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Meleady, Laura

University of British Columbia

As the innate immune cell of the central nervous system, microglia play critical roles in the development and homeostasis of the brain. Microglia are active surveyors of their microenvironment and respond to pathogens through release of pro-inflammatory cytokines, phagocytosis of diseased or dying cells, and release of reactive nitrogen species. How microglia dynamically shift patterns of gene expression to respond to environmental cues is regulated by epigenetic mechanisms including the transcriptional activating modification histone acetylation. Histone acetylation disrupts chromatin contacts to increase accessibility of DNA to transcription factors and machinery at genomic regulatory regions including promoters (marked by H3K9ac) and enhancers (marked by H3K27ac). Dynamic regulation of histone acetylation is performed by ‘writer’ histone acetyltransferase (HAT) and ‘eraser’ histone deacetylase (HDAC) enzymes. Growing evidence supports therapeutic benefits of HDAC inhibition on dampening inflammatory processes, including Hdac3 inhibition in peripheral macrophages. This research aims to interrogate a possible immunomodulatory role of Hdac3 using an immortalized mouse microglial model through changes in global and promoter specific histone acetylation, gene expression, and microglia-relevant functions. To test this hypothesis, we utilized the BV2 microglia cell line combined with pre-treatment of a Hdac3-specific inhibitor (RGFP966) or pan-Hdac inhibitor (SAHA) followed by an immune stimulation of lipopolysaccharide (LPS). BV2 microglia treated with increasing durations of LPS (1, 3, 6, 24 hours) showed increased global histone acetylation of both H3K27 and H3K9, as well as increased expression of pro-inflammatory cytokines Il-6, Tnfa, Il-1b. Hdac inhibition of BV2 microglia resulted in increased global H3K27ac, indicating successful inhibition of Hdac activity. Furthermore, Hdac3-specific inhibition induced baseline increases in expression of pro-inflammatory cytokines Il-1b and Tnfa, as well as anti-inflammatory gene targets Cxcl16, Il-10, Arg1, and Tlr4. A subset of these genes were hyper-induced by Hdac3 inhibition after LPS stimulation. Functionally Hdac3-inhibition reduced LPS-induced nitric oxide production, suggesting an anti-inflammatory polarization of the microglia. Additionally, at 24 hours Hdac3 inhibition induced phagocytosis to similar levels as LPS treated microglia. Together, these results affirm Hdac3 as a valuable target for microglia immunomodulation, with possible therapeutic impacts such as dampening the release of neuroinflammatory reactive nitrogen species.

Text

2023-06-30

Attribution-NonCommercial-NoDerivatives 4.0 International

http://hdl.handle.net/2429/81763

Biochemistry and Molecular Biology

University of British Columbia

UBCV

http://creativecommons.org/licenses/by-nc-nd/4.0/