n–3 PUFA dietary supplementation inhibits proliferation and store-operated calcium influx in thymoma cells growing in Balb/c mice

The antitumor effect of daily individual administration of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) (2 g/kg body weight) in Balb/c mice bearing a transplantable thymoma was investigated. Mice received oleic acid (control group), EPA and DHA ethyl esters starting 10 days before tumor inoculation. Analysis of phospholipid composition of neoplastic cell revealed that EPA and DHA levels were significantly increased (63 and 22% increase) after EPA and DHA treatments, respectively. Conversely, decreased levels of arachidonic acid were found in both cases (19 and 24% decrease in EPA and DHA groups, respectively). EPA and DHA delayed the appearance of macroscopic ascites (100% of animal, from 7 to 28 days), prolonged animal survival (100% of animal, from 22 to 32 and 33 days, respectively) and reduced the percentage of proliferating tumor cells detected by immunostaining of proliferation cell nuclear antigen (PCNA) (80 and 85% decrease, respectively). Moreover, the regulatory effects of these dietary n–3 fatty acids on the influx of Ca2+, activated by depletion of intracellular stores with thapsigargin (Tg), were investigated. By using a Ca2+-free/Ca2+-reintroduction protocol and Fura-2 as fluorescent indicator of intracellular free Ca2+([Ca2+]i), we observed that EPA and DHA treatments markedly decreased Tg-induced rise in [Ca2+]i (49 and 37% decrease, respectively). This effect was related to the inhibition of the store-operated Ca2+ influx, as confirmed also by Mn2+ influx experiments. The inhibitory action of EPA and DHA on the store-operated Ca2+ influx could explain, at least in part, their antitumoral activity, as this Ca2+ mobilization pathway appears to be involved in the cell signaling occurring in non-excitable cells to evoke many cellular processes, including cell proliferation. —Calviello, G., P. Palozza, F. Di Nicuolo, N. Maggiano, and G. M. Bartoli. N–3 PUFA dietary supplementation inhibits proliferation and store-operated calcium influx in thymoma cells growing in Balb/c mice

Synthetic PreImplantation Factor (sPIF) reduces inflammation and prevents preterm birth.

Preterm birth (PTB) is the leading cause of neonatal morbidity and mortality and spontaneous PTB is a major contributor. The preceding inflammation/infection contributes not only to spontaneous PTB but is associated with neonatal morbidities including impaired brain development. Therefore, control of exaggerated immune response during pregnancy is an attractive strategy. A potential candidate is synthetic PreImplantation Factor (sPIF) as sPIF prevents inflammatory induced fetal loss and has neuroprotective properties. Here, we tested maternal sPIF prophylaxis in pregnant mice subjected to a lipopolysaccharides (LPS) insult, which results in PTB. Additionally, we evaluated sPIF effects in placental and microglial cell lines. Maternal sPIF application reduced the LPS induced PTB rate significantly. Consequently, sPIF reduced microglial activation (Iba-1 positive cells) and preserved neuronal migration (Cux-2 positive cells) in fetal brains. In fetal brain lysates sPIF decreased IL-6 and INFγ concentrations. In-vitro, sPIF reduced Iba1 and TNFα expression in microglial cells and reduced the expression of pro-apoptotic (Bad and Bax) and inflammatory (IL-6 and NLRP4) genes in placental cell lines. Together, maternal sPIF prophylaxis prevents PTB in part by controlling exaggerated immune response. Given the sPIF`FDA Fast Track approval in non-pregnant subjects, we envision sPIF therapy in pregnancy

Identification of Novel Putative Urinary Markers of Endometriosis by High-Resolution Quantitative Proteomics

Endometriosis is a chronic gynecological inflammatory disease characterized by the presence of functional endometrial glands..

Synthetic PreImplantation Factor (PIF) prevents fetal loss by modulating LPS induced inflammatory response

Maternal control of inflammation is essential during pregnancy and an exaggerated response is one of the underlying causes of fetal loss. Inflammatory response is mediated by multiple factors and Toll-like receptors (TLRs) are central. Activation of TLRs results in NALP-3 mediated assembly of apoptosis-associated speck-like protein containing a CARD (ASC) and caspase-1 into the inflammasome and production of pro-inflammatory cytokines IL-1\u3b2 and IL-18. Given that preventing measures are lacking, we investigated PreImplantation Factor (PIF) as therapeutic option as PIF modulates Inflammation in pregnancy. Additionally, synthetic PIF (PIF analog) protects against multiple immune disorders. We used a LPS induced murine model of fetal loss and synthetic PIF reduced this fetal loss and increased the embryo weight significantly. We detected increased PIF expression in the placentae after LPS insult. The LPS induced serum and placenta cytokines were abolished by synthetic PIF treatment and importantly synthetic PIF modulated key members of inflammasome complex NALP-3, ASC, and caspase-1 as well. In conclusion our results indicate that synthetic PIF protects against LPS induced fetal loss, likely through modulation of inflammatory response especially the inflammasome complex. Given that synthetic PIF is currently tested in autoimmune diseases of non-pregnant subjects (clinicaltrials.gov, NCT02239562), therapeutic approach during pregnancy can be envisioned

Low-molecular-weight heparins induce decidual heparin-binding epidermal growth factor–like growth factor expression and promote survival of decidual cells undergoing apoptosis Nicoletta Di

Objective: To evaluate the effects of low-molecular-weight heparins (LMWHs) on decidual heparin-binding epidermal growth factor–like growth factor (HB-EGF) expression/secretion and on TNF-a–induced decidual apoptosis. Design: Experimental study. Setting: Department of Obstetrics and Gynecology, Universita Cattolica del Sacro Cuore, Rome, Italy. Patient(s): Cultures of primary decidual cells isolated from human term placenta. Intervention(s): The effects of LMWHs (tinzaparin and enoxaparin) on decidual HB-EGF expression and secretion were investigated by Western blot analysis and ELISA, respectively. TNF-a–induced decidual apoptosis was evaluated by annexin V staining, terminal deoxynucleotide transferase– mediated dUTP nick-end labeling (TUNEL) assay, and caspase activities. Main Outcome Measure(s): Decidual HB-EGF expression/secretion and apoptotic rate induced by TNF-a were investigated. Result(s): Tinzaparin enhanced decidual HB-EGF expression and secretion. TNF-a reduced the number of viable cells by inducing apoptosis. Simultaneous addition of LMWHs (primarily tinzaparin) blocked the increase in annexin V– and TUNEL-positive cells and reduced the amount of caspase activities. Conclusion(s): Both LMWHs induced a significant increase in decidual HB-EGF expression/secretion and reduced TNF-a–induced decidual apoptosis. Tinzaparin demonstrated higher efficacy. (Fertil Steril 2012;97:169–77. 2012 by American Society for Reproductiv

Effect of Low Molecular Weight Heparins (LMWHs) on antiphospholipid Antibodies (aPL)-mediated inhibition of endometrial angiogenesis

Antiphospholipid syndrome (APS) is an autoimmune disorder characterized by vascular thrombosis and/or pregnancy morbidity in the presence of circulating antiphospholipid antibodies (aPL). Different pathogenic mechanisms for aPL-mediated pregnancy failure have been proposed. In particular a direct effect of aPL on both maternal and fetal side of the placental tissue has been reported, since their reactivity with \u3b22-glycoprotein I (\u3b22GPI) makes them adhere to trophoblast and human endometrial endothelial cell (HEEC) membranes. \u3b22GPI can be recognized by aPL that, once bound, interfere with both trophoblast functions and with the HEEC differentiation.APS patients can be successfully treated with Low Molecular Weight Heparin (LMWH). Recent reports suggest that LMWH acts through mechanisms alternative to its well known anticoagulant effect, because of its ability to bind \u3b22GPI. In our previous studies, we showed that LMWH is able to reduce the aPL binding to trophoblasts and restore cell invasiveness and differentiation. So far, however, no study has described its effects on endometrial angiogenesis.The aim of our research was to evaluate whether two LMWHs, tinzaparin and enoxaparin, have an effect on the aPL-inhibited endometrial angiogenesis. This prompted us to investigate: (i) in vitro HEEC angiogenesis through a Matrigel assay; (ii) VEGF secretion by ELISA; (iii) matrix metalloproteinase-2 (MMP-2) activity by gelatin zymography; (iv) Nuclear Factor-\u3baB (NF-\u3baB) DNA binding activity by colorimetric assay; (v) STAT-3 activation by a sandwich-ELISA kit. Furthermore, using an in vivo murine model we investigated the LMWHs effects on angiogenesis.We demonstrated that the addition of LMWHs prevents aPL-inhibited HEEC angiogenesis, both in vitro and in vivo, and is able to restore the aPL inhibited NF-\u3baB and/or STAT-3 activity, the VEGF secretion and the MMPs activity.The demonstration of a beneficial role for LMWHs on the aPL-inhibited HEEC angiogenesis might provide additional mechanisms whereby this treatment protects early pregnancy in AP

Adipokines: New Emerging Roles in Fertility and Reproduction

Adipose tissue is a specialized endocrine and paracrine organ producing specific factors called adipokines. It is well known that adipokines balance is fundamental to prevent obesity, metabolic syndrome, and cardiovascular diseases. During the last years, new roles of adipokines have been emerging in the field of fertility and reproduction. Although the literature is still quite controversial, this review serves to resume current knowledge on this topic. Alterations in adipokine levels or in their mechanism of action are associated with fertility impairment and pregnancy diseases, as well as with obesity, metabolic syndrome, and cardiovascular diseases. Normal levels of adipokines are fundamental to maintain integrity of hypothalamus-pituitary-gonadal axis, regular ovulatory processes, successful embryo implantation, and physiologic pregnancy. More efforts are needed to understand the mechanisms and to the extent to which adipokine changes are involved in the impairment of fertility and pregnancy outcome, to find possible medical treatments. Target Audience: Obstetricians & Gynecologists, Family Physicians Learning Objectives: After completion of this educational activity, the obstetrician/gynecologist should be better able to demonstrate current knowledge in the research field of adipokines in fertility and reproduction; evaluate the central role of metabolism balance in good pregnancy outcome; and apply new perspectives of studies