Myometrial contraction is a central feature of labor. Although a link between infection and preterm labor is widely accepted, surprisingly little is known about the mechanisms coupling infection-induced inflammation to myocyte contractile machinery. This study explores the myocyte response to pathogen-derived ligands in vitro. The pregnant human myometrial cell line PHM1-41 and primary cultured uterine myocytes responded to Toll-like receptor (TLR) ligands, including the bacterial wall component LPS, which at 100 ng/ml increased contraction of cells embedded within collagen gels over 72 h compared to PBS. LPS-treated myocytes secreted inflammatory mediators, including prostaglandin F2α, the cytokines TNF-α and IL-6, and a range of chemokines. The contractile response to LPS required TLR4 signaling and was independent of prostaglandin synthesis. Neutralizing TNF-α had no effect on LPS-mediated contraction; however, the Rho-associated protein kinase (ROCK) inhibitors Y-27632 (10 μM) and GSK-269962 (50 nM) both abrogated the contractile response. The finding of LPS-mediated contraction was supported by a 1.38 ± 0.072-fold (mean±SE) increase in myosin light-chain phosphorylation 48 h post-treatment, assessed by in-cell Western blot analysis. Together, these data suggest that, in addition to modulating the local inflammatory environment, pathogen-derived ligands may directly promote myometrial contractility via Rho/ROCK signaling, thus contributing to preterm labor-mediated preterm birth.

Both term and preterm parturition are characterized by an influx of macrophages and neutrophils into the myometrium and cervix, with co-incident increased peripheral blood monocyte activation. Infection and inflammation are strongly implicated in the pathology of preterm labour (PTL), with progesterone considered a promising candidate for its prevention or treatment. In this study, we investigated the effect of monocytes on myometrial smooth muscle cell inflammatory cytokine production both alone and in response to LPS, a TLR4 agonist used to trigger PTL in vivo. We also investigated the effect of monocytes on myocyte contraction. Monocytes, isolated from peripheral blood samples from term pregnant women, were cultured alone, or co-cultured with PHM1-41 myometrial smooth muscle cells, for 24 h. In a third set of experiments, PHM1-41 myocytes were cultured for 24 h in isolation. Cytokine secretion was determined by ELISA or multiplex assays. Co-culture of monocytes and myocytes led to synergistic secretion of pro-inflammatory cytokines and chemokines including IL-6, IL-8 and MCP-1, with the secretion being further enhanced by LPS (100 ng/ml). The synergistic secretion of IL-6 and IL-8 from co-cultures was mediated in part by direct cell–cell contact, and by TNF. Conditioned media from co-cultures stimulated contraction of PHM1-41 myocytes, and the effect was inhibited by progesterone. Both progesterone and IL-10 inhibited LPS-stimulated IL-6 and IL-8 secretion from co-cultures, while progesterone also inhibited chemokine secretion. These data suggest that monocytes infiltrating the myometrium at labour participate in crosstalk that potentiates pro-inflammatory cytokine secretion, an effect that is enhanced by LPS, and can augment myocyte contraction. These effects are all partially inhibited by progesterone.