Decidual Inflammation Drives Chemokine-Mediated Immune Infiltration Contributing to Term Labor.

Infiltration of maternal peripheral leukocytes into the uterine tissues is a critical event occurring before, during, and after term labor (TL). In this article, we investigate the contribution of uterine smooth muscle (myometrium) and pregnant endometrium (decidua) to the inflammatory process during human TL. We hypothesize that labor-related physiological inflammation is orchestrated by uterine-secreted cytokines, which dually activate the uterine vascular endothelium and maternal leukocytes to promote their adhesion and infiltration into the uterus. Using Luminex and ELISA assays, we examine a full range of cytokines (45 proteins) in media conditioned by primary decidual and myometrial cells from TL and term not in labor (TNL) women. The effect of conditioned media on the activation of human uterine microvascular endothelial cells was measured by qPCR and on peripheral leukocytes by flow cytometry. Transendothelial migration of calcein-labeled primary leukocytes toward media was assessed by fluorometry. Stromal decidual cells secrete significantly higher levels of multiple cytokines compared with myometrial cells (p < 0.05) and significantly more cytokines during TL than TNL. These cytokines activate uterine microvascular endothelial cells through the upregulation of cell adhesion molecule VCAM-1 and peripheral leukocytes by upregulation of CD11b. Furthermore, multiple cytokines secreted from the TL decidua and myometrium significantly increase migration of granulocytes, monocytes, and lymphocytes compared with TNL (p < 0.05), which was blocked by a broad-spectrum chemokine inhibitor (FX125L). These data reveal the critical role for decidual- and myometrial-secreted cytokines in the activation of inflammatory pathways leading to labor. We suggest that these pathways represent targets for therapeutic intervention during preterm labor.

Differential expression of myometrial AP-1 proteins during gestation and labour.

Preterm labour (PTL) is a leading cause of perinatal mortality and postnatal morbidity. Contractions of the uterine muscle (myometrium) that determine the onset of labour depend on the expression of contraction-associated proteins (CAPs, i.e. connexin43) regulated by dimeric AP-1 transcription factors. Here, we examined subcellular (by immunoblotting) and tissue expression (by immunohistochemistry) of myometrial AP-1 proteins (cJUN, JUNB, JUND, cFOS, FOSB, FRA1, FRA2) throughout gestation and TL in different species (mouse, rat and human). To identify the critical AP-1 members associated with preterm birth, we studied their expression in mouse model of 'infectious' (LPS-induced) and 'sterile' (RU486-induced) PTL. We found that (1) myometrial AP-1 composition is preserved in vivo between different species (rodents and human) indicating that Fos/Jun heterodimer (i.e. FRA2/JUND) may be indispensable for labour initiation. (2) Our in vivo study using murine models of gestation shows that there is a similarity in the myometrial AP-1 protein composition during TL and pathological PTL of different aetiology suggesting the involvement of similar molecular machinery in the induction of labour. (3) This study is first comprehensive protein analysis of seven AP-1 members in human labouring versus non-labouring myometrium, showing their cellular expression and tissue distribution in relation to labour status.

Isolation of Primary Human Decidual Cells from the Fetal Membranes of Term Placentae.

The decidua, also known as the pregnant endometrium, is a critically important reproductive tissue. Decidual cells, comprised mainly of decidualized stromal cells and immune cells, are responsible for the secretion of hormonal and inflammatory factors which are critical for successful blastocyst implantation, placental development and play a role in the initiation of labor at term and preterm. Many pregnancy complications can arise from perturbations of a fine balance of different cell populations comprising decidua. Alterations in the proportion of specific decidual cell types may disrupt these crucial processes and increase the risk of developing serious complications of pregnancy, such as embryo implantation failure, intrauterine growth restriction, preeclampsia and preterm labor. The protocol outlined here demonstrates a cost and time effective method for the isolation of primary human decidual cells collected from the fetal membranes of term placentae. By combining enzymatic digestion and gentle mechanical disruption of the decidual tissue, a high yield of decidual cells was obtained with virtually no chorion contamination. Importantly, isolated decidual cells were characterized (stromal cells (55-60%), leukocytes (35%), epithelial (1%) or trophoblast (0.01%) cells) and maintained high viability (80%) which was confirmed by multicolor imaging flow cytometry assay. This protocol is specific to the decidua parietalis and can be adapted to first and second trimester placentae. Once isolated, decidual cells can be used for a multitude of experimental applications aiming to understand the role of different decidual cell sub-populations in pregnancy complications.