Determining the role of the glucocorticoid receptor in phagocyte pro-inflammatory responses.

About This Grant

Project Summary/Abstract Circulating levels of cortisol, the body's stress hormone, increase during stresses like infections, which then dampens immune responses preventing excessive tissue damage. Synthetic glucocorticoids that structurally mimic cortisol are potent immunosuppressive and anti-inflammatory drugs given to patients with inflammatory conditions and to prevent transplant rejection. Patients undergoing prolonged glucocorticoid therapies are prone to opportunistic infections. The discovery of a safe glucocorticoid therapy that is immunosuppressive yet immune-active enough for the host to fight opportunistic pathogens has been challenging due to our lack of understanding of glucocorticoid biology in immunity. While the anti-inflammatory mechanism of synthetic glucocorticoid activity is well-described, increasing evidence suggests immune-enhancing properties of these drugs. However, ii is not fully known if endogenous cortisol-glucocorticoid receptor signaling regulates pro-inflammatory mechanisms. Cortisol levels are tightly regulated in the body and within cells; in addition to circulating levels, cells can autonomously regulate intracellular levels of cortisol, thereby regulating glucocorticoid receptor-mediated signaling. The overarching goal of this proposal is to identify the mechanisms that endogenous glucocorticoid signaling regulates in macrophage and neutrophil responses to pathogens. The larval zebrafish is an ideal host to tackle these questions. Optical and genetic accessibility of zebrafish larvae allows for the investigation of molecular mechanisms modulating dynamic host-pathogen interactions in a live organism. Importantly, glucocorticoid receptor mutant zebrafish are viable. Using zebrafish larvae as a model, I propose to test how glucocorticoid receptor activation affects macrophage- and neutrophil-mediated clearance of fungal-derived zymosan particles. Our preliminary data suggests that glucocorticoid receptor mutation causes aberrant zymosan clearance by macrophages and neutrophils. In line with recent findings by others, this data suggests that endogenous glucocorticoid receptor signaling promotes pro-inflammatory responses. In the first aim, I will determine the cellular mechanisms affected by glucocorticoid receptor mutation that result in defective zymosan clearance. I will use CRISPR/Cas9 to generate glucocorticoid receptor mutant larvae and use high-resolution live imaging of injected larvae. In the second aim, I will use a Gal4/UAS inducible system to conditionally overexpress an enzyme to decrease cortisol levels, specifically in macrophages or neutrophils, to test how cell-specific glucocorticoid receptor activation affects zymosan clearance. Overall, the findings from these aims will broaden our understanding of how glucocorticoid receptor signaling regulates phagocyte response to pathogens. Long-term, this information will contribute to the development of novel glucocorticoid therapies with minimal risk for opportunistic infections.