component of this current (INa,L) is present at baseline but increases in response to heightened adrenergic challenge and has been correlated with fatal forms of congenital and acquired arrhythmia. The majority of work on Nav1.5 has focused on molecular pathways for positive regulation of INa,L. However, less is known about the pathways for negative regulation. These pathways are critical and can be targeted to alter the pathogenic INa,L in potentially fatal forms of arrhythmia. My previous work identified a key pathway for INa,L regulation via the B56α regulatory subunit of protein phosphatase 2A (PP2A). My previous and current data support that: 1) B56α co-localizes with Nav1.5 in the heart at the intercalated disc, along with ankyrin-G and CaMKIIδ, 2) PP2A activity is regulated by the specific regulatory subunit (B56α) and 3) activation of PP2A through modulation of B56α is sufficient to regulate Nav1.5 activity and resist the increase in INa,L following adrenergic challenge. Our work illustrates a key role of B56α in INa,L regulation as well as the potential for targeting this pathway in arrhythmias. However, the field has been limited by two key issues: 1) the lack of selective and inducible in vivo models to selectively impact key PP2A regulatory proteins, and 2) the lack of molecules that selectively target PP2A activity in vivo. For this proposal, we have generated two novel tools to identify the impact of PP2A-B56α on cardiac action potential and arrhythmias in vivo. First, we have created the first in vivo model to both constitutively as well as inducibly impact local PP2A regulatory subunit (B56α) function, and thus cardiac PP2A activity. This model has the impact in a field where work was limited to in vitro studies or non-selective activation or protein reduction. Second, we have identified a new molecule to directly test the in vivo impact of PP2A activation in disease. Our collaborators at Ohio State have synthesized a new molecule and have shown that this molecule increases PP2A activity. Our preliminary data support our central hypothesis that PP2A, via the B56α regulatory subunit, regulates the intercalated disc Nav1.5 to counteract CaMKII-dependent phosphorylation in disease. Further, we hypothesize that molecules that selectively modulate the PP2A-B56α holoenzyme will impact altered INa,L in heart disease. Combining molecular, biochemical, pharmacological, patch clamping and Ca2+ imaging approaches, we will: 1) define the functional role of PP2A-B56α signaling complex in primary myocytes, 2) determine the effects of PP2A-B56α regulation on myocyte excitability, Ca2+ handling and cardiac function and 3) define the in vivo impact of molecular activation of PP2A via B56α modulation in disease. Thus, this proposal will generate critical information and will provide fundamental and clinically-relevant data for INa,L regulation and arrhythmias.

NIH Research Projects · FY 2025 · 2024-07

Project Summary Mobile CT scanners are routinely used in the neuro intensive care unit (ICU) for critically ill patients to avoid morbidity and adverse events associated with patient transport. The image quality of mobile CT is inferior to a fixed MDCT in terms of image noise, spatial resolution, soft tissue contrast, and susceptibility to artifacts from beam hardening, motion, metallic implants, and truncation. Reduced image quality may compromise care and necessitate transport to a fixed scanner. For example, on a mobile CT scanner, it is difficult to diagnose small infarcts or hemorrhage, or to differentiate between intracranial hemorrhage and contrast extravasation after endovascular processes. In this project, we will leverage the benefits from an FDA-approved mobile photon counting CT (PCCT), as well as deep learning-based image reconstruction algorithms to improve the image quality of the mobile PCCT to or beyond that of a fixed scanner. The multi-spectral and high-resolution features of the mobile PCCT will be explored and combined with deep learning algorithms for noise and artifacts reduction. In this project, the following aims will be investigated to achieve our goal to match the image quality of a mobile PCCT to a fixed scanner: (1) We will develop low-dose, high-resolution deep learning-based reconstruction algorithms to reduce the noise and improve gray-white matter contrast in the mobile PCCT; (2) We will develop methods for material decomposition with reduced noise amplification and spectral optimization to overcome beam hardening artifacts and achieve discrimination between calcium/contrast/hemorrhage; (3) We will develop deep learning-based algorithms for image artifacts correction to tackle artifacts that are more frequent on a mobile CT, including motion, truncation, and metal artifacts; (4) To validate the methods, the optimized mobile PCCT images will be compared with fixed CT images by trained radiologists.

NSF Awards · FY 2024 · 2024-07

As the ultimate arbiter of crucial legal disputes, the U.S. Supreme Court occupies a pivotal position in American democracy. However, relatively few cases make it to the Court. In any given year, the Court receives thousands of petitions asking it to review lower court decisions, of which less than one hundred are granted review, or writ of certiorari. The discretionary authority to choose its cases endows the Court with substantial influence in shaping national public policy. Given the statistical rarity of a case receiving a formal decision by the U.S. Supreme Court, and the unrepresentative nature of the petitions that justices choose among, it is critical to understand not just the decisions the Court makes on the merit, but also the density and diversity of issues and interests competing for the Court’s consideration. How does the supply of certiorari petitions shape the justices’ behaviors? How do external actors, such as interest groups and the media, shape the Court’s docket? How does the Court’s selection of petitions granted review represent the plurality of public interests? To answer these and related questions, the research project endeavors to collect, categorize and analyze a host of important features in the writ of certiorari process. Recognizing the seminal importance of the agenda-setting stage, the research intends to provide a better understanding of judicial decision-making and judicial processes as well as the essential role of courts in American democracy. The research enhances existing theories of judicial behavior, with a specific focus on the early stage of decision-making in the United States Supreme Court: the decision to grant review, or writ of certiorari. The research collects and analyzes case features from all writ of certiorari petitions— that is, all lower court cases where the losing party, dissatisfied with the outcome, appeals to the US Supreme Court. The study catalogs—for the first time—the geographic origin, temporal distribution, and issue areas of cases the Court is asked to review. It also examines the role played by external actors in shaping the Court’s agenda, including the media and interest groups, as well as the extent to which the Court’s final selection of cases is consistent with the expectations of the plurality of interests in society. This multifaceted investigation inquires into how these factors collectively impact the decision-making process of the justices, leading to both empirical and theoretical contributions in the study of judicial decision-making. First, it provides new data on writs of certiorari petitions that are comprehensive over time and space, and across issues. Second, the project presents a unique perspective on the courts more generally by contributing to theories that are built around informational cues, and by using computational social science methods to test how the selection of cases impacts judicial outcomes. Third, it seeks to refine and extend machine learning algorithms for legal text analysis of cert petitions and external actors, potentially paving the way and setting new standards for data-driven research in judicial politics. Finally, it supplements the quantitative analyses with interviews of former Supreme Court clerks and amicus-filing entities to provide a richer perspective on how the justices sort through the large number of petitions received every year, and how the Court selects the final set of cases to review. Through the research, and, importantly, the creation of a dataset that categorizes a host of features of all cert petitions to the Court, this project will provide judicial, interest group, media scholars and the public with new insights on the workings of the judicial system. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

NSF Awards · FY 2024 · 2024-07

Biomolecular condensates, formed through liquid-liquid phase separation (LLPS) and related phase transitions, represent a novel mechanism to organize diverse cellular processes in biological systems. These dynamic structures play pivotal roles in fundamental processes such as growth, development, and stress responses. Currently, the standard approach for investigating biomolecular condensation involves microscopy-based techniques extensively applied in individual cells and in vitro settings. However, tools are lacking to explore condensates within the complex environments of tissues, organs, and whole organisms. The goal of this EAGER project is to develop innovative biosensors to probe condensate dynamics with expanded spatiotemporal resolution in multicellular plants at the organismal level. The success of this innovative tool will greatly extend the timescale of detecting condensates from minutes to several days, providing a more comprehensive understanding of condensate dynamics over longer periods. The broader impacts of this interdisciplinary research extend to education, diversity and science communication. The project will also contribute to the training of scientists at the undergraduate, doctoral, and postdoctoral levels. Cellular compartmentalization is a fundamental process in all living systems to control biochemical reactions in space and time. In addition to the canonical membrane-enclosed organelles, biomolecular condensates assembled through phase transitions create another layer of cellular compartments without biological membranes. To date, cell-based assays have been extensively employed to understand the function of condensates. Significant knowledge gaps exist in delineating the contribution of condensates to physiologically relevant processes in vivo, particularly in multicellular organisms such as plants and mammals. This gap is in part due to a dearth of tools to detect biomolecular condensation at the organismal level. Microscopy-based methods are powerful to monitor condensates’ behavior in individual cells but do not support the throughput and sensitivity in organs, tissues, and organisms. This work is guided by a recent discovery by the PI of a new LLPS-mediated host defense mechanism in Arabidopsis thaliana, wherein plants assemble biomolecular condensates mobilized by a group of immune GTPases, Guanylate-Binding Protein-Like (GBPLs) proteins, to combat infection. This project aims to develop novel genetically encoded biosensors as a proxy for GBPL condensate formation at the organismal level over a long period of time. The successful development of the tool will benefit both plant and animal research communities and holds significant promise in uncovering more fascinating insights into how these membraneless compartments contribute to cellular dynamics. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

NIH Research Projects · FY 2025 · 2024-07

PROJECT SUMMARY Over 250,000 angioplasties, stents and open bypasses are performed each year to open occluded vessels for preservation of life and limb, yet over half of these will fail in under 5 years due to restenosis. The underlying pathology of restenosis is cellular proliferation that re-narrows the vessel lumen. Restenosis increases the cost and complications of re-intervention, as well as risking cardiac ischemia, stroke and limb loss. Current treatment involves a balloon coated with an anti-proliferative drug that is delivered to the vessel wall by direct contact. Drug coated balloons (DCB) present several limitations including: 1) Technical limits to the type and amount of drug that can be adhered to a balloon 2) Much of the drug is lost simply by inserted the DCB into the bloodstream. 3) The occlusive nature of DCB limits the time it can transfer the drug without distal ischemia. The overarching problem is the inability to deliver high intensity intravascular therapeutics, while minimizing losses to the circulation that might otherwise be costly or toxic at the systemic level. A Retrievable Drug Delivery Stentgraft (RDDS) is a novel, dumbbell shaped stent covered in polymer. A center lumen preserves distal perfusion, while an isolated outer chamber isolates the vascular target. The RDDS would infuse drug only after the target has been isolated by the stent outer chamber, with aspiration of unused drug prior to stent removal, thereby reducing systemic losses and toxicity. The design of the RDDS could accomodate larger quantities of drug and also liquid agents. By preserving distal perfusion, the RDDS can offer longer incubation of the drug with the vessel wall, contrasting to occlusive DCB that otherwise risk distal ischemia. The RDDS bears some important distinctions to current permanent drug eluting stents, which are at risk of circulatory drug loss, should not be used at vascular branches or areas of anatomic flexion, incur thrombosis over poorly re-endothelialized stent struts and carry traditional risks of a permanent implant. By contrast, the RDDS is removed after used by sheath advancement to collapse the stent. As a result, the RDDS is more comparable to DCB since it delivers a therapeutic without leaving an implant. This study will first compare the RDDS to DCB in a bioreactor, as well as a porcine model of neointimal hyperplasia, using assays such as quantitative mass spectrometry of both drug delivery and drug losses, histologic assessment of arterial healing and restenosis. We hypothesize that the RDDS will reduce circulatory losses of the anti-restenotic agent paclitaxel (PXL), increase the amount of PXL delivered to the vessel wall, allow doses of PXL that are not possible with DCB, increase the duration of PXL exposure to the vessel wall, and reduce intimal hyperplasia more effectively than DCB. More broadly, the beneficial effect of many other intravascular therapeutics is limited by their systemic toxicity. A vehicle to focus drugs to a specified region (vascular wall or vascular bed) could improve outcomes while reducing complications in a variety of medical conditions (e.g. chemotherapy, vasoactive agents, gene vectors, and immunomodulatory agents).

NIH Research Projects · FY 2024 · 2024-07

Project Summary/Abstract Elevated expression of KIF20A, a mitotic kinesin required to promote cytokinesis, the final step of cell division, is a near-universal feature of human cancer. Increased KIF20A levels are associated with poor prognosis in multiple tumor types and correlate with chromosomal instability (CIN). Cancer cells are sensitive to loss of KIF20A function via both small molecule (KIF20Ai) and CRISPR-mediated KIF20A knockout (KIF20A-KO) whereas RNAi-mediated depletion of KIF20A is generally tolerated and has not informed us of the enzyme’s role in CIN. We find that increasing KIF20Ai concentrations leads to a dose-dependent loss of viability that correlates with an increase in mitotic duration caused by defective chromosome congression. While perturbing mitosis is a clinically relevant therapeutic strategy, current agents target microtubules and induce neurotoxicity, which limits the utility of these agents and precludes their use in tumors of the central nervous system. Glioblastoma (GBM; isocitrate dehydrogenase [IDH]-wild-type) is an aggressive, highly proliferative brain tumor with limited treatment options. Radiotherapy is central to standard of care treatment for GBM. However, refractory cells ultimately lead to recurrence and highlight the need for new strategies. We and others have shown that perturbing mitosis is a vulnerability of GBM cells and have demonstrated overexpression of KIF20A in these tumors. Indeed, we find that patient-derived models of GBM are sensitive to KIF20Ai. In contrast, non-transformed cells have low sensitivity to KIF20Ai. In addition, as mitotic cells are highly sensitive to radiotherapy, the mitotic arrest induced by KIF20Ai offers the potential for radiosensitizing these tumors, enhancing the current standard of care. Based on these observations and our preliminary data, we hypothesize that cancer cells utilize a previously underappreciated activity of KIF20A to promote efficient mitotic progression and prevent CIN. Furthermore, we postulate that blocking this function has therapeutic potential. The objective of this proposal is to further characterize the importance of KIF20A activity in early mitosis and investigate the impact of KIF20Ai as a potential new therapeutic strategy for GBM. Two Aims are proposed; 1) Define the requirement for KIF20A in early mitosis by establishing the impact of KIF20A-KO on early mitosis, identifying the essential KIF20A function, and examining factors that dictate KIF20Ai sensitivity, 2) Determine the impact of KIF20Ai on tumor growth by comparing KIF20Ai to radiation and testing KIF20Ai as a radiosensitizer in orthotopic GBM PDX models. The successful completion of this work will provide rationale to further explore 1) the cell biology of KIF20A and KIF20Ai, to further our understanding of CIN, and identify cellular characteristics and mechanisms that dictate sensitivity to KIF20Ai, and 2) the preclinical impact and translational potential of KIF20Ai as a non-microtubule- targeting anti-mitotic strategy, both as a single agent and combination strategy in GBM and other tumor types.

NIH Research Projects · FY 2025 · 2024-06

PROJECT SUMMARY/ABSTRACT: Early life interactions between commensal microbes and the developing immune system have formative effects on human health and susceptibility to chronic inflammatory disease. Previous studies from the host lab and other groups have demonstrated that our microbial symbionts can tune skin-resident T cell function, especially during the neonatal window. However, comparatively little is known about how commensals influence cutaneous myeloid immunity in early life. Dr. Dhariwala has found that commensal microbes influence the composition and function of the myeloid compartment in neonatal skin. Specifically, commensal microbes drive the accumulation of a population of Ly6ChiMHC-IIlo inflammatory monocytes in neonatal skin. These cells have traditionally been studied as primary mediators of inflammation in response to infection or for their ability to fill vacant tissue myeloid niches. However, Dr. Dhariwala’s preliminary studies demonstrate a previously unexplored regulatory immune imprinting role for neonatal skin monocytes. To dissect the molecular mechanisms that underpin this phenomenon, we will employ a combination of transgenic mouse models, high throughput assays like mass cytometry and single cell RNA sequencing along with novel ex vivo translational tools. Studies proposed in this application will (Aim 1) Dissect the mechanism(s) by which commensal microbes influence monocyte recruitment and function in neonatal skin, (Aim 2) Elucidate the mechanism(s) by which neonatal monocytes contribute to skin immune homeostasis and (Aim 3) Functionally elucidate the role of neonatal monocytes in human skin. Results from this cross-disciplinary approach will benefit our understanding of early immune development and lay the groundwork for next generation therapies for chronic inflammatory diseases. The proposed research and training plan will build on Dr. Dhariwala’s strengths in the fields of microbiology, immunology and translational science. Leveraging tools he has established in the host lab, such as mass cytometry, ex vivo functional assays in human skin and manipulation of skin-resident myeloid cell populations in murine models, along with studies like metabolomics proposed in this application will not only elevate the project but also advance his training. His scientific and career progress will be well supported by a strong mentorship team including Drs. Tiffany Scharschmidt, Clifford Lowell, Michael Rosenblum and Peter Turnbaugh. Additionally, through presentations at local and international conferences along with formal and informal training at UCSF Dr. Dhariwala will advance his transition to independence. Promising preliminary data, a strong training plan and an excellent training environment will allow Dr. Dhariwala to carve out a niche for himself and establish an independent research program studying the influence of commensal microbes on myeloid immune development in neonatal barrier tissues.

NIH Research Projects · FY 2025 · 2024-06

Abstract Idiopathic Pulmonary Fibrosis (IPF) is an age-related and progressive lung disease characterized by loss of the normal alveolar architecture and accumulation of scarring tissue replacing the functional lung parenchyma. One of the main characteristics is the loss of alveolar epithelial type II stem cells (AT2), with the concurrence appearance of transitional AT2 cells. This aberrant alveolar epithelial cell co-expresses genes of AT2, AT1, and airway epithelial cells simultaneously. Although transient AT2 epithelial cells are enriched in areas of severe fibrosis, the mechanisms involved in remodeling and lung fibrosis remain poorly understood. Our preliminary data show that under normal conditions, AT2 actively metabolizes long fatty acids by fatty acid oxidation (FAO). As the mitochondrial membrane is impermeable to acyl-CoA FA, the outer membrane mitochondrial protein, carnitine palmitoyltransferase 1A/1B (CPT1a/CPT1b), catalyzes transport step of the lipid metabolism been the limiting rate of this pathway. In the lung, in AT2 from aging and IPF lungs, there is a decrease in the expression of the CPT1a with reduced levels of Acetyl CoA, an FAO metabolite. To determine if FAO is essential for the repair responses, we developed CPT1a deficient cell lines and a conditional AT2-CPT1a deficient mouse. Using these tools, we have found that: 1) expression and activity of CPT1a in AT2 cells is required for the protection against injury-induced lung fibrosis, 2) deficiency of CPT1a enhances markers of transitional AT2 stem cells, with markers of TGFβ1 activation, and cellular senescence, 3) CPT1a regulates TGF- β1 signaling by acetylation and degradation of the negative regulator Smad7, and 4) deficiency of CPT1a engages histone hypoacetylation in lung epithelial cells. These observations have led to the hypothesis that FAO regulates AT2 differentiation by epigenetic reprogramming via modulation of acetyl-CoA levels. A deficiency of FAO triggers AT2 differentiation into a transient aberrant AT2 without reaching a terminal AT1 differentiation, decreasing the resilience against lung fibrosis. To evaluate this hypothesis, we propose the following aims: Aim 1. To examine the hypothesis that acetyl-CoA FAO generated in AT2 cells regulates epigenetically and transcriptionally AT2 differentiation. Acetyl CoA generated in the mitochondria via FAO is an essential metabolite linking metabolic and nuclear programs influencing AT2 differentiation. Aim 2. To test the hypothesis that AT2 CPT1a expression confers resilience against the development of lung fibrosis. Our preliminary data show defective FAO in aging AT2 cells, and deficiency of CPT1a/FAO in AT2 increases susceptibility to tissue remodeling associated with pulmonary fibrosis. Completion of these aims will enhance our understanding of the role of CPT1a and FAO in the wound healing process and age-related mechanisms of resilience to disrepair and fibrosis.

NSF Awards · FY 2024 · 2024-06

Despite an increase in interest in STEM degrees, the number of students completing STEM degrees has decreased. Additionally, time to degree has increased for students who complete STEM degrees. For students majoring in engineering and physics, in particular, inefficient mathematics preparation for introductory physics sequences has been cited as one of the barriers to students completing their degrees and completing their degrees on time. The overarching goal of this NSF INCLUDES Network Connector: Transforming Introductory Physics Sequences (TIPS) to Support All Students (Physics TIPS) is to create a network of people and departments that are redesigning introductory physics sequences to better support all accepted students - regardless of math preparation by reducing course-specific mathematics requirements and incorporating quantitative literacy in the physics sequences. This will be accomplished by 1) documenting and disseminating models of successful introductory physics sequences designed for students with varying levels of math preparation, 2) evaluating the effectiveness of redesigned introductory physics sequences, and 3) developing curricular materials that focus on quantitative literacy in physics contexts that can be easily adapted in introductory physics courses of various styles. The goals of this project align with the vision of NSF INCLUDES to innovate and facilitate collaborative approaches to broaden participation in STEM. The Alliance will collaborate with and build upon the efforts of the NSF INCLUDES STEM CORE Alliance and the NSF INCLUDES PUSH Alliance to mitigate the impact of math preparation as a barrier to entering and succeeding in STEM courses. The Physics TIPS Network leverages the work of researchers and instructors at three universities that have successfully modified/developed a new introductory physics sequence to support students at all levels of math readiness. These courses reduce mathematics requirements, incorporate quantitative literacy, and offer additional contact hours to support integration of physics and math content. To better understand these existing efforts, project members will conduct a mixed methods study investigating the efficacy of the alternative introductory sequence. The quantitative aspects of the study include descriptive statistics (DWF rate comparison between traditional and transformed sequences), analysis of course performance in sophomore/junior courses (a t-test comparison between students who took the traditional sequence and those who took the transformed sequence), and a two-way ANOVA to determine the success rate for first generation college students versus those that are not first generation college students (as determined by degree completion and type). Qualitative inquiry will investigate student experiences in the transformed sequences, engineering identity after taking the transformed courses, and reflections on the course experience post-graduation. To expand efforts already determined to be successful, this project includes implementation of similar courses within 3 additional physics departments and the development of curricular materials to support quantitative literacy. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

NIH Research Projects · FY 2024 · 2024-06

SUMMARY The goal of this proposal is to enable a comprehensive modernization of an aging system in the shared zebrafish research facility in Bevis Hall at the Ohio State University. The modernization proposal will align the capabilities of the zebrafish facility with modern demands on zebrafish science and increase capabilities to allow affiliated researchers to consider experiments that were previously unattainable. The zebrafish facility in Bevis Hall houses the fish of three core- and five additional users. The research supported by this facility spans neural development, heart regeneration, systems neuroscience, cancer research and nutrition. The renovation will allow automatic monitoring and control of critical water parameters and automatic feeding. These advances will increase research reproducibility, reduce waste, and dramatically shorten generation time. The expected reduction in size and growth variability of adult and juvenile zebrafish will allow for new research avenues in these stages. The decrease in time from hatching to sexual maturity of zebrafish will increase the amount of generated transgenic and mutant fish available to researchers at The Ohio State University and beyond. The proposed modernization will also centralize water filtration and eliminate food-waste, reducing the environmental footprint of the facility. Increasing the reproducibility of experiments through better controlled feeding and water parameters will allow reducing the amount of animals used in experiments.

NSF Awards · FY 2024 · 2024-06

This research project focuses on enhancing the way vital information is delivered to smart mobile devices—such as smartphones and tablets. With the advancement of technology, there is a growing necessity for these devices to receive various types of information (like images, videos, and texts) instantly and effectively. One promising approach to achieving this is through the use of Geospatial Digital Twins (GDT), which are digital models of physical environments. GDTs are becoming increasingly important as they allow for real-time updates and interactions, making them invaluable for various applications such as monitoring, maintenance, and emergency response. Traditionally, data for GDTs has been collected through automated systems like distributed sensor devices, satellites and drones. However, these methods have limitations, especially when it comes to updating data quickly and covering hard-to-reach areas. To overcome these challenges, this project will develop a novel approach that involves the community through “human-in-the-loop” strategies. This means using crowd-sourced data, where people provide real-time updates to digital models. This method not only promises to enhance the accuracy and timeliness of the information but also to allow discovery of new information. The project has the potential to revolutionize how we interact with and understand our physical world, potentially making this work a cornerstone for further scientific and educational advancements. The project will also play an important role in education, integrating research findings into university curricula and offering unique learning opportunities for students, including students from underrepresented groups. The goal of this project is to establish an intellectual foundation for building a real-time crowd-sourced GDT. To achieve this goal, we will work toward a fundamental understanding of crowd-sourced multi-modal information collection and processing to account for the underlying human incentives and human-machine integration, which underpin the foundation of crowd-sourced GDT. In this project, we will investigate the design of crowd-sourced GDT to ensure timely, truthful, and unbiased imagery data collection from the crowd. Our efforts will be organized around four tightly integrated research thrusts: 1) ensuring crowd-sourced data freshness for a GDT; 2) integrating crowd-sourced data for real-time GDT updates; 3) guaranteeing truthful reporting in crowd-sourced data collection; and 4) mitigating self-reinforcing bias in crowd-sourced GDT updates. Collectively, this project will result in new tools for optimization and control that directly contribute to real-time crowd-sourced GDTs. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

NIH Research Projects · FY 2025 · 2024-06

Project Summary/Abstract Despite exciting advances, lung cancer remains the leading cause of cancer-related deaths in the United States. A critical gap in our understanding is how lung cancer cells become metastatic and resistant to our treatments. Historically, lung cancer research has focused on genetic drivers as the main culprits of disease. But recent, large DNA and RNA sequencing studies have shown that genetic changes account for only 50% of lung cancer progression, strongly suggesting there are unidentified non-genetic mechanisms of disease progression. The PI’s preliminary data has identified that propionate metabolism—a metabolic pathway previously not known to play a role in cancer—is dysregulated in non-small cell lung cancer (NSCLC). Indeed, their data show that methylmalonic acid (MMA), a key metabolic byproduct of propionate metabolism that can promote epithelial-to- mesenchymal transition (EMT), is elevated in lung cancers and in the serum of patients with metastatic disease. Moreover, the PI has identified that MMA can activate and bind to G-protein coupled receptors, uncovering a novel signaling axis. His preliminary data also shows that MMAB, a key regulatory gene of propionate metabolism, is underexpressed in NSCLC and knocking down MMAB promotes EMT and drug resistance in lung cancer cells. These studies have led to the central hypothesis that propionate metabolism dysregulation promotes lung cancer progression and drug resistance in NSCLC and is an important non-genetic mechanism of disease progression. The goal of the proposed studies are to understand i) the mechanism by which MMA signals ii) the effect of MMAB loss in NSCLC and iii) the in vivo effects of MMA in NSCLC using mouse models. Successful completion of the proposed work will improve our understanding of a metabolic pathway in lung cancer that is poorly understood and potentially identify a new pathway that could be targeted therapeutically. This proposal describes a five-year training program to launch an independent research career focusing on mechanisms of NSCLC drug resistance and metastasis. The candidate, Dr. Bobak Parang, is a medical oncology fellow at Weill Cornell Medicine who has been training to become a physician-scientist for over ten years. He has outlined a career development plan that will build on his strong background in cancer biology and clinical oncology by providing rigorous training and new skills in molecular biology, metabolomics, and mouse models of lung cancer. He will perform the proposed studies under the mentorship of Dr. John Blenis, a world renowned expert in molecular biology, cancer metabolism, and cellular signaling in the rich institutional environment of Weill Cornell Medicine. With the mentorship team he has assembled, his strong clinical and scientific background, and his training plan, Dr. Parang is ideally positioned to develop into an independent laboratory- based physician scientist with R01 funding.

NIH Research Projects · FY 2025 · 2024-06

Project Summary/Abstract HEV is an emerged zoonotic pathogen. It infects nearly 20 million people annually, resulting in approximately 70,000 deaths yearly (2). In the U.S. the virus is believed to be endemic to swine herds with the virus making its way into the commercial food supply (3, 4). Genotype 3 HEV is increasingly being seen as a pathogen afflicting immunocompromised populations including those with HIV, cancer, and solid organ transplant (SOT) recipients (5-9) and mortality can reach up to 30% in pregnant women (10) . Currently chronic HEV patients are treated with reduction of immunosuppression risking organ rejection in SOT patients or with ribavirin therapy. There have been reported incidents of ribavirin failure both during and post treatment leading to poor patient outcomes and ribavirin is contraindicated during pregnancy (11-13). Additionally, single nucleotide variants (SNVs) have been identified from ribavirin treatment failure that enhance HEV replication (14). New therapeutics that specifically target HEV, are amenable to treating at risk populations, and prevent complications such as organ rejection while combatting the emergence of new ribavirin resistant HEV strains are essential to improving outcomes in HEV patients. Emerging technologies such as CRISPR/Cas13 which can specifically target and degrade viral RNA along with liver targeting nanoparticles may be an ideal tool to treat HEV infections. Our proposal will test rationally designed CRISPR RNA targets for efficacy using a newly developed cell culture system from our laboratory followed by studies on efficient liver targeting of the therapeutic using cutting edge lipid nanoparticle technologies in a natural host pig model. If successful, these new technologies could be placed into clinical trials as new therapies to combat HEV infection.

NIH Research Projects · FY 2025 · 2024-06

PROJECT SUMMARY Clostridioides difficile infection (CDI) is a major cause of healthcare-related mortality and a significant public health burden in the US. C. difficile (Cd) remains a persistent cause of morbidity and mortality in healthcare settings, in part because many patients are asymptomatic for CDI yet colonized with Cd. These patients outnumber CDI patients, can transmit Cd and progress to CDI (especially in the context of antibiotic exposure). Importantly, the clinical outcomes in Cd-colonized patients exists on a spectrum influenced both by the microbial community and the virulence of the Cd strain. Critically, conventional animal models of CDI do not recapitulate microbiome and pathogen variation seen in asymptomatically-colonized patients, thus requiring the development of novel animal models to study this patient population. The long-term goal of this research is to identify opportunities for novel therapeutic intervention or pathogen surveillance by better understanding and predicting Cd-associated clinical outcomes. The objective of this proposal is to use clinically-relevant animal models to 1) investigate the extent to which commensal microbiota protect the host from diverse Cd strains, 2) predict microbiome vulnerabilities to antibiotic-induced CDI, and 3) identify microbiome features that synergize with prebiotic administration. The central hypothesis of this work is that the composition of the commensal microbiota plays a central role in determining host disease severity. The specific aims of this proposal are to: 1) investigate the in vivo role of commensal Eubacteriaceae in microbiome-based protection against Cd infection and 2) identify microbiome correlates of antibiotic-induced CDI and prebiotic synergy. The proposal will use microbiome-humanized models of Cd colonization/infection that integrate both clinically-represented Cd strains and patient-derived bacterial communities to understand the microbiota's impact on host inflammation, community metabolism, and Cd proliferation. This research will spur the development of innovative treatment and diagnostic approaches to mitigating CDI. Dr. Dantas will oversee the project, provide direct mentoring on statistical modeling of multi-omics data, and help Dr. Fishbein's transition to independence through support of networking strategies. Dr. Fishbein has prepared a Scholarly Advisory Committee along with other Significant Contributors with expertise in host-pathogen interactions, gut microbiome-pathogen dynamics, and intestinal inflammation. The training plan combines primary mentorship, committee interactions, formal coursework (at the University and externally), and seminar/conference presentations to expand Dr. Fishbein's technical and conceptual foundations in the microbiome field. This award will facilitate Dr. Fishbein's acquisition of independent funding, enabling her transition to an independent research program at a research-intensive academic institution.

NIH Research Projects · FY 2026 · 2024-06

With the emergence of SARS-CoV-2 variants with mutations in the Spike protein, there remains an urgent need for vaccines that are both effective against variants and that generate long-lived mucosal immunity. Generation of durable cell-mediated and humoral immunity is critical for optimal naturally occurring and vaccine-induced protection against respiratory pathogens, including SARS-CoV-2, and includes IFN-γ and IL-17 producing tissue- resident memory T (TRM) cells, T follicular helper (TFH) cells, germinal center (GC) and memory B cells, that contribute to the production of pathogen-specific neutralizing antibodies. Most currently approved vaccines are adjuvanted with alum, which is a strong adjuvant that elicits TH2 skewed cellular and humoral responses, associated with short-lived immunity to intracellular respiratory pathogens. Experimental adjuvants that generate TH1 and TH17 driven systemic and mucosal responses, provide effective and long-lived protection against infection. Bordetella Colonization Factor A (BcfA) is an adjuvant that elicits strong TH1 and TH17 responses and has the unique ability to attenuate the detrimental TH2 responses primed by alum. Polyfunctional IL-21 and IFN-γ (TFH1 cells) or IL-21 and IL-17 (TFH17 cells) cells are important for generation of effective antibodies against viral respiratory pathogens. The TH1/TH17 skewing properties of BcfA may promote the differentiation and function of these specialized TFH cell populations. Mucosal vaccination is a more effective means of generating tissue-resident memory that is not generated by parenterally administered alum-adjuvanted vaccines. A prime-pull regimen (systemic priming and intranasal booster) generates mucosal responses to vaccines containing TH1/TH17 skewing adjuvants and provides superior protection. We will test the overarching hypothesis that a BcfA/alum-adjuvanted subunit SARS CoV-2 vaccine containing S, M and N proteins, delivered via a heterologous prime-pull immunization regimen will reduce SARS-CoV-2 infection of the mouse respiratory tract and elicit long-lived systemic and mucosal TH1/TH17 driven immune responses.

NIH Research Projects · FY 2026 · 2024-06

PROJECT SUMMARY With the increasing awareness of distinct disease outcomes during polymicrobial infections, there is an urgent need for mechanistic studies to better define microbial interactions. This application seeks to understand community interactions between Pseudomonas aeruginosa and Staphylococcus aureus, which are model organisms to study polymicrobial interactions. They are two of the most common pathogens causing chronic infections in the lungs of people with cystic fibrosis (CF), medical devices, and wounds, suggesting co- existence in vivo. Indeed, chronic co-infections caused by these organisms are more problematic than mono- species infections. However, the relationship between P. aeruginosa and S. aureus is intriguingly complicated. These organisms often compete in vitro but are frequently co-isolated from the same clinical samples. We contribute to the studies of anti-S. aureus mechanisms by proposing a novel role for the P. aeruginosa exopolysaccharide Psl in antagonizing S. aureus growth and interactions with S. aureus protein A (SpA). Studies proposed in aim 1 will utilize several complementary strategies to define mechanisms underlying P. aeruginosa Psl-mediated interactions with S. aureus. In aim 2, we will investigate a unique cooperative behavior associated with P. aeruginosa and S. aureus. This involves production of 2-heptyl-4-hydroxyquinoline N-oxide (HQNO) by P. aeruginosa, and staphyloxanthin (STX), a yellow pigment synthesized by the S. aureus crt operon. STX can promote S. aureus resistance to oxidative stress and neutrophil-mediated killing. We found that STX production in S. aureus, either as surface-grown macrocolonies or planktonic cultures, was elevated when exposed to sub-lethal concentrations of HQNO. When subjected to hydrogen peroxide, human neutrophils, or during in vivo infections, P. aeruginosa survival was significantly higher when mixed with wild- type S. aureus, in comparison to P. aeruginosa cultured alone or with an S. aureus crt mutant deficient in STX production. Therefore, the focus of aim 2 will be to define the pathways and biological relevance of HQNO- mediated induction of STX, resulting in resistance to host ROS during infection. Finally, we discovered that a prior infection of both human- and murine-derived macrophages with S. aureus induces tolerance and prevents responsiveness to subsequent infection. This is due to significant metabolic rewiring of the macrophage innate immune response during primary infection. Thus, aim 3 will use primary macrophages and in vivo animal models to investigate how a S. aureus infection leads to metabolic reprogramming and define how this leads to epigenetic changes and subsequent innate immune gene silencing. P. aeruginosa and S. aureus interactions are clearly multifaceted and likely mediated by bacterial factors, spatial distribution during infection, nutrient availability, and the host microenvironment. These proposed studies will contribute to the understanding of complex polymicrobial interactions and lay the foundation for strategies to modulate these pathways for better control of mixed-species infections.

NIH Research Projects · FY 2026 · 2024-06

Excessive binge alcohol intake is a well-recognized risk factor for atrial fibrillation (AF), the most common arrhythmia with a high morbidity and mortality, yet current therapies have suboptimal effectiveness. [While aging is an unavoidable AF risk factor, alcohol as a secondary stressor exacerbates AF risk in the aging population.] Clinical data suggest that one-third of all new-onset AF cases are related to alcohol intoxication and alcohol abuse brings a high AF risk even in people without co-existing cardiovascular diseases. It is a classic concept that enhanced inflammation contributes to alcohol-caused organ damage, which could also lead to AF. However, the ineffectiveness of the anti-inflammation therapies in AF patients has demonstrated the urgent need to understand the detailed underlying mechanisms of inflammation-associated AF and explore novel anti-AF therapeutic targets. The goal of this proposal is to fill this knowledge gap by establishing a previously unrecognized crosstalk between the pro-inflammatory signaling pathways and the stress kinase JNK2 in AF pathogenesis. In human donor atria, binge alcohol exposure increased pro-inflammatory TNFα and IL1β-NLRP3 signaling along with activated JNK2. And TNFα or IL1β significantly enhanced diastolic SR Ca2+ leak and triggered activities (Ca2+ waves and delayed after depolarizations), while either JNK2 or NLRP3 inhibition effectively alleviated those triggered activities, suggesting a crosstalk between JNK2 and pro-inflammatory signaling. However, we found that only TNFα, but not IL1β, activates cJNK2. Yet, TNFα is known to upregulate IL1β signaling, which could explain the involvement of NLRP3 in TNFα-induced triggered activities. But why JNK2 is not influenced by IL1β yet is critically involved in IL1β-NLRP3-mediated Ca2+ mishandling remains completely unknown. [This proposal is thus aimed to establish a novel and potentially paradigm-shifting and translationally important link between the JNK2-NLRP3 nexus (as a pathological nodal point; it has never been revealed before) and a complex web of co-existing pro-inflammatory pathways in AF pathogenesis and most importantly, testing therapeutic potentials of targeting this pathological nexus as a novel anti-AF approach.] Our multi-discipline team will use a unique diversified approach (molecular biochemistry to single channel/myocytes/whole-heart electrophysiology in animal models/human donor hearts) to systematically address two Specific Aims 1) Determine the underlying mechanisms of the JNK2-NLRP3 feedback loop in atrial Ca2+ mishandling; 2) Establish the functional contribution of JNK2 and/or pro-inflammatory signaling pathways in binge alcohol-evoked arrhythmic activities and AF pathogenesis. The scientific premise of this proposal is strong because it integrates important functional measurements and fundamental mechanistic studies along with appropriate alternative approaches. [Cardiac specific interventions (in vivo atrial gene transfer & genetic modification & target-specific pharmacological inhibition) that limit cJNK2 and/or NLRP3-inflammasome activity will be used as proof-in-principal studies to prove the JNK2-NLRP3 feedback loop in AF pathogenesis and shed new light on future development of anti-AF therapeutic strategies and new drug discovery.]

NIH Research Projects · FY 2025 · 2024-06

Project Summary/Abstract: Heart failure (HF) is a leading cause of morbidity and mortality worldwide, with projected numbers continually rising, mandating a need for novel therapeutic approaches. A common feature in the development of HF is hypertrophic growth of cardiac myocytes and associated remodeling of the size, dimensions, and function of the heart. Pathologic hypertrophy initially occurs as an adaptive response, leading to increased width of individual myocytes and causing concentric growth characterized by thickened heart walls, reduced wall strain, and maintained function. Left unchecked, this hypertrophic growth becomes maladaptive and reorients to growth along myocyte length, causing relative wall thinning, heart dilation, and declining function leading to HF. We currently have a poor understanding of the mechanisms which govern this transition, yet, limited observations where adaptive growth is preserved shows resistance to HF development. Therefore, this proposal seeks to identify the fundamental mechanisms underlying adaptive and maladaptive hypertrophic growth and investigate targeted interventions to maintain and/or restore the adaptive state for HF prevention. This proposal will address the critical distinction that not all pathologic hypertrophy is adverse and that preserving the adaptive, concentric state is therapeutically advantageous in response to chronic stress. Preliminary data has implicated a role for the phospho-regulation of the transcription factor STAT3 in mediating this transition. In particular, phosphorylation of the serine residue 727 on STAT3 was revealed as a critical target with dramatic influence over concentric/eccentric growth. Therefore, our central hypothesis is that STAT3 Ser727 phosphorylation is directly responsible for the induction of gene programs which drive adaptive versus maladaptive hypertrophy and represents a therapeutic target in HF treatment. The approach will be to: 1) Determine the molecular mechanism linking STAT3 Ser727 phospho-regulation to hypertrophic orientation. 2) Define novel gene targets and pathways which tune cardiac myocyte growth and hypertrophy. Specifically, this approach will address altered STAT3 transcriptional activity dependent on Ser727 phosphorylation through ChIP-seq and RNA-seq to identify gene programs which enact concentric/eccentric states. 3) Lastly, we will test novel therapeutic strategies to support adaptive cardiac remodeling during pathologic hypertrophy in vivo to assess effectiveness in HF prevention. Overall, we anticipate that these data will expand our understanding of HF remodeling, delineate the nature of adaptive, concentric hypertrophy, and reveal novel therapeutic opportunity in HF. Furthermore, characterization of STAT3 phospho-regulation and transcriptional activity will provide significant pathophysiologic insight to numerous other disease states such as cancer, fibrosis, inflammation, and immune signaling where STAT3 activity has been implicated.