Research Abstract
Activation of human monocytes with a yeast-derived pathogen associated molecular pattern, β-glucan
Nandita Bose, Megan L. Griggs, Lindsay R. Wurst, Christine M. Dudney, Michael E. Danielson, Nathaniel T. Theoharis, Richard M. Walsh, Myra L. Patchen and John P. Vasilakos, Biothera, Eagan, MN, USA, 55121.
2008 Keystone Symposia’s conference on Innate Immunity: Signaling Mechanisms.
β-glucan, an abundant polysaccharide in fungal pathogens, serves as a pathogen-associated molecular pattern (PAMP) recognized by a variety of immune cell pattern recognition receptors. Imprime PGGTM (Imprime PGG) is a soluble, purified, yeast β 1,3/1,6 glucan with broad immunomodulatory effects, including activation of innate immune functions such as oxidative burst activity. Many previous studies have focused on mouse macrophages or human neutrophils and implicated different receptors in mediating β-glucan-induced oxidative burst activity, including Dectin-1, CR3, and lactosylceramide. These studies have used soluble as well as particulate β-glucan preparations, some even containing impurities such as TLR2 agonists. Thus, lack of a purified, soluble β-glucan reagent has confounded deciphering the mechanism of β-glucan-induced oxidative burst. Furthermore, few studies have evaluated the mechanism of purified β-glucan-induced oxidative burst in human peripheral blood mononuclear cells (PBMC), a repertoire of innate immune effector cells expressing the reported β-glucan receptors. Here we have characterized the oxidative burst induced by β-glucan in human PBMC and evaluated the receptors and signaling mechanisms involved in this activity. Human PBMC were stimulated with various concentrations of Imprime PGG, and oxidative burst activity was measured using a cytochrome c assay. Inhibition of the oxidative burst reaction by superoxide dismutase (SOD) showed that the primary reactive oxygen species (ROS) measured was superoxide anion (O2-). Monocyte depletion from the PBMC significantly reduced O2-production, indicating that monocytes were the key players in producing the oxidative burst in response to Imprime PGG stimulation. Alteration of oxidative burst activity following blocking of various putative β-glucan receptors, and after inhibition of putative β-glucan signaling pathways, such as Syk, PI3K, Src family kinases, and Akt kinase will also be presented.