Temple Univ Of The Commonwealth

NIH Research Projects · FY 2025 · 2022-09

PROJECT SUMMARY Asthma pathogenesis involves structural and functional changes in resident airway cells including airway smooth muscle (ASM) cells. Understanding the cellular and molecular mechanisms involved in altered homeostasis in airway cells provides opportunities to develop newer and effective anti-asthma drugs. Ubiquitination is an evolutionarily conserved and highly regulated homeostatic process that ensures turnover of key signaling intermediates. Alterations in E3 ligases have been implicated in human pathologies including cancer, aging, neurodegeneration. This proposal is based on our solid preliminary data derived from human ASM cells and mouse models using molecular and pharmacological approaches that suggest: A) an E3 ligase Ring Finger Protein 145 (RNF145) is expressed in the human lung and ASM cells, with expression significantly increased in fibrotic/remodeled airways and ASM cells derived from asthmatics; B) silencing or pharmacological inhibition of RNF145 reduces ASM cell proliferation and extracellular matrix secretion through regulation of PI3 kinase and AMPK signaling, respectively; C) profibrotic mediator transforming growth factor (TGF) b1 treatment upregulates expression of RNF145 in ASM cells; D) inhibition of RNF145 reduces ECM secretion in vitro, cellular ATP levels, and restores AMPK signaling; E) over-expression of RNF145 drives both ASM cell growth and ECM synthesis independent of mitogen or profibrotic agent; and E) inhibition of RNF145 prevents house dust mite (HDM)- induced airway hyperresponsiveness, inflammation and markers of airway remodeling. Our data are supported by clinical findings from collaborator Dr. Frey who shows a single nucleotide polymorphism in RNF145 (rs10076782) is associated with neonatal lung function, more respiratory symptoms and wheezing at school age. ASM cells play a pivotal role in the regulation of lung function, and increased ASM mass and hypercontractility are associated with asthma symptoms. However, the role of E3 ligase RNF145 in ASM cells or in asthma is not known. RegulomeDB analysis suggests that RNF145 regulates signaling molecules involved in metabolic, cell cycle, matrix production, cell fate and oxidative stress pathways, all of which play a role in asthma pathogenesis. Therefore, we hypothesize that RN145 plays a key role in regulating ASM functions such that inhibition of RNF145 will prevent development of the allergen-induced asthma phenotype. Studies using ASM cells obtained from healthy and asthmatic lung donors will delineate the role of RNF145 in ASM cell proliferation (Aim 1) and extracellular matrix production (Aim 2) and establish the cellular and molecular pathways regulated by RNF145. To further establish the role of RNF145 in airway functions and in asthma pathogenesis, we will use wild type and global and smooth muscle-specific conditional RNF145 knockout mice subjected to allergen challenge (Aim 3). We will also test the effectiveness of a pharmacological inhibitor of E3 ligase, SMER in murine model of asthma. Collectively, findings from the proposed studies will provide novel insights into an important and previously unrecognized role of RNF145 in regulating ASM functions and in the pathogenesis of asthma.

NIH Research Projects · FY 2024 · 2022-09

PROJECT SUMMARY / ABSTRACT The expanding visceral fat depots (VAT) of overweight and obese individuals experience dysfunction of the microcirculation with hypoxia and abnormal infiltration of leukocytes. Infiltrating immune cells and hypoxic adipocytes cause a systemic low-grade inflammation that is considered now causative of cardiovascular disease and metabolic disorders. Thus, the incidence of cardiometabolic disorders (hypertension, kidney disease, and insulin resistance) are becoming epidemic in the ever-growing overweight population of the USA. The precise mechanisms by which weight gain alters microvascular responses in VAT remain poorly understood. Furthermore, it has not been determined whether impaired microvascular responses of VAT are the initiating cause of cardiometabolic dysfunction and disease. We have identified a novel adipokine, IL-19, expressed in the adipose tissue of humans and mice. IL-19 is a newly discovered Th2-interleukin which has been recently studied for its protective vascular effects in the setting of large vessels disease, such as atherosclerosis and ischemia reperfusion injury. Preliminary data obtained in the Co-PI's laboratories strongly suggest that IL-19 regulate the homeostasis of adipose tissue microcirculation in diet-induced obesity. Accordingly, the overarching hypothesis of this dual-investigator application is that IL-19 is a unique interleukin that maintains cardiometabolic homeostasis by controlling angiogenesis and inflammatory responses in VAT depots. The long-term goals of this project are: 1) to understand the mechanisms through which IL-19 regulates vascularity and inflammation in VAT; and 2) to mechanistically correlate these microvascular actions of IL-19 to whole-body insulin signaling. Toward these goals, we will utilize knockout and transgenic mouse technology along with physiological, cellular, molecular, and biochemical techniques. We anticipate that the results of this work will advance our understanding of the integrated mechanisms that initiate and maintain VAT dysfunction and related cardiometabolic disorders in the overweight/obese organism.

NIH Research Projects · FY 2025 · 2022-09

ABSTRACT Understanding diagnose, on signaling well-known for its essential activities promoting cell proliferation during development. suggest important roles of YAP as a cell-fate determinant in early embryogenesis. Accordingly, our ongoing studies using human Embryonic Stem Cells (hESCs) 2D- models of gastrulation reveals that YAP is essential for the correct specification of the three germ-layers. The YAP KO-derived human 2D-gastruloids display expanded mesoderm and endoderm layers (ME) and reduced ectoderm layer, compared to WT. NODAL signaling homeostasis is essential for normal gastrulation; active NODAL cues are needed for ME differentiation. However, efficient NODAL inhibition is needed for the acquisition of an ectodermal potential. Our data suggest that the gastrulation phenotype in YAP KO cells is associated with an overly active NODAL signaling. The NODAL gene codifies for the ligand that activates the intracellular Smad2.3 signaling. Interestingly, our transcriptomic and epigenomic analysis suggest that YAP is needed to repress the well-characterized Proximal Epiblast Enhancer (PEE) of the NODAL gene during the exit of pluripotency toward an ectodermal fate. Our data further suggest that YAP restricts the chromatin development the human embryo will lead to discovery therapeutic tools to treat and prevent birth defects. The overarching goal of this proposal is to advance our knowledge the signaling and epigenetic mechanisms that regulate the process of human gastrulation. The Hippo- effector YAP1 (YAP) is However, increasing evidences the of the of accessibility of the NODAL locus, essential for gene repression. These general specification”. findings form the premise of our hypothesis: “Epigenetic regulation NODAL signaling by YAP is crucial for correct germ-layer We will address this idea in the following Aims. In Aim1 of , we will analyze the role of YAP in human 3D-gastruloids. We hypothesize that the ability of human epiblast cells to undergo three-dimensional gastrulation, and organization in an anteroposterior axis is regulated by YAP. To test this idea, we will apply a state-of-art synthetic human embryology model to study gastrulation in 3D. We also hypothesize that YAP- repression of the PEE of the NODAL gene is essential to balance pluripotency and ectoderm differentiation. To address this idea, we will investigate enhancer function and chromatin structure by combining cutting-edge CRISPR and genome-sequencing approaches (Aim2). Findings from Aim 1 and 2 will inform about human- specific regulatory roles of YAP that occur during axial organization. Finally, we will utilize a conditional YAP KO mouse to investigate the role of YAP in the differentiating epiblast, in vivo (Aim3). Our findings will inform on new mechanisms that regulate the activity of NODAL signaling and help to decipher the role of YAP during gastrulation.

NIH Research Projects · FY 2024 · 2022-08

Project Summary Transgender women (TW) are at significantly higher risk of getting HIV, yet research indicates they are less likely to use pre exposure prophylaxis (PrEP) and continue to have misconceptions about its utility. Trans- inclusive communication, delivered by trusted peers, is a potentially useful strategy to address this gap, but reaching a large number of TW can be difficult. The SIS (Sisters Influencing Sisters) study will use preliminary findings from a NIMH funded R21 that utilized commercial marketing techniques called perceptual mapping and vector message modeling to develop and concept test TW specific PrEP messages. Using the Gender Affirmation Framework and the EPIS Implementation Framework as guides, we propose to embed these tested PrEP messages and materials in the SIS intervention and pilot it using TW social influencers (SIs) via Instagram. Specific aims of the research are: Aim 1: Design a community-led, social-media based PrEP messaging training for TW SIs. We will recruit a Community Advisory Board (CAB) who will consult on all aspects of the study, including the development of an SI training and recruitment of SIs, tools/measures, dissemination of findings, and input on a future trial. Aim 2: Refine PrEP messaging materials, design the SIS intervention modules, and identify and train TW SIs. We will finalize PrEP messaging materials, engaging trans artists to refine images and convert to social media formats. With the help of our CAB, we will identify and engage SIs on social media from two intervention locations (San Francisco Bay, Philadelphia) and two comparison locations (New York and Los Angeles). Intervention SIs will be trained on the SIS intervention content as well as delivery of clinical referrals and resources to their communities. Aim 3: Aim 3: Pilot test the SIS intervention to collect feasibility, acceptability, and preliminary efficacy data to inform a full trial. We will implement standardized core components of the Instagram-based intervention, evaluate dissemination and reach of the PrEP messages, and monitor and evaluate engagement with the materials. To assess promise of efficacy, we will identify a cohort of 120 followers (80 intervention, 40 comparison) to complete baseline, mid and end of intervention surveys to track changes and compare levels of PrEP knowledge and PrEP perceptions. We will also pilot remote collection of hair samples to monitor PrEP uptake and use in a subsample of participants (n=30; 5 each SI) and do in-depth intervention implementation evaluation. We expect that findings will significantly contribute to our understanding of how to communicate with a diverse group of TW, how to leverage social media as a powerful communication channel, and how to disseminate information to increase PrEP awareness and uptake by addressing the unique HIV prevention needs and concerns of TW.

NIH Research Projects · FY 2025 · 2022-07

Natural selection on genetic variation is the ultimate cause of adaptation and the formation of new species. All studies of natural selection at the genomic level require a control group: a set of variants or substitutions that have not been subject to selection. The class of variants that, by far, are the most widely used for this purpose are synonymous changes within protein coding genes. Because these variants are interspersed with nonsynonymous variants, they are a natural control group. However, overwhelming evidence that synonymous changes are often not strictly neutral has accumulated over the past three decades. In many contexts the clear evidence for selection provides investigators with ways to identify which synonymous changes are most likely to be affected by selection and which are more likely to be strictly neutral. What is missing are ways to include this information, on classified synonymous changes (neutral and selected), within quantitative evolutionary models that enable a broad range of inferences about where, when, and on what selection operates. The proposed research is based on a new class of evolutionary models (Multiple Synonymous Substitution or MSS models) that include both selected and neutral synonymous changes in estimates of evolutionary change. These models can be fitted to homologous sequence alignments to infer relative rates of amino-acid replacement changes, and selected synonymous changes, to strictly neutral synonymous changes. These types of (dN/dS) analyses go back to the origins of evolutionary genetics, and today they remain among the most commonly used tools. Our hypothesis is that the current definition of what constitutes neutrally evolving sites is importantly wrong in that it biases the estimation of selective forces (by underestimating the rates of neutral changes), and that improved estimates without that bias will provide significantly better and more accurate understanding of the action of natural selection in a broad array of evolutionary contexts. We will develop and test a new class of substitution model that uses multiple sets of synonymous substitutions (MSS models), and we will apply them to a diverse set of taxa sampled from across life's kingdoms. Estimated MSS models, discovered using a genetic algorithm that searches over the space of all possible MSS models, will be validated using internal controls, based on neutral model predictions of substitution rate speed and constancy, and using external controls, based on predicted covariates of population genomic and functional effects. We will revise an array of widely-used comparative and population genetics techniques to take advantage of MSS models, and develop new approaches that seek to identify selection on synonymous changes. Our models will be applied to a wide taxonomic range of genomic data and made available as a part of popular software package HyPhy and web application Datamonkey.

NIH Research Projects · FY 2026 · 2022-07

SARS-CoV-2, the virus that causes COVID-10, impacts multiple organ systems including the central nervous system. Neurological symptoms are seen in about one third of COVID-19 patients and the virus has been found in brain tissue. SARS-CoV-2 uses angiotensin converting enzyme 2 (ACE2) located on cell surfaces for entry into cells, and the spike protein of the virus is the key recognition element for ACE2. ACE2 is expressed by endothelial cells of the cerebral vasculature that comprise the blood-brain barrier (BBB), the main barrier to entry of pathogens and toxins into the CNS. Recently, SARS-CoV-2 spike protein has been shown to alter BBB function using in vitro model systems. Many drugs of abuse, including psychostimulants, negatively impact BBB function as well. Therefore, it is likely that SARS-CoV-e infection in the presence of psychostimulants could result in greater damage ot the BBB, further increasing barrier permeability and resulting in CNS injury. Thus, investigation of the mechanisms underlying the interactions of SARSCoV- 2 and stimulants on the BBB is needed.

NIH Research Projects · FY 2026 · 2022-07

PROJECT SUMMARY Early life experiences can have profound and long-lasting consequences on health trajectories. Conditions, including low resources, have been identified as important factors that influence the development of psychiatric illnesses, including substance use disorders (SUD). In this proposal, a rat model of early life scarcity will be combined with behavioral assays relevant to SUD risk behavioral phenotypes to better understand the neural and molecular mechanisms that involved in the effects of early life resource scarcity on reward processing. Each brain region contains highly heterogenous cell populations that include different neuronal subtypes as well as glia. Accounting for the heterogeneity and differences in cell types is essential to improving our understanding of the impact of early life exposures on the brain and on motivated behavior. In this proposal, the influence of early life scarcity on adult reward processing and motivation will be characterized in male and female rats using state-of-the-art behavioral approaches where rats are tested for their motivation to earn drug (opioid) or natural (social and sucrose) rewards. Our preliminary data indicate sex- and reinforcer-specific effects of early scarcity. This work will be expanded here, and in some of the experiments, rats will choose between two available reinforcers. Given that interventions for SUD involve social reinforcers, these results could inform research aimed at developing new prevention and treatment strategies. To better identify factors that mediate the effects of early scarcity on motivated behavior, we will delineate molecular changes in the nucleus accumbens—a central hub in the brain that is critical for motivated and reward-related behaviors— and causally link them to behavior. To this end, we will perform cell-type specific assays of gene expression and chromatin remodeling, an epigenetic process that regulates the expression of genes. Lastly, the proposal will examine the impact of early life scarcity on the electrophysiological properties of two major neuron subtypes in the nucleus accumbens, delineating cell type-specific physiological changes induced by altering the early environment. Collectively, this proposal leverages cutting-edge behavioral, molecular, and physiological approaches to provide a better understanding of the neurochemical and intracellular pathways affected by early life scarcity that drive changes in motivated behavior. Importantly, the proposed experiments will determine sex- and cell-type specific mechanisms by which early life scarcity alters the substance use trajectory, identifying potential targets for improving therapeutics and prevention of SUDs.

NIH Research Projects · FY 2024 · 2022-06

PROJECT SUMMARY Influenza is a persistent viral disease worldwide, with high case counts leading to deaths and losses in productivity yearly. In temperate regions of the world, such as the United States, influenza appears in annual wintertime cycles, with the influences of changing seasons creating predictable patterns of incidence, despite the fact that different subtypes and strains of the virus predominate during each season. In tropical regions, distinct seasonal patterns are not well established for influenza, suggesting that annual or climatic forces may not dictate transmission in these areas. The tropics include many resource-limited areas of the world where respiratory disease research is underfunded and influenza vaccination is not prioritized. Transmission patterns of influenza can be described through the use of mathematical models, which incorporate known biological mechanisms, such as infectivity, recovery, and duration of immunity following infection, in order to describe the dynamics, or population-level behaviors, of the virus. Models can incorporate cyclic patterns of influenza dynamics, which can include both seasonal cycles aligning with a calendar year as well as nonannual periodic cycles; models can also incorporate irregular epidemics, though this is more challenging to model and less commonly seen in current literature. Irregular epidemics can be viewed as “regime switches,” which describes transitions between endemic time periods with consistently low incidence and epidemic time periods with short periods of elevated incidence. This study aims to investigate potentially cyclic or acyclic dynamics of influenza in the tropics, using Vietnam as a case study. The first aim seeks to define patterns describing cyclic or acyclic dynamics and then use a mathematical model to show under what conditions acyclic dynamics can exist. Influenza characteristics that could allow acyclic dynamics potentially include duration of immunity following infection, cross-immunity, and population structure. The second aim seeks to use the developed model as well as influenza data from Vietnam to determine whether influenza dynamics follow cyclic or acyclic patterns. Comparing these results to identical analyses using data from temperate regions influenza data will allow us to compare the strength of annual cycles between tropical and temperate regions. The third aim seeks to identify characteristics of influenza virus, population patterns, or external factors that may be able to explain the cyclic or acyclic patterns observed in influenza dynamics in Vietnam. This will show what characteristics of influenza or external factors provide the strongest predictions of future influenza incidence in Vietnam. Uses of the study results include better informed and appropriately-timed vaccination strategies for influenza in Vietnam. The main points of novelty in this study lie in the explicit testing for cyclic or acyclic patterns in the tropics, which is less commonly investigated, and a mechanistic modeling approach to identify the potential causes of non- annual behavior of influenza transmission in the tropics.

NIH Research Projects · FY 2026 · 2022-04

Neonatal Brachial Plexus Palsy (NBPP) is a complication of childbirth that can result in significant long-term sequelae. NBPP associated injuries often include cases of spinal cord injury (SCI) and are very poorly understood. Our long-term research goals are to develop both prevention and treatment strategies for NBPP. Consequently, objectives of this study are to use our novel and unique clinically-relevant neonatal piglet model: a) To identify brachial plexus (BP) strains and forces that lead to morphological and functional SCI by investigating the effects of BP stretch on acute pathology and functionality within the spinal cord (SC), b) To investigate clinically-relevant systemic biomarkers for early diagnosis of SCI during NBPP, and c) To develop computational models of maternal pelvis and fetus that predict risk of NBPP and associated SCI during complicated NBPP delivery scenarios. Based on our preliminary work, our central hypothesis is that moderate to severe BP stretches will transmit forces to SC resulting in SC tissue damage, which are also observable as high expression of systemic injury biomarkers in the serum and cerebrospinal fluid, and suppression or loss of motor neuron excitability. Our innovative approach is aimed at developing new knowledge of acute NBPP injury mechanisms and report SCI tolerance values of BP strains and forces. These data will enable novel clinical tools for diagnosis and prevention, as well as suggest targets for early clinical interventions for NBPP. We will test the overall hypothesis through the following independent specific aims: 1) To determine whether moderate to severe BP stretches will lead to SCI as evident by acute SCI markers in situ, functional loss in motor neurons and systemic acute SCI biomarkers, and 2) To identify delivery maneuvers and NBPP risk factors that lead to abnormal BP strains and forces, increasing the likelihood of associated SCI. The expected outcomes of this work are the first ever data on: 1) morphological and functional injury outcomes that help determine SCI tolerance values of BP strains and forces during BP stretch, 2) molecular biomarkers that can enable early diagnosis of SCI, and 3) developing highly biofidelic computational models for NBPP prediction and obstetric training. The results will also have an important positive impact, because, they lay the groundwork to develop a new class of targeted clinical interventions.

NIH Research Projects · FY 2026 · 2022-04

Abstract: This project is designed to identify a noninvasive and highly effective gene delivery strategy to target specific neural cells in the CNS and to validate this technology by delivering regenerative molecules to mammals with CNS injury. We will determine whether systemic delivery of our newly engineered AAV9 vectors can transduce most target CNS cells and whether noninvasive delivery of the genes that target neuronal intrinsic and extrinsic factors can promote robust axon regeneration and functional recovery in rodents with spinal cord injury (SCI). A major challenge in neuroscience research is to deliver target genes to specific types of neural cells widely distributed in CNS. Engineered AAV9 vectors usually show limited efficacy after intravenous (IV) injection by transducing cells only in some CNS regions. We thus created new AAV9 vectors that include multiple features of engineered AAV9 capsids, aiming to develop highly efficient BBB-crossing AAV9 vectors (HEBC-AAV9) that can transduce most target CNS cells after IV injection. In Aim 1, we will study efficiency of our novel HEBC-AAV9-GFP vectors for selectively transducing each type of neural cells (neurons, astrocytes, oligodendrocytes, and microglia) in several strains of adult mice. To solve a crucial issue in neuroscience research with this technology, in Aim 2 we will develop a regenerative therapy for SCI by systemic delivery of genes to target neuronal let-7 miRNA. After SCI, severed axons fail to regenerate partly because of reduced intrinsic growth capacity of mature neurons. Many genes are known to control the growth ability of mature neurons, but none have been translated to clinical use. The best targets are probably those with potential to impact multiple genes. Among them, let-7 is important for regulating age-dependent decline in axon regeneration in worms. In Aim 2, we propose to use unique HEBC-AAV9-synapsin vectors to target neurons selectively for inducing expression of let-7 inhibitor, lin28, and lin41, aiming to promote robust regeneration of multiple axon tracts by enhancing growth capacity of mature neurons in SCI rodents. Chondroitin sulfate proteoglycans (CSPGs) generated by glial scars strongly suppress axon extension and are major extrinsic molecular targets for treating CNS injury. Our lab designed small peptides to block functions of CSPG receptor LAR, PTPσ, and PTPδ by targeting their critical activity domains and demonstrated their high efficiency for promoting axon regrowth. In Aim 3, we will induce astrocytic expression of secreted 3 peptides for each of LAR, PTPσ, and PTPδ with HEBC- AAV9-GFAP vectors, aiming to promote robust axon regeneration after SCI by targeting extrinsic CSPGs alone or combined with intrinsic let-7 signals. Our new viral vectors should provide a powerful tool for gene delivery in CNS and for developing effective regenerative therapies for SCI and other neurological disorders.

NIH Research Projects · FY 2026 · 2022-02

Project Summary/Abstract Spinal cord injury (SCI) is a life-changing event that sets in motion profound alterations in motor output, sensory processing, and reflex activity. Immediately following injury there is a depression of motor activity mediated by a lack of descending drive. In the weeks and months following injury, a reorganization of spinal neurons (both motoneurons and interneurons) occurs in response to this altered milieu. This includes changes in the excitability and reorganization of remaining pathways through strengthening of latent connections and neuronal sprouting. SCI induced changes in the excitability of spinal motoneurons have a profound effect on their discharge characteristics and ultimately on the generation of muscle force for functional movements. The reorganization of spinal interneurons is less well understood, even though the overwhelming majority of synaptic drive to motoneurons is mediated by spinal interneurons. Therefore, this project will explore how spinal interneurons regulate the discharge of spinal motoneurons. We will combine intraspinal microelectrode arrays with arrays on the muscle to record the individual discharges of spinal interneuron and motor unit populations. On these data, we will bring to bear advanced statistical modeling to electrophysiology dissect the organization of spinal neurons through the quantification of the strength and directional effects of these excitatory/inhibitory connections. We will use this approach to quantify the organization of spinal neurons in the intact cord and following either chronic or acute SCI. Further, we will explore the effects of neuromodulation by targeting specific serotoninergic receptor subtypes. The expected outcome of this work is a new understanding of the function of the mammalian spinal cord and the capabilities for reorganization of spinal neurons following both chronic and acute spinal transections. This approach will be a platform for characterizing the synaptic input from spinal interneurons to motoneurons and how this ultimately produces movement. Our combined motor unit and interneuron approach will directly quantify the neural substrate underlying motor unit discharge patterns and will provide a strong basis for motor unit discharge patterns to be a detailed biomarker for quantifying the state of spinal interneurons in humans with SCI.

NIH Research Projects · FY 2026 · 2022-01

PROJECT SUMMARY/ABSTRACT Secondary bacterial infection following influenza (super-infection) can lead to cytokine storm (an overexuberant immune response) that often leads to pneumonia and death in patients. Our work focuses on the molecular mechanisms by which the immune system returns to homeostasis after microbial infections. Taking a holistic, systems approach, we investigated the inflammatory responses during a single (influenza or Staphylococcus aureus) and super-infection (influenza/S. aureus). We conducted transcriptional and lipidomic analyses in samples from a mouse super-infection model. Our lipidomic analysis was focused on eicosanoids because they play critical roles in inducing and resolving inflammation. When compared to single infections, we discovered an overproduction of a subset of eicosanoids during super-infection. These lipids (anti-inflammatory CYP450 lipid mediators, primarily DHET) can activate the nuclear receptors and transcription factors PPARa and PPARg. During influenza single infection, moderate induction of CYP lipids (primarily EET) during the resolution phase allows for appropriate anti-inflammatory responses to promote the return to homeostasis. We hypothesize that while EET promotes the physiological resolution of inflammation after microbial infections, DHET produced at an aberrant level during super-infection leads to the alteration in macrophage polarization and inhibition of bacterial clearance. The failure to control the bacterial pathogen amplifies the immune signals to recruit additional immune cells which eventually cause irreversible tissue damage. We will take the following approaches during single and super-infection to investigate the effects of the eicosanoid-PPAR axis on the inflammatory response. First, we will determine the effects of perturbing the eicosanoid-PPAR axis on the resolution or amplification of inflammation during single and super-infection. We will use chemical inhibitors in combination with genetic models to determine whether the animals will be protected from or succumb to disease during single and super- infection. We will determine the lipidomic profiles to assess the specific effects of the inhibitors have on the eicosanoid metabolism networks. We will also determine the bacterial/viral loads, cellularity, pathohistology, and targeted transcriptional profiling of macrophages. Second, we will determine the mechanism by which eicosanoid-activated PPARα/γ modulates immune signaling, macrophage polarization and immune metabolism in vitro. Macrophage polarization (classically or alternative activated) can amplify or resolve inflammatory responses. We will determine the potency of different CYP450 metabolites to activate PPARα/γ within mouse and primary human macrophages. We will determine how eicosanoids (CYP450 metabolites) affect the immune signaling, macrophage polarization, and lipid metabolism. Interestingly, While the induction of inflammation has been the subject of active investigation, the mechanisms underlying the resolution of inflammation have been elusive. By gaining insights into the resolution of inflammation during single and super-infection, we will develop novel therapeutic targets for infection- and immune-related human diseases.

NIH Research Projects · FY 2024 · 2021-09

The identification of the Apolipoprotein L-1 or APOL1 gene as an indicator of increased likelihood of developing chronic kidney disease has sparked a debate about point-of-care genetic testing for potential living kidney donors. While integrating genetic testing into the medical evaluation of potential living donors benefits patients, ensuring an informed, deliberative decision making process is paramount. The proposed study is premised on a large body of research demonstrating that traditional SDM processes and interventions fail to meet the needs of ethnic populations. Because SDM requires an informed patient to engage in the decision making process, novel methods are needed to assure a full, accurate understanding of risks, benefits, implications and alternate options. We propose a participatory approach to developing a decisional support tool for African American potential donors as they consider genetic testing as part of their medical evaluation. Specifically, this mixed-method study will conduct the formative research needed to develop and refine the tool. Up to 6 focus group interviews will explore potential donors’ preferences for shared decision making and understanding of the risks, benefits and implications of genetic testing in the context of live kidney donation (Aim 1). The findings will inform the design of a brief quantitative survey assessing potential donors’ decision-making preferences, communication style, and knowledge of and attitudes toward genetic testing, kidney transplantation and living donation. The survey (N=500) will be the first to include the Best-Worst Scaling methodology to assess the level of importance potential donors place on the risks and benefits of genetic testing as well as trade offs between risks and benefits donors are willing to make (Aim 2). A draft of the tool will be created, using the Ottowa Framwork as guide, to support African American patients and their families making decisions about both living donor kidney transplants and genetic testing in this context. Additional modifications will be made based on the findings of the focus group interviews and completed surveys. The tool will then be refined iteratively over four rounds of testing in simulated consultations with standardized providers and four rounds with a Delphi panel of transplant professionals (Aim 3). A Stakeholder Committee, comprised of transplant professionals and previous and potential living donors, will provide ongoing feedback and guidance across all study phases. In a subsequent R01, the fully-vetted, newly developed tool will be implemented and tested in clinical settings using a randomized controlled design. If successful, the tool will optimize care for the thousands of African American potential living kidney donors evaluated in the United States annually.

NIH Research Projects · FY 2025 · 2021-09

PROJECT SUMMARY/ABSTRACT Pediatric obesity is considered an epidemic with striking disparities by socioeconomic status (SES). There is therefore a pressing need to identify novel, modifiable risk factors for obesity prevention and treatment in children, particularly those who are at greater risk. Compelling evidence that is rooted in circadian biology and our understanding of circadian rhythms, and is supported by emerging work with rodents and adults suggests that in addition to focusing on how much we engage in eating, activity and sleep behaviors, it may also be important to understand the timing and consistency with which we engage in these behaviors (i.e., behavioral rhythms) for obesity risk reduction. Mounting evidence demonstrates that behavioral rhythms that are aligned with underlying circadian rhythms, such as sleeping during the biological night and eating during the biological day are associated with improved metabolic processes and weight regulation. Although less is known regarding how behavioral rhythms may optimize weight regulation in children, findings from related lines of pediatric research support those with rodents and adults and highlight the potential importance of behavioral rhythms for excess weight gain prevention. Understanding the role of behavioral rhythms in weight regulation for children from lower SES backgrounds may be particularly important given that they may be at greater risk for disruptions to behavioral rhythms due to economic adversity and neighborhood disadvantage, placing them at increased risk for excess weight gain. Further, understanding associations between SES, behavioral rhythms and weight regulation in young school-aged children focuses on a time period when a number of health behaviors that carry through adolescence are shaped and thus may represent an opportune time for optimizing weight regulation. The present study therefore proposes to enroll 176 children 5-8 years old from diverse SES backgrounds into a 16-month observational study. Children will complete five 10-day assessments across the study (baseline, 4, 8, 12, and 16 months) during which the following will be measured: timing and consistency of sleep (via actigraphy), timing and distribution of eating (via 24-hour dietary recalls), timing and consistency of physical and sedentary activities (via accelerometry and self-report), timing of the circadian clock (dim light melatonin onset; DLMO), measures of the home and neighborhood environment, and anthropometrics (height, weight, adiposity). The primary aims of this proposed work are to assess: a) how behavioral rhythms affect body mass index (BMI) trajectories in children, and b) how SES affects behavioral rhythms and thus BMI trajectories. Secondary aims will assess how circadian factors (i.e., circadian phase and circadian phase angle) and the household and neighborhood environments affect behavioral rhythms. Exploratory aims will also assess the relative influence of behavioral, circadian and environmental factors on children's BMI trajectories, including potential moderation of associations between behavioral rhythms and BMI trajectories by circadian phase and phase angle.

NIH Research Projects · FY 2025 · 2021-09

PROJECT SUMMARY This R01 application is in response to PAS-18-915 entitled “HIV/AIDS High Priority Drug Abuse Research”. Our project will investigate the molecular and functional interactions between HIV and morphine in regulation of alternative pre-mRNA splicing of the opioid receptor M1 (OPRM1) with implications for enhanced opioid dependence observed in the people with HIV (PWH). Clinically used opioids, such as morphine, as well as illicit drugs, such as heroin, activate OPRM1 that is a member of the G protein-coupled receptor (GPCR) family. OPRM1 pre-mRNA undergoes extensive alternative splicing events. To date, 21 isoforms of the human OPRM1 with alternative C-terminal and/or N-terminal regions and 17 isoforms of the rat OPRM1, have been identified. Given the importance of these regions in G protein-coupled receptor (GPCR) signaling, differential regulation of OPRM1 isoforms would have functional consequences. However, characterization of OPRM1 signaling is generalized, and only one isoform (MOR1) has been extensively studied. Our preliminary data suggest that expression of splicing regulatory protein SRSF1 and alternative splicing of MOR-1X is preferentially induced in neuronal cells exposed to morphine. Interestingly, our results also revealed that alternative splicing and expression of MOR-1X isoform is induced in postmortem brain tissues obtained from the PWH. These results suggested that HIV and morphine may impact OPRM1 alternative splicing and synergistically induce MOR-1X isoform expression. The mutually exclusive exon X of OPRM1 pre-mRNA is incorporated into the mature mRNA transcript following exon 3 in the MOR-1X mRNA transcript. This insertion results in substantially longer C-terminal tail having new motifs potentially binding with several cellular kinases that is unique to the MOR-1X isoform. We recently reported that glial cells infected with HIV-1 release Nef protein captured within the extracellular vesicles (Nef-EVs) which are readily taken up by neurons. We further assessed possible role of Nef-EVs and two other HIV secretory proteins, Tat and gp120, in alternative splicing of MOR-1X. Interestingly, while recombinant Tat and gp120 had no visible effects, treatment of neurons with Nef-EVs caused a comparable induction in MOR-1X alternative splicing as did treatment with morphine. Our preliminary results also revealed that co-treatment of neurons with Nef-EVs and morphine synergistically induced MOR-1X alternative splicing. Additionally, alternative splicing of MOR-1X was induced in brain regions involved in the reward pathways of the F344 rat that is the control strain of HIV-1Tg rat that has an additive induction with the exposure to morphine. Taken all together, we hypothesize that HIV-1 contributes to the increased rate of opioid dependence in the PWH by amplifying the rate of MOR-1X alternative splicing induced by morphine. Our proposed studies will reveal a novel synergistic interaction between morphine and HIV on alternative splicing of OPRM1 pre-mRNA leading to preferential expression of MOR-1X isoform with implications in physical dependence of morphine.

NIH Research Projects · FY 2025 · 2021-09

Project Summary/Abstract Nucleases are a class of enzyme that hydrolyze nucleic acid substrate in a variety of cellular processes. Their activity is a requirement for the maintenance of genomic integrity in DNA replication, repair, and recombination. Understanding nuclease regulation and specificity in these processes is critical for modeling these fundamental pathways and human diseases linked to dysregulation. This includes Lynch syndrome, a cancer predisposition syndrome linked to colorectal and endometrial cancers, Huntington’s disease, and infertility. Nucleases unique to bacteria are also potential targets for antibiotics and a complete understanding of nuclease biochemistry paves the way for the discovery of new drug targets and exploiting nucleases as new biotechnological tools and therapeutic agents. Characterizing and developing novel nucleases is a future area of interest for my research program. We will use proteins in DNA repair processes to model nuclease activity and determine regulation and specificity steps. Using the tractable DNA mismatch repair pathway which spellchecks newly replicated DNA, we will identify how inherently nonspecific nucleases can be given specificity. Proteins in this process have been co-opted for meiotic recombination and also play a role in the regulation of trinucleotide repeat expansions indicating that the associated nuclease activity is modular. This work addresses the origins of this co-option and provides missing mechanistic detail for how all of these pathways communicate substrate specificity to nucleolytic sites. In bacteria, homologous recombination is a method for acquiring antibiotic resistance. MutS2, a homolog to mismatch recognition complexes, has been implicated in several bacteria as being involved in this pathway. Its mechanisms of action and whether it follows paradigms established by canonical mismatch repair proteins are not clear and are addressed by mechanistic work described here. The nuclease domain of MutS2 is found throughout biology as a fusion to proteins with diverse specificities and functions. We will determine the modularity of this nuclease domain, how it achieves specificity by other domains, and test its potential for adaptation as a gene editing tool. We will also investigate the specificity and regulatory mechanisms of the newly discovered NucS protein which is multi-functional, and is implicated in multiple DNA repair processes in archaea and mycobacteria, including Mycobacterium tuberculosis, the bacteria that causes tuberculosis. This will provide key evidence for adaptation processes of organisms that utilize NucS in DNA repair processes. Our work will provide an underpinning for complex mechanistic models that can be ultimately used to detect and develop therapeutics for human disease. In addition, this work provides general insight into how nucleases are regulated and will guide future studies in other cellular pathways.

NIH Research Projects · FY 2025 · 2021-09

PROJECT ABSTRACT The overall goal of this K01 proposal is to provide Gina Tripicchio, PhD, MSEd, with the training and mentorship to establish an independent program of research focused on innovative behavioral interventions to improve dietary intake in high-risk adolescents. Added sugar (AS) is a prime target for dietary intervention in adolescents; it contributes excess calories with no nutritional benefits, and adolescents consume more AS than any other age group. High-risk adolescents (e.g., racial/ethnic minorities from low-income communities), experience disproportionately higher rates of obesity and diet-related chronic diseases yet remain understudied. A key limitation in developing efficacious trials for this high-risk group is the dearth of information about key factors associated with AS intake. To address this gap, this project aims to characterize key drivers of AS intake in high-risk adolescents using ecological momentary assessment (EMA). EMA allows for the examination of contextually specific dietary influences in real-time, while reducing recall bias and participant burden. This rich information can be used to guide the development of a tailored, salient behavioral intervention. Additionally, an objective stable isotope biomarker of AS has recently emerged and can be used to address key limitations of self-reported outcomes in dietary interventions. Two studies will be employed to meet the aims of this project. Study 1 (years 1-2), will collect EMA from 40 adolescents and examine the feasibility and acceptability of the δ13C biomarker in the target sample. Study 2 (years 3-5), will implement a pilot RCT to test a contextually tailored, technology-enriched intervention in a sample of 70 adolescents, 12-16 years of age, from a low-income community in north Philadelphia. A 3-month behavioral intervention, including group-based sessions, interactive text messaging and parent support, will aim to reduce AS intake compared to an educational comparison group. Changes in AS, as measured by the δ13C biomarker, will be assessed as the primary outcome and total changes in diet quality will be examined as a secondary outcome. A rigorous training plan will complement the proposed research, facilitated by mentors who are experts in nutrition, eating behavior, digital interventions, statistics, and behavioral trials in pediatric populations. Specific training objectives include: 1) ecological momentary assessment, 2) multi-level statistical modeling, 3) assessment of objective nutritional biomarkers and 4) designing and implementing behavioral interventions to address disparities. Temple University's Center for Obesity Research and Education is an ideal setting for this work as it provides extensive research resources and access to a population of diverse, high-risk youth. This K01 will provide novel information about AS intake in high-risk adolescents and test a targeted, tailored intervention to reduce consumption. Findings will support the application of a fully powered R01 trial. This award holds significant potential for public health impact and will enable Dr. Tripicchio to establish a rigorous program of research to improve dietary intake and reduce diet-related chronic disease risk in vulnerable adolescents.

NIH Research Projects · FY 2025 · 2021-09

PROJECT SUMMARY With widespread public concern about the increasing presence of smartphone and social media (SSM) technologies in the lives of American youth,1–3 there is a conspicuous need for investment in rigorous scientific work that will inform our understanding of the psychological precursors of SSM habits, and on the impacts that these habits may have on subsequent development and everyday functioning. While a foundational literature is starting to form, research on the origins and outcomes of SSM habits remains skeletal, and two basic questions are still unanswered: 1) Are the correlates of digital media habits stable across development, or do they vary with age? 2) In what direction do these relationships ensue – do observed correlations reflect individual differences that presage subsequent variation in SSM habit formation/intensity, or, do they reflect the impacts of SSM habits on consequent brain/psychological development and everyday functioning? The present study aims to refine our answers to these key questions through a multi-methodological (behavior, neuroimaging, self-reports, ecological momentary assessment) cross-sequential investigation of the individual differences factors that prospectively predict SSM habit formation and of the outcomes associated with intensification of such habits across development. Through an initial cross-sectional assay of children, adolescents, and young adults, we will investigate whether the relations between psychological/brain functioning and SSM behaviors vary as a function of age. In a subsequent longitudinal phase, we will concurrently track trajectories of psychological/brain maturation, everyday functioning (e.g., academic outcomes, psychological and physical wellbeing), and SSM engagement, in order to shed light on the temporal chain of processes linking mental functioning and SSM usage. Our investigative team is extremely well positioned to execute the proposed research, which will draw heavily from the theoretical framing and methods that have propelled our previous work, and which will recruit from active developmental cohorts who have participated in related recent studies implemented in our labs. Based on guiding neurodevelopmental theories, we hypothesize that individual differences in self-regulatory control, reward and sensation seeking, reactivity to social inputs, and risk-taking propensity will be differentially predictive of usage patterns at varying points in development, and that these relationships will be shown to have bidirectional interactions with intensifying SSM behaviors, with important implications for everyday functioning.

NIH Research Projects · FY 2025 · 2021-09

7. Project Summary/Abstract Adolescence is an “age of risk” for the emergence of first onset of bipolar spectrum disorders (BSD). Despite their prevalence and public health significance, major unanswered questions exist regarding the mechanisms involved in vulnerability to BSDs. BSDs are associated with hypersensitivity to reward and elevated reward- related brain function. However, research has not yet tested whether chronically high reward responsivity (RR) or increases in RR development during adolescence, beyond baseline RR, predicts first onset of BSD. A separate literature documents circadian rhythm disruption in BSDs, and social rhythm disruption (SRD) can trigger BSD episodes. Yet, research has not tested whether baseline circadian dysregulation, chronic social and circadian rhythm disruptions, or increases in these rhythm disruptions during adolescence predict onset of BSD. Further, circadian and reward approaches to BSDs mostly have proceeded in parallel. However, we and others have proposed integrated reward-circadian models of BSDs based on evidence the two systems influence each other and interact to affect mood functioning. When dysregulated, reward and circadian system signaling may combine to form a positive feedback loop, whereby dysregulation in one system exacerbates dysregulation in the other. This proposal is the first systematic test of a novel, integrated reward-circadian model for first onset of BSD. We will use an innovative biobehavioral high-risk design to examine bidirectional relationships between multiple indices and domains (monetary, social) of RR and multiple indices of social and circadian rhythms and their joint prediction of first onset of BSD and increases in bipolar symptoms. Three hundred twenty 14-16 year old participants (Ps) will complete a prospective 3-year longitudinal study. Ps with no prior BSD will be selected along the entire dimension of self-reported RR, with oversampling at the high tail of the dimension in order to increase the likelihood of BSD onsets. At Times 1-6, every 6 months, Ps will complete assessments of reward-relevant and SRD life events and self-report and diagnostic assessments of bipolar symptoms and episodes. Yearly, at Times 1, 3, and 5, Ps also will complete self-report measures of circadian chronotype (morningness-eveningness) and social rhythm regularity, a salivary dim light melatonin onset (DLMO) procedure to assess circadian phase, self-report, behavioral, and neural (fMRI) assessments of monetary and social RR, and a 7-day EMA period. During each EMA period, Ps will complete continuous measures of sleep/wake and activity (actigraphy) and 3 within-day (morning, afternoon, evening) measures of life events coded for reward-relevance and SRD, monetary and social reward responsivity, positive and negative affect, and hypo/manic and depressive symptoms. The fMRI scan and DLMO procedure will occur on the day before the start of each EMA period, excluding weekends. This proposal is an innovative integration of research on reward and circadian signaling in understanding first onset of BSD in adolescence. It has the potential to facilitate reward and social/circadian rhythm interventions to treat, and ideally prevent, BSD.

NIH Research Projects · FY 2025 · 2021-09

PROJECT SUMMARY One common pathologic hallmark of Alzheimer's disease (AD) and related tauopathies is the abundant presence of abnormal aggregates of highly phosphorylated tau protein in the brain parenchyma. Because the etiology of the vast majority of these cases (late-onset) is not known, currently there are no effective preventative measures or therapeutic approaches against them. Considering the increasing number of individuals affected by these disorders there is a sense of urgency to identify the mechanisms responsible for the onset and development of their neuroptahologic phenotype. Recently, small non-coding RNAs, also called microRNAs (miRNAs), have emerged as important post- transcriptional master regulators of several key cellular processes involved in neurodegeneration. Consistent data in the literature showed that miRNAs are dysregulated in AD and related tauopathies. However, since these studies typically assessed a single time point in the disease evolution they did not address whether the changes antecede or follow the onset of the pathology. Using an unbiased approach, in our preliminary studies we discovered an age-dependent increase in the expression levels of a specific miRNA, miRNA22-3p, in the hippocampus of a relevant mouse model of tauopathy at an early stage of its phenotype. Importantly, we confirmed this observation also in post-mortem human tauopathy brain tissues when compared with age-matched healthy controls. Additional studies revealed that this miRNA directly modulates pathologic tau accumulation. The hierarchical hypothesis of this research program is that miRNA22- 3p directly contributes to the tauopathy pathogenesis through regulation of specific targets involved in tau metabolic pathways, and that the modulation of its level represents a new therapeutic approach for the treatment of these diseases. To test our hypothesis, we will over-express miRNA22-3p in a relevant tauopathy mouse model (Specific Aim 1) and down-regulate miRNA22-3p expression levels in the same model (Specific Aim 2). In order to establish its cellular source and contribution to the tau phenotype, we will generate tauopathy mouse models with cell- specific miRNA22-3p deficiency (Specific Aim 3). In all these models we will assess the effects and mechanisms of manipulating this specific miRNA level on tau neuropathologic phenotype and cognitive functions. Overall the long-term goal of our research proposal is to establish the functional role that miRNA22-3p plays in the pathophysiology of AD and related tauopathies, and ultimately by identifying its biological targets to develop novel and viable therapeutic tools and strategies against these devastating diseases.

NIH Research Projects · FY 2025 · 2021-08

While antiretroviral therapy (ART) has dramatically reduced HIV disease morbidity and mortality, it has failed to eliminate viral reservoirs. Interruption of treatment leads to activation of latent virus and rebound viremia within weeks. Novel strategies are urgently needed to eradicate latent infections and enhance the immune system leading to sustained, durable control of viral rebound following the cessation of ART. In response to RFA-AI- 20-035 Martin Delaney Collaboratories for HIV Cure Research, we now submit the application entitled "CRISPR for Cure." The overarching goal of this program is to use genome editing mediated by CRISPR to enhance immune responses and directly ablate HIV proviruses. We have assembled a collaborative team of highly accomplished basic and translational scientists working in tandem with community stakeholders and a small biotechnology company to develop CRISPR-based therapies to directly target the HIV provirus and to enhance immunological responses. The research program is comprised of three highly interactive research foci (RF) that will utilize interdisciplinary, innovative and collaborative research approaches with community and government input. RF1 will use next generation sequencing and novel barcoded viruses to define the HIV reservoir and the impact of epigenetic mechanisms on proviral rebound. In RF2, we will enhance effector NK and CTL cell function and killing and limit viral spread by target cells using innovative genome editing strategies. RF3 will create and test the next generation of inducible, multiplex CRISPR with increased specificity, potency and safety for delivery by CD4 tropic lymphoid AAV9 for eradication of HIV-1 proviral DNA in animal models whose immune cells are modified in RF2 and assess the possibility of both a universal and personalized CRISPR in eliminating replication competent virus in vivo. In addition to the shared focus on CRISPRs technology, the Collaboratory will undertake a highly integrated experimental agenda through the shared use of barcoded viruses in humanized mice and unique support MISTRG humanized mice that differentially human hematopoietic stem and progenitor cell maintenance and myelopoiesis;rhesus macaques infected with a novel SIV barcoded virus; ex vivo clinical samples from a well characterized cohort and the use of adenoviruses to efficiently deliver CRISPRs to an in vivo humanized animal model carrying cells from patient-derived PBMCs. The outcome of this comprehensive and multidisciplinary program by the “CRISPR for Cure” the Collaboratory, will accelerate the use of gene editing strategies towards eradication of HIV infection from body or sustained viral remission following cessation of antiretroviral therapy.With resources available from our private sector partner, we will be well positioned for further GMP manufacturing development, and future initial clinical investigations.

NIH Research Projects · FY 2026 · 2021-07

Project Summary The lack of regular physical activity (PA) in over 290,000 individuals with spinal cord injury (SCI) in the United States (US) is an ongoing health crisis. This lack of activity has potentially devastating consequences because low levels of PA in people with SCI elevates the risk of mortality due to cardiovascular diseases, diabetes, and lung disease. Furthermore, low levels of PA in individuals with SCI have been associated with secondary conditions such as pain, fatigue, weight gain, and deconditioning. Regular PA and exercise-based interventions have been linked with improved outcomes and healthier lifestyles among those with SCI. Sensor- based activity monitors can assess PA and exercise interventions by quantifying wheelchair movement, movement of the individual, and physiological changes. However, these monitors do not provide real-time, tailored feedback and recommendations that might help individuals with SCI increase their PA levels in the community. The overarching goal of this proposal is to evaluate a sensor-enabled, just-in-time adaptive intervention (JITAI) strategy to increase and sustain PA levels among individuals with SCI in their communities. The long-term goal of this research is to effectively integrate a mobile health JITAI with existing PA intervention programs to motivate health-related behavior change in individuals with SCI. A primary objective of this proposal is to extend our pilot work to evaluate the integration of a JITAI with a web-based 14-week PA intervention program from the National Center on Health, Physical Activity and Disability (Aim 1). We hypothesize that the integration of web-based PA intervention program with JITAI will result in significantly higher PA levels over 14 weeks compared to the standard we-based PA intervention program alone. A secondary objective of this study is to extend existing algorithms that use commercial wearable technology to robustly detect PA behaviors to facilitate the delivery of tailored just-in-time actionable feedback and PA recommendations for individuals with SCI (Aims 3 and 4). The integration of the JITAI, which provides feedback and PA recommendations due to sensor-based assessments of PA, with the standard web-based PA intervention program will be tested via a clinical trial that combines a randomized controlled trial and a micro-randomized trial. Our team includes investigators with expertise in SCI research, mobile health, PA tracking, and behavioral change interventions. The proposed study will yield novel insights about JITAIs and JITAIs combined with more traditional, web-based PA intervention programs, which will help researchers design engaging PA interventions for individuals with disability in the community that may improve their health and quality of life.

NIH Research Projects · FY 2024 · 2021-07

PROJECT SUMMARY Patients with chronic obstructive pulmonary disease often show airway epithelial remodeling including basal cell hyperplasia, and goblet cell and squamous cell metaplasia. Such pathologic changes profoundly affect the outcome of respiratory infection as airway epithelium plays a crucial role in defining the innate and adaptive immunity in the lungs. Airway basal cells are the specialized stem cells and regenerate functional mucociliary- differentiated airway epithelium upon injury. The fact that COPD patients show airway epithelial remodeling indicate dysregulated repair mechanisms in airway basal cells. Our research suggests that quercetin, a natural polyphenol reverses airway epithelial remodeling in a mouse model of COPD. Our preliminary studies indicate that quercetin reprograms dysregulated repair pathways in COPD basal cells leading to regeneration of normal airway epithelium. We conducted transcriptomic analysis of airway basal cells from healthy non-smokers and COPD subjects and COPD basal cells treated with quercetin. Results from this microarray indicated dysregulation of genes involved in tissue development and epithelial differentiation in COPD cells. Intriguingly, the topmost differentially regulated genes in both these pathways are genes involved in lung morphogenesis HOXA1 and HOXB2. In normal basal cells, HOXA1 is highly expressed in basal cells, while HOXB2 expression increased at two weeks of culturing and correlated with polarization of cells, a prerequisite step in differentiation. COPD basal cells showed significantly reduced expression of both HOXA1 and HOXB2 and quercetin treatment increased expression of both genes. Based on these observations, we will examine a novel hypotheses that quercetin via modulation of HOXA1 and HOXB2 corrects the dysregulated repair mechanisms, thus improving immune responses to respiratory infections and lung function in COPD. In Specific Aim 1, we will determine the role of HOXA1 and HOXB2 in the regeneration of airway epithelium, and whether quercetin corrects the dysregulated repair mechanism in COPD by modulation of these HOX genes. In Specific Aim 2, we will examine the molecular mechanisms by which quercetin-induced HOXA1 and HOXB2 participates in the regeneration of airway epithelium. In Specific Aim 3, we will examine whether quercetin-induced HOXA1 and HOXB2 participate in limiting exaggerated innate immune responses to rhinovirus infection and prevent progression of lung disease in COPD. Finally, we will confirm whether quercetin treatment reduces airway epithelial remodeling in COPD patients and correlate with the expression of HOXA1 and HOXB2. Completion of these studies will provide important insight into airway epithelial regeneration and the mechanisms by which quercetin reduces airway epithelial remodeling in COPD.

NIH Research Projects · FY 2025 · 2021-07

Candida albicans exists as a commensal yeast in healthy people while it can cause mucosal and systemic fungal infections including oropharyngeal candidiasis (OPC) in immunocompromised individuals and diabetics. C. albicans has the ability to sense environmental iron as a signal with the help of its signaling pathways, such as Mitogen Activated Protein Kinase (MAPK) Cek1, to modulate gene expression in response to extrinsic iron levels. Recent evidence has shed light on the role of different iron levels between free iron rich gut and free iron deplete blood, allowing C. albicans to choose between a commensal or virulent lifestyle in these respective niches. Nothing is known about how variations in iron levels within the oral cavity will influence virulence during OPC. Our preliminary data shows that iron chelation in murine OPC causes significant reduction in virulence, while iron supplementation leads to greater fungal load in the tongue tissue of the infected mice. We also show how iron communicated with various signaling pathways including C. albicans MAPK Cek1 to modulate the fungal cell wall (CW), thereby affecting response to antifungals (by changing CW component levels) and host immune attack (by affecting exposure of immunogenic β-glucan in the CW). The main goal of this project is to understand the mechanisms behind iron-mediated changes in C. albicans CW that in turn effect the outcome of infection during OPC. Besides, in vitro experiments to address this goal, we will use our murine OPC model and mice will be repleted or depleted in systemic iron levels with iron supplementation (with iron dextran) and iron chelation (with iron chelator Deferasirox), respectively. We will also study mucosal colonization by C. albicans in Caenorhabditis elegans worms with human-like iron disorders, to evaluate fitness of fungal virulence traits directly in host with varying iron levels. Our overall hypothesis is that iron communicates with various signaling pathways to modulate fungal CW, in turn affecting hyphal morphogenesis, drug sensitivity, and susceptibility to host immune response, in OPC. We will test our hypothesis using three specific aims: 1) Define iron-induced signaling mechanisms that modulate C. albicans β-glucan exposure, levels of CW components, and hyphal morphogenesis, 2) Determine how iron affects C. albicans CW remodeling and hyphal morphogenesis during murine OPC to impact antifungal drug susceptibility, and 3) Evaluate how changes in host iron levels modulate innate immune defense mechanisms against C. albicans in vitro, during murine OPC, and in C. elegans. This work will provide insights into how varying iron levels in susceptible populations will affect the outcome of C. albicans infection during OPC.

NIH Research Projects · FY 2025 · 2021-07

The biomedical workforce research needs of our society have increased in size and diversity. We have the responsibility to help train the next generation of scientists, not only to feed the research needs of academia and industry, but to train inspiring college science faculty, effective research administrators, science policy makers, medical writers and scientific editors, business and financial analysts, science consultants, big bioscience data managers as well as other future essential biomedical research jobs. Thus, our graduates will need to be equipped with abilities for rigorous critical thinking, develop an interdisciplinary perspective, learn to analyze complex sets of data, develop complex research approaches utilizing complex equipment while maintaining the mental flexibility to keep up with the fast pace of biomedical research exploration and application. The overall goal of this predoctoral program is to offer exceptional interdisciplinary training opportunities in the broad area of Molecular and Cellular Biology and Genetics with special interest in cell signaling, epigenetics and genome maintenance. Members of this interdisciplinary program include faculty from the Lewis Katz School of Medicine (LKSOM), who are mentors in the Cancer Biology and Genetics (CBGN) and the Molecular and Cellular Biosci- ence (MCBS) concentration clusters. While there are many traditional programs that offer training in each of these three areas, this program will provide students with the tools and perspectives needed to consider how these three highly disease-relevant fields intersect. The focus for this training proposal was chosen primarily to reflect the presence of multiple leaders in these fields in the CBGN and MCBS clusters. This proposal will support research training opportunities in the labora- tories of 36 Assistant, Associate and full Professors. Mentors in this program bring strength in multidisciplinary programs in molecular and cellular biology, biochemistry, biophysics, genetics and epigenetics, using state of the art approaches in confocal and TIRF microscopy, mass spec proteomics, structural biology, molecular mod- eling, bioinformatics, high throughput screens, genomics, and epigenomics using a variety of animal and cellular model systems and human samples. Mentors in this program have a long tradition in the training of both graduate students and postdoctoral fellows with very high academic standards and will receive additional training in mod- ern evidence-based mentoring practices. This program will further provide a broader understanding of disease mechanisms and will develop in the trainees an in-depth understanding of the clinical challenges associated with their research interest area. Upon graduating, we are confident that this unique interdisciplinary training experi- ence will make our students not only highly competitive for future positions in academia, industry, science college education, research administration, science policy and advocacy, technology transfer, science and medical writ- ing and scientific editing, business and financial analysis, science consulting, big bioscience data managing and a broader array of future biomedical research jobs.