Browsing by Author "Szymanski, Megan"

Now showing 1 - 4 of 4
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    A reporter Oropouche virus expressing ZsGreen from the M segment enables pathogenesis studies in mice
    (American Society for Microbiology, 2024) Gunter, Krista; Omoga, Dorcus; Bowen, James M.; Gonzalez, Lorimar Robledo; Severt, Sydney; Davis, Mackenzie; Szymanski, Megan; Sandusky, George; Duprex, W. Paul; Tilston-Lunel, Natasha L.; Microbiology and Immunology, School of Medicine
    Oropouche fever caused by Oropouche virus (OROV) is a significant zoonosis in Central and South America. Despite its public health significance, we lack high-throughput diagnostics, therapeutics, and a comprehensive knowledge of OROV biology. Reporter viruses are valuable tools to rapidly study virus dynamics and develop neutralization and antiviral screening assays. OROV is a tri-segmented bunyavirus, which makes generating a reporter virus challenging, as introducing foreign elements into the viral genome typically affects fitness. We previously demonstrated that the non-structural gene NSm on the OROV medium (M) segment is non-essential for replication in vitro. Taking advantage of this, we have now generated a recombinant OROV expressing fluorescent protein ZsGreen in place of NSm. This reporter OROV is both stable and pathogenic in IFNAR-/- mice and provides a powerful tool for OROV pathogenesis studies and assay development.IMPORTANCEEmerging and reemerging infectious agents such as zoonotic bunyaviruses are of global health concern. Oropouche virus (OROV) causes recurring outbreaks of acute febrile illness in the Central and South American human populations. Biting midges are the primary transmission vectors, whereas sloths and non-human primates are their reservoir hosts. As global temperatures increase, we will likely see an expansion in arthropod-borne pathogens such as OROV. Therefore, developing reagents to study pathogen biology to aid in identifying druggable targets is essential. Here, we demonstrate the feasibility and use of a fluorescent OROV reporter in mice to study viral dynamics and pathogenesis. We show that this reporter OROV maintains characteristics such as growth and pathogenicity similar to the wild-type virus. Using this reporter virus, we can now develop methods to assist OROV studies and establish various high-throughput assays.
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    Limited Approaches for Genomic Studies in Rapid Postmortem Tissue Collection
    (IntechOpen, 2023) Sandusky, George; Yard, Michael; Szymanski, Megan; Emmert , Lydia; Valadares, Vivian; Biology, School of Science
    Rapid postmortem tissue collection has been shown to have increasing use for molecular and genetic profiling. Although research on human tissue has been conducted for many years, modern molecular assays have significantly higher sensitivity and specificity than those used in the past. Higher grade tissue specimens are now required for the extraction of macromolecules as a result of this. In fact, these studies have paved the way for multiple postmortem tissue collection studies such as COVID-19, brain, skin, and small cell lung cancer. Best practices for tissue collection have also been developed by a number of professional organizations, including the National Institutes of Health Office of Biorepositories and the International Society of Biological and Environmental Repositories (ISBER). These guidelines can be used to establish procedures for tissue collection.
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    Loss of Dnmt3a impairs hematopoietic homeostasis and myeloid cell skewing via the PI3Kinase pathway
    (The American Society for Clinical Investigation, 2023-05-08) Palam, Lakshmi Reddy; Ramdas, Baskar; Pickerell, Katelyn; Pasupuleti, Santhosh Kumar; Kanumuri, Rahul; Cesarano, Annamaria; Szymanski, Megan; Selman, Bryce; Dave, Utpal P.; Sandusky, George; Perna, Fabiana; Paczesny, Sophie; Kapur, Reuben; Pediatrics, School of Medicine
    Loss-of-function mutations in the DNA methyltransferase 3A (DNMT3A) are seen in a large number of patients with acute myeloid leukemia (AML) with normal cytogenetics and are frequently associated with poor prognosis. DNMT3A mutations are an early preleukemic event, which - when combined with other genetic lesions - result in full-blown leukemia. Here, we show that loss of Dnmt3a in hematopoietic stem and progenitor cells (HSC/Ps) results in myeloproliferation, which is associated with hyperactivation of the phosphatidylinositol 3-kinase (PI3K) pathway. PI3Kα/β or the PI3Kα/δ inhibitor treatment partially corrects myeloproliferation, although the partial rescue is more efficient in response to the PI3Kα/β inhibitor treatment. In vivo RNA-Seq analysis on drug-treated Dnmt3a-/- HSC/Ps showed a reduction in the expression of genes associated with chemokines, inflammation, cell attachment, and extracellular matrix compared with controls. Remarkably, drug-treated leukemic mice showed a reversal in the enhanced fetal liver HSC-like gene signature observed in vehicle-treated Dnmt3a-/- LSK cells as well as a reduction in the expression of genes involved in regulating actin cytoskeleton-based functions, including the RHO/RAC GTPases. In a human PDX model bearing DNMT3A mutant AML, PI3Kα/β inhibitor treatment prolonged their survival and rescued the leukemic burden. Our results identify a potentially new target for treating DNMT3A mutation-driven myeloid malignancies.
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    Stromal heterogeneity may explain increased incidence of metaplastic breast cancer in women of African descent
    (Springer Nature, 2023-09-14) Kumar, Brijesh; Khatpe, Aditi S.; Guanglong, Jiang; Batic, Katie; Bhat-Nakshatri, Poornima; Granatir, Maggie M.; Addison, Rebekah Joann; Szymanski, Megan; Baldridge, Lee Ann; Temm, Constance J.; Sandusky, George; Althouse, Sandra K.; Cote, Michele L.; Miller, Kathy D.; Storniolo, Anna Maria; Nakshatri, Harikrishna; Surgery, School of Medicine
    The biologic basis of genetic ancestry-dependent variability in disease incidence and outcome is just beginning to be explored. We recently reported enrichment of a population of ZEB1-expressing cells located adjacent to ductal epithelial cells in normal breasts of women of African ancestry compared to those of European ancestry. In this study, we demonstrate that these cells have properties of fibroadipogenic/mesenchymal stromal cells that express PROCR and PDGFRα and transdifferentiate into adipogenic and osteogenic lineages. PROCR + /ZEB1 + /PDGFRα+ (PZP) cells are enriched in normal breast tissues of women of African compared to European ancestry. PZP: epithelial cell communication results in luminal epithelial cells acquiring basal cell characteristics and IL-6-dependent increase in STAT3 phosphorylation. Furthermore, level of phospho-STAT3 is higher in normal and cancerous breast tissues of women of African ancestry. PZP cells transformed with HRasG12V ± SV40-T/t antigens generate metaplastic carcinoma suggesting that these cells are one of the cells-of-origin of metaplastic breast cancers.