7 Likewise, some miRNAs are found less expressed in choriocarcinoma cells than in normal trophoblast, which

suggests a role in carcinogenesis.8 We focused on five miRNAs previously published to correlate with tumor grade, to be implicated in pregnancy, or to be related with members of the signaling intracellular cascade of LIF. For instance, miR-141, belonging to the miR-200 cluster, is found upregulated in nasopharyngeal and ovarian carcinomas in comparison with normal tissues and correlates with poor prognosis.9,10 As biological marker, levels of miR-141 are increased in plasma from pregnant women.11 Also, expression check details of miR-9 may serve as a biomarker, which correlates with tumor grade and metastatic status in breast and cervical cancer.12,13 Its inhibition results in increased levels of phospho-STAT3 in embryonic stem cells.14 Among the miRNAs selected for the present investigation, to date, miR-21 is the most extensively studied. Because of its over-expression in at least six different solid cancers (lung, stomach, prostate, colon, pancreas, and

breast), it has been considered an oncomir (reviewed in15). MiR-21 can be induced by STAT3.7 Mir-93 seems to be related with the trophoblast response EPZ6438 to hypoxia as it is upregulated in hypoxic trophoblast cells.16 MiR-93 shares some features with miR-141 and miR-21 as they all are expressed in human embryonic stem cells, but their effects in cell maintenance or differentiation seem to be dissimilar. While miR-93 expression remains similar also in adult tissue, miR-141 attenuates differentiation and miR-21 expression intensifies it.17–20 Finally, we selected let-7g, a member of one of the currently most important miRNA families (let-7), which is aberrantly expressed in human cancer.21 Let-7g and also miR-21 were expressed in vitro as well as in vivo via STAT3 activation after IL-6 stimulation.22 Although the LIF-induced STAT3 activation in trophoblastic cells seems to be crucial for many cell functions, thus far, the LIF-induced miRNA expression in these cells has not yet been investigated.

Therefore, in the present study, we aim to analyze the kinetics of the expression Y-27632 2HCl of miR-9, miR-21, miR-93, miR-141, and let-7g after LIF treatment in JEG-3 cells. Being the most affected, influence of miR-141 on proliferation has been analyzed by its experimental over-expression and silencing. JEG-3 (DSMZ, Braunschweig, Germany) is an adherent human choriocarcinoma cell line preserving several trophoblast-like capacities including production of pregnancy-related hormones and cytokines. JEG-3 cells cultures were performed at 106 cells/175 cm2 flask and maintained under standard conditions (37°C, 5% CO2, humid atmosphere) in Ham’s F-12 Nutrient Mixture with l-glutamine (Gibco, Paisley, UK) supplemented with 10% heat-inactivated fetal calf serum (FCS; Gibco) and 1% penicillin/streptomycin antibiotic solution (Gibco).