Regulation of sPLA2-IID in Human Decidua: Insights Into the Complexity of the Prostaglandin Pathway in Labor.

Prostaglandins are implicated in the labor process, yet the precise role and regulation of the prostaglandin pathway remains to be elucidated. The first step in the pathway is cleavage of membrane phospholipids by phospholipase A2 (PLA2). Previous work demonstrated upregulation of secretory PLA2 (sPLA2)-IIA with labor in human myometrium, and recent evidence shows that there are numerous PLA2 isoforms. The present study investigates the potential of additional sPLA2 isoforms during pregnancy and labor. Real-time reverse transcriptase-polymerase chain reaction (RT-PCR), Western blotting, and immunohistochemistry were used to determine sPLA2 expression and localization. Results show the presence of sPLA2-IID in amnion, chorion, placenta, decidua, and myometrium. Expression of sPLA2-IID in decidua was significantly decreased in term labor compared to nonlabor patients, whereas no significant labor-associated changes were observed in other gestational tissues. Secretory PLA2-IID was localized within chorion fibroblasts, placenta trophoblasts, decidual cells, and in myometrial smooth muscle cells. In primary decidual cell cultures, interleukin (IL) 10 (IL-10) increased sPLA2-IID messenger RNA (mRNA) expression, while IL-1ß had no effect on sPLA2-IID mRNA expression. In conclusion, decreased expression of sPLA2-IID in the decidua at labor indicates that it is unlikely to contribute to increased prostaglandin production during labor. However, increased expression of sPLA2-IID, induced by IL-10, suggests that sPLA2-IID may play an important anti-inflammatory role at the maternal-fetal interface. Nevertheless, precise functions of sPLA2-IID within the human uterus remain to be determined.

Development and validation of primary human myometrial cell culture models to study pregnancy and labour.

BACKGROUND: The development of the in vitro cell culture model has greatly facilitated the ability to study gene expression and regulation within human tissues. Within the human uterus, the upper (fundal) segment and the lower segment may provide distinct functions throughout pregnancy and during labour. We have established primary cultured human myometrial cells, isolated from both upper and lower segment regions of the pregnant human uterus, and validated them for the purpose of studying human pregnancy and labour. The specific objectives of this study were to monitor the viability and characterize the expression profile using selected cellular, contractile and pregnancy associated markers in the primary cultured human myometrial cells. Labour has been described as an inflammatory process; therefore, the ability of these cells to respond to an inflammatory stimulus was also investigated. METHODS: Myometrial cells isolated from paired upper segment (US) and lower segment (LS) biopsies, obtained from women undergoing Caesarean section deliveries at term prior to the onset of labour, were used to identify expression of; α smooth muscle actin, calponin, caldesmon, connexin 43, cyclo-oxygenase-2 (COX-2), oxytocin receptor, tropomyosin and vimentin, by RT-PCR and/or immunocytochemistry. Interleukin (IL)-1ß was used to treat cells, subsequently expression of COX-2 mRNA and release of interleukin-8 (CXCL8), were measured. ANOVA followed by Bonferroni's multiple comparisons test was performed. RESULTS: We demonstrate that US and LS human myometrial cells stably express all markers examined to at least passage ten (p10). Connexin 43, COX-2 and vimentin mRNA expression were significantly higher in LS cells compared to US cells. Both cell populations respond to IL-1ß, demonstrated by a robust release of CXCL8 and increased expression of COX-2 mRNA from passage one (p1) through to p10. CONCLUSIONS: Isolated primary myometrial cells maintain expression of smooth muscle and pregnancy-associated markers and retain their ability to respond to an inflammatory stimulus. These distinct myometrial cell models will provide a useful tool to investigate mechanisms underlying the process of human labour and the concept of functional regionalization of the pregnant uterus.

Prostaglandin E2 represses interleukin 1 beta-induced inflammatory mediator output from pregnant human myometrial cells through the EP2 and EP4 receptors.

Inflammatory mediators, including prostaglandins, cytokines, and chemokines, are strongly implicated in the mechanism of human labor, though their precise roles remain unknown. Here we demonstrate that interleukin 1 beta (IL-1beta) significantly increased the expression and release of interleukin-8 (CXCL8), monocyte chemotactic protein-1 (CCL2), and granulocyte macrophage colony-stimulating factor (CSF2) by primary human myometrial cells. However, this effect was repressed by prostaglandin E(2) (PGE(2)). As PGE(2) can activate four distinct PGE(2) receptors (EP(1), EP(2), EP(3), and EP(4)) to elicit various responses, we sought to define the EP receptor(s) responsible for this repression. Using selective EP receptor agonists and a selective EP(4) antagonist, we show that PGE(2) mediates the repression of IL-1beta-induced release of CXCL8, CCL2, and CSF2 via activation of the EP(2) and EP(4) receptors. The use of siRNA gene-specific knockdown further confirmed a role for both receptors. Real-time RT-PCR demonstrated that EP(2) was the most highly expressed of all four EP receptors at the mRNA level in human myometrial cells, and immunocytochemistry showed that EP(2) protein is abundantly present throughout the cells. Interestingly, PGE(2) does not appear to reduce mRNA expression of CXCL8, CCL2, and CSF2. Our results demonstrate that PGE(2) can elicit anti-inflammatory responses via activation of the EP(2) and EP(4) receptors in lower segment term pregnant human myometrial cells. Further elucidation of the EP receptor-mediated signaling pathways in the pregnant human uterus may be beneficial for optimizing the maintenance of pregnancy, induction of labor or indeed treatment of preterm labor.

Knowledge translation: principles and practicalities for trainees within interdisciplinary health research teams.

Within a dynamic health research environment with trends toward increasing accountability, governments and funding agencies have placed increased emphasis on knowledge translation (KT) as a way to optimize the impact of research investments on health outcomes, research products and health service delivery. As a result, there is an increasing need for familiarity with the principles of KT frameworks and components of KT strategies. Accordingly, health research trainees (graduate students and post-doctoral fellows) must be supported to enhance their capacity to understand KT principles and the practicalities of implementing effective KT practices.In this paper, the unique opportunities and challenges that trainees within an interdisciplinary research team encounter when they begin to understand and apply constructive and relevant KT practices are considered. Our commentary is based on trainee experiences within the Preterm Birth and Healthy Outcomes Team (PreHOT), an interdisciplinary research team.