Hypereosinophilic (HES) syndrome is an umbrella term encompassing several disease subsets that affects humans and veterinary species, ultimately resulting in >1,500 eosinophils/uL circulating in the blood documented over six-months. This eventually culminates in end-organ infiltration and increased patient morbidity and mortality. In mice where the gene Map3k14 encoding NF -kB inducing kinase (NIK) is knocked out, a HES-like syndrome develops that is dependent on Th2 cells and cytokines. NIK is the upstream regulator of the noncanonical NF-kB pathway and is involved in lymphoid organ development, B cell lymphopoiesis, and myelopoiesis. In addition to regulating the noncanonical NF-kB pathway, NIK is also involved in regulation of kB dimers of the canonical NF-kB pathway and can function independent of NF-kB signaling by regulating lipid and glucose metabolism, mitochondrial, and RIP1 binding to influence cell survival and death. Despite previous studies performed in the Nik-/- model, the mechanisms underlying eosinophil development, plasticity, and fitness in conjunction with the bone marrow and splenic microenvironments have not been fully elucidated.
In the present work, we reviewed current data exploring the influence of the noncanonical NF-kB pathway and NIK specifically on the development of acute myeloid leukemias (AMLs) and Myelodysplastic Syndrome (MDS) with a focus on how these mechanisms might induce subvariants of HES. We next examined the effect of NIK loss on eosinophilopoiesis within hematopoietic tissues in vivo and in various cell culture environments in vitro via cytology, histology, flow cytometry, FACS, positive cell selection, MTT assay, BrDU assay, and protein microarray analysis. Overall, our findings suggest that NIK influences eosinophil maturation, proliferation, metabolism, survival, and potentially plasticity in vivo and in vitro under different environmental conditions and Th2 cytokine influence. NIK loss was also associated with altered free and bound TNFR1 levels on day 13 in vitro. TNFR1 acts upstream of RIP1 and suggests that these differences may be due to NF-kB independent functions of NIK. Overall, these results provide further insight into the potential mechanisms underlying eosinophilopoiesis in the Nik-/- murine model. This information may prove useful in discovering new treatment options underlying subvariants of HES in both human and veterinary patients. / Doctor of Philosophy / A less well-known albeit important white blood cell (WBC) is the eosinophil. It is essential for combating parasitic infections but is also involved in allergic responses. Hypereosinophilic Syndrome (HES) is an umbrella term encompassing a variety of diseases that affects human and veterinary patients. It results in an overproduction of eosinophils not associated with parasitic infections or allergic responses. Although several variants of the disease exist, diagnosing subsets of HES poses a diagnostic challenge and can impact patient care and prognosis long term.
Mice carrying a genetic deletion (Map3k14) encoding a kinase known as NF-kB inducing kinase (NIK) develop HES-like syndrome; these mice are hereafter referred to as Nik-/- mice. HES-like syndrome in Nik-/- mice develops secondary to NIK loss in Th2 lymphocytes (another type of WBC). NIK is an upstream regulator of the noncanonical NF-kB pathway that influences WBC, lymph node, and spleen development. NIK also regulates canonical NF-kB molecules and can function independent of NF-kB signaling by impacting fat and glucose metabolism, binding to mitochondria, and interacting with a kinase known as RIP1 to regulate cell death or survival. Despite previous work studying HES-like syndrome in Nik-/- mice, the specifics of the bone marrow and eosinophil development in response to Th2 cells have not been fully characterized.
In the present work, we reviewed data exploring the noncanonical NF-kB pathway and NIK specifically in the development of WBC cancers and how this might manifest as HES. We then studied eosinophil development in the bone marrow, spleen and in culture for both Nik-/- and wild-type mice by assessing cell and tissue morphology, cell surface marker expression, response to Th2 signaling molecules, as well as cell maturation, death, metabolism, proliferation, and cytokine production. Our findings suggest that NIK is essential for eosinophil growth and survival. We also noted differences in a molecule known as TNFR1 in Nik-/- cultures on day 13. This molecule acts upstream of RIP1, suggesting an NF-kB independent function of NIK in regulating eosinophil maturation in response to Th2 molecules. This information may prove useful in discovering new treatments for HES in human and veterinary patients.
| Identifer | oai:union.ndltd.org:VTETD/oai:vtechworks.lib.vt.edu:10919/118652 |
| Date | 22 April 2024 |
| Creators | Trusiano, Briana Lynn |
| Contributors | Biomedical and Veterinary Sciences, Allen, Irving Coy, Zimmerman, Kurt L., Dervisis, Nikolaos, Luo, Xin, LeRoith, Tanya |
| Publisher | Virginia Tech |
| Source Sets | Virginia Tech Theses and Dissertation |
| Language | English |
| Detected Language | English |
| Type | Dissertation |
| Format | ETD, application/pdf |
| Rights | Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International, http://creativecommons.org/licenses/by-nc-sa/4.0/ |
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