Browse
Recent Submissions
Item Pharmacodynamic Gene and Pathway Profiling of INPP5D Modulation Across Models(2025-07-25) Matharu, Roma; Soni, Disha; Lin, Peter; Caballero-Floran, Isaac; Lamb, Bruce; Rangel-Barajes, Claudia; Oblak, AdrianAlzheimer’s disease (AD) is a progressive neurodegenerative disorder marked by the accumulation of extracellular amyloid-β plaques and intracellular neurofibrillary tau tangles. Microglia respond to these pathological features but often become functionally impaired as the disease advances. INPP5D (also known as SHIP1), a phosphatase broadly expressed in hematopoietic cells and restricted to microglia within the brain, plays a key inhibitory role in regulating microglial activity. It acts through receptors such as TREM2, Dectin-1, and FcγR via the PI3K/AKT signaling axis. Preclinical studies suggest that inhibiting INPP5D enhances microglial function and reduces AD-related pathology. To advance INPP5D-targeted therapies, it is essential to characterize the downstream transcriptional effects of inhibition. Identifying consistent gene expression changes across experimental models can help establish pharmacodynamic markers and clarify biological pathways to guide therapeutic development. We investigated transcriptomic alterations associated with INPP5D inhibition using four models: (1) primary mouse microglia from Inpp5d haplodeficient versus wild-type mice (in vitro), (2) TAD32-treated versus vehicle-treated primary microglia (in vitro), (3) TAD32-treated versus vehicle-treated C57BL/6J mice (in vivo), and (4) INPP5D-targeting siRNA versus scrambled RNA- treated C57BL/6J mice (in vivo). Gene expression was profiled using Nanostring’s neuroinflammation and glial panels. Differentially expressed genes (DEGs) were compared across models to identify shared molecular signatures, and enrichment analysis was performed. Despite a limited overlap of individual DEGs, we observed consistent enrichment of key biological pathways related to phagocytosis (including FcγR signaling, lysosomal function, and complement activation), inflammatory regulation (TLR, IL-17, and TNF signaling), and PI3K/AKT signaling. These findings suggest that INPP5D inhibition elicits conserved functional effects across diverse experimental contexts. Our findings identify convergent transcriptional and biological responses to INPP5D inhibition that may serve as pharmacodynamic indicators. These results support the utility of INPP5D modulation in therapeutic development, offering potential biomarkers for drug selection and efficacy monitoring across preclinical models.Item Wound Healing Post-Gingival Damage: Evidence for the Mesenchymal-to-Epithelial Transition in the Oral Cavity(2025-07-25) Thompson, Tyler; Walton, Carson; Syed, Sofia; Yuan, XueBackground: Oral cancers are particularly aggressive due to their propensity for early invasion into adjacent lymphatic and vascular structures. Consequently, a significant proportion of cases are diagnosed at advanced stages, underscoring the need to further elucidate mechanisms of metastasis. Given the established parallels between wound healing and tumor progression, we utilized a gingival injury model to investigate cellular migratory dynamics that may inform oral cancer metastatic behavior. Methods: K5/GFP transgenic mice were injected with Tamoxifen to induce GFP expression in Keratin 5-positive epithelial cells, followed by surgical extraction of the first molar to simulate oral tissue injury. Tissue was collected and processed for immunohistochemistry. Co-staining for Keratin 5 and GFP was used to track epithelial cell lineage, while additional co-staining with mesenchymal and microenvironmental markers was performed to further characterize cellular identity and tissue architecture. Results: Select regions within intact gingival epithelium lacked Tamoxifen-induced GFP signal, suggesting a non-epithelial cell lineage, with this pattern further exacerbated by tooth extraction surgery. The integrity of the basement membrane was maintained in GFP-negative regions as evidenced by ITGB1 staining. Keratin 17 expression in both GFP-positive and GFP-negative regions highlighted that GFP loss was not stress induced. Notably, Vimentin and Collagen I showed highly specific expression within GFP-negative zones, suggesting the epithelial cells present were mesenchymal in origin. Furthermore, WNT5a, known to promote the Mesenchymal-to-Epithelial Transition (MET), was prominently expressed in these GFP-negative areas, indicating its potential role in facilitating mesenchymal cell integration into the epithelium. Conclusion: The detection of mesenchymal markers within epithelial regions, along with the absence of GFP in these infiltrating cells, provides evidence for MET in oral mucosa. Our data indicate this cellular transition in oral mucosa may contribute to the highly metastatic nature of oral cancers by facilitating cellular plasticity and invasion processes.Item Modeling Inflammaging of the Bone Marrow Microenvironment Stimulates Multiple Myeloma Associated Stem Cells(2025-07-24) Thaker, Krishi; Wilburn, Piper; Kurihara, Noriyoshi; Suvannasankha, Attaya; Quinlan, Conner; Marinković, MilošBackground: Multiple Myeloma (MM) is a malignancy of mature plasma cells, primarily affecting those over 65, with a 5-year survival rate of ~62%. Aging alters the bone marrow (BM) microenvironment, including remodeling of the extracellular matrix (ECM), which may promote tumor-supportive functions of mesenchymal stromal cells (MSCs) and hematopoietic stem cells (HSCs). However, determining the impact of aging-related matrix remodeling on MM onset/progression remains challenging due to the absence of microphysiological systems that preserve the tumorigenic phenotype of MM cells in vitro. We hypothesize recapitulating aged BM ECM may overcome these limitations by providing a physiologically relevant framework that sustains MM cell behavior and models the tumor-supportive microenvironment. Additionally, interleukin-6 (IL-6), an inflammatory cytokine elevated in both the inflammaging BM microenvironment and MM, preferentially expands MM-associated MSCs without affecting HSCs. Methods: Cryopreserved tumor-associated MSCs and HSCs were cultured on tissue culture plastic (TCP), young ECM (≤25 y/o), or aged ECM (≥60 y/o), in either α-MEM or IMDM media, with some cultures supplemented with IL-6. HSC cluster formation was observed, and cells were analyzed using cytospin with Giemsa-May-Grünwald staining. Results: MM-derived HSCs, cultured on aged ECM, exhibited myelopoietic skewing. In contrast, young ECM with IL-6 promoted MSC expansion, suggesting a synergistic effect in developing patient-derived stromal models. Adherent MSCs were harvested and expanded on TCP for ECM generation. Conclusion: ECM-based microphysiological systems offer a promising platform for scalable, in vitro systems that recapitulate aging- and MM-associated remodeling of the BM microenvironment. Understanding how aged ECM drives myeloid skewing may reveal therapeutic targets to delay/prevent MM progression. The observed myeloid bias in MM-derived HSCs cultured on aged ECM may reflect physiologically relevant disruptions in immunoregulatory cell populations. Future work will test whether ECM from MM patient-derived MSCs preserves tumor-specific phenotypes to advance translational models for MM drug discovery.Item The Impact of Manual Correction on Automated Hippocampal Volumetry in a Young Athletic Cohort with Variable Level of Exposure to Head Trauma(2025-07-25) Kamiotis, Demetrios; Yang, Ho-Ching; Takieldeen, Yomna; Shahid, Salman; Wu, Yu-ChienIntroduction: An estimated 1.6 to 3.8 million people suffer from sport-related concussion annually, yet the long-term effects of concussion and repetitive head injury exposure are not fully understood. Given the potential risk of future neurodegeneration, the hippocampus has emerged as a key region of interest. While post-mortem studies demonstrated marked hippocampal atrophy in athletes from high-impact sports, more recent in-vivo research reveals more nuanced findings as they rely heavily on imaging and segmentation accuracy. Freesurfer, a widely used tool for automated hippocampal segmentation, has been shown to overestimate the volume, especially in older populations. In this study, we evaluated the accuracy of Freesurfer’s segmentation against manual refinement in a cohort with head impact exposure. Method: Structural MRI data (T1W and T2W) of the CARE-SALTOS Integrated (CSI) project was used for hippocampal segmentation using FreeSurfer (7.4.1). Twenty participants (age range: 29 to 32) were sex-matched and grouped into high-exposure (n=10, including 5 football players) and low-exposure cohorts (n=10). The hippocampal volumes were compared between initial automatic segmentations and subsequent manually refined delineations by a trained medical professional using paired t-tests. Results: Our results showed significant differences in the hippocampal volume between automatic segmentation and after manual refinement. By visual inspection, FreeSurfer tends to overestimate hippocampal segmentation volumes as generated masks extend into lateral ventricular spaces and include choroid plexus. In both groups, hippocampal volumes significantly decreased after manual refinement (Left Hippocampus, -788.81±245.30mm3, p=1.15E-11; Right Hippocampus, -649.60±246.83mm3, p=3.59E-10). Conclusion: Based on our results, we conclude that while automatic segmentation is a feasible and fast method for large datasets, it requires subsequent manual refining to ensure data accuracy by avoiding volumetric overestimation and potential bias from adjacent ventricular areas.Item Association Between Non-Alcoholic Fatty Liver Disease and Atherosclerotic Cardiovascular Disease-Associated Acute Myocardial Infarction and Ischemic Stroke Using Real World Data(2025-07-25) Rusche, Jonathan; Lomada, Uday; Wu, Steven; Seetharam, GattadahalliBackground/Objectives: ASCVD-related conditions are the leading cause of death in individuals with NAFLD/nonalcoholic steatohepatitis (NASH). Despite this, the current 10-year ASCVD risk assessment tools do not consider a patient’s NAFLD/NASH history. The aim of this study is to examine the association between NAFLD/NASH and ASCVD-associated myocardial infarction and ischemic stroke, and to reinforce existing evidence showing that NAFLD/NASH patients are at greater risk for ASCVD-associated cardiac events. Methods: We used de-identified patient data from the IU School of Medicine–Evansville RWEdataLab (CRC/Sidus Insights) National Real-World Cardiology database. Patients with diagnoses of NAFLD or NASH who later developed ischemic stroke (IS) or myocardial infarction (MI) were identified using ICD-9&10 codes. Odds ratios were calculated to estimate the likelihood of MI/IS in patients with NAFLD/NASH. Results: Of 3,396,429 patients in the database we identified 674 patients with NAFLD/NASH and MI/IS. In 424 patients, the odds of MI/IS after NAFLD/NASH diagnosis were significantly higher (OR 1.648; 95% CI: 1.489–1.823) compared to patients without NAFLD/NASH, with females at significantly greater odds (OR 1.983; 95% CI: 1.730–2.273) than males (OR 1.366; 95% CI: 1.176−1.587). Patients with type 2 diabetes mellitus (T2DM) and NAFLD/NASH had increased odds of subsequent MI/IS (OR 2.328; 95% CI: 2.015-2.689) compared to those without NAFLD/NASH. Additionally, T2DM females had significantly greater odds of MI/IS (OR 2.861; 95% CI: 2.358-3.470) than T2DM males (OR 1.873; 95% CI: 1.508-2.327) following NAFLD/NASH diagnosis. Conclusions/Impact: Our results support an association between NAFLD/NASH and ASCVD-related MI/IS suggesting NAFLD/NASH may be an important early marker of cardiovascular risk. One limitation of this research includes 4-year binning of patient age. Furthermore, NAFLD/NASH is progressive, and diagnosis underrepresents true incidence in the population analyzed. Nonetheless, these results may support revising current ASCVD risk calculators to include NAFLD status, improving risk stratification and encouraging earlier preventive interventions.Item Hematological and inflammatory biomarkers associated with pain in sickle cell disease(2025-07-24) Clay, Abbie; Carpenter, Caitlyn; Houran, Lina; Debats, Candice; Pucka, Andrew; Wang, YingHematological and Inflammatory Biomarkers Associated with Pain in Sickle Cell Disease Background and Objective: Pain is the hallmark of sickle cell disease (SCD). Clinically, SCD is characterized by a steady-state phase with chronic pain and a vaso-occlusive crisis (VOC) phase marked by unpredictable, severe pain episodes often requiring hospitalization and opioids. VOCs, caused by the adhesion and obstruction of sickled red blood cells in microvasculature, are among the most debilitating manifestations of the disease. VOCs have four clinically defined phases: prodromal, initial, established, and resolving. We observed abnormal elevation of inflammatory biomarkers in SCD compared to healthy controls at baseline and changes throughout the VOC stages. The goal of this exploratory project of our clinical trial (ClinicalTrial.gov ID: NCT05045820) is to evaluate the inflammatory biomarker-based immunological mechanism associated with pain and acupuncture analgesia in SCD. Methods: Peripheral blood samples are collected at baseline and at each VOC phase. Hematological testing (complete blood count with white blood cell differential, hemoglobin electrophoresis, and reticulocyte count) is performed while serum, plasma, and peripheral blood mononuclear cells are simultaneously isolated, aliquoted, and stored for future analyses. Inflammatory markers are assessed with immune assays and cell activation analyses. Patient-reported outcome measures are collected using validated instruments to assess pain, neuropathic symptoms, physical dysfunction, emotional well-being, and quality of life. Samples and data are collected before, during, and after the treatment. Results: Our published results have demonstrated significant differences in patient-reported outcome measures and hematological and inflammatory biomarkers in SCD as compared to healthy controls. Additional analytical work is ongoing. Conclusion and Potential Impact: Published and preliminary data demonstrate robust clinical outcomes of acupuncture treatment and biomarker-based mechanistic findings related to inflammation in SCD. This project has strong clinical relevance as it investigates biomarker-based mechanisms of pain in SCD and aims to identify therapeutic targets to improve pain management and quality of life.Item Optimization of a RAG Model Chatbot for Obstetric Medical Education(2025-07-25) Knight, Kenzie; Pape, Kelsey; Kim, Esther; Rouse, Caroline; Hanson, David; McKenzie, Rachel; Shanks, AnthonyBackground/Objective: In fast-paced clinical settings like labor and delivery, quick access to accurate, evidence-based guidance is critical. Traditional methods of sharing protocols can be inefficient, especially for trainees managing complex scenarios. Advances in artificial intelligence (AI) and large language models (LLMs) offer the potential to transform how critical clinical decision-making tools are disseminated, making them more accessible, responsive, and integrated into real-time learning. Our objective was to develop our own chatbot to facilitate quick retrieval and enhanced learning. Methods: The AI chatbot was developed by IUSM researchers using Microsoft Copilot Studio and made accessible via Microsoft Teams. The AI chatbot was built on a retrieval-augmented generation (RAG) framework, leveraging a fine-tuned LLM. The base LLM was augmented with domain-specific documents, including IUSM protocols – developed by the Maternal-Fetal Medicine division - which incorporate evidence-based practices and provide a consistent management strategy for OB conditions. The RAG architecture was configured to retrieve contextually relevant documents from the collection of protocols in response to user queries. Results: The chatbot retrieves information from the protocols based on a user’s query and generates a relevant answer with reference to the sources. Upon testing the chatbot, we learned that the chatbot was unable to retrieve information from figures, flowcharts, and diagrams. Therefore, figures were manually annotated and narrated, enabling the model to process and retrieve relevant visual information during inference. With other adjustments, the chatbot is able to read all aspects of the protocols and synthesize accurate answers to clinical questions. Conclusion and Potential Impact: We optimized an RAG model AI chatbot for clinical education, demonstrating that existing educational material can be used to activate a chatbot to serve clinical needs with sufficient oversight to ensure clinical accuracy. Future directions include evaluating its impact on clinical decision making and student perception of the learning environment once the chatbot is more uniformly adopted.Item Targeting Ref-1 in cancer: Progression of advanced generation Ref-1 inhibitors through the drug discovery pipeline(2025-07-24) Ball, Anthony; Gampala, Silpa; Peil, Jacqueline; Grese, Tim; Wikel, Jim; Kelley, Mark; Fishel, MelissaAPE1/Ref-1 (Apurinic endonuclease1 / Redox factor-1, referred to as Ref-1) is a multifunctional protein with both DNA repair and redox activities. The redox activity of Ref-1 activates transcription factors such as HIF1a, STAT3, and NFkB that are involved in cancer cell survival and proliferation. Targeting Ref-1 with specific inhibitors represents a promising therapeutic strategy across multiple cancer types. This study focuses on screening and characterizing advanced-generation Ref-1 inhibitors to identify potential candidates for further preclinical development. As part of this drug discovery pipeline, we evaluated four advanced generation compounds and compared them to second-generation compound, APX2009. We selected malignant peripheral nerve sheath tumor (MPNST) cells for evaluation because these tumors often exhibit elevated oxidative stress and redox signaling, making them more dependent on Ref-1 activity. Using MPNST cells, I measured inhibition of downstream Ref-1 target, NFκB and cytotoxicity after treatment. NFκB activity was measured via a luciferase-based reporter assay and two viability assays were compared: Alamar Blue and Methylene Blue. Luciferase assays further confirmed on target inhibition of Ref-1–regulated NFκB activity. Both cell viability assays demonstrated dose-dependent reduction in cellular viability, confirming compound efficacy. Among the compounds tested, WGK2105 exhibited high potency with an IC₅₀ of ~2 µM. WGK2110 and WGK2111 demonstrated similar potency and Ref-1 inhibition when compared to APX2009, while WGK2092 required significantly higher concentrations and will not be prioritized for further development. These findings support the potential for discovery of more potent Ref-1 inhibitors and identify multiple promising candidates for preclinical advancement. Future efforts will assess metabolic stability in mouse and human liver microsomes to determine in vivo suitability. In vivo efficacy studies and full pharmacokinetic profiling (ADME) will follow for top candidates. Comparative analysis of active versus inactive compounds will inform structure activity relationship (SAR) development, guiding next-generation inhibitor design. These efforts are key steps toward Investigational New Drug (IND) applications and clinical trial readiness.Item Infection Rates of Initial and Exchanged Peripherally-Inserted Central Catheters in Pediatric Patients(2025-07-25) Walker, Charlie; Pfaff, SeanBackground/Objective: Peripherally-inserted central catheters (PICCs) act as modes of long-term access to the bloodstream in patients who would otherwise need frequent needle sticking to obtain intravenous access. PICCs are beneficial because they allow for efficient access to the bloodstream to administer medications such as chemotherapy, antibiotics, total parenteral nutrition, and pain control through a single long-term access point. This allows for less pain and discomfort for patients (especially pediatric patients), minimizes waste, and allows for convenient access to the bloodstream for delivery of medications or sampling of blood for testing which allows for better long-term care. However, PICC replacement procedures are associated with increased risk of introducing infections and thrombosis. We hypothesize that patients who do not receive PICC exchanges will have lower rates of infection than those who do receive an exchange. Methods: For this quality improvement retrospective study, we examined one year worth of PICC placement/exchange procedures in pediatric patients at a single academic hospital through their electronic medical records in Cerner. Infections were documented by positive blood cultures at the PICC site within 30 days of the procedure and were corroborated through clinical documentation to ensure the PICC was the origin of the infection, rather than just a site of infection spread. Results: In the year 2024, pediatric patients displayed significantly higher infection rates in PICC exchanges at 1.97%, compared to 0.40% in initial PICC placements (p=0.040). Clinical Implications and Conclusion: The data from this investigation supports previously identified infection rates in the pediatric patient population. The results from this project will help support and guide future guidelines regarding clinical decision making in pediatric patients who require long-term intravenous access.Item Red Cell Distribution Width as a Prognostic Indicator of Mortality in Extracorporeal Life Support(2025-07-25) Jones, Jacob; Morton, John; Mohammed, AsimBackground Red cell distribution width (RDW) is a component of routine inpatient blood work that has been indicated as a potential prognostic mortality indicator for various conditions. Extracorporeal membrane oxygenation (ECMO) patients have a high mortality rate, and ECMO does not have easily obtainable prognostic measurements. RDW values may be a potential prognostic factor for ECMO patients and improve clinical decision making. Methods 142 patient charts (70 survivors, 72 non-survivors) placed on ECMO care at Lutheran Hospital met inclusion criteria. RDW values were collected at six specific times during care along with peak RDW value and survivorship outcome. Patients were stratified into high (>14.5%) and low (≤14.5%) RDW groups for each time. Relative risk (RR) and odds ratio (OR) were used to analyze survivorship outcome between the groups. Results Only several RR and OR calculations yielded results that indicated differences in mortality risk and odds and were statistically significant. RDW at 24Hr after ECMO initiation showed a RR of 1.72, 95% CI 1.06-2.76, p<.05 and OR of 2.68, 95% CI 1.24-5.80, p<.05. RDW at ECMO termination and discharge had an OR of 3.88, 95% CI 1.20-12.55, p<.05. Discussion Results showed an association between high RDW value and increased risk and odds of mortality at 24Hr after ECMO initiation. Results also indicated increased odds of mortality at ECMO termination and discharge. These preliminary results are promising for further analysis and encourage the feasibility of potentially using RDW as a prognostic mortality indicator during ECMO treatment. Conclusions Further analysis must be conducted on the samples collected during the study, but these preliminary results demonstrated feasibility of RDW as a prognostic biomarker for clinical decision making with ECMO survivorship outcomes.