Stat3 and Socs3 expression patterns during murine placenta development.

Signal transducers and activators of transcription 3 (Stat3) has been identified as an important signal transducer in the invasive phenotype of the trophoblasts cells in in vitro studies. However, the in situ distribution and patterns of expression of this molecule in trophoblast cells during the development of the placenta are still under-elucidated. Mice uteri of gestational ages between 7 and 14 days of pregnancy (dop) were fixed in methacarn and processed with immunoperoxidase techniques for detection of Stat3 and its phosphorylation at serine (p-ser727) residues, as well as the suppressor of cytokine signaling 3 (Socs3) expression. Stat3 was observed at 7 through 9 dop in both the antimesometrial and mesometrial deciduas, while continued immunoreactivity between 10 and 13 dop was seen only in the mesometrial decidua. In the placenta, Stat3 was detected in the cytotrophoblast cells of labyrinth and giant trophoblast cells between 10 and 14 dop. Immunoreactivity for Stat3 was also seen in trophoblast cells surrounding the maternal blood vessels. On days 10 and 11 of pregnancy, p-ser727 was detectable in the mesometrial decidua and in giant trophoblasts, while during 12-14 dop in the spongiotrophoblast region. In addition, Socs3 was immunodetected in maternal and placental tissues, principally in the giant trophoblast cells during the whole period of the study. The present in situ study shows the distribution of Stat3, its serine activation and Socs3 in different maternal and fetal compartments during murine placental development, thus further supporting the idea that they play a role during physiological placentation in mice. 

PP007. Effects of STAT1 suppression on ERK1/2 in trophoblastic cells.

INTRODUCTION: Migration and trophoblast invasion are controlled functionally along with the active participation of cytokines and growth factors. Two important intracellular signaling pathways are the Janus kinase/signal transducer and activator of transcription (JAK-STAT) and extracellular regulated kinase1/2 (ERK1/2). These pathways have been associated with the regulation of gene expression, cellular proliferation, differentiation, angiogenesis, embryo development and invasion in tumor and trophoblast cells. OBJECTIVES: The aim of our study is to characterize and analyze the regulation and crosstalks of STAT1 and ERK1/2 in trophoblast cells and the identification of activating cytokines. METHODS: The trophoblast derived cell line HTR-8/svneo and a choriocarcinoma cell line (JEG-3) were stimulated with interleukin-6 (IL-6), IL-11, granulocyte-macrophage colony-stimulating factor (GMC-SF), leukemia inhibitory factor (LIF) or oncostatine M (OSM). The the expression and phosphorylation of STAT1(tyr705) and ERK1/2 were analyzed by gel electrophoresis and Western blotting. Expression of STAT1 was inhibited by administration of 50µM fludarabine (2-fluoro-ara-AMP) for 2, 4, 8, 24, 48 or 72h or by using small interfering RNA (siRNA). The full activation of STAT1 was assessed by using an STAT1 DNA-binding assay. Finally, proliferation and invasion assays were performed (Grant Deutscher Akademischer Austausch Dienst A/10172477). RESULTS: LIF and OSM induce STAT1 and ERK1/2 phosphorylation in HTR-8 and JEG-3 cells. Fludarabine inhibits the so induced phosphorylation of STAT1 when administered 48 or 72h before stimulation. Simultaneously, ERK phosphorylation increases. In contrast, silencing of STAT1 by application of specific siRNA induces reduction of ERK1/2 phosphorylation. Fludarabine reduces STAT1 DNA-binding capacity. LIF and OSM increase proliferation. Silencing of STAT1 slightly decreases invasiveness of analyzed cells. CONCLUSION: STAT1 in trophoblast cells can be activated by placental cytokines. Suppression of STAT1 by fludarabine or siRNA influences activity of ERK1/2 which indicates a crosstalk between both pathways. Current studies will clarify the reason for the different effects on ERK1/2 in trophoblastic cells.

PP124. Trofoblastic invasion: The role of subunits STAT1 and STAT3.

INTRODUCTION: The trophoblastic migration/invasion are controlled by cytokines and growth factors that use intracellular pathways of signal to promote the regulation of gene expression, proliferation, cells differentiation, angiogenesis and embryonic development. The most important mediator of cytokine in trophoblastic invasion is the Janus-Kinase/signal transducer and activator of transcription (JAK/STAT). STATs are amino acids, compounds of 700-850 variable long-chain with isoforms α and ß and molecular weight between 83-113kDa. The role of these factors in the pregnancy set up may contribute to adopt interventions that could contribute to prophylaxis and/or treatment of abnormalities in the course of gestation when installed early. OBJECTIVES: Search on database the role of STAT in the process of trophoblastic invasion with emphasis on subunits STAT1 and STAT3. METHODS: This is a review performed on PubMed database. Have been included Studies found from 1992 (the year of discovery of STATs) until July 2011, without language restriction. The descriptors were: "Signal transducers and activators of transcription "and" Trophoblast". In the end we excluded bibliographical review. RESULTS: Five of the six selected papers studied the role of STAT3 in the physiology of the trophoblastc invasion process. One of them, indirectly by selection process of lactobacilli of vaginal flora endogenous, during change of vaginal pH on pregnancy, altering the release of greater or lesser number of Interleukin-10 which modulates the activation JAK/STAT. Among them, one of the study refers to involvement of STAT1 in the immunomodulation of interface fetus-mother. CONCLUSION: STAT3 is directly involved in the process of trophoblast invasion either in its endometrium adherence to, angiogenesis, invasion and regulation of invasion. And STAT1 is involved in immunomodulation through its suppression by trophoblast STAT utron. Several soluble factors that are generally present in the decidua, especially hepatocyte growth factor, granulocyte macrophagocytic-colony stimulating factors, interleukin-6, interleukin-11 and inhibition leukemia factor , which have been described by using the JAK-STAT activating STAT1 and STAT3 for intracellular signaling and from this process may influence the invasion trophoblast.

Interleukin regulation of asymmetric antibody synthesized by isolated placental B cells.

PROBLEM: Protecting antibodies against trophoblast surface molecules were previously described. Here we analysed the synthesis of asymmetric IgG by placental B-lymphocytes. METHOD OF STUDY: B cells were isolated from human term placenta and cord blood, stimulated with anti-CD40 IgG and cocultured with transfected Fcgamma R-expressing mice Ltk-fibroblast. Interleukin-4, IL-6, IL-10, IL-11 and IL-13 were added to cultures for 14 days. Asymmetric IgG were assessed in culture supernatants by concanavalin A (Con A) fixation and enzyme-linked immunosorbent assay. RESULTS: When IL-6 was added to the cultures, the percentages of asymmetric IgG synthesized by placental B cells were: IL-6: 29 +/- 10; IL-6 + IL-10: 24 +/- 7; IL-4 + IL-10 + IL-6: 38 +/- 9. The last combination induced the highest increase in the asymmetric IgG synthesis as compared with control (19 +/- 10%, P < 0.05). Additionally, placental B cells synthesized more asymmetric IgG than umbilical cord blood B-lymphocytes (P = 0.0015). CONCLUSIONS: Isolated placental B-lymphocytes synthesized asymmetric IgG in response to Th2 interleukins, more notably IL-6 in combination with IL-4 and IL-10. The in vitro increase of protective asymmetric IgG synthesis in response to Th2-cytokines support the hypothesis that a local Th2-switch is beneficial for pregnancy outcome.