Validation and application of a multiplex immunoassay for the detection of PGE2 induced inflammatory mediators of human decidual macrophages (92.15)

Abstract

Abstract A multiplex bead immunoassay was developed for the simultaneous quantitation of 8 inflammatory mediators including 6 cytokines (IL-1 β, IL-4, IL-6, IL-8, TNFα and IFNγ) and 2 eicosanoids (PGE2, and TXB2). The sensitivities of the cytokine assays are less than 5 pg/mL with the exception of IL-4 (19.9 pg/mL). The sensitivity of the PGE2 and TXB2 assays at 80% B/Bo are 64.2 and 65.9 pg/mL for PGE2 and TXB2 respectively. The antimicrobial actions of human decidual macrophages (DMs) isolated from the upper reproductive tract were characterized in the assay. Decidual macrophages are innate immune cells known to defend the upper reproductive tract from infection. PGE2 is a potent regulator of innate immunity in the reproductive tract during pregnancy and/or in response to infection. CD14+ DMs were incubated with bacteria, peptidoglycan or LPS and analyzed for the expression of the inflammatory mediators detected in the MultiBead(tm) multiplex immunoassay. The DMs produced PGE2 in response to bacteria, peptidoglycan or LPS with increased production of IL-8, TNFα, IL-6 and IL-1 β. The addition of 1 μM PGE2 decreased expression of TNFα and IL-1β and increased the expression of IL-6. The DMs produced PGE2 in response to inflammatory stimuli and modulated the expression of cytokines and eicosanoids in vitro. We conclude that PGE2 is involved in immunomodulation that may be relevant when PGE2 levels are increased during pregnancy and infection.