University Of Illinois At Chicago

NIH Research Projects · FY 2025 · 2021-07

Project Summary MicroRNAs (miRNAs) are a family of small non-coding RNA molecules that downregulate the expression of their gene targets. Both computational and experimental studies have shown that thousands of human protein-coding genes are regulated by miRNAs, and miRNAs are master regulators of many important biological processes. Changes in miRNA expression can have profound biological impacts, leading to a variety of human diseases. Due to their critical roles in gene expression regulation, functional characterization of miRNAs has become one of the most active research fields in biology in recent years. Despite rapid progress in miRNA research, one major obstacle in this field is the lack of robust computational and experimental methods for functional miRNA analysis. Funded by this NIGMS R01 grant over the past ten years, we have made significant progress and published many methods on miRNA and gene expression studies. The methods we established have been widely adopted by the scientific community. In particular, we have developed a miRNA target prediction tool, miRDB, which has quickly become a major bioinformatics resource and has been cited by thousands of publications. Moreover, we have developed innovative experimental methods to manipulate the expression levels of miRNAs. Built on our recent progress, we propose to perform combined computational and experimental analyses to continue to develop cutting-edge technologies for miRNA studies. Our work is expected to exert a significant impact in the miRNA field by providing valuable resources for functional miRNA analysis.

NIH Research Projects · FY 2024 · 2021-07

Acoustic communication is crucial for social interactions in many species, including humans. Understanding the neural underpinnings that govern the production and processing of communication sounds is paramount to advance the fields of auditory neuroscience and social behavior. Studies investigating speech and sound processing in humans have mostly implemented non-invasive methods, leaving a gap in knowledge about underlying neural mechanisms. My project bridges this gap by exploiting scientific advantages of echolocating bats, mammals that produce and process a rich repertoire of acoustic signals, to investigate the circuits that contribute to the discrimination of complex sounds that carry different meanings. Bats are social mammals with well-developed audio-vocal systems and produce ultrasonic vocalizations for navigation and social communication, providing a distinct opportunity to study the pathways, molecules and brain regions, which enable complex sound processing. Aim 1 combines behavior and neurophysiology to investigate the specific acoustic features of communication calls that are key to evoke behavioral responses and the neural systems involved in sound discrimination. Aim 2 combines psychophysical, neurophysiological, and pharmacological inactivation methods to study the midbrain-amygdala circuit’s role in mediating discrimination of sounds that show overlap in spectro-temporal features but carry different semantic content. Aim 3 investigates circuit phenomena in a social context by combining neurophysiological recordings and targeted pharmacological inactivation in freely interacting bats. The overarching hypothesis of this research program is that social- emotional processing of auditory stimuli through a midbrain-amygdala circuit mediates the discrimination of sounds that carry different meaning. The significance of this project resides in the extraordinary scientific opportunities to bridge studies of auditory behaviors, single neuron recordings, circuit dissection and computational modeling in a mammalian model. This work will contribute key new knowledge of natural sound processing mechanisms in mammals that could inform a deeper understanding of human auditory communication disorders. This supplement will enable the training of Zaria George to carry out Aim 3 of the project. The University of Illinois at Chicago offers an outstanding environment to conduct the independent portion of this project, as it provides access to world class research facilities, seminars and workshops offered by the Laboratory of Integrative Neuroscience; along with an extraordinary network of colleagues and collaborators.

NIH Research Projects · FY 2025 · 2021-06

Summary: Tissue repair is a complex process that involves a delicate temporal balance between inflammatory and regenerative mechanisms. In health, initial inflammatory events are replaced by regenerative processes in a coordinated manner. This sequence is disrupted in diseased states and complex injuries. The goal of regenerative medicine is to reestablish this balance by preventing chronic/aberrant inflammation and promoting repair and regeneration in a tissue-specific manner. While stem cell and growth factor therapies have been explored for this purpose traditionally, recent studies highlight the immunomodulatory and protective functions of mesenchymal stem cell derived extracellular vesicles (MSC EVs). Although MSC EVs possess versatile properties, to engage tissue-specific pathways and fit the goals of precision medicine with translational relevance, MSC EVs have to be engineered for enhanced pathway-specific functionality and delivered on site in a spatially and temporally controlled manner. In this proposal, leveraging our preliminary results and our expertise in EV biology, immunology and bone biology, we hypothesize that: Spatiotemporal control of immunomodulatory and regenerative pathways can be achieved by selective incorporation of Functionally Engineered EVs (FEEs)in 3D printed scaffolds. Using bone regeneration as a model system, we will test this hypothesis in three specific aims. In aim 1, we will generate functionally engineered EVs (FEEs) that target specific osteoinductive and immunomodulatory pathways. In aim 2, we will develop a photocrosslinkable alginate-based delivery system with EV carrier and release motifs for spatial localization and temporally controlled delivery of the FEEs developed in aim 1. In aim 3, we will utilize 3D printing technology to print defined structures encapsulating the FEEs for spatially and temporally controlled biphasic delivery in vivo. This system will be tested in a rat calvarial defect model. From the proposed studies, we will develop a platform technology that can impact the field of regenerative medicine beyond the craniofacial and musculoskeletal systems.

NIH Research Projects · FY 2025 · 2021-06

PROJECT SUMMARY Chronic alcohol exposure is associated with the development of tolerance to alcohol’s aversive properties, which serve to limit drinking. In contrast, the response to alcohol’s rewarding properties, which promote drinking, remain unchanged. The brain undergoes a host of neuroadaptive changes as a result of chronic alcohol exposure but the neural mechanisms underlying this tolerance are unknown. Impaired signaling in circuits encoding aversion is a likely candidate mechanism. The rostromedial tegmental nucleus (RMTg) is characterized for its involvement in aversion including signaling the aversive properties of ethanol. The prelimbic (PL) medial prefrontal cortex (mPFC) shares important functional similarities with the RMTg including facilitation of aversion learning and regulating the behavioral response to aversive stimuli. The neighboring infralimbic (IL) subregion of the mPFC exerts opposing effects to PL mPFC by facilitating extinction of aversive responding. Our work has uncovered a dense projection from the mPFC to the RMTg that spans both PL and IL subregions. Our findings from experiments investigating the function mPFC inputs to the RMTg support a role for these circuits in regulating aversive responding. These data lead us to hypothesize that subregion- and circuit-specific dependence-induced plasticity in RMTg-projecting mPFC neurons facilitates chronic tolerance to ethanol’s aversive properties. The aims described in the current proposal will use innovative, circuit-specific strategies to test this hypothesis. Aim 1 will use in vivo fiber photometry to measure changes in calcium signal in RMTg-projecting PL and IL mPFC inputs during the development of tolerance. Aim 2 will use closed-loop in vivo optogenetics to dissect the effects of bidirectional manipulation of PL and IL mPFC inputs to the RMTg on measures of chronic tolerance to ethanol’s aversive properties. Aim 3 will use whole-cell patch-clamp slice electrophysiology and a virally- mediated intersectional approach to identify dependence-induced changes in synaptic and structural plasticity underlying tolerance to ethanol’s aversive properties. The results of these studies will provide crucial insight into the circuit-specific neural mechanisms that contribute to uncontrolled alcohol drinking. In doing so, our findings have the potential to uncover new therapeutic targets for the treatment of alcohol use disorder.

NIH Research Projects · FY 2024 · 2021-06

ABSTRACT Coronary artery calcium (CAC) provides predictive information for risk of coronary heart disease (CHD) events beyond that gained from traditional cardiovascular disease (CVD) risk factors. Hispanics/Latinos have lower prevalence of CAC than whites, even after accounting for differences in CVD risk factors. They also have lower rates of CVD mortality and longer life expectancies despite higher rates of risk factors and adverse socioeconomic conditions (the ‘Hispanic Paradox’), lending credence to the existence of Hispanic/Latino-specific CVD protective mechanisms. Little data currently exist on the presence/absence and extent of CAC and CT- derived plaque volume, density, and distribution in diverse Hispanics/Latinos, and their associations with sociocultural characteristics and lifestyle factors that have been associated with CVD in the Hispanic Community Health Study/ Study of Latinos (HCHS/SOL) and other cohorts. Another critical knowledge gap is whether epigenetic mechanisms, i.e., DNA methylation, alone or in conjunction with genetic factors, act as an important biological link between such risk or protective factors and subclinical/clinical CVD in Hispanics/Latinos. The proposed study aims to generate critical data on subclinical coronary atherosclerosis (CAC and CT-derived plaque volume, density, and distribution) for Hispanic/Latino adults from all major heritage groups in the US (Mexican, Puerto Rican, Cuban, Dominican, and Central/South American) and examine relationships with sociocultural factors, lifestyle factors, traditional CVD risk factors, and epigenomic patterns. This study will be conducted ancillary to the ongoing HCHS/SOL, which examined 16,415 Hispanic/Latino men and women ages 18-74 years at Visit 1 (2008-2011) from 4 US communities (San Diego, CA; Chicago, IL; Bronx, NY; and Miami, FL) and 11,623 participants (81.4% follow-up rate) at Visit 2 (2014-2017). The proposed study will capitalize on available HCHS/SOL data and will assess CAC absence/presence and features in conjunction with ongoing HCHS/SOL Visit 3 (2020-2023) in 4,109 participants ages >45 years (mean age 60.5 years, an optimal time window for CAC detection) and free of known history of CVD. Longitudinal examination of epigenetic biomarkers will be conducted using stored blood DNA samples from HCHS/SOL Visits 1 and 2 for a subset of participants (50% of participants who undergo scanning, n=2,054). The proposed study will address critical knowledge gaps in a highly cost-effective manner by capitalizing on the established HCHS/SOL infrastructure. Examination of these factors in this heterogeneous, well-characterized cohort, has the potential to generate novel and insightful epidemiological, etiologic, and mechanistic information beyond that gained from previous studies, which can help to identify novel preventive targets and inform development of precision prevention/treatment strategies that may benefit all racial/ethnic groups.

NIH Research Projects · FY 2025 · 2021-06

PROJECT SUMMARY Despite the significant burden that viral infections continue to impose on human health, effective therapeutics and vaccines are still unavailable for many viral pathogens. A stronger understanding of the mechanisms by which the immune system senses and controls viral infections will enable the development of new strategies for their prevention and treatment. Interferons (IFN) are critical mediators of the immune response to viral infections, but excessive amounts can lead to chronic inflammation and autoimmunity. The production of IFN is therefore tightly regulated. Replicating viruses produce RNAs that are detected within cells by RNA sensors, RIG-I and MDA5. RIG-I detects viruses such as Sendai virus (SeV) and vesicular stomatitis virus (VSV), whereas MDA5 is essential for IFN induction to control picornaviruses such as Theiler’s murine encephalomyelitis virus (TMEV), and the Calicivirus, murine norovirus (MNoV). Once activated, these sensors initiate a common signaling cascade that leads to the transcriptional induction of IFN and establishes an anti-viral environment. Although the Linear Ubiquitin Chain Assembly Complex (LUBAC) has been shown previously to inhibit IFN induction by RIG-I, our studies have shown that LUBAC subunit, HOIL1, is essential for IFN induction by MDA5 in murine dendritic cells and fibroblasts, and to control MNoV infection in vivo. Furthermore, we have found that the E3 ubiquitin ligase activity of HOIL1 is essential for IFN induction during TMEV infection of fibroblasts. However, the mechanism by which HOIL1 E3 ligase activity regulates MDA5-dependent IFN induction, and the biological consequences of ubiquitination by HOIL1 are unknown. We hypothesize that HOIL1 and LUBAC are recruited selectively to the MDA5 signaling pathway, wherein HOIL1 catalyzes ubiquitination of one or more signaling molecules in the pathway to facilitate signal transduction and transcription of IFN genes. First, we will determine the location of the blockade in the MDA5 signaling cascade in HOIL1 E3 ligase mutant cells, and identify other LUBAC protein domains that regulate MDA5 signaling and IFN induction. Second, we will use both candidate and unbiased biochemical approaches to identify HOIL1-interactors and ubiquitinated proteins during MDA5 signaling. Last, we will test whether HOIL1 E3 ligase activity differentially regulates IFN induction and viral pathogenesis in vivo during MNoV, SeV, VSV and TMEV infection of mice, and will use cell type-specific knock-out mice to identify the cell types that require HOIL1 to induce IFN and control MNoV persistent infection in vivo. We expect our studies to identify a novel mechanism of IFN regulation, thereby revealing potential therapeutic targets for viral and IFN-mediated autoimmune diseases.

NIH Research Projects · FY 2025 · 2021-06

The proposed work focuses on (a) understanding how natural multimetallic clusters operate as active sites in metalloenzymes, and (b) using synthetic multimetallic clusters for productive chemical synthesis. Many metalloenzymes feature multimetallic clusters as active sites for catalyzing multielectron/multiproton transformations of gaseous small molecules, including cases of environmental significance to human health like atmospheric regulation of carbon dioxide and nitrous oxide. In many cases, these multimetallic clusters have unusual compositions and geometries, and so the chemical reactivity patterns have not been mapped out in laboratory settings. This proposal aims to achieve that for several multimetallic active sites, including the tetrametallic active site of nitrous oxide reductase and the heterobimetallic active site of carbon monoxide dehydrogenase, using synthetic model studies based on the PI’s established expertise in this area. Additionally, lessons about how these natural multimetallic clusters operate will be applied to develop synthetic multimetallic catalysts for C-C and C-X bond-forming reactions of relevance to pharmaceutical synthesis. The benefits of developing this catalytic paradigm for synthetic organic chemistry include emergence of reactivity and selectivity patterns that are complementary to established mononuclear catalyst systems, as well as the advancement of a bio-inspired tool to design earth-abundant metal catalysts that improve the sustainability of synthetic technologies in the pharmaceutical industry.

NIH Research Projects · FY 2025 · 2021-06

Summary Diabetes mellitus is a major cause of morbidity and mortality in the United States and training the next generation of researchers is essential for developing new strategies to prevent and improve the treatment of diabetes. This T32 program will provide closely mentored, individualized, in-depth training in an interactive and supportive, multidisciplinary environment that fosters collaboration between disciplines and strongly promotes recruitment and retention of underrepresented minorities at the University of Illinois at Chicago (UIC). The largest university in the Chicago area with annual research expenditures of approximately $335 million, and recognized as a Minority and Hispanic Serving Institution, UIC provides outstanding opportunities for training in basic, clinical, translational, and behavioral and community-based research targeting diabetes and access to cutting edge technologies. PhD, MD and trainees in other clinical disciplines will be highly selected based on demonstrated research interest, accomplishments and potential, engagement in research, and the accomplishment of milestones (identification of research mentors, preparation of a preliminary project proposal and IDP) will be required prior to joining the T32 program. Individualized training and career development plans are developed at the outset with input from multidisciplinary mentoring teams that identify appropriate goals, needs and milestones. Trainees will receive training in grant writing, research integrity and ethics and methods to enhance research reproducibility, and participate in research seminars, journal clubs and work-in-progress meetings, and, when appropriate, complete graduate level courses pertinent to their research and career goals, including degree conferring programs at the Masters level (e.g. MS in Clinical and Translational Science, Health Informatics, etc). The development and submission of an individual NRSA or other career development application will be a major goal and training experience during the T32. The progress of trainees will be closely monitored and feedback from trainees on mentoring and the research program reviewed annually. Additional oversite and guidance will be provided by Internal and External Advisory Committees. The success of this program will be evaluated based on the research and academic success of its trainees. This program is designed to provide outstanding training in research related to diabetes in a vibrant, diverse and scientifically responsible environment at a major research institution in Chicago.

NIH Research Projects · FY 2025 · 2021-05

Abstract /Summary: Streptococcus pneumoniae (pneumococcus) is a major global bacterial human pathogen, causing ~1 million deaths annually worldwide, due to pneumonia, sepsis, and meningitis. Two strategies are used to combat such infections. Antibiotics can often cure such infections, and vaccines are used to reduce the circulating populations of the most dangerous serotypes. However, both strategies are failing at an increasing rate. Antibiotic resistant strains are continually arising and spreading globally; vaccination effectiveness is also under challenge, as serotypes not targeted by current vaccine formulations are continually arising and rapidly replacing the targeted ones. The cause of these failures is transfer of multiple foreign genes into the bacteria, but the mechanisms that create the new infectious and resistant strain types are unclear. Transfer events are of two types, named as micro- and macro-recombination events. The micro events, involving dozens to several thousands of base pairs, are consistent with the known properties of gene transfer by transformation in pneumococcus. However, more significant events involve transfer of multiple blocks of tens of thousands of nucleotides, sometimes all from a single donor strain. These macro-recombination events were difficult to reconcile completely with any known mechanism of gene transfer - whether conjugation, transduction, or transformation. This project would use microfluidics to create numerous small chambers (droplets) within which attacker- target interactions can be studied and characterized for the first time at both the cellular and molecular levels, both by identifying the participant cells and by tracing all gene exchange events at full genome scale and 200-bp resolution. Medical Relevance. Most pathogenic streptococci share the mechanism of gene transfer by natural genetic transformation. Genetic transformation is an important path for genetic flexibility in pneumococcus, where it is documented as key to vaccine escape and creation and spread of new drug-resistance genes. Because Streptococcus pneumoniae is a model organism for the study of DNA uptake, this work on the mechanism that transfers unexpectedly large blocks of genes between strains or species will have broad impacts on understanding and targeting the many similar peptide regulated gene exchange systems among Gram positive bacteria that are often associated with the ability of these bacteria to cause disease.

NIH Research Projects · FY 2025 · 2021-05

Project Summary The purpose of this grant is to study a model of mental health navigation for African American and Latinx children in high poverty urban communities focused on reducing key parental attitudinal barriers to care. Reducing persistent racial and ethnic disparities in children’s mental health is a national priority and patient navigation is a highly promising approach that is rarely used in children’s mental health services. The study will examine the effectiveness of two types of navigators: paraprofessionals (PP) who have strong community knowledge, and case managers (CM) who are formally trained. The study will examine specific mechanisms of navigator effectiveness in children’s mental health and compare the two types of navigators to provide a rigorous test of the proposed mechanisms. The knowledge gained from this application may be important to reducing disparities and employing the workforce best suited to navigation in the community mental health system. Two community boards, one focused on identifying factors important to supporting navigators at the agencies, and the other focused on implications for state and federal policy, will meet annually with the goal of identifying key findings with the potential to influence local, state, and national priorities for children’s mental health.

NIH Research Projects · FY 2025 · 2021-05

Title: Mechanism of CX3CR1+ macrophage-mediated resolution of eosinophilic allergic lung inflammation Abstract: Recent studies show that tissue-resident macrophages participate in not only the initiation of inflammation but also in the resolution and prevention of local inflammation. To precisely determine the subsets of macrophages engaged in resolving lung inflammation and gain insight into their functions, we adopted new techniques of mass cytometry and single-cell RNA-seq (sc-RNA-seq) to analyze human and mouse macrophages in the lung. Our supporting data showed that alveolar macrophages (AMs) are phenotypically diverse and highly dynamic in response to allergen challenge. Based on the sc-RNA-seq data, AMs can be clustered into a few groups at a steady status. Among these groups, a subset of CX3CR1-expressing AMs (CX3CR1+ AMs) are unique in terms of their phenotype and patterns of gene expression, compared to classical resident AMs which are CX3CR1 negative. In patients with allergic asthma and a mouse model of asthma, we found that CX3CR1+ AMs are markedly increased in BAL by allergen challenge. The CX3CR1+ AMs express not only the macrophage but also eosinophil markers such as human Siglec-8. Further investigation with the CX3CR1- reporter and Epx-cre (a.k.a. Eo-cre) reporter mice reveals that CX3CR1+ macrophages engulf eosinophils at a steady state and in allergic lung inflammation. Depletion of CX3CR1+ macrophages in mouse models resulted in spontaneous tissue eosinophilia at a steady status and prolonged tissue eosinophilia in allergic lung inflammation. Based on this data, we hypothesized that the newly recruited CX3CR1+ AM subset promote the clearance of tissue eosinophils and facilitates the resolution of allergic lung inflammation. In aim 1, we will focus on the cellular dynamics of CX3CR1+ macrophages in allergic lung inflammation. Regarding the molecular mechanism of CX3CR1+ mediated-eosinophil clearance, we examined the potential ligands for CX3CR1 – CX3CL1 and CCL26. We discovered that CCL26 plays a key role in activating CX3CR1+ macrophages, whereas CX3CL1 is indispensable. Our sc-RNA-seq data revealed that CX3CR1+ AM subset is the sole source of the transcript of C1q - a key molecule for efferocytosis. In in-vitro setting, CCL26 triggers CX3CR1+ macrophages to secrete C1q in a CX3CR1 receptor-mediated manner. Furthermore, C1q and CCL26 are increased in BAL by allergen challenge in patients with allergic asthma. This data suggests CCL26 activates CX3CR1+ macrophages to facilitate efferocytosis via C1q secretion. In aim 2, we will examine the detailed mechanisms of CX3CR1+ macrophage activation through CCL26-mediated C1q secretion. Finally, we will extend the study to translational human research using the IRB-approved protocol for the segmental provocation with an allergen to evaluate the human relevance of the above proposed experiments. The proposed study is based on our strong supporting data on the new roles of CX3CR1+ macrophages in the resolution of allergic lung inflammation. This study will lead to a better understanding of the resolution process of allergic asthma.

NIH Research Projects · FY 2025 · 2021-05

ABSTRACT This application proposes to examine a novel social epigenetic mechanism for lung cancer: Reducing lung cancer by decoding the link between neighborhood exposure and biological responses (RECODE). We will examine the relationships between exposure to neighborhood stress, smoking, inflammatory responses, and epigenetic changes in protein arginine methyl transferases (PRMT6) that increase the risk of developing lung cancer. Our preliminary studies demonstrated that smoking induces increased expression of PRMT6 in the lung epithelium. We also showed that overexpression of PRMT6 triggers spontaneous lung tumors in mice. We argue that the increased overexpression of PRMT6 may explain a higher rate of lung cancer. While smoking is a key contributing factor for lung cancer, the frequency and amount of cigarette smoking are not necessarily higher among population groups with higher lung cancer rates, which suggests that other factors are responsible for lung cancer risk. Neighborhood stress exposure may be responsible for a higher rate of lung cancer. In particular, individuals living in neighborhoods with high levels of crime/violence are exposed to chronic stress, which may intensify the epigenetic changes for lung cancer. We hypothesize that exposure to neighborhood violence increases biophysical inflammatory responses, which exacerbate the path between smoking, PRMT6 overexpression, and lung cancer. To examine the proposed epigenetic mechanism of lung cancer, first, we will test the independent effect of smoking and exposure to neighborhood violence, and the interaction of the two risks on PRMT6 expression using retrospective tissue samples of lung cancer cases (Aim 2). Second, we will test the effect of exposure to violence on an inflammatory response (hair cortisol) and lung cancer screening outcomes by conducting a prospective survey and data collection (Aim 3). Finally, we will build multilevel, context-specific lung cancer risk profiles (Aim 1) that take into account not only individual behavioral risk (smoking), but neighborhood stress (exposure to violence), physiological inflammatory responses (increased cortisol), and molecular changes (PRMT6 overexpression). To develop such risk profiles, we utilize a synthetic population to establish accurate counts of all individuals within census tracts with sociodemographic, behavioral, and neighborhood risk profiles. The strength of RECODE is its innovative approach to unveiling a social epigenetic mechanism of lung cancer risk. RECODE has the potential to transform understanding of multilevel risks of lung cancer and improve the national lung cancer screening guidelines to reflect stress exposure and epigenetic changes that may increase the risk of developing lung cancer.

NIH Research Projects · FY 2025 · 2021-05

Abstract In the three major intracellular trafficking pathways–exocytosis, endocytosis and autophagy–proteins and membranes are secreted, internalized or shuttled for degradation, respectively. These pathways are regulated by the conserved Ypt/Rab GTPases that when activated by nucleotide exchangers (GEFs), recruit their effectors to membranes. These effectors are machinery components that mediate vesicular transport steps, from vesicle formation, through motility and targeting, to fusion. I have worked in the Ypt/Rab field since its inception and contributed to the formulation of principles that underlie their mode of action. These include ideas that they function in “GTPase modules”, which contain GEF/s, a GTPase, and effector/s, to organize pathway- or step-specific membrane microdomains. While mechanisms underlying Ypt/Rab function are currently known, questions regarding pathway and step coordination remain open. We propose that Ypt/Rab GTPases coordinate intracellular trafficking at three levels: Coordination of multiple pathways, integration of transport steps into whole pathways, and coordination of vesicular transport sub-steps of individual transport steps. The proposed research relies on our recent findings using yeast as a model system, and we will continue using yeast due to its smaller proteome that results in a much smaller interactome, which is important for exploring the following coordination issues: Multiple pathways coordination: I propose that Ypt/Rabs coordinate autophagy with secretion and endocytosis at two intersections. In the first, Ypt1/Rab1 is required for the beginning of secretion and autophagy in the context of two different GTPase modules. In the second intersection, merging of endocytosis and late autophagy is regulated by a shared Vps21/Rab5 GTPase module. Here, we will determine whether cells prioritize certain pathways under different environmental conditions and how such a priority is promoted. Integration of transport steps: Here, we will address two major questions: First, how do Ypts regulate the beginning of a pathway, especially when a single GTPase functions in the context of two different modules? Second, what are the specific mechanisms by which Ypts coordinate early and late steps in secretion and autophagy? Coordination of vesicular transport sub-steps: We will explore late steps of the secretory and autophagy pathways, for which members of the GTPase modules are known, and ask how effectors that function sequentially are recruited. We will use classical and molecular genetics combined with cell biology and biochemistry approaches to address these questions. An efficient and well-coordinated network of cellular trafficking pathways is important for all the systems of the human body, and even a minor defect can result in a severe disease. Ypt/Rabs in general were implicated in a spectrum of acquired and inherited diseases, and those we study were associated with cancer and neurodegeneration. Finally, we recently showed the value of yeast modeling in understanding how a conserved protein variant causes a neurodevelopmental disorder.

NIH Research Projects · FY 2025 · 2021-03

Cancers that depend on the spatial location of the disease affect all ethnicities and age groups, accounting for significant mortality and therapy-related side effects. In one instance, over 50,000 new cases of head and neck squamous carcinomas are diagnosed each year in the United States, leading to large, rich repositories of patient data. For each of these cases, oncologists need to anticipate survival, oncologic, and toxicity outcomes associated with treatment strategies in order to select a treatment which balances efficacy and toxicity. However, despite the wealth of data available, in the clinic decision support for cancer treatment is rudimentary and incorporates only a handful of patient characteristics, largely due to a lack of computational methodology and tools. We propose to construct a novel statistical and computational methodology for longitudinal and personalized treatment decisions over time, with specific application to head and neck cancer therapy planning. Simultaneous incorporation of complex factors---such as radiation dose location with respect to radiosensitive organs or patient reported side effects affecting quality of life---into treatment decisions over the course of cancer therapy requires the development of novel methodology. This methodology is revolutionary in that it is the first in the field to include both imaging and nonimaging data, while taking into account large-scale biological and clinical correlates. The approach is innovative through its leverage of big data repositories and through its unique blend of computational modeling principles from bioengineering and computer science. These methods allow us to incorporate diverse data types and model competing outcomes. From a clinical perspective, this integrative approach is novel in the field of cancer therapy. The resulting clinical decision support methodology will mark a significant advance in biomedical computing because it will be able to identify, for the first time, actionable timepoints for therapy and toxicity modification, based on a patient’s characteristics and quality of life indicators. The empirically-derived treatment decision support methodology developed in this project has the potential to directly improve the standard of care and the quality of life of surviving patients with a grave, often fatal and debilitating illness.

NIH Research Projects · FY 2026 · 2021-03

PROJECT SUMMARY/ABSTRACT A major challenge of targeting metabolism for cancer therapy is pathway redundancy, where multiple sources of critical nutrients can diminish the effects of metabolic therapies. An example of this can be found in recent attempts to target the serine synthesis pathway for cancer therapy, where the abundance of serine available to be taken up from the circulation has hampered the success of inhibitors of serine biosynthesis. This places a premium on pursuing strategies of limiting pathway redundancy if we wish to successfully target serine and other critical metabolic pathways for cancer therapy. We have taken the approach of analyzing human tumor gene expression data to identify scenarios where pathway redundancy is limited due to lineage-dependent gene expression, thereby creating potential vulnerabilities. Using this approach, we have found that the two major lineages of breast tumors—luminal and basal—express vastly different levels of PSAT1 (phosphoserine aminotransferase 1), the gene encoding the second enzyme of the serine synthesis pathway. Luminal breast cancer cells, which express extremely low levels of PSAT1, are unable to activate the serine synthesis pathway even when extracellular serine is completely absent. As a result, they are entirely dependent on exogenous serine for proliferation and survival. This is in contrast to basal breast cancer cells, which are able to synthesize serine and proliferate in the absence of extracellular serine. Mechanistically, this serine auxotrophy appears to be due to luminal-specific methylation of the PSAT1 gene. Based on this data, we have developed the hypothesis that lineage-specific epigenetic silencing of the PSAT1 gene induces serine auxotrophy in luminal breast tumors and makes them vulnerable to inhibition of serine uptake. In this proposal, we will 1) determine whether luminal breast tumors are sensitive to dietary serine starvation in vivo, 2) define the mechanism of PSAT1 suppression in luminal tumors, and 3) identify and characterize serine transporters as potential pharmacological targets of this vulnerability. While luminal breast cancer patients initially have a favorable prognosis due to the utility of endocrine therapies, over half of all patients eventually develop resistance to these therapies and undergo relapse. As a result, over half of all breast cancer fatalities are due to luminal breast cancer, making this an area of significant unmet clinical need. The experiments described in this proposal have the potential to identify new therapeutic options for patients with advanced luminal breast cancer.

NIH Research Projects · FY 2025 · 2021-02

Abstract The voltage-gated Hv1 proton channel is a member of voltage-gated ion channel family, it plays a key role in the acid extrusion from excitable and non-excitable cells and regulate pH homeostasis in a variety of cell types. The Hv1 proton channel provides charge and pH compensation during the respiratory burst of the phagocyte NADPH oxidase and controls production of reactive oxygen species in phagocytes, it mediates proton efflux at the pulmonary alveolar cell membrane and acidifies excessively alkaline airway surface liquid in the airway cells. Hv1 is found to be a sperm flagellar regulator of intracellular pH, and play a crucial role in sperm capacitation. In addition, Hv1 activity is required for acid extrusion to shape action potentials in snail neurons. Studies have shown that biophysical properties and functions of the Hv1 channel are not identical in a variety of tissues, the distinct manifestations of native voltage-gated proton channels in different cell types indicate the existence of modulatory partners modulating the Hv1 proton channel's function. We recently identified a family of transmembrane protein as the Hv1 channel interactor. We discovered that the presence of the transmembrane protein alters the Hv1 channel's activity and modulates the channel's voltage-dependent gating. The goal of this proposal is designed to determine the effects of the transmembrane protein on the Hv1 channel physiology, biophysics, and pharmacology. The proposed research will establish the physiological relevance of Hv1 channel regulation by transmembrane modulators and shed light on novel voltage-gated ion channel regulatory mechanisms.

NIH Research Projects · FY 2025 · 2021-02

ABSTRACT We have discovered a new mechanism for inflammation on the ocular surface. First, we discovered the presence of neutrophil extracellular traps (NETs) on the ocular surface of dry eye disease (DED) patients and subsequently, the presence of citrullinated proteins and anti- citrullinated protein antibodies (ACPAs). ACPAs not only cause ocular surface disease, but also stimulate formation of NETs, thus creating a self-perpetuating cycle of chronic inflammation on the ocular surface. We present an innovative pathophysiological concept: DED is characterized by breach of self-tolerance towards citrullinated antigens with generation of autoantibodies (ACPAs) and NETs could represent a source of citrullinated antigens fueling the ACPA autoimmune response over the ocular surface. We performed a first-in-human pilot clinical trial and showed that ocular surface immune globulin (OSIG) eye drops, formulated from pooled human immune globulin products (IVIG), were safe and efficacious in treating DED patients. Our findings shift the current paradigm that focuses on T-cell mediated inflammation as central to the pathophysiology of DED to also include autoimmune inflammation that is driven by post- translational modifications in self-proteins (citrullination) and autoantibodies (ACPAs). The purpose of this R24 application is to produce preclinical data that supports a commercial Investigational New Drug (IND) application to the U.S. Food and Drug Administration (FDA). The longer-term goal is for testing the efficacy of OSIG eye drops in clinical trials and positioning OSIG eye drops as a new topical biologic immunotherapy for DED patients. Therefore, in Aim A, we seek to develop an optimum OSIG eye drop formulation using Quality by Design (QbD) principles, test critical quality attributes (CQAs) and GMP manufacture clinical supplies. In order to use OSIG ophthalmic formulation in human, an FDA IND is a prerequisite, therefore, in Aim B, we propose to perform pre-clinical in vitro and in vivo toxicology and efficacy studies using the OSIG ophthalmic formulation to meet regulatory requirements for IND studies. In Aim C, we propose to perform a clinical study to identify DED subtypes, clinical presentations and patient characteristics that are most associated with ACPAs or NETs, hence most likely to show therapeutic benefit with OSIG therapy. To successfully achieve these three Aims, we have established a highly qualified, multi-disciplinary team that is experienced in ophthalmic drug development, and FDA processes and regulations. If the aims of this grant proposal are successfully achieved, we will be one-step closer to introducing the first immunotherapy for ocular surface diseases into clinical practice.

NIH Research Projects · FY 2025 · 2021-01

ABSTRACT. Colorectal cancer (CRC) is associated with multiple risk factors including, obesity, low fiber diets, and diets high in animal protein and saturated fat (SFat). African Americans (AAs) have a higher prevalence of these risk factors and they have the highest incidence of CRC and related mortality. These multiple risk factors are also linked to higher circulating and fecal bile acids (BA) and a shift in BA amino acid conjugation from glycine to taurine. These BA-related changes can alter the composition, structure, and metabolic activity of the gut microbiota, fostering conditions for gut bacteria to expand and metabolize taurine-conjugated BAs to genotoxic hydrogen sulfide (H2S) and the tumor promoter, deoxycholic acid (DCA); a colonic milieu conducive to the formation of CRC. We have shown that the abundance of H2S-producing bacteria is significantly higher in the colon of AAs compared to non-Hispanic whites and is a defining feature among AA CRC cases implicating these bacteria as contributors to CRC development in a race-dependent manner. Moreover, the microbial difference is associated with higher intake of SFat and animal protein in AAs, providing a pivotal intervention target. We hypothesize that targeting the BA-gut microbiome axis to suppress abundance, growth and metabolic activity of H2S and DCA producing bacteria through diet and weight loss (WL) may reduce CRC risk, especially among AAs. A Mediterranean Diet (MedDiet), a largely plant-based dietary pattern, is relevant to CRC prevention and microbial production of anti-cancer metabolites in observational studies. A MedDiet can shift BA metabolism as shown in primates and when combined with calorie restriction, shows superior adherence and weight control in humans, given its palatability. To date, no studies have tested in an RCT the effects of a MedDiet alone (Med- A), WL through lifestyle intervention (WL-A) or a calorie-restricted MedDiet for WL (WL-Med) on the BA-gut microbiome axis and its relevance to CRC prevention among AAs. Our multidisciplinary team combining expertise in psychology, nutrition, microbiology, molecular cell biology, computational biology, medicine and biostatistics, propose to conduct a four-arm RCT in which 200 obese AAs, 45-75 years old complete one of the following 8-month interventions: Med-A, weight stable; WL-A, calorie restriction with no diet pattern change; WL- Med; or Control. We will use samples and data collected at baseline, mid-study (month-4) and post-intervention to compare the effects of the interventions on 1) Concentration and composition of circulating and fecal BAs; 2) Gut microbiota and metabolic function; and 3) Gene expression profiles of exfoliated intestinal epithelial cells. Our approach is strong given our multidisciplinary team, use of evidence-based lifestyle interventions, and sophisticated –omics analyses to examine crosstalk between diet/WL, gut microbiome, and host intestinal physiology. If successful, this study could have profound public health impact on CRC risk among AAs and other high-risk populations, that would translate into timely dissemination opportunities.

NIH Research Projects · FY 2025 · 2021-01

Project Summary The ability to heal is essential for human health. The remarkable wound healing capacity of humans (and indeed of all animals) is a complicated process. Genomic studies suggest that more than 5000 mRNA transcripts change expression patterns during wound healing, and more than a dozen cell types participate. Despite the large number of cells and molecules that are involved, many aspects of the complexity of wound healing are not well understood. Systems approaches have established the breadth of genes involved in healing, and have compared gene profiles in different types of wounds. Yet many questions about the networks and interactions within wounds are still unanswered. The research proposed here couples novel quantitative approaches with basic biologic research to examine several questions related to the complexity and diversity of the events that compose tissue repair. The research plan addresses three separate but complementary questions. Question 1 asks - What are the regulatory pathways that underlie the differential, site specific healing that is seen in skin versus oral mucosa? Studies by us and others have shown that wounds in the oral mucosa exhibit faster re-epithelialization, reduced inflammation, a better-developed angiogenic response, and less scar formation as compared to skin. Oral mucosal and skin wounds also have distinctive transcriptomes. The central concept underlying Question 1 is that key transcription factors are responsible for the differential healing seen in these two tissues. The research approach uses state-of-the-art algorithms and in vivo experiments to discover and validate the transcription factors (and their networks) that distinguish oral mucosal and skin healing. Question 2 asks - What is the level of redundancy in healing wounds? This question explores the robustness of healing by assessing molecular redundancy. Redundancy has been posited to exist in wounds as a “fail-safe” mechanism, insuring that wound healing proceeds even if some key elements are functionally inactivated. The underlying concept for Question 2 is that significant gene compensation occurs in wounds when specific genes are deleted. The approach uses genetically deficient (“knockout”) mice to examine the extent of redundancy in healing wounds. Question 3 asks- Can quantitative models be used to predict wound healing outcomes? Our ongoing collaboration utilizes a novel computational modeling framework called the dynamic cellular finite-element model (DyCelFEM) to develop a model of epithelial repair that is predictive of healing responses. The research plan extends the model to include additional features of wound healing, such as angiogenesis. When completed, the model can be used to test the effect of perturbations of single or multiple factors on healing outcomes. This advanced model will be a powerful tool that can contribute to our understanding of both the pathophysiology of chronic wounds and the development of therapeutics. Taken together, the three elements of the research plan address how wound healing is governed at a network level, and will uncover critical features that regulate the ability to heal.

NIH Research Projects · FY 2025 · 2021-01

PROJECT SUMMARY/ ABSTRACT Mesenteritis is an inflammatory disorder of mesenteric tissues. It includes mesenteric lipodystrophy (mesenteric fat necrosis), mesenteric panniculitis (chronic mesenteric fibrosis) and mesenteric lymphadentis (mesenteric lymph node inflammation). It is diagnosed by an abdominal computed tomography (CT) scan; however, the cause of mesenteritis remains unknown. Emerging clinical evidence shows that mesenteritis is associated with gut inflammation such as IBD and Chron's Disease. As innate myeloid cells, macrophages are distributed throughout the whole organism and they play crucial roles in mediating tissue inflammation. Based on our supporting data, we identified macrophage populations in serous membrane of gut mesentery (termed membrane-associated macrophages). Thus, this research proposal seeks to address fundamental questions regarding the tissue specific role of membrane-associated macrophages in steady state and during mesenteritis induced gut inflammation. These include how membrane- associated macrophages are reprogrammed by local niches and gut inflammation. To better describe the role of these macrophages and its mechanism, we performed genetic studies of membrane-associated macrophages that provided insight into their roles in steady state and gut inflammation. These preliminary data results have led us to pursue to the following specific aims: (1) Determine distinct homeostatic functions of membrane-associated macrophages and local signals that shape their tissue specification; (2) Determine the role of membrane- associated macrophages during mesenteritis induced by gut inflammation. We will perform lineage tracing, cell ablation, transcriptomic analysis of membrane-associated macrophages and in vivo live imaging to understand the role and dynamic interactions of macrophages with local environment in steady state and during mesenteritis induced by gut inflammation. Ultimately, we hope our studies will lead to the discovery of therapeutic targets to prevent mesenteritis as well as gut inflammation.

NIH Research Projects · FY 2024 · 2020-09

Summary The Mesenchymal Stromal Cell (MSC) Secretome Study is designed to assess the safety of allogeneic bone marrow-derived MSC secreted factors on the ocular surface, and also obtain a preliminary observation on 1) epithelial healing; 2) development of scarring; and 3) final visual acuity; with the objective of improving clinical outcomes in significant non-healing corneal epithelial disease. To achieve these goals, the MSC Secretome Study will include a Phase 1 safety and maximum tolerated dose study (Specific Aim #1) and a Phase 2a double- masked vehicle-controlled study to assess preliminary evidence of efficacy while identifying patient subgroups most likely to benefit from the treatment (Specific Aim#2).

NIH Research Projects · FY 2024 · 2020-09

This project’s long-term goal is to elucidate how anti-VEGF (i.e. acute neutralization of VEGF (vascular endothelial growth factor)) stops leakage of blood vessels that are chronically exposed to excess VEGF. Anti- VEGF’s ability to curb blood vessel permeability has improved and/or sustained the quality of life of patients afflicted with pervasive, incurable diseases such as proliferative diabetic retinopathy (PDR), diabetic macular edema (DME) and neovascular age-related macular degeneration (nAMD). Such clinical observations demonstrate anti-VEGF’s efficacy, and beg the question of how anti-VEGF calms pathological blood vessels. We posit that chronically elevated VEGF alters gene expression in the endothelium of blood vessels in a way that relaxes the endothelial barrier, and that anti-VEGF reverses such changes. Aim 1. Test the hypothesis that anti-VEGF acts transcriptionally to close the barrier. Our preliminary data reveal that prolonged exposure to VEGF changes the expression of many genes, including those that encode known regulators of the endothelial cell barrier. These discoveries support our work hypothesis that anti-VEGF acts transcriptionally to close the barrier. This hypothesis will be tested as follows. The initial, RNAseq-based phase will compare the gene expression profile in cells that have or have not been treated with anti-VEGF and thereby identify anti-VEGF differentially expressed genes (DEGs). Candidates DEGs will be prioritized based on their known function, and then their contribution to VEGF-/anti- VEGF-mediated control of barrier function will be determined. We will identify genes that anti-VEGF depends on to close the endothelial cell barrier that has been breached with VEGF. Aim 2. Investigate the mechanism of action of anti-VEGF on patient-derived retinal endothelial cells. Aim 1 will be done with primary human retinal endothelial cells (HRECs) from a healthy adult donor. Aim 2 will be a repeat of aim 1, except using PRECs, retinal endothelial cells isolated from pathological blood vessels that develop in patients with PDR. This is the target cell type of anti-VEGF therapy. Determining the mechanism of action of anti-VEGF in these cells will provide clinically relevant information. Aim 3. Determine how anti-VEGF calms pathological blood vessels in patients. In aim 3 we will learn how anti-VEGF acts in patients by comparing the gene expression profile in freshly isolated endothelium from pathological blood vessels of treatment naïve and anti-VEGF-treated patients. In addition, we will compare the results from all 3 aims to determine which of the anti-VEGF-mediated effects that occur in patients are faithfully modeled by anti-VEGF treatment of cultured PRECs (aim 2) or HRECs (aim 1). This project will unveil the molecular mediators of anti-VEGF’s therapeutic benefit. Such information will remove current roadblocks to developing biomarkers and alternatives to anti-VEGF, which are needed to address the needs of patients afflicted with a variety of blinding conditions such as DME, PDR and nAMD.

NIH Research Projects · FY 2024 · 2020-09

Nearly 100 people die every day from a prescription opioid overdose in the United States (US). Over-reliance on opioids for these with chronic pain is one of the factors that led to this crisis. Pain, both acute and chronic, that is so severe that it requires opioids to attempt to keep to tolerable levels, is a constant companion to the 100,000 people in the United States, mostly of African or Hispanic background, and millions more worldwide living with sickle cell disease (SCD). Pain is SCD’s hallmark symptom, and the leading cause for almost 200,000 annual emergency department (ED) admissions, and is the leading cause of hospitalization, with estimated annual health care costs in the US of $2.4 billion. We will conduct a hybrid type 1 effectiveness implementation trial to assess the effectiveness of acupuncture and guided relaxation on 360 people with SCD while observing and gathering information on implementation in three health systems: University of Illinois Hospital & Health Sciences System, University of Florida Health, and Duke University Health Systems. Each serves a large population with SCD, uses EPIC as their electronic health record, and has a Clinical and Translational Science Award (CTSA), which will help speed the translation of discovery into improved patient care. UG3 1-year Planning Phase: Year 1 comprises milestone-driven planning to prepare the three health systems for the subsequent pragmatic clinical trial (UH3). During the UH3 Implementation Phase, our 3-arm, 3- site randomized controlled trial will follow a quantitative SMART design. A pragmatic trial that evaluates adaptive interventions where our guided relaxation and acupuncture interventions responds to patients’ characteristics and evolving pain status. We rely on the Consolidated Framework for Implementation Research (CFIR) to plan, execute, and evaluate associated implementation processes. The use of complementary and integrative (CIH) therapies by those with SCD to reduce pain, opioid use, and enable themselves to better cope with their pain is well known, but there are few studies that evaluate the effectiveness of these therapies, and none that also evaluates the implementation across multiple health care systems and patient populations as this study will.

NIH Research Projects · FY 2025 · 2020-09

The Type 1 Diabetes in Acute Pancreatitis Consortium (T1DAPC) was established by the National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK) as a multicenter and multidisciplinary program to investigate the incidence, etiology, and pathophysiology of diabetes following acute pancreatitis (AP). The Chicago Clinical Center (CCC) of the T1DAPC is an interdisciplinary clinical-translational program led by the University of Illinois Chicago (UIC) and includes Northwestern University as a satellite site. The CCC is well-integrated into the T1DAPC and has made significant contributions to its objectives. It provides considerable leadership in the organization, participates in Committees and Working Groups, and contributes to the development and execution of ancillary studies and publications. Specifically, the CCC has been the leading center in enrollment of AP patients into the Diabetes Related to Acute Pancreatitis and its Mechanisms (DREAM) study, a prospective longitudinal observational clinical study of the T1DAPC. Our proposal leverages our large AP cohort and our multidisciplinary team with complementary expertise in pancreatology, diabetes, gut microbiome, metabolome, artificial intelligence (AI), and diet, with a proven track record of collaboration. We propose the following two specific aims to meet the goals of RFA-DK-25-017 in the next five years of funding. Aim 1. Continue to contribute to the assembly of the DREAM cohort, longitudinal follow-up of its study participants, and completion of its ancillary studies. Aim 2. Define the mechanisms of AP-driven diabetes by proposing the following three ancillary studies: Aim 2.1. Determine the association of gut microbiome and metabolome with the development of AP-driven diabetes, Aim 2.2. Develop novel, precise, reliable, and trustworthy artificial intelligence (AI) algorithms for early prediction of AP-driven diabetes, and Aim 2.3. Define the association of social vulnerability with the development of AP-driven diabetes. We are well-positioned to execute these specific aims with our interdisciplinary expertise, well-established clinical and research infrastructure, extremely productive patient enrollment profile, streamlined sample processing, and transfer. In addition, we will continue to contribute to the success of the T1DAPC with our existing leadership roles, active participation in T1DAPC Committees and Working Groups, and strong collaborations within the T1DAPC.

NIH Research Projects · FY 2024 · 2020-09

Sex steroids, including estrogens and androgens, play an important role in regulating energy homeostasis. Brain sex steroid signaling is required for normal body weight maintenance. We previously showed that estrogen receptor α (ERα) neurons in the medial amygdala (MeA) stimulate physical activity and energy expenditure to decrease body weight in both males and females. It suggests that the estrogen/ERαMeA circuit constitutes part of a previously undefined brain metabolic signaling in both males and females. Interestingly, the MeA has high levels of other three key components of testosterone/estrogen signaling, including an essential enzyme for estrogen synthesis (aromatase; Aro), and key mediating receptors for testosterone/estrogen signaling (androgen receptor, estrogen receptor α and β; AR, ERα and ERβ). These data raise the possibility that circulating testosterone directly binds to AR or is aromatized by Aro to estradiol, which then binds to ERα or ERβ to exert metabolic functions. We hypothesize that the neurosteroid testosterone/estrogen signaling pathways in the MeA interact to maintain normal energy homeostasis. To test this, three mutant mice will be generated to have each of these three components deleted specifically in the MeA neurons, respectively. These mouse strains (both males and females) will be used to determine the physiological roles of these three components in maintaining energy homeostasis in different sexes. The functional interactions between these components and the sex hormones will also be examined. Results from these studies will advance our current understanding of body weight control and the development of obesity in general. Further, our studies may narrow down the brain regions and hormone/receptors that are critical for the regulation of energy balance, which may serve as targets for the development of new anti-obesity strategies.