University Of Nebraska Medical Center

NIH Research Projects · FY 2025 · 2024-09

Project Summary This application describes the research plan and program of study leading to a PhD as part of a combined MD- PhD Program at the University of Nebraska Medical Center and the Boys Town National Research Hospital. The central goal of this research plan is to identify how dehydroepiandrosterone (DHEA), testosterone, and estradiol modulate the development of cognitive systems serving selective attention and working memory processing. Briefly, the pubertal transition period is marked by significant increases in these hormones and substantial developmental advancements in higher-order cognitive functions, including working memory, selective attention, and others. However, on the other hand, the incidence of psychiatric disease also increases during puberty, and is accompanied by greater sex differences in disease prevalence. It has been suggested that this emergence of psychiatric illness during puberty might be associated with the drastic hormonal changes that mark this developmental stage, but the net impact of these hormonal changes on brain function and cognition is poorly understood. This F30 fellowship proposal will examine the sex and region-specific effects of DHEA, testosterone, and estradiol on working memory and selective attention processing using an advanced dynamic functional mapping approach based on magnetoencephalography (MEG), and includes a rigorous training and career development plan for a future physician-scientist in the areas of pediatric psychiatry and developmental cognitive neuro- science. The specific aims are (1) to examine the unique effects of pubertal hormones in predicting performance on selective attention and working memory tasks and (2) to identify the relationship between sex steroid hormone levels, neural oscillatory dynamics, and behavior during attention and working memory tasks. The trainee is a minority MD/PhD student in his 2nd year of PhD study who has already distinguished himself through two first- authored manuscripts in top journals, participation and presentation in scientific conferences, and a strong history of academic achievement. In sum, this application for an F30 fellowship will use MEG, advanced source reconstruction methods, neural oscillatory analyses, and hair-sampled hormone analysis to delineate the relationship between pubertal hormone levels, brain dynamics, and cognitive development. The program of study is designed to support this research plan through rigorous training by an internationally known expert in MEG and dynamic brain mapping who has a strong record of mentoring trainees, as well as an expert collaborator in pubertal hormone analysis and interpretation, with the primary goals of developing and expanding the research and clinical skills of the applicant to produce a physician-scientist who is well equipped and prepared for the next stage of his career.

NIH Research Projects · FY 2024 · 2024-09

Scientific Abstract As the leading cause of cancer death in the United States, lung cancer accounts for about 20% of all cancer deaths. While there are two major types of lung cancer (i.e., 80%~85% for non-small cell lung cancer (NSCLC) and 10%~15% for small cell lung cancer (SCLC)), each type of lung cancer has multiple distinct subtypes characterized by morphological, molecular, and genetic alterations. Identifying lung cancer subtypes can facilitate downstream risk stratification and tailored treatment design. While various conventional methods like morphological analysis, computed tomography (CT) and imaging techniques, cytogenetic analysis, immunophenotyping, or molecular profiling have been used for lung cancer subtype identification, they are usually costly, time-consuming, labor-intensive, and sometimes inaccurate. Recent progress has witnessed the application of next generation sequencing (NGS) for identifying lung cancer subtypes, but they are limited to bulk NGS data, or single omics data only. With tons of omics data being generated within and beyond the Common Fund data sets (e.g., GTEx and HuBMAP), we hypothesize that integration of single-cell and bulk multi-omics data including genomics, transcriptomics, and epigenetics data will significantly facilitate subtype-specific biomarker discovery and boost the accuracy of lung cancer subtype identification. To address these concerns, we propose to develop an integrated machine learning (ML) framework for accurate and cost-effective lung cancer subtype identification by combining single-cell and bulk multi-omics data within and beyond Common Fund data sets. To achieve this, two specific aims are undertaken. Aim 1, to establish a gene- signature-transfer ML model that leverages large-scale bulk and single-cell transcriptomics data within and beyond Common Fund data sets for lung cancer subtype identification. Besides identifying well-annotated lung cancer subtypes, we will also explore novel lung cancer subtypes by detecting rare cell types from large-scale single cell data, from which cluster-specific and rare-cell-type specific gene signatures can be transferred to the bulk transcriptomics data for improving performance of lung cancer subtype identification. Aim 2, to develop a multi-omics integration framework to systematically combine single-cell and bulk multi-omics data (including genomics, transcriptomics, epigenomics) to further boost lung cancer subtype identification. Our model is flexible to tackle cases when only partial or incomplete multi-omics data are available for new patients. We believe successful completion of this study will have direct impacts on improving downstream lung cancer risk stratification, facilitating diagnosis and prognosis, and optimizing treatment selection. We also expect that our proposed framework in this study can be customized and extensible to identifying subtypes of other types of cancer.

NIH Research Projects · FY 2025 · 2024-09

Abstract Temporomandibular Joint (TMJ) degeneration is a painful and debilitating disease of the synovial joint, affecting over 20 million people in the United States. Obesity, advancing age and mechanical loading are greatest risk factor for many diseases including TMJ degeneration. TMJ degeneration significantly impair the quality of life by causing acute and chronic pain, thus making this disease a global health issue and a financial burden of epidemic proportion. The incidence of the TMJ degeneration is expected to rise substantially as the prevalence of obesity has risen dramatically over the last two-decades. As there is no effective treatment for the TMJ degeneration in an obese and aged individual, there is an unmet clinical need for an effective approach to treat TMJ degeneration. The current proposal seeks to address this unmet clinical challenge using a highly innovative approach of targeting p21 high cells both genetically and therapeutically for the treatment of TMJ degeneration. Our overarching hypothesis is that p21 high cells plays a central role in obesity, age-related and mechanical loading induced TMJ degeneration and targeted elimination of the p21 high cells may alleviate TMJ degeneration. To test this hypothesis, we will define: (1) the role of p21 high cells in osteochondral tissue in obesity and aging induced TMJ degeneration. Using a novel inducible p21-cre ERT2 X td-Tomato mice we will examine the effects of p21 high cells on the homeostasis of the osteochondral tissues of the TMJ in obesity and aging. (2) the pathophysiological mechanism by which p21high cells causes TMJ degeneration. Using the novel mice model (p21-cre ERT2 X td-Tomato X Rela flox/flox), we will examine the effect NF-κB pathway on the senescent cells in TMJ degeneration. 3) To define the role of p21high cells in traumatic TMJ injury and repair. p21-cre ERT2 X td-Tomato mice will be subjected to traumatic injury to TMJ and we will genetically clear the p21 high cells after traumatic injury to see whether we can alleviate TMJ-OA or repair the damaged cartilage. A combination of mechanical, immunohistochemical, molecular biology and histomorphometric techniques coupled with novel genetic mice models will be used to study the proposed specific aims. The proposed project has the immense potential to reveal new regulatory pathways that controls homeostasis of the osteochondral tissues of the TMJ in an obese and aged individual and to open new insight on understanding the disease mechanism and developing therapeutic interventions.

NIH Research Projects · FY 2025 · 2024-09

PROJECT SUMMARY The CDC estimates that 10% of women between the ages of 15 to 19 test positive for Chlamydia trachomatis and 50-70% of these infections are asymptomatic. This increases the risk of widespread transmission and untreated infections, which can lead to pelvic inflammatory disease or infertility in a significant portion of women of childbearing age. Hence, there is a great need to identify strategies to reduce/prevent transmission and limit infections to the primary site of inoculation. Within the host cell, infectious elementary bodies (EBs) differentiate into non-infectious reticulate bodies (RBs) in a pathogen-specified organelle termed the chlamydial inclusion. During later stages of chlamydial development that precede egress, RBs will undergo secondary differentiation to form new EBs. The success of Chlamydia as a pathogen is owed to its obligate intracellular lifestyle and successful completion of its developmental cycle within the inclusion. The size of the inclusion enlarges over the course of chlamydial development to accommodate the increased number of organisms and at later stages, the inclusion can occlude much of the cytoplasm and dislocate organelles to the periphery of the host cell. Yet, the host cell remains relatively unstressed, which allows Chlamydia to promote a “silent” infection that does not attract unwanted attention from the host immune system. We hypothesize that one mechanism of maintaining a silent infection is the multiple pathways from which Chlamydia draw nutrients. Early in infection, the inclusion will intercept free amino acids from the lysosome and form membrane contact sites with the endoplasmic reticulum. During mid- to late- cycle, the inclusion intercepts post-Golgi exocytic vesicles to obtain sphingomyelin and cholesterol and recruits transferrin to the periphery of the inclusion via the slow-transferrin recycling pathway. A previous study in the laboratory demonstrated that the clathrin adaptor molecule, PICALM, localizes to the chlamydial inclusion. PICALM plays a central role in maintaining cholesterol homeostasis and regulates transferrin recycling. siRNA knockdown of PICALM in chlamydial infected cells increased lipid trafficking and transferrin trafficking to the inclusion. These data suggest that PICALM may function during chlamydial infection to limit host cell stress by maintaining a balance in nutrient trafficking within the cell. Current data in the field have linked PICALM to managing trafficking in the trans- Golgi, as part of its role in recycling pathways. Our data also indicate that PICALM may function in part to maintain Golgi structure and play a role in intra-Golgi trafficking—a previously undescribed finding. For this current proposal, we hypothesize that Chlamydia manipulates PICALM activity in the Golgi and slow-transferrin recycling pathways to support host cell homeostasis. A better understanding of how Chlamydia manipulates these pathways may be useful towards improving efficiency of delivery of novel anti-chlamydial compounds and/or defining novel mechanisms of previously undefined intersections of subcellular pathways.

NIH Research Projects · FY 2025 · 2024-09

The goal of the application is to define the interaction among superoxide dismutase 1 (SOD1), adenosine monophosphate-activated protein kinase (AMPK), and catalase in modulating liver injury induced by ethanol exposure. Additionally, there is a strong link between alcohol-associated liver disease (ALD) and adipose tissue dysfunction. Although SOD1 is known to inhibit the progression of ALD, very little is known regarding mechanisms involved in mediating the hepatoprotective effects of SOD1. Thus, there is an urgent need to define mechanisms by which SOD1 modulates ALD and to develop novel therapies to ameliorate this alcohol-related morbidity. Central to our hypothesis is that SOD1-mediated redox regulation of AMPK and catalase inhibits exosome biogenesis and/or promotes ethanol clearance, thereby attenuating ALD, and that a novel mannose receptor-targeted SOD1 nanoparticles (Man-Nano), is an effective agent in attenuating ALD. The studies will employ genetic, molecular, and pharmacological approaches to examine the interplay among SOD1, AMPK signaling, and catalase in modulating ALD and to demonstrate the effectiveness of Man-Nano in ameliorating ALD. We hypothesize that SOD1 increases intracellular H2O2 thereby increasing AMPK/catalase activities in hepatocytes and/or adipocytes, which contribute to the protective effects of SOD1 against ALD. We will test this hypothesis by manipulating levels of SOD1 in hepatocytes and adipocytes as presented in the following three aims. In the first aim, we will use hepatocyte-specific SOD1 knock-out mice to determine the role and mechanisms of hepatocyte SOD1 in modulating ethanol-induced liver injury. Studies in Aim 2 will employ adipocyte-specific SOD1 knock-out mice to determine the role and mechanisms of adipocyte SOD1 in modulating ethanol-induced adipose tissue and liver injury. In our last aim, we will determine the efficacy and mechanism of action of Man-Nano in ameliorating ethanol-induced liver and adipose tissue injury. Overall, this project will provide critical information regarding the novel mechanism(s) by which SOD1 attenuates ALD. Completion of this project will also yield insight into the therapeutic impact of a novel antioxidant-based therapy (i.e. Man-Nano) in ameliorating ethanol-induced organ injury.

NIH Research Projects · FY 2025 · 2024-09

No change from original submission

NIH Research Projects · FY 2025 · 2024-09

Abstract Excessive oxidative stress has been shown to be a potent pathophysiological mediator in multiple disease states. Cardiac and peripheral sympathetic nerve activity is increased in chronic heart failure, in part, due to augmented neural inflammation and oxidative stress. Importantly, several major antioxidant enzymes and molecules are decreased in the cytosol and mitochondria of pre-sympathetic neurons in chronic heart failure. A major regulator of these antioxidant molecules is the transcription factor, Nrf2. Previous studies from our laboratories have shown a reduction in Nrf2 in heart failure. Genetic knock down of Nrf2 in the rostral ventrolateral medulla of normal mice results in significant sympatho-excitation and hypertension. On the other hand, overexpression of Nrf2 results in sympatho-inhibition in mice with chronic heart failure along with a reduction in oxidative stress and an increase in several antioxidant enzymes. In order to determine if this mechanism plays a role in cardiac arrhythmogenesis in the post myocardial infarction (MI) chronic heart failure model we propose here to evaluate the role of Nrf2 in the stellate ganglia, a source of major sympathetic outflow to the heart. Three specific aims are proposed. Specific Aim 1: We will determine the time-course changes in neural inflammation-Oxidative stress-Nrf2 signaling in the SG post MI. We will evaluate message and protein for the key molecules involved in maintaining redox homeostasis in the SG, namely the Nrf2/Keap1 system and its target antioxidant enzymes, including NQO1, HO1, Catalase, SOD and glutathione peroxidase. We will also assess neural inflammation in the SG by measuring macrophage infiltration and pro-inflammatory cytokines. Correlations will be made between levels of Nrf2 and cardiac sympathetic tone across a wide range of cardiac function. Specific Aim 2: We will determine if selective knockdown and overexpression of Nrf2 in the SG alters cardiac sympathetic tone in normal and CHF animals by altering ganglionic neuronal excitability. Using Nrf2 and Keap1 floxed mice we will evaluate antioxidant enzyme expression, redox homeostasis, inflammation and arrhythmia incidence. These molecular alterations induced by Nrf2 will be evident in SG cell action potential generation. Specific Aim 3: We will determine if pharmaceutic manipulation of Nrf2 signaling in the SG by local SG delivery of micelle conjugated dynamic hydrogels with encapsulation of two Nrf2 activators (i.e., Curcumin and sulforaphane) reduces arrhythmogenesis and improves cardiac function in the post-MI rats. We hypothesize that the redox status and electrophysiological activity of SG neurons in CHF rats will be partially restored following treatment with Nrf2 activators. Furthermore, these effects will be abolished or attenuated in mice with stellate ganglia Nrf2 deficiency. We believe that this proposed research will lay a solid scientific foundation for developing a new therapies for the patients with CHF.

NIH Research Projects · FY 2024 · 2024-09

Compound heterozygous mutations of the DHCR7 gene give rise to a developmental disorder known as Smith- Lemli-Opitz syndrome (SLOS). The loss of DHCR7 function in these patients disrupts the cholesterol biosynthesis pathways, resulting in two hallmarks of the disorder: abnormally low cholesterol/desmosterol levels and high concentrations of the immediate precursor 7-DHC. 7-DHC is the most reactive lipid known to date, and 7-DHC derived oxysterols are toxic for both neurons and glia. The few current treatments for SLOS are symptomatic, and largely inefficient. We are focusing on finding a SLOS treatment by which 7-DHC levels can be reduced and toxicity can be counteracted while conserving residual cholesterol biosynthesis. Our high throughput screening of 727 compounds with a history of use in human clinical trials revealed that 40 of these compounds decreased 7-DHC formation in Dhcr7-deficient N2a cells. Following up on initial findings in vitro and in vivo we found that three compounds were particularly effective in reducing toxic 7-DHC levels – ziprasidone (ZIP), valproic acid (VPA) and hydroxyzine (HYZ) – both in human dermal fibroblasts of SLOS patients and in SLOS mouse models. These three compounds act at different sites of post-lanosterol biosynthesis. Our central hypothesis is that postnatal treatment with ZIP, VPA and/or HYZ will improve SLOS sterol profile, improving the neurochemical disruptions seen in a SLOS transgenic mouse model, and establishing a rational basis for the therapeutic use of one of these medications in SLOS patients. We also propose that the beneficial effects of ZIP, VPA and/or HYZ will be summative or synergistic when combined with a Vitamin E (VIT-E) rich diet. Finally, in an exploratory high-throughput screening we will attempt to identify additional, natural compounds that could also counteract the rise and effects of 7-DHC without suppressing residual sterol biosynthesis. We anticipate that the biological findings we obtain will provide a framework for follow-up clinical trials on SLOS patients. Furthermore, these medications could also be beneficial for treatment of other inborn errors of sterol metabolism.

NIH Research Projects · FY 2025 · 2024-09

Project Summary Youth electronic cigarette (e-cigarette) use is a public health concern. Many youths are eager to quit vaping, and our prior study estimated that 67.3% of adolescent vapers reported past-year quit attempts, with an average of 5.3 times past-year quit attempts. However, the overall success of vaping cessation remains low, and a majority of adolescent vapers had unassisted quit attempts. Meanwhile, current e-cigarette use is significantly more prevalent among high school than middle school students, and over 90% of frequent adolescent vapers are those aged 15-18 years old. It is imperative to develop age-appropriate vaping cessation interventions with a focus on older adolescents to address their specific needs, thus curbing the high vaping prevalence among this vulnerable population. The family environment is essential for youth growth, and parental support has long been documented as a critical protective factor for youth substance use. In 2022, nearly 1 in 6 adolescent vapers seeking assistance for quitting had already sought advice from their parents or caregivers, and more adolescent vapers had asked for advice from their parents than using a mobile app or text messaging program. Peer coach has been a successful strategy in tobacco and other substance cessation programs, and connecting parents of current adolescent vapers with the community of parents who have successfully helped their kids quit vaping can be an effective intervention to improve the intervention fidelity and effectiveness. Grounded on the Social Influence model and the Social Cognitive Theory, this feasibility study aims to develop and test a Parents- helping-Parents for youth Vaping Cessation (PhP-VX) program by providing parent-facilitated youth vaping cessation intervention. In Aim 1, our multidisciplinary team will partner with Parents Against Vaping e-cigarettes (PAVe), a national advocacy and education organization with nearly 14,000 race/ethnic diverse parent supporters, to advance the translation of epidemiological research on youth vaping and inputs from an advisory board (i.e., parents and adolescents) into a multifaceted vaping intervention that provides online training sessions and weekly parent-to-parent support. In Aim 2, We will conduct a feasibility pilot test with 100 dyads recruited through pediatric EHR systems, with half receiving the “PhP-VX” program and the other half in the control group for 3 months, with assessments conducted at baseline, 3-month, and 6-month follow-up. In Aim 3, we will explore the effectiveness of the intervention with saliva cotinine-validated past 7-day point prevalence abstinence as the primary outcome. This study is novel for using EHR to identify and enroll adolescent e-cigarette users and incorporating PhP peer support into the youth vaping cessation program. This feasibility study is highly significant in addressing youth vaping cessation in partnership with a robust community-based organization. The success of this R34 study will prepare us for a future R01 pragmatic trial to test the efficacy of PhP-VX through EHR and parent support in multiple pediatric clinics. 1

NIH Research Projects · FY 2025 · 2024-09

Abstract Due to the widespread use of antiretroviral therapy, HIV replication is effectively controlled in infected individuals. However, viral reservoirs form early post-infection, posing a persistent challenge to complete virus eradication. It has been suggested that sex differences may influence the establishment of HIV reservoirs, with variations between women and men attributed to socio-economic and biological factors, a phenomenon further compounded by opioid use. Established knowledge indicates that women are more susceptible to HIV acquisition than men due to robust estrogen receptor signaling. Despite this, women tend to exhibit lower viral loads and more robust antiviral responses than men, potentially contributing to differences in the size of viral reservoirs. Moreover, opioid abuse remains a significant problem among HIV-infected individuals, with common opioids like morphine implicated in differentially augmenting SIV/HIV persistence within the Central Nervous System (CNS) and periphery. The overarching goal of this proposal is to explore the role of sex differences in the establishment and maintenance of HIV/SIV reservoirs in the context of substance use. Preliminary studies using SIV-infected rhesus macaques as a model of HIV infection demonstrate that chronic use of morphine modulates SIV reservoirs differentially. Higher reservoir sizes are observed in CNS-derived microglial cells compared to peripheral PBMC and lymph nodes in animals exposed to morphine, as opposed to control animals. However, systematic studies are needed to document these differences comprehensively. To address this question, we plan to utilize age/sex-matched rhesus macaques and unique reagents prepared in our laboratory. This involves employing a well-established model of the SIVmac251 virus, which has been previously used to generate preliminary data, demonstrating efficient replication in both the CNS and periphery. The proposed research aims to delineate the role of sex factors in seeding and establishing viral reservoirs (Aim 1). Subsequently, we will determine the location, abundance, and persistence of viral reservoirs in the context of opioid dependence. Modeling simulations will be performed to understand the differences between establishing reservoirs in males and females (Aim 2). These studies are expected to reveal whether sex differences play a significant role in establishing reservoirs in people living with HIV infection (PLWH) in the setting of opioid usage. This information will be valuable for designing effective HIV cure strategies.

NIH Research Projects · FY 2025 · 2024-08

PROJECT SUMMARY The trp operon in several bacterial species is regulated at multiple levels that include transcription, translation, post-translation, and enzyme activity. The recent discovery of a distinct attenuation mechanism of C. trachomatis trp operon, which we termed “trans”-attenuation because it involved a trans-acting factor, the tryptophan- biosensing YtgR repressor, points to the significance of maintaining stringent regulation of expression of the tryptophan salvage genes. Regulation of trp operon by the TrpR repressor and attenuator is thought to be a check on the relatively high metabolic cost of tryptophan biosynthesis, with transcription only induced when tryptophan levels are low. The biosynthesis of tryptophan is an energetically costly process due to its requirement for D-erythrose-4-phosphate, phosphoenolpyruvate and ATP for chorismate biosynthesis, as well as glutamine, phosphoribosyl pyrophosphate and serine for the conversion of chorismate to tryptophan. It is not clear if this applies to C. trachomatis, which only requires one step – serine and indole condensation reaction to form tryptophan. Instead, we propose an alternative model, whereby dysregulated transcription of the tryptophan synthase genes trpB and trpA leads to serine overutilization, diverting this amino acid away from other serine- requiring metabolic pathways, such as phosphatidylserine (PS) biosynthesis. The additional YtgR-mediated regulation of trpBA expression ensures equitable distribution of serine between tryptophan and PS production. The iron-dependent regulation by YtgR is also proposed to be essential for optimal expression of trpBA in tissue environment that support indole-producing microbiome, such as the stratified squamous epithelium of the female lower genital tract. During epithelial cell differentiation, the gene for transferrin receptor, TFRC is progressively downregulated. We previously reported that the slow transferrin receptor recycling pathway is how C. trachomatis acquires iron. Therefore, TFRC downregulation would subject the pathogen to gradual iron starvation. Noteworthy is the location of the microbiome to the uppermost differentiated layers of the squamous epithelium; and YtgR through its iron-responsiveness could be how Chlamydia senses its proximity to indole- producing microbiota. We propose that this underpins YtgR/IGR importance to C. trachomatis. The trans-attenuation function of YtgR on the trp operon relies on a triple tryptophan motif (WWW) that renders translation of the precursor protein YtgCR sensitive to tryptophan-limiting growth conditions. We obtained evidence that there are several genes that are transcriptionally induced when tryptophan is at low levels, but are not regulated by the tryptophan-responsive TrpR repressor. Therefore, there must be another transcriptional regulator responsible. We propose that YtgR fulfills this role, which would mean a more global role for this repressor in regulating transcriptional adaptation strategies in times of tryptophan limitation.

NIH Research Projects · FY 2025 · 2024-08

ABSTRACT There is a high incidence of neurological disorders (HAND) in middle-aged people living with HIV infection (PLWH). To date, the molecular causes for diverse array of HAND remain poorly understood, and specific pharmacological interventions to manage HAND remain elusive. The working hypothesis for this R21 is “that the reactive glycolytic byproduct methylglyoxal (MG) is triggering the multitude of early-onset neurological disorders by exerting varying actions of different cell types in the brain”. The aims are (1) define pathobiological trajectory of early-onset neurological disorders in relation to MG, Glo-1, microvascular leakage, and inflammation in HIV-1 infected Hu-mice with and with anti-retroviral treatment, and (2) show that lowering MG with novel, cell- penetrant arginine-rich cyclic peptides will blunt neurological disorders in HIV-infected Hu-mice with and without anti-retroviral treatment. Data generated from this R21 will establish for the first time a link between the elevated levels of the glycolysis byproduct MG, microvascular leakage, impaired synaptic transmission, and neurological disorders, in our clinically relevant HIV-1 infected Hu-mice model. Preliminary structure-activity relationship studies will also provide novel insights into the use of arginine-rich cyclic peptides to scavenge MG and attenuate these early-onset HAND.

NIH Research Projects · FY 2025 · 2024-08

PROJECT SUMMARY/ABSTRACT Migrant agricultural workers and their families face formidable challenges in accessing healthcare and community resources in public health emergencies due to geographic, cultural, linguistic, and social isolation from community institutions and resources. Our research team has partnered with migrant families in rural Nebraska to improve access to COVID-19 testing, healthcare, and community response resources by implementing the Mobile Health for Migrant Health (mHealth-4-Mhealth) program. This program collaborates with state and local health departments and the Title IC Nebraska Migrant Education Program and seeks to mitigate direct and indirect negative effects of the COVID-19 pandemic by providing mHealth-guided decision support for at-home antigen testing and infection management coupled with mHealth-assisted socioeconomic challenges screening and response protocols to connect families to community resources and aid. Our experience to date highlights the feasibility and acceptability of mHealth-based public health interventions to engage rural communities in public health responses and facilitate access to community and health systems resources. The research objective of this study is to evaluate the impact of mHealth-4-Mhealth program implementation on migrant families’ utilization of COVID-19 testing, treatment, and related healthcare resources by achieving three specific aims. Aim #1: Determine the effectiveness of self-directed decision support via an mHealth tool and human-assisted systems navigation for engaging rural migrant families in COVID-19 (1a) testing and (1b) treatment. Aim #2: Identify socioeconomic and other implementation factors at participant, household, and community levels associated with program adoption among rural migrant families. Aim #3: Determine the cost- effectiveness of implementing mHealth and systems navigation to increase COVID-19 (3a) testing and (3b) treatment among rural migrant families. We will conduct a prospective, cluster-randomized clinical trial among Nebraska rural migrant families. Households will be randomized to one of three study arms to first receive free at-home antigen test kits for varying periods of baseline observation followed by addition of our mHealth tool and then the tool combined with human-assisted systems navigation. A fourth study arm will enroll mHealth-4-mHealth families who already received all program components. Primary outcomes will be COVID-19 test and antiviral therapy utilization. Secondary outcomes will include COVID-19 vaccination and unplanned healthcare utilization. We will analyze these outcomes across intervention states. For Aim 2 we will assess the association of socioeconomic and other implementation factors that may contribute to rural migrant families’ use (adoption) of program interventions. For Aim 3 we will determine the incremental cost-effectiveness of program implementation using the mHealth tool with or without systems navigation. Results will provide evidentiary support to inform public health policy, including development of future household-based surveillance and response programs aimed at increasing rural community resiliency.

NIH Research Projects · FY 2025 · 2024-08

Abstract The advancement of combinational antiretroviral therapy (cART) converts the life of people living with HIV (PLWH) to manageable diseases. However, due to the persistence of viral reservoirs, a cure for HIV remains elusive. That implies we need to eliminate and/or reduce the size of the viral reservoirs to achieve a cure. It has been reported that Neonates/infants whose immune system is dominated by naïve T cells with a limited number of antigen-specific memory T cells have a small HIV reservoir compared to adults. Chronological aging leads to the clonal expansion of effector memory T cells with simultaneous loss of naïve T cells, resulting in a Naïve- memory imbalance. In CNS, with aging, the blood-brain barrier (BBB) leakiness keeps increasing, and chronic activation of microglia/perivascular macrophages occurs. Additionally, aging-associated epigenetic changes in immune cells result in chronic inflammation in older people. How all these factors impact HIV reservoirs in older PLWH remains understudied and has significant implications for developing HIV cure research strategies for aged PLWH. Therefore, this proposal aims to understand how age-associated alterations in composition, epigenetic state, and functionality of immune cells alter the HIV reservoirs using SIV-infected, ART-treated young vs. aged rhesus macaques. We will study the age-associated changes in (1) DNA methylation pattern, (2) SIV integration site, and (3) in composition/functionality of T-cell and monocyte/ macrophages/microglia subsets from PBMC, lymph nodes, rectal tissue, and brain in young vs. aged macaques. Subsequently, we will measure the SIV reservoirs in all these cell types. We anticipate that understanding the aging-associated changes in immune cells at viral reservoir sanctuaries will help to provide a more informed approach to designing HIV cure research interventions for older people living with HIV.

NIH Research Projects · FY 2025 · 2024-08

Project Summary: PGC-1 family proteins (PGC-1α, PGC-1β, and PPRC1) are transcriptional coactivators that act as central hubs to coordinate diverse cellular inputs to promote the transcription of genes that regulate metabolism to maintain homeostasis. Transcriptional coactivator Peroxisome Proliferator-Activated Receptor Gamma Coactivator 1 β (PGC-1β) is over-expressed in colorectal cancers (CRC) with K-Ras mutations and promotes the survival of tumor cells. PGC-1 family proteins lack intrinsic enzymatic activity and function by facilitating interactions between transcription factors, epigenetic modifiers, and transcription initiation machinery. To identify the protein-protein interactions required by PGC-1β to promote gene expression we immunoprecipitated PGC- 1β and identified binding partners by mass spectrometry. Our data reveal that Estrogen-Related Receptor α (ERRα) and Host Cell Factor 2 (HCF2) are PGC-1β binding proteins that we propose are required for PGC-1β to regulate the transcription of genes that promote CRC survival. Our long-term goal is to inhibit CRC growth by blocking PGC-1β signaling and we propose that can be achieved by preventing PGC-1β from signaling through either ERRα or HCF2. In Aim 1, we will determine the mechanism by which phosphatase PGAM5 modifies PGC-1β to increase ERRα transcription. In Aim 2, we will identify PGC-1β-HCF2 regulated genes and determine the requirements for HCF2 signaling. In Aim 3, we will test the loss of the PGC-1β, the loss of the PGC-1β-ERRα interaction, the loss of the PGC-1β-HCF2 interaction, and the loss of both interactions in vivo using orthotopic submucosal injections in immunodeficient mice receiving Crispr-Cas9-modified patient-derived tumor organoids establish from liver metastases.

NIH Research Projects · FY 2025 · 2024-08

In an era of improved survival due to modern antiretroviral therapy (ART), liver disease has become a major cause of morbidity and mortality among HIV-infected persons causing death in 15–17% of HIV-infected patients. Whereas the rate of alcohol abuse is approximately 4.6% in the general U.S. population, 8% of HIV-infected individuals in U.S. cohorts can be classified as heavy drinkers. Despite the great progress in the development of efficient and less liver-toxic ART, current ART regimens do not eliminate all symptoms associated with HIV- induced pathology in various organs, including the liver. HIV-induced liver fibrosis is a frequent event in people living with HIV (PLWH), and it is further potentiated by alcohol use disorders (AUD). Previously, we have shown that hepatocytes exposed to HIV and ethanol express HIV RNA and HIV proteins and secrete the highest levels of extracellular vesicles (EVs/exosomes). These EVs may program the activation of non-parenchymal liver cells, including hepatic stellate cells (HSC) to promote the fibrosis progression. However, it is not clear which components of exosome cargo are responsible for HSC activation. Here, we explore the possibility that liver fibrogenic effects are induced by HIV proteins contained in hepatocyte-derived EVs. In this regard, we will focus on HIV proteins post-translationally modified with ubiquitin-like ISG15, the product of interferon-stimulated genes (ISGs) that promotes protein ISGylation to reduce toxicity of HIV proteins and decrease its release with EVs. In preliminary experiments, we demonstrated that protein ISGylation is suppressed in HIV-infected hepatocytes treated with ethanol metabolites and the engulfment of these exosomes by HSC induced their pro-fibrotic activation. These data allowed us to generate the hypothesis: Ethanol metabolism sorts of HIV proteins to exosomes via suppression of protein ISGylation in hepatocytes. These exosomes with non-ISGylated HIV protein cargo are internalized by hepatic stellate cells (HSC) to induce pro-fibrotic activation. This hypothesis will be tested in two Specific Aims: Aim1: To study the effects of ethanol metabolism on HIV protein ISGylation in hepatocytes and subsequent release of HIV proteins with exosomes; Aim 2: To elucidate the effects of exosomes derived from HIV-ethanol exposed hepatocytes on pro-fibrotic activation of hepatic stellate cells (HSC). Here, we will use multiple in vitro and in vivo approaches. Our experimental results will offer strong translatable potential into clinical practice by clarifying one of the mechanisms of HIV-alcohol-induced liver fibrosis development, which requires therapeutic interventions additional to ART to prevent liver disease progression.

NIH Research Projects · FY 2026 · 2024-07

PROJECT SUMMARY The majority of patients with Opioid Use Disorder (OUD) report sleep problems such as insomnia. Sleep disruption commonly occurs during opiate withdrawal and has been considered as a major factor that contributes to relapse. However, little is known about the underlying mechanisms, and the circuits/neurons/genes that are involved. One of major component of both the opioid withdrawal and arousal circuitry resides in the monoaminergic nuclei—the locus coeruleus (LC), which is known to abundantly express µ-opioid receptors, mediates opioid withdrawal symptoms, and promote wakefulness. In the proposed project, we will combine activity-dependent labelling and intersectional strategies to specifically target the wake-active populations of LC noradrenergic (LCNE-Wake) neurons and determine whether they are critical for morphine withdrawal-induced sleep disturbances. In Aim 1, using cutting-edge systems neuroscience tools, we will determine whether spontaneous withdrawal from chronic morphine administration disrupts the neuronal activity and membrane properties of LCNE-Wake neurons, and functionally and molecularly characterize these dysfunctional neurons. In Aim 2, we will further evaluate the functional role of major presynaptic inputs of LC, gigantocellular reticular nucleus (Gi) of medulla in withdrawal-induced sleep disturbances, test whether activation of GABAergic neurons in Gi can sufficiently mitigate the dysregulated sleep, and lastly map the spatial transcriptomic signatures of Gi-LC neurobiological pathway during morphine withdrawal. Successful completion of these aims will advance our understanding of the circuit mechanisms underlying OUD-associated sleep disturbances, potentially leading to the development of effective therapies.

NIH Research Projects · FY 2025 · 2024-07

Project Abstract: Investigating the Function of Bacterial Cytoskeletal Elements in the Division and Growth of the FtsZ-less Chlamydia Chlamydia is an obligate intracellular bacterial pathogen that causes a range of serious diseases in humans. In developed countries, Chlamydia trachomatis is the leading cause of bacterial sexually transmitted infections (STI). In developing countries, C. trachomatis is not only a significant cause of STI, but it is also responsible for the primary cause of infectious preventable blindness, trachoma. The major concern of chlamydial infections is that they are often asymptomatic and undiagnosed, which can lead to chronic sequelae. These include pelvic inflammatory disease, tubal factor infertility, and reactive arthritis for C. trachomatis. Chlamydial diseases remain a significant burden on health care systems around the world. In adapting to obligate intracellular growth, Chlamydia has significantly reduced its genome size and eliminated genes from various pathways as it relies on the host cell for its metabolic needs. One such “essential” gene that Chlamydia has eliminated is FtsZ, the central organizer of the division septum during binary fission. A major focus of the lab is to understand how Chlamydia divides. Previously, we have demonstrated that Chlamydia divides by an MreB-dependent polarized division mechanism that relies, in part, on the unique N-terminal domain of chlamydial MreB. This pathogen also alternates between different functional and morphological forms during its normal growth, also referred to as its developmental cycle. These observations, combined with its obligate intracellular dependence, make Chlamydia a difficult, but fascinating, organism with which to work. Nonetheless, recent development of genetic tools to mechanistically study chlamydiae has significantly enhanced our understanding of this pathogen. The current application goals are designed to leverage prior work investigating the function of chlamydial cytoskeletal elements. Goal 1: We will investigate how MreB is maintained at the division septum after its initial recruitment to a specific site on the bacterial surface as well as the contributions of various phospholipids to this process. We will also determine factors that contribute to how daughter cell outgrowth occurs during the budding process. Goal 2: We will investigate a novel bactofilin cytoskeletal element we identified that functions in maintaining cell size. We seek to understand its function in regulating cell size in Chlamydia by interrogating specific domains necessary for its effects and identifying interaction partners that contribute to cell size. We hypothesize that, given the absence of FtsZ in these organisms and their use of MreB in its place, the mechanisms of cell size control will be unique in these bacteria. Consequently, our results may lead to the identification of Chlamydia- unique targets for antimicrobial development. This will limit the impact of broad spectrum antibiotics on the microbiome during treatment of chlamydia infections. More fundamentally, findings from the proposed research will enhance our understanding of the evolutionary strategies bacteria employ to regulate a critical physiological function.

NIH Research Projects · FY 2024 · 2024-07

ABSTRACT Hematopoiesis produces ~300 billion blood cells daily. Hematopoietic stem and progenitor cells (HSPCs) give rise to lineage-committed progenitors (e.g., the megakaryocyte-erythrocyte progenitor (MEP)), which differentiate into mature cell types. Understanding mechanisms required for HSPC and MEP fate determination has important implications for pathologies such as leukemia, chronic anemia, and myelodysplastic syndrome (MDS), as all can be caused by mutations and misregulation of HSPC and/or MEP activities. I identified that the sterile alpha motif protein-1 (SAMD1) protein is expressed highly in HSPCs and MEPs. My preliminary data indicates that SAMD1 promotes Kit signaling, impairs erythropoiesis and HSPC activity, and modulates the expression of multiple genes required for hematopoiesis/erythropoiesis, including GATA2. In other contexts, SAMD1 acts as a transcriptional co-repressor with LSD1 to control transcription. I hypothesize that SAMD1 directs transcription and signaling during hematopoiesis. To test this hypothesis, I will use in vivo and ex vivo loss-of-function approaches. First, I will perform bone marrow transplantation experiments in mice to examine the requirement of Samd1 for HSPC activity. Second, using CRISPR-Cas9, I will knock out human SAMD1 in CD34+ hematopoietic progenitors and test SAMD1-dependent changes to the progenitor phenotype (using colony and cell signaling assays). Finally, I will establish the role of SAMD1 in modulating GATA2 gene expression using the human umbilical cord-derived erythroid progenitor-2 (HUDEP2) cell line. These experiments will determine whether and how SAMD1 expression is needed to promote HSPC and MEP cell phenotypes, including progenitor potential, cell signaling, and transcription. Linking Samd1 function to signaling and transcriptional mechanisms opens the door to translational avenues for studying the contribution of Samd1 in hematologic pathologies. The University of Nebraska Medical Center, the parent institution of this research proposal, is rapidly developing as a biomedical research leader. The scientific environment in which the proposed studies are to be performed will directly contribute to the probability of the project’s success. UNMC is an intellectual and collaborative environment with ample resources to facilitate training and career development. The training plan includes learning fundamental skills in the field of hematopoiesis, including hematopoietic stem cell transplantation. With the presentations, career workshops, and writing opportunities this fellowship will provide, the career goal of becoming an expert in the field of hematopoiesis working in an academic environment will be achieved by the applicant.

NIH Research Projects · FY 2025 · 2024-07

Project Summary: PARP inhibitors-based synthetic lethal therapy, such as BMN673 (Talazoparib), shows great efficacy for BRCA mutated (or homologous recombination repair (HRR)-deficient) cancers in preclinical and clinical studies, but demonstrates limited efficacy for majority of cancer types that are HRR proficient. To improve the PARPi efficacy and extend their therapeutic application, additional combination approaches are necessary. 5-azacytidine (AZA) is a DNA methylation inhibitor, which shows increasing clinical use for epigenetic therapy of solid tumors. AZA has been reported to enhance PARPi response through trapping large DNMT-PARP1 complexes to DNA break sites and enhancing formation of cytotoxic double-strand breaks (DSBs). Based on our preliminary data and data from others, AZA may also affect DNA repair, inducing HRR defects and sensitizing tumors to PARPi. Moreover, we hypothesize that combination of AZA with BMN673 may enhance the tumor immune response through generation of novel neoantigens and upregulation of MHC class I molecules. However, the effectiveness of this combination therapy is limited by the poor bioavailability of the two drugs and the challenge of codelivery to tumors due to the distinct physiochemical properties of BMN and Aza. In our preliminary studies, we developed a small sized nanocarrier that could selectively accumulate in the tumors and effectively penetrate to the core of experimental tumors, importantly, it could efficiently load both hydrophilic AZA and hydrophobic agent BMN673 and was more effective in suppressing tumor growth in PARPi-insensitive NSCLC tumor models at relatively lower doses. We propose to further optimize the BMN/PAZA formulation (Aim 1) and evaluate their in vivo pharmacokinetics, biodistribution and tumor penetration efficiency using tumor models that closely mimic human NSCLC (Aim 2). We will also investigate the therapeutic efficacy and the underlying mechanism (Aim 3). These studies will address the issues of off-target toxicity of AZA and PARPi, and provide a promising and safe strategy to expand the clinical use of synthetic lethal therapy to both HRR-deficient and HRR-proficient cancer patients.

NIH Research Projects · FY 2025 · 2024-07

The American Society for Rickettsiology (ASR) is the primary society for scientists studying Rickettsiales, obligate intracellular bacteria that are significant cause of morbidity and mortality world-wide. The 33rd Meeting of the AmericanSociety for Rickettsiology (ASR) will be held July 13-16, 2024, in Williamsburg, Virginia, with the 34th meeting to be held in 2026 at a location to be determined. In addition to the 2024 and 2026 meetings, ASR will participate in the 2025 International Conference on Rickettsia and Rickettsial Diseases in Lograno, Spain, by sponsoring an oral session and travel awards for trainees and young investigators. The annual meetings of the ASR aim to 1) provide the highestquality scientific program to attract outstanding researchers in academic, governmental, and private sectors and 2) promote scientific opportunities for trainees and early career investigators to present and discuss their work in a collegial, yet scientifically rigorous environment. The 33rd and 34th meetings will feature a balance of oral presentations from established/junior investigators and trainees, intermingled with posters from submitted abstracts. To increase crossdisciplinary research, oral sessions will feature a Plenary Lecture from an accomplished investigator outside the field of rickettsia. Members of the ASR Scientific Committee will select oral (short) presentations based on: (i) the quality and originality of the research; and (ii) representation from a wide range of institutions and geographic regions. In all activities, we will provide networking opportunities for students, postdoctoral fellows, and early career investigators to engage with established scientists. Mailings, e-posters, and social media advertisement will highlight the availability of travel awards for attendees. A website will feature the schedule, information and registration materials for the meeting, and ASR will offer optional financial assistance for participants with family care needs. The meeting venue includes on-site childcare options and is accessible to all guests. Approximately two hundred attendees representing early career investigators, mid-career scientists, established researchers, and clinicians from academia, government, and private-sector laboratories are anticipated to attend. There is no equivalent meeting that focuses exclusively on rickettsial diseases in the United States of America.

NIH Research Projects · FY 2026 · 2024-07

Project Summary: Currently, there are over 40 million people living with HIV-1 worldwide. Although combination antiretroviral therapy (cART) has improved the health and lifespan of millions of those living with HIV-1, long-term use of antiretrovirals coupled with limited penetration of some of these medications into the CNS, is often accompanied with neurocognitive impairments in at least 50% of infected individuals, commonly referred to as NeuroHIV. Additionally, while cART can successfully suppress peripheral viremia, persistence of cytotoxicity viral protein such as Transactivator of transcription (Tat) in tissues such as the brain, remains a significant concern. Adding further complexity is the co-morbidity of drug use disorder in those infected with HIV-1, especially that of opiates. While the direct combined effects of HIV Tat and Opiates involving astrocytic cytokines and chemokines on neuronal injury are well studied, how morphine exacerbates aberrant glial-neuronal responses to Tat & HIV-1, involving shuttling of microRNAs and reprogramming the neurons to a synaptically injured phenotype in is an unmet need in the field and warrants investigation. Our preliminary findings suggest in human primary astrocytes/ A172 cells exposure to HIV Tat mediated the induction and release of miRNA-7 and exposure to morphine mediated the upregulation and release of miR-23a in extracellular vesicles (EVs). Furthermore, neuronal uptake of Tat and Morphine stimulated astrocyte EVs (containing miR-7 and 23a) resulted in exacerbated synaptic injury compared to uptake by EVs with individual miR. These findings are backed by strong in vivo data demonstrating morphine-mediated potentiation of synaptodegeneration induced by HIV Tat and also, reversal of synatic injury by anti-miR. Taken together, the scientific hypothesis of this proposal is that HIV Tat/HIV-1 & morphine co- operate to mediate exacerbated synaptic alterations involving: a) HIV Tat/ HIV-1-mediated release of astrocytic EV miR-7 that downregulates its targets (NLGN2, GRIN2A, PBX3), & b) Morphine-mediated release of astrocytic EV miR-23a downregulates its targets (NRGN, PCLO, NLGN1), in neurons. We will test the efficacy of engineered, astrocyte specific AAVs expressing short hairpin RNA for -miR7 & -23a to restore HIV-1 Tat & morphine-mediated synaptic impairments in two complementary in vivo rodent models of NeuroHIV: a) Inducible brain Tat transgenic & b) infectious humanized mice administered opiates along with validation in archival macaque brain tissues. These studies will make a significant contribution to our understanding of glial-neuronal crosstalk, and the role of miRNAs in HIV and opiate-mediated synaptic neurodegeneration and will set a stage for a future development of RNA therapeutics for these comorbid conditions.

NIH Research Projects · FY 2025 · 2024-07

The MD-PhD program at the University of Nebraska Medical Center (UNMC) trains physician scientists to improve health through scientific advances and evidence-based approaches. Through doubling the program size since 2007, increasing NIH-funded F30/F31 fellowship awards from 2 to 19, maintaining time-to-degree that is better than the national average, improving anatomy test scores over 20 percentage points, and holding attrition below 10%, we have documented training success. Over a third of our trainees earn extramural fellowships (42%) and most hold student leadership roles. UNMC is ranked in the top 10 nationally for primary care training and rural practice and has research strengths in cancer biology, infection and inflammation, neuroscience, cardiovascular disease, and aging. The program curriculum provides career development that fosters critical thinking, adult learning, challenging paradigms, and using data to inform research and care. We foster a positive culture of training for students and focus on trainee well-being and professional activities. The program objectives are: (1) Recruit trainees committed to advance biomedical research; (2) Integrate medical and research training; (3) Promote productivity of students in advanced research areas; (4) Develop a cohort of mentors who promote student development; (5) Support service learning, community care, and development of clinical skills; (6) Develop student transferable and leadership skills through principles and practice; (7) Maintain appropriate time-to-degree for student success; and (8) Foster retention by preparing students for research and career setbacks. UNMC and our partner institutions have infrastructure for clinical and translational research through the Great Plains IDeA CTR and the Nebraska Pediatric Clinical Trials Unit. In this proposal, we propose the Nebraska Medical Scientist Training Program (MSTP) to support physician scientist training and request two funded training slots in the first year and four per year after that. Our program would be among the few MSTP programs based in an IDeA state. In this funding period, we will increase assessment sophistication, enhance alumni engagement, build our mentorship training, and provide leadership skills to all trainees. We will leverage the requested T32 funding to advance our program and change medicine.

NIH Research Projects · FY 2026 · 2024-07

Project Summary: According to the Centers for Disease Control and Prevention, HIV infection and drug abuse are intertwined epidemics that limit adherence to combination antiretroviral therapy (cART) and contribute to the worsening of HIV-associated neurocognitive disorders referred to as "NeuroHIV." Despite the ability of cART to significantly decrease viremia, the brain remains a reservoir of low-level HIV replication with the accumulation and persistence of cytotoxic viral proteins, including the HIV Transactivator of transcription (Tat). Methamphetamine is a highly addictive synthetic stimulant commonly abused by HIV-infected people worldwide. It is generally established that HIV/HIV proteins and methamphetamine activate inflammasomes in central nervous system cells, particularly microglia. However, there is a knowledge gap in our understanding of how HIV/HIV proteins and methamphetamine contribute to activating astrocyte-specific inflammasomes – NLRP6, leading to neuroinflammation, which is the focus of this study. The long-term goal is to identify the molecular mechanism(s) involved in HIV and methamphetamine-mediated astrocyte activation, which can set the stage for developing novel therapeutic targets to alleviate neuroinflammation associated with NeuroHIV. This proposal uses in vitro, ex vivo, and in vivo approaches to identify the molecular mechanism(s) involved in HIV Tat and methamphetamine-mediated astrocyte-specific NLRP6 inflammasome and its role in PANoptosis-mediated neuroinflammation. The central hypothesis is that HIV Tat-mediated astrocyte activation and neuroinflammation in NeuroHIV involve ZBP1/NLRP6-mediated PANoptosis. The hypothesis will be tested with two specific aims, and the outcome of this study will set the stage for developing novel therapeutics to dampen HIV and drug abuse- mediated neuroinflammation in NeuroHIV. Overall, this R01 application aims to determine how exposure to HIV Tat and methamphetamine induces the astrocyte-specific inflammasome, NLRP6, and its role in PANoptosis- mediated neuroinflammation using in vitro, ex vivo, and in vivo approaches.

NIH Research Projects · FY 2025 · 2024-07

Abstract: This proposal meets a critical need to strengthen the foundations of basic science by developing a program to train students in biochemistry and molecular biology. UNMC has outstanding translational research in cancer, neuroscience and other clinically relevant foci, all of which rely heavily on training in foundational biochemistry and molecular biology. Despite this, we do not yet have a training program designed to bolster this crucial fundamental area of research. The overall goal of our proposal is to enhance training in basic biochemistry and molecular biology through the funding of 9 predoctoral students, 3 of which will come from internal UNMC support. The program will rely on a carefully vetted list of experienced and motivated mentors, as well as new mentors who will work in tandem with more experienced mentors. The program will select highly qualified and diverse predoctoral students and provide them with outstanding training in scientific rigor, critical thinking, and the bench skills necessary to succeed in academic or biomedical industry research. Training will not be limited to research skills, but will provide exposure to a wide range of careers, and to develop the requisite skills in professionalism, leadership, organization, time management, teaching, oral presentations, scientific writing, etc. The proposed training program includes formal didactic courses, including a key discussion-based course in critical thinking in biochemistry and molecular biology directed by the PI of this proposal, as well as journal clubs, seminars, oral and poster presentations and other opportunities for scientific discourse. The program is designed for students to complete their training within a 5-year window and to produce an outstanding cadre of graduates that will contribute significantly to the Nebraskan and US scientific workforce.