Elevated eosinophil counts in the circulation and/or tissues is considered a clinical feature and biomarker of several chronic airway diseases including asthma. As such, many therapeutic biologics for asthma developed within the past decade target eosinophil recruitment to and accumulation in the airways to mixed success. Although the nature of adhesive interactions and directional migration of eosinophils has been well studied, there remains a lack of comprehensive understanding regarding the components which modulate eosinophil movement from the blood into respiratory tissues that impacts the efficacy of these clinical studies; therefore, continued research in this area may reveal novel therapeutic targets and ultimately improve clinical outcomes of patients with eosinophilia-mediated diseases.
The Janssen lab serendipitously discovered that the mere perfusion of standard media without pharmacological additives over human eosinophils in vitro induced the release of intracellular calcium (Ca2+) reminiscent of chemokine-induced Ca2+ release well documented in the literature. The central focus of my doctoral research was to characterize this novel phenomenon of the perfusion-induced calcium response (PICR), and to determine its physiological role in the eosinophil extravasation process to inflamed tissue sites.
In our first research objective, we optimized a protocol of eosinophil isolation directly from whole blood with emphases on maximizing population purity and yield efficiency while minimizing cell activation that could potentially interfere with secondary functional assays. For our latter two studies, we utilized real-time fluorescent confocal microscopy and immunofluorescence staining to investigate the PICR. We observed that the latency to the PICR post-perfusion was significantly shorter in eosinophils subjected to physiological rates of shear stress, suggesting a temporal-regulatory function of eosinophil mechanosensitivity. Furthermore, the disruption of the PICR via pharmacological inhibitors significantly reduced eosinophil motility by increasing the latency to cytoskeletal rearrangements (flattening onto substrate-coated surfaces, formation of membrane protrusions that explore the environment) necessary for cell migration out of the vasculature. Detailing the role of eosinophil sensitivity to the mechanical trigger of fluid shear stress expands upon the current paradigm of eosinophil recruitment and will contribute to the development of clinical strategies. / Dissertation / Candidate in Philosophy
| Identifer | oai:union.ndltd.org:mcmaster.ca/oai:macsphere.mcmaster.ca:11375/26328 |
| Date | January 2021 |
| Creators | Son, Kiho |
| Contributors | Janssen, Luke, Medical Sciences (Division of Physiology/Pharmacology) |
| Source Sets | McMaster University |
| Language | English |
| Detected Language | English |
| Type | Thesis |
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