Louisiana Research Day Program Book 2025

Biomedical Research: Section 1

Biomedical Research: Section 1

A. Madi Grote, BS, OMS-III; J. Annelise Hayward, BS, OMS-III; Riva Kelly, BS, OMS-III; Caroline Naquin, BS, OMS-II; K Adam Morrow, PhD; Melissa EH Lipsmeyer, MS, PhD VCOM-Louisiana 2 INDOLE-3-PROPIONINC ACID MODULATES INSULIN SIGNALING IN ENDOMETRIOSIS CELLS IN VITRO

Context: Pseudomonas aeruginosa is an organism known to cause nosocomial pneumonia that can progress to sepsis and acute lung injury, especially in immunocompromised patients (Balzcon et. al. 2024, Stevens et. al. 2014). Following recovery from this infection, patients have elevated death rates and manifest various morbidities such as cognitive deficits (Balzcon et. al. 2023). Through a type 3 secretion system (T3SS) P. aeruginosa introduces exoenzymes into host cells, including exoenzyme Y (ExoY) during infection. ExoY-dependent production of cAMP activates protein kinase A (PKA) causing the hyperphosphorylation of endothelial tau, a multifunctional microtubule-associated protein (MAP) found in neuronal and endothelial cells. The phosphorylation of tau consequently impairs microtubule assembly (Baas and Qiang 2019, Morrow et. al. 2015). This cytotoxic tau can enter circulation making it a target of study for the diffuse effects seen following a nosocomial pneumonia infection (Balzcon et. al. 2023). Furthermore, tau has other properties beyond acting as a simple stabilizer of microtubules. Tau has been indicated in Alzheimer’s disease along with amyloid beta (A β ), a multifunctional protein Alexis Ross OMS-II; Marco Taveras, OMS-II; Rebekah Morrow, PhD VCOM-Louisiana 3

EXAMINING TAU EXPRESSION AND PRESENCE

Context: Endometriosis is a benign gynecological condition characterized by the presence of endometrial-like tissue outside the uterus, often causing chronic pain and infertility. Recent studies have demonstrated an increased risk of developing endometriosis in metabolic conditions derived from insulin dysregulation such as Type 1 diabetes, gestational diabetes and metabolic syndrome. Disturbances in the gut-microbiome have been linked to both endometriosis development and metabolic disorders by altering circulating bacterial derived metabolites, yet precise mechanisms remain unknown. Indole-3-propionic acid (IPA), a gut-bacteria-derived metabolite that has functions in other female reproductive tissues and pathologies, has previously been shown to modulate insulin signaling and glucose homeostasis. Objective: The objective of this study is to determine if IPA can modulate insulin signaling endometriotic cells which may ultimately alter numerous cell processes that may contribute to the severity of the disease.

Methods: For these experiments, we utilized the endometriotic epithelial cell line 12Z. To examine the effects of IPA on insulin signaling, cells were cultured for 24 or 72 hours in the presence or absence of 100nM insulin (hyperinsulinemic levels) and either 1uM (physiologic levels) or 5uM (supraphysiological levels) IPA. Alterations in insulin signaling we determined on whole cell lysates by western blot analysis following standard protocols. Results: Western blot analysis demonstrated that IPA was able to modulate a few insulin responsive genes such as pAKT, FOXO1/3, SPARC and PPAR- γ in a time and dosage dependent manner. The majority of the effects were seen at the 72hr timepoint. Other insulin responsive genes such as p-mTOR were not impacted by IPA treatment. Conclusions: These initial experiments reveal that IPA is capable of modulating insulin signaling in endometriotic cells. This offers a new area of investigation in understanding how metabolic disturbances involving hyperinsulinemia and the gut microbiome influence the pathogenesis of endometriosis. Further experiments investigating

how IPA alters insulin-induced changes in endometriosis cell migration, proliferation and extracellular matrix dynamics are underway.

produced during infection that has been shown to be an innate immune effector (Balzcon et. al. 2024, Kent et. al. 2020). The interrelationship of tau and A β suggests that unphosphorylated tau could be implicated in the antimicrobial activity exhibited by A β . Objective: The focus of this experiment is to confirm the presence or lack of tau protein in rat endothelial cells with previously introduced plasmids encoding for a tau knockout (KO), 1N4R tau isomer, and Ser-214-Ala tau mutant which cannot be phosphorylated. Methods: Starting with cell lines previously listed, we plan to conduct RNA isolations, PCRs, and western blots on the tau variants: 1N4R, Ser-214-Ala, and tau KO using an empty vector as the control. This will allow us to quantify the translation and subsequent transcription of tau into a functional protein from the pulmonary vascular endothelial cells (PVEC). Anticipated Results: We expect to find tau mRNA and protein in 1N4R and Ser-214-Ala; however, in the tau KO it should not be present and have similar results to our control, an empty

vector. We hope to confirm that our tau variants are being actively transcribed and subsequently translated into a functional tau protein as expected. Conclusions: This confirmatory experiment hopes to aid in future investigations into the connection between phosphorylated endothelial tau and the downstream effects found in patients following a nosocomial pneumonia infection by ensuring the suitability of these variants for future experimental use.

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2025 Research Recognition Day