Prostaglandin E2 receptors differentially regulate the output of proinflammatory cytokines in myometrial cells from term pregnant women

Abstract

Prostaglandin (PG) E₂ plays critical roles during pregnancy and parturition. Emerging evidence indicates that human labour is an inflammatory event. We sought to investigate the effect of PGE₂ on the output of proinflammatory cytokines in cultured human uterine smooth muscle cells (HUSMCs) from term pregnant women and elucidate the role of subtypes of PGE₂ receptors (EP₁, EP₂, EP₃ and EP₄). After drug treatment and/or transfection of each receptor siRNA, the concentrations of inflammatory secreting factors in HUSMCs culture medium were detected by the corresponding ELISA kits. The results showed that, PGE₂ increased interleukin 6 (IL-6) and tumor necrosis factor alpha (TNFα) output, decreased chemokine (c-x-c motif) ligand 8 (CXCL8) output in a dose-dependent manner, but had no effect on IL-1β and chemokine (c-c motif) ligand 2 (CCL-2) secretion of HUSMCs. EP₁/EP₃ agonist 17-phenyl-trinor-PGE₂ stimulated IL-6 and TNFα whilst suppressing IL-1β and CXCL8 output. The effects of 17-phenyl-trinor-PGE₂ on IL-1β and CXCL8 secretion were remained whereas its effect on IL-6 and TNFα output did not occur in the cells with EP₃ knockdown. The stimulatory effects of 17-phenyl-trinor-PGE₂ on IL-6 and TNFα were remained whereas the inhibitory effects of 17-phenyl-trinor-PGE₂ on IL-1β secretion was blocked in the cells with EP₁ knockdown. Either of EP₂ and EP₄ agonists stimulated IL-1β and TNFα output, which was reversed by EP₂ and EP₄ siRNA, respectively. The inhibitors of phospholipase C (PLC) and protein kinase C (PKC) blocked EP₁/EP₃ modulation of TNFα and CXCL8 output. PI3K inhibitor LY294002 and P38 inhibitor SB202190 blocked 17-phenyl-trinor-PGE₂-induced IL-1β and IL-6 output, respectively. The inhibitors of adenylyl cyclase and PKA prevented EP₂ and EP₄ stimulation of IL-1β and TNFα output, whereas PLC and PKC inhibitors blocked EP₂- and EP₄-induced TNFα output but not IL-1β output. Our data suggest that PGE₂ receptors exhibit different effects on the output of various cytokines in myometrium, which can subtly modulate the inflammatory microenvironment in myometrium during pregnancy.