Date of Award
Master of Science (MS)
Gout is a chronic inflammatory disease caused by the phagocytosis of monosodium urate monohydrate (MSU) crystals by monocytes/macrophages resulting in downstream expression and production of interleukin-1 beta (IL-1β) and chemokines. The activation of monocytes by MSU crystals involves the priming of monocytes with danger signals e.g. lipopolysaccharide (LPS) or soluble uric acid (UA), crystal phagocytosis and subsequent NLRP3 inflammasome activation and conversion of pro-IL-1β to active IL-1β. Protein-phosphatase-2A (PP2A) is a serine/threonine phosphatase that plays an important role in cell growth and inflammation. The prodrug Fingolimod (FTY720) and its phosphorylated active metabolite (p-FTY720) activate intracellular PP2A. We hypothesized that monocyte activation by MSU crystals is mediated by a reduction in intracellular PP2A activity and restoring PP2A activity reduces MSU-induced inflammation in monocytes. We aimed to investigate the role of PP2A in regulating monocyte priming and activation by MSU crystals and evaluate whether intracellular PP2A activation exerts an anti-inflammatory effect in MSU-stimulated monocytes. Human THP-1 monocytes were primed with a combination of UA and LPS. MSU stimulation was performed for 4-6 hours and MSU crystal phagocytosis, PP2A activity, IL-1β expression and production were studied in primed and unprimed monocytes. We performed PP2A knockdown in THP-1 monocytes and evaluated the impact of PP2A attenuation on IL-1β expression and production in unprimed THP-1 monocytes. Time-dependent intracellular PP2A activation in response to FTY720 or p-FTY720 treatments was studied and we evaluated the impact of p-FTY720 treatment on IL-1β expression and production in MSU stimulated human monocytes. Priming with UA+LPS increased MSU phagocytosis and IL-1β expression and production in monocytes. This effect was associated with a reduction in intracellular PP2A activity. PP2A knockdown increased IL-1β expression and production. FTY720 and p-FTY720 increased intracellular PP2A activity in monocytes. p-FTY720 treatment reduced IL-1β expression and production in UA+LPS pre-treated monocytes following MSU stimulation mediated by an increase in PP2A activity with no alteration in PP2A gene expression. In summary, UA and LPS enhanced MSU phagocytosis, expression and production of IL-1β via a reduction in PP2A activity. Pharmacological restoration of PP2A activity exerted an antiinflammatory effect. We conclude that PP2A is a novel therapeutic target for gout treatment.
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ElSayed, S. Role of Protein Phosphatase-2A in Regulating Monocyte Activation by Soluble and Crystalline Uric Acid in Gout. [master’s thesis]. Irvine, CA: Chapman University; https://doi.org/10.36837/chapman.000150