Dissertation
Mechanisms of neutrophil nuclear hypersegmentation and consequences of Helicobacter pylori induced subtype differentiation
University of Iowa
Doctor of Philosophy (PhD), University of Iowa
Summer 2021
DOI: 10.17077/etd.005865
Abstract
Neutrophils are heterogeneous and can exhibit considerable phenotypic and functional plasticity. In keeping with this, other laboratory members discovered previously that Helicobacter pylori infection induces N1-like subtype differentiation of human PMNs that is notable for profound nuclear hypersegmentation. Herein, I utilized biochemical approaches and confocal and super-resolution microscopy to gain insight into the underlying molecular mechanisms. Sensitivity to inhibition by nocodazole and taxol indicated that microtubule dynamics were required to induce and sustain hypersegmentation. Super-resolution Stimulated Emission Depletion (STED) imaging demonstrated that microtubules were significantly more abundant and longer in H. pylori infected PMNs. Dynein activity was also required, and enrichment of this motor protein at the nuclear periphery was enhanced following H. pylori infection. In contrast, centrosome splitting did not occur, and lamin B receptor abundance and ER morphology were unchanged. Finally, analysis of STED image stacks revealed that nuclear volume increased markedly prior to the onset of hypersegmentation and that nuclear size was differentially modulated by nocodazole and taxol in the presence and absence of infection. Taken together, our data define a new mechanism of hypersegmentation that is mediated by microtubules and dynein and as such advance understanding of processes that regulate nuclear morphology. Similarly, I discovered that PKC inhibitors induced hypersegmentation in neutrophils and microtubules were also involved. Confocal microscopy illustrated that MTs were apparently more abundant too in PMNs treated with PKC inhibitors.To start exploring the functional consequences of the N1-like phenotype, I studied the effect of exposing PMNs or N1-like PMNs to T cells and measured T cell proliferation and CD45RA expression after CD3/28 stimulation. H. pylori infected neutrophils inhibited T cell proliferation, but they also increased CD45RA expression in CD8+ T cells. In addition, I visualized interesting relationships between T cells and neutrophils and H. pylori infected neutrophils and extensive H. pylori growth in the presence of PMNs and T cells that provided clues to the mechanisms that might explain why CD45RA is upregulated in CD8+ T cells in the presence of H. pylori.
Details
- Title: Subtitle
- Mechanisms of neutrophil nuclear hypersegmentation and consequences of Helicobacter pylori induced subtype differentiation
- Creators
- Stephanie Lee Silva-Del Toro
- Contributors
- Lee-Ann Allen (Advisor)Jon Houtman (Committee Member)Steven Varga (Committee Member)Prajwal Gurung (Committee Member)Richard Roller (Committee Member)
- Resource Type
- Dissertation
- Degree Awarded
- Doctor of Philosophy (PhD), University of Iowa
- Degree in
- Immunology
- Date degree season
- Summer 2021
- DOI
- 10.17077/etd.005865
- Publisher
- University of Iowa
- Number of pages
- xix, 137 pages
- Copyright
- Copyright 2021 Stephanie Lee Silva-Del Toro
- Grant note
- Funding sources, National Institutes of Health, NIAID R01 AI119965 and VA Merit Review Grant 1I01BX002108 awarded to L-AHA as well as a National Science Foundation predoctoral fellowship #1546595 awarded to SLS-DT. In addition, I received a year of support from the Immunology training grant T32. Sloan Foundation also contributed to my training
- Comment
- This thesis has been optimized for improved web viewing. If you require the original version, contact the University Archives at the University of Iowa: https://www.lib.uiowa.edu/sc/contact/
- Language
- English
- Description illustrations
- color illustrations
- Description bibliographic
- Includes bibliographical references (pages 128-137).
- Public Abstract (ETD)
Neutrophils are the most abundant white blood cell in humans and the first responders to bacterial infections. Helicobacter pylori is a type of bacteria that infects half of the people in the world and causes chronic infection which can lead to stomach inflammation (gastritis), ulcers and even cancer. These serious diseases are mediated mainly by neutrophils. Thus, it is essential to understand how neutrophils respond to H. pylori infection to understand the processes that can lead to ulcers and cancer.
Neutrophil appearance, lifespan, and capability as part of the immune system change upon H. pylori infection, and this includes a very dramatic alteration of the nucleus of the cell. My project demonstrated that the nucleus of infected neutrophils doubles its volume upon infection and that the cell’s skeleton (microtubules) plays an important role in changing nuclear size and shape. Finally, I explored the consequences of H. pylori infected neutrophils when exposed to T cells. T cells are part of long-lasting immunity, and they can be controlled by neutrophils under certain conditions. Here I demonstrated that T cells cannot proliferate in the presence of H. pylori. The experiments and data shared in this dissertation will help understand host-pathogen interactions better and how neutrophils respond to H. pylori infection.
- Academic Unit
- Immunology Graduate Program
- Record Identifier
- 9984124269002771
Metrics
46 File views/ downloads
81 Record Views