The selective progesterone receptor modulator-promegestone-delays term parturition and prevents systemic inflammation-mediated preterm birth in mice.

BACKGROUND: Progesterone, acting via its nuclear receptors called progesterone receptors, promotes myometrial relaxation during pregnancy, and suspension of this activity triggers labor. We previously found that 20α-hydroxysteroid dehydrogenase causes a local withdrawal of progesterone in the term and preterm myometrium by converting the progesterone into an inactive form before it accesses the progesterone receptors. OBJECTIVE: We hypothesized that a selective progesterone receptor modulator called promegestone, which is not metabolized by 20α-hydroxysteroid dehydrogenase, would sustain progesterone receptor signaling and prevent/delay term labor and preterm labor in mice. STUDY DESIGN: In the term labor mouse model, promegestone (0.2 mg/dam) or a vehicle were administered subcutaneously in timed-pregnant CD-1 mice at gestational days 15, 16, and 17 (term gestational days, 19.5). In the inflammation preterm labor model, pregnant mice received promegestone or a vehicle on gestational days 15, 16, and 17, which was 24 hours before, immediately before, and 24 hours after systemic bacterial endotoxin (50 µg intraperitoneal; lipopolysaccharide group) or vehicle (saline) administration. The maternal and fetal tissues were collected on gestational day 16 6 hours after lipopolysaccharide±promegestone injection and at term gestational day 18.75. The protein levels of 10 cytokines were measured by multiplex immunoassay in maternal plasma and amniotic fluid. Myometrial, decidual, and placental messenger RNA levels of multiple cytokines and procontractile proteins were evaluated by real-time polymerase chain reaction and confirmed by immunoblotting. RESULTS: Promegestone prevented term labor and maintained mice pregnancy postterm >24 hours. The litter size and fetal weights were not different from the controls. Promegestone prevented systemic bacterial-endotoxin-induced preterm labor in 100% of the mice, blocked uterine contractions, significantly inhibited all systemic inflammation-induced myometrial cytokines, and partially inhibited decidual and placental inflammation. Promegestone did not prevent bacterial-endotoxin-induced fetal toxicity. CONCLUSION: Promegestone a selective progesterone receptor modulator that binds progesterone receptors with high affinity and is not metabolized by 20α-hydroxysteroid dehydrogenase could completely suppress term parturition and systemic bacterial-endotoxin-induced preterm birth in mice. We suggest that such selective progesterone receptor modulators may represent a potential therapeutic approach to the prevention of preterm labor in women at high risk of preterm birth.

Pro-inflammatory signals induce 20α-HSD expression in myometrial cells: A key mechanism for local progesterone withdrawal.

Metabolism of progesterone (P4) by the enzyme 20α hydroxysteroid dehydrogenase (20α-HSD) in myometrial cells is postulated to be a mechanism for P4 withdrawal, which occurs concomitant to uterine inflammation (physiologic or infection-induced) and associated activation of transcription factors: NF-кB and AP-1, common to term and preterm labour. We found that 20α-HSD protein is significantly increased in human myometrium during term labour, and in mouse uterus during term and preterm labour. Treatment of human myometrial cells with the pro-inflammatory mediators, lipopolysaccharide (LPS, mimicking infection) and 12-O-tetradecanoylphorbol-13-acetate (TPA, mimicking inflammation), induced 20α-HSD gene expression and increased 20α-HSD protein abundance. LPS treatment decreased P4 release into the culture medium and resulted in up-regulation of GJA1 in the hTERT-HM cells. The NF-кB /AP-1 transcription factors mediated effects of LPS and TPA on 20α-HSD gene transcription. Both pro-inflammatory stimuli induced 20α-HSD promoter activity in LPS/TPA-treated cells which was significantly attenuated by inhibition of NF-кB (JSH: 20 µM) or AP-1 signalling (T5224: 10 µM). Deletion of NF-кB consensus sites abrogated LPS-mediated promoter induction, while removal of AP-1 sites reversed the TPA-mediated induction of 20α-HSD promoter. We conclude that inflammatory stimuli (physiologic or pathologic) that activate NF-кB or AP-1 induce 20α-HSD transcription and subsequent local P4 withdrawal resulting in up-regulation of GJA1 and activation of myometrium that precedes labour.

Expression of severe acute respiratory syndrome coronavirus 2 cell entry genes, angiotensin-converting enzyme 2 and transmembrane protease serine 2, in the placenta across gestation and at the maternal-fetal interface in pregnancies complicated by preterm birth or preeclampsia.

BACKGROUND: Although there is some evidence that severe acute respiratory syndrome coronavirus 2 can invade the human placenta, limited data exist on the gestational age-dependent expression profile of the severe acute respiratory syndrome coronavirus 2 cell entry mediators, angiotensin-converting enzyme 2 and transmembrane protease serine 2, at the human maternal-fetal interface. There is also no information as to whether the expression of these mediators is altered in pregnancies complicated by preeclampsia or preterm birth. This is important because the expression of decidual and placental angiotensin-converting enzyme 2 and transmembrane protease serine 2 across gestation may affect the susceptibility of pregnancies to vertical transmission of severe acute respiratory syndrome coronavirus 2. OBJECTIVE: This study aimed to investigate the expression pattern of specific severe acute respiratory syndrome coronavirus 2 cell entry genes, angiotensin-converting enzyme 2 and transmembrane protease serine 2, in the placenta across human pregnancy and in paired samples of decidua and placenta in pregnancies complicated by preterm birth or preeclampsia compared with those in term uncomplicated pregnancies. STUDY DESIGN: In this study, 2 separate cohorts of patients, totaling 87 pregnancies, were included. The first cohort was composed of placentae from first- (7-9 weeks), second- (16-18 weeks), and third-trimester preterm (26-31 weeks) and third-trimester term (38-41 weeks) pregnancies (n=5/group), whereas the second independent cohort included matched decidua and placentae from pregnancies from term uncomplicated pregnancies (37-41 weeks' gestation; n=14) and pregnancies complicated by preterm birth (26-37 weeks' gestation; n=11) or preeclampsia (25-37 weeks' gestation; n=42). Samples were subjected to quantitative polymerase chain reaction and next-generation sequencing or RNA sequencing for next-generation RNA sequencing for angiotensin-converting enzyme 2 and transmembrane protease serine 2 mRNA expression quantification, respectively. RESULTS: In the first cohort, angiotensin-converting enzyme 2 and transmembrane protease serine 2, exhibited a gestational age-dependent expression profile, that is, angiotensin-converting enzyme 2 and transmembrane protease serine 2 mRNA was higher (P<.05) in the first-trimester placenta than in second-trimester, preterm birth, and term placentae (P<.05) and exhibited a negative correlation with gestational age (P<.05). In the second cohort, RNA sequencing demonstrated very low or undetectable expression levels of angiotensin-converting enzyme 2 in preterm birth, preeclampsia, and term decidua and in placentae from late gestation. In contrast, transmembrane protease serine 2 was expressed in both decidual and placental samples but did not change in pregnancies complicated by either preterm birth or preeclampsia. CONCLUSION: The increased expression of these severe acute respiratory syndrome coronavirus 2 cell entry-associated genes in the placenta in the first trimester of pregnancy compared with those in later stages of pregnancy suggests the possibility of differential susceptibility to placental entry to severe acute respiratory syndrome coronavirus 2 across pregnancy. Even though there is some evidence of increased rates of preterm birth associated with severe acute respiratory syndrome coronavirus 2 infection, we found no increase in mRNA expression of angiotensin-converting enzyme 2 or transmembrane protease serine 2 at the maternal-fetal interface.

Overexpression of miR-210-3p Impairs Extravillous Trophoblast Functions Associated with Uterine Spiral Artery Remodeling.

Hsa-miR-210-3p has been reported to be upregulated in preeclampsia (PE); however, the functions of miR-210-3p in placental development are not fully understood, and, consequently, miR-210-3p's role in the pathogenesis of PE is still under investigation. In this study, we found that overexpression of miR-210-3p reduced trophoblast migration and invasion, extravillous trophoblast (EVT) outgrowth in first trimester explants, expression of endovascular trophoblast (enEVT) markers and the ability of trophoblast to form endothelial-like networks. In addition, miR-210-3p overexpression significantly downregulated the mRNA levels of interleukin-1B and -8, as well as CXC motif ligand 1. These cytokines have been suggested to play a role in EVT invasion and the recruitment of immune cells to the spiral artery remodeling sites. We also showed that caudal-related homeobox transcription factor 2 (CDX2) is targeted by miR-210-3p and that CDX2 downregulation mimicked the observed effects of miR-210-3p upregulation in trophoblasts. These findings suggest that miR-210-3p may play a role in regulating events associated with enEVT functions and its overexpression could impair spiral artery remodeling, thereby contributing to PE.

Low molecular weight heparin promotes transcription and release of placental growth factor from endothelial cells.

Circulating levels of placental growth factor (PlGF) are significantly reduced in women who develop preeclampsia. Low molecular weight heparin (LMWH) has been shown to acutely elevate circulating PlGF levels in pregnant women at increased risk of preeclampsia. The objective of the current investigation was to determine the mechanisms by which LMWH mediates the extracellular release of PlGF from endothelial cells. Cultured human aortic endothelial cells (HAECs) and human umbilical vein endothelial cells (HUVECs) were exposed to LMWH; PlGF transcription, translation, mobilization, and secretion were then assessed. LMWH significantly increased the release of PlGF from both HAECs and HUVECs. LMWH treatment promoted a significant increase of PlGF-1 mRNA expression in HAECs, accompanied by the intracellular transport and release of PlGF into the conditioned media. LMWH-mediated release of PlGF from HAECs was not directly mediated by extracellular mobilization, synthesis, or stability of PlGF mRNA/protein. LMWH exposure promotes the release of PlGF from endothelial cells through the upregulation of PlGF-1 mRNA expression. Stimulation of circulating PlGF levels by LMWH may be an important mechanism by which LMWH could reduce the risk of preeclampsia or minimize disease severity.NEW & NOTEWORTHY There are few therapeutic options available for the prevention of preeclampsia, a serious hypertensive disorder of pregnancy. Women who subsequently develop preeclampsia exhibit significantly reduced circulating levels of the proangiogenic placental growth factor protein. Low molecular weight heparin (LMWH) has previously been investigated as a preventative therapy against the development of preeclampsia; however, its mechanism of action is not known. The current study determined that LMWH promotes the transcription and release of placental growth factor protein from endothelial cells, providing a mechanistic basis by which LMWH could reduce the risk of preeclampsia or minimize disease severity.

MicroRNA-218-5p Promotes Endovascular Trophoblast Differentiation and Spiral Artery Remodeling.

Preeclampsia (PE) is the leading cause of maternal and neonatal morbidity and mortality. Defects in trophoblast invasion, differentiation of endovascular extravillous trophoblasts (enEVTs), and spiral artery remodeling are key factors in PE development. There are no markers clinically available to predict PE, leaving expedited delivery as the only effective therapy. Dysregulation of miRNA in clinical tissues and maternal circulation have opened a new avenue for biomarker discovery. In this study, we investigated the role of miR-218-5p in PE development. miR-218-5p was highly expressed in EVTs and significantly downregulated in PE placentas. Using first-trimester trophoblast cell lines and human placental explants, we found that miR-218-5p overexpression promoted, whereas anti-miR-218-5p suppressed, trophoblast invasion, EVT outgrowth, and enEVT differentiation. Furthermore, miR-218-5p accelerated spiral artery remodeling in a decidua-placenta co-culture. The effect of miR-218-5p was mediated by the suppression of transforming growth factor (TGF)-ß2 signaling. Silencing of TGFB2 mimicked, whereas treatment with TGF-ß2 partially reversed, the effects of miR-218-5p. Taken together, these findings demonstrate that miR-218-5p promotes trophoblast invasion and enEVT differentiation through a novel miR-218-5p-TGF-ß2 pathway. This study elucidates the role of an miRNA in enEVT differentiation and spiral artery remodeling and suggests that downregulation of miR-218-5p contributes to PE development.

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.

Glucocorticoids modulate multidrug resistance transporters in the first trimester human placenta.

The placental multidrug transporters, P-glycoprotein (P-gp, encoded by ABCB1) and breast cancer resistance protein (BCRP, ABCG2) protect the foetus from exposure to maternally derived glucocorticoids, toxins and xenobiotics. During pregnancy, maternal glucocorticoid levels can be elevated by stress or exogenous administration. We hypothesized that glucocorticoids modulate the expression of ABCB1/P-gp and ABCG2/BCRP in the first trimester human placenta. Our objective was to examine whether dexamethasone (DEX) or cortisol modulate first trimester placental expression of multidrug transporters and determine whether cytotrophoblasts or the syncytiotrophoblast are/is responsible for mediating these effects. Three models were examined: (i) an ex-vivo model of placental villous explants (7-10 weeks), (ii) a model of isolated first trimester syncytiotrophoblast and cytotrophoblast cells and (iii) the BeWo immortalized trophoblast cell line model. These cells/tissues were treated with DEX or cortisol for 24 hour to 72 hour. In first trimester placental explants, DEX (48 hour) increased ABCB1 (P < .001) and ABCG2 (P < .05) mRNA levels, whereas cortisol (48 hour) only increased ABCB1 mRNA levels (P < .01). Dexamethasone (P < .05) and cortisol (P < .01) increased BCRP but did not affect P-gp protein levels. Breast cancer resistance protein expression was primarily confined to syncytiotrophoblasts. BeWo cells, when syncytialized with forskolin, increased expression of BCRP protein, and this was further augmented by DEX (P < .05). Our data suggest that the protective barrier provided by BCRP increases as cytotrophoblasts fuse to form the syncytiotrophoblast. Increase in glucocorticoid levels during the first trimester may reduce embryo/foetal exposure to clinically relevant BCRP substrates, because of an increase in placental BCRP.

Cytoplasmic mislocalization of p27 and CDK2 mediates the anti-migratory and anti-proliferative effects of Nodal in human trophoblast cells.

p27(Kip1), a cyclin-dependent kinase (CDK) inhibitor, is a multi-functional protein that regulates various cellular activities. Trophoblast proliferation, migration and invasion are some of the key processes of placental development. We have recently reported that Nodal, a member of the transforming growth factor-ß (TGF-ß) superfamily, inhibits human trophoblast cell proliferation, migration and invasion. In the present study, we investigated the mechanism by which Nodal regulates trophoblast activities. We found that Nodal increased p27 mRNA and protein levels by enhancing their stability. Interestingly, Nodal signaling also induced nuclear export of p27 and CDK2. Cytoplasmic translocation of p27 induced by Nodal requires p27 phosphorylation at S10. In addition, Nodal enhanced the association of p27 with CDK2, CDK5 and a microtubule-destabilizing protein, stathmin, and induced stathmin phosphorylation at S25 and S38. Furthermore, Nodal increased tubulin stability as revealed by immunofluorescent staining of acetylated tubulin. Finally, silencing of p27 reversed the inhibitory effect of Nodal on trophoblast cell proliferation, migration and invasion. Taken together, our findings revealed a novel function of simultaneous p27 and CDK2 cytoplasmic mislocalization in mediating growth-factor-regulated cell proliferation, migration and invasion.

MicroRNA-378a-5p promotes trophoblast cell survival, migration and invasion by targeting Nodal.

Nodal is a member of the transforming growth factor-ß superfamily that plays crucial roles during embryogenesis. Recently, we have reported that Nodal inhibits trophoblast cell proliferation, migration and invasion, but induces apoptosis in the human placenta. In this study, we examined the regulation of Nodal by microRNAs. In silico analysis of Nodal 3'UTR revealed a potential binding site for miR-378a-5p. In luciferase reporter assays, we found that miR-378a-5p suppressed the luciferase activity of a reporter plasmid containing Nodal 3'UTR but this suppressive effect was completely abolished when the predicted target site was mutated. Western blot analysis showed that miR-378a-5p decreased whereas anti-miR-378a-5p increased Nodal protein levels. These results indicate that miR-378a-5p targets Nodal 3'UTR to repress its expression. Stable transfection of the miR-378a-5p precursor, mir-378a, into HTR8/SVneo cells enhanced cell survival, proliferation, migration and invasion. Transient transfection of mature miR-378a-5p mimic, and to a lesser extent, siRNA targeting Nodal, produced similar effects. However, anti-miR-378a-5p inhibited cell migration and invasion. In addition, overexpression of Nodal reversed the invasion-promoting effect of miR-378a-5p. Furthermore, miR-378a-5p enhanced, whereas anti-miR-378a-5p suppressed, the outgrowth and spreading of extravillous trophoblast cells in first trimester placental explants. Finally, miR-378a-5p was detected in human placenta throughout different stages of gestation and in preterm pregnancies, placental miR-378a-5p levels were lower in preeclamptic patients than in healthy controls. Taken together, these findings strongly suggest that miR-378a-5p plays an important role in human placental development by regulating trophoblast cell growth, survival, migration and invasion, and that miR-378a-5p exerts these effects, at least in part, through the suppression of Nodal expression.

Beneficial effect of TNF-α inhibition on diabetic peripheral neuropathy.

BACKGROUND: Tumor necrosis factor-α (TNF-α) is an important inflammatory factor produced by activated macrophages and monocytes and plays an important role in the pathogenesis of diabetic peripheral neuropathy (DPN). To evaluate the effect of TNF-α signaling suppression and the potential of TNF-α in the treatment of DPN, a recombinant human TNF-α receptor-antibody fusion protein (rhTNFR:Fc) was used. We focused on the pathophysiology of the sciatic nerve and examined the expression of myelin basic protein (MBP) under DPN status with or without TNF-α inhibition. METHODS: The DPN rat model was generated by intraperitoneal injection of streptozotocin and by feeding with a high-fat, high-sugar diet. The nerve conduction velocity (NCV) in sciatic nerve of rat was monitored over a period of four weeks. The histopathological changes in nerve tissue were examined through traditional tissue histology and ultrastructure transmission electron microscopy (TEM). The expression of MBP was examined through western blot analysis. RESULTS: The DPN induced rats showed significant signs of nerve damage including lower NCV, demyelination of nerve fibers, disorganization of lamellar and axonal structures, and decreased expression of MBP in the nerve tissue. The inhibition of TNF-α in the DPN rats resulted in a significant recovery from those symptoms compared to the DPN rats. CONCLUSIONS: Our study demonstrates that TNF-α plays a key role in the pathogenesis of DPN and its inhibition by rhTNFR:Fc can prove to be a useful therapeutic strategy for the treatment of and/or prevention from DPN symptoms.

Adenovirus-delivered angiopoietin-1 treatment for phosgene-induced acute lung injury.

CONTEXT: Exposure to phosgene can result in an acute lung injury, leading to pulmonary edema and even death. Angiopoietin-1 (Ang1) is a critical factor for vascular stabilization due to its ability to reduce endothelial permeability and inflammation. OBJECTIVE: In this study, the histopathological changes of the lungs after exposure to phosgene and the effect of Ang1 treatment were examined. MATERIALS AND METHODS: Rats were exposed to phosgene gas at 8.33 g/m³ for 5 min. Ang1 overexpressing rats were established by an intravenous injection of adenovirus-Ang1 (Ad/Ang1). The histological changes of the lung were examined by Haematoxylin-Eosin (H&E) staining and fluorescence microscopy. The inferior lobe was used for the determination of the ratio of wet weight to dry weight of the lung. The concentration of cytokines in the serum and bronchoalveolar lavage fluid was determined by enzyme-linked immunosorbent assay. RESULTS: The pathological analysis showed signs of inflammation and edema, evident from a significant increase in the number of leukocytes in bronchoalveolar lavage fluid and the ratio of wet to dry weight of the lungs. The lung injury induced by phosgene was markedly reduced after the injection of Ad/Ang1. The increase of IL-1ß and IL-17 and decrease of vascular endothelial growth factor in the serum and bronchoalveolar lavage fluid of phosgene-exposed animals were abolished by the administration of Ad/Ang1. DISCUSSION AND CONCLUSIONS: Ang1 has the beneficial effects on phosgene-induced lung injury. The adenovirus-delivered Ang1 may have the potential as a novel approach for the treatment of the acute lung injury caused by phosgene gas inhalation in humans.

Progesterone control of myometrial contractility.

During pregnancy, the primary function of the uterus is to be quiescent and not contract, which allows the growing fetus to develop and mature. A uterine muscle layer, myometrium, is composed of smooth muscle cells (SMCs). Before the onset of labor contractions, the uterine SMCs experience a complex biochemical and molecular transformation involving the expression of contraction-associated proteins. Labor is initiated when genes in SMCs are activated in response to a combination of hormonal, inflammatory and mechanical signals. In this review, we provide an overview of molecular mechanisms regulating the process of parturition in humans, focusing on the hormonal control of the myometrium, particularly the steroid hormone progesterone. The primary reason for discussing the regulation of myometrial contractility by progesterone is the importance of the clinical problem of preterm birth. It is thought that the hormonal mechanisms regulating premature uterine contractions represent an untimely triggering of the normal events occurring during term parturition. Yet, our knowledge of the complex and redundant hormonal pathways controlling uterine contractile activity leading to delivery of the neonate remains incomplete. Finally, we introduce recent animal studies using a novel class of drugs, Selective Progesterone Receptor Modulators, targeting progesterone signaling to prevent premature myometrial contractions.

Nodal signals through activin receptor-like kinase 7 to inhibit trophoblast migration and invasion: implication in the pathogenesis of preeclampsia.

Trophoblast cell invasion into the uterus is an essential process for successful pregnancy, and shallow invasion of trophoblasts into the maternal decidua is linked to preeclampsia. We have reported that Nodal, a member of the transforming growth factor-ß superfamily, acts through activin receptor-like kinase 7 (ALK7) to inhibit trophoblast proliferation and to induce apoptosis. In this study, we examined the spatial and temporal expression patterns of Nodal and ALK7 in human placenta from normal and preeclamptic pregnancies and investigated whether Nodal regulated trophoblast migration and invasion. Nodal and ALK7 were detected in villous and extravillous trophoblast cell populations in early gestation, and their levels were strongly up-regulated in preeclamptic placenta. Overexpression of Nodal or constitutively active ALK7 decreased cell migration and invasion, whereas knockdown of Nodal and ALK7 had the opposite effects. In placental explant culture, treatment with Nodal inhibited trophoblast outgrowth, whereas Nodal small-interfering RNA strongly induced the expansion of explants and the migration of extravillous trophoblast cells. Nodal stimulated the secretion of tissue inhibitor of metalloproteinase-1 and inhibited matrix metalloproteinase (MMP)-2 and MMP-9 activity. These findings suggest that the Nodal/ALK7 pathway plays important roles in human placentation and that its abnormal signaling may contribute to the development of preeclampsia.

Reprint of: Myometrial activation: Novel concepts underlying labor.

Term labour is a state of physiological inflammation orchestrated by multiple uterine tissues (both fetal and maternal). This physiological inflammation preceding and accompanying labour onset is characterized by an increase in cytokine and chemokine secretion by the fetal membranes, as well as uterine tissues (i.e., decidua and myometrium). Pro-inflammatory cytokines and chemokines activate circulating maternal peripheral leukocytes as well as the uterine vascular endothelium to permit leukocyte infiltration into the uterus. This inflammatory milieu, in the absence of infection, is required for the initiation of labour as the uterine-infiltrated leukocytes secrete matrix metalloproteinases to induce fetal membrane rupture and cervical ripening as well as various labour mediators, which promote contractions of the myometrium. Myometrial activation at term and the onset of labour contractions are directly related to the changes in the ovarian/placental hormone progesterone and its downstream mediators (i.e., the progesterone receptors, PRA/B), which are also critical for maintenance of pregnancy. Our recent data provides direct evidence in support of local and functional P4 withdrawal in the uterine muscle (myometrium) via the activator protein-1 (AP-1) mediated pathway. This review outlines known mechanisms regulating activation of human labour, including progesterone and cytokine signaling. Understanding of the molecular mechanism of myometrial activation and labour onset could facilitate the development of new therapeutics for high-risk pregnant women to prevent premature uterine activation and preterm birth.

Myometrial activation: Novel concepts underlying labor.

Term labour is a state of physiological inflammation orchestrated by multiple uterine tissues (both fetal and maternal). This physiological inflammation preceding and accompanying labour onset is characterized by an increase in cytokine and chemokine secretion by the fetal membranes, as well as uterine tissues (i.e., decidua and myometrium). Pro-inflammatory cytokines and chemokines activate circulating maternal peripheral leukocytes as well as the uterine vascular endothelium to permit leukocyte infiltration into the uterus. This inflammatory milieu, in the absence of infection, is required for the initiation of labour as the uterine-infiltrated leukocytes secrete matrix metalloproteinases to induce fetal membrane rupture and cervical ripening as well as various labour mediators, which promote contractions of the myometrium. Myometrial activation at term and the onset of labour contractions are directly related to the changes in the ovarian/placental hormone progesterone and its downstream mediators (i.e., the progesterone receptors, PRA/B), which are also critical for maintenance of pregnancy. Our recent data provides direct evidence in support of local and functional P4 withdrawal in the uterine muscle (myometrium) via the activator protein-1 (AP-1) mediated pathway. This review outlines known mechanisms regulating activation of human labour, including progesterone and cytokine signaling. Understanding of the molecular mechanism of myometrial activation and labour onset could facilitate the development of new therapeutics for high-risk pregnant women to prevent premature uterine activation and preterm birth.

Breast Cancer Resistance Protein (BCRP/ABCG2) Inhibits Extra Villous Trophoblast Migration: The Impact of Bacterial and Viral Infection.

Extravillous trophoblasts (EVT) migration into the decidua is critical for establishing placental perfusion and when dysregulated, may lead to pre-eclampsia (PE) and intrauterine growth restriction (IUGR). The breast cancer resistance protein (BCRP; encoded by ABCG2) regulates the fusion of cytotrophoblasts into syncytiotrophoblasts and protects the fetus from maternally derived xenobiotics. Information about BCRP function in EVTs is limited, however placental exposure to bacterial/viral infection leads to BCRP downregulation in syncitiotrophoblasts. We hypothesized that BCRP is involved in the regulation of EVT function and is modulated by infection/inflammation. We report that besides syncitiotrophoblasts and cytotrophoblasts, BCRP is also expressed in EVTs. BCRP inhibits EVT cell migration in HTR8/SVneo (human EVT-like) cells and in human EVT explant cultures, while not affecting cell proliferation. We have also shown that bacterial-lipopolysaccharide (LPS)-and viral antigens-single stranded RNA (ssRNA)-have a profound effect in downregulating ABCG2 and BCRP levels, whilst simultaneously increasing the migration potential of EVT-like cells. Our study reports a novel function of BCRP in early placentation and suggests that exposure of EVTs to maternal infection/inflammation could disrupt their migration potential via the downregulation of BCRP. This could negatively influence placental development/function, contribute to existing obstetric pathologies, and negatively impact pregnancy outcomes and maternal/neonatal health.

Physical interaction of STAT1 isoforms with TGF-ß receptors leads to functional crosstalk between two signaling pathways in epithelial ovarian cancer.

BACKGROUND: The signal transducer and activator of transcription (STAT) and transforming growth factor-ß (TGF-ß) signaling pathways play important roles in epithelial ovarian cancer (EOC). However, the mechanism of crosstalk between two pathways is not completely understood. METHODS: The expression of STAT1 protein was detected by tissue microarray and immunoblotting (IB). The interaction of STAT1 isoforms with TGF-ß receptors was confirmed by immunoprecipitation and IB. The effect of TGF-ß signaling on STAT1 activation was examined in EOC and non-tumorous HOSEpiC cells treated with TGF-ß1 in the presence or absence of the inhibitor of TGF-ß type I receptor. The gain-of-function and loss-of-function approaches were applied for detecting the role of STAT1 on EOC cell behaviours. RESULTS: The high level of STAT1 was observed in patients with high-grade serous EOC. STAT1 expression was higher in ovarian cancer cells than noncancerous cells. TGF-ß1 activated the STAT1 pathway by inducing the phosphorylation of STAT1α on S727 residue. The full-length STAT1α and the truncated STAT1ß directly interacted with TGF-ß receptors (ALK1/ALK5 and TßRII), which was mediated by TGF-ß1. STAT1α and STAT1ß blocked the activation of the TGF-ß1 signaling pathway in EOC cells by reducing Smad2 phosphorylation. STAT1 overexpression induced EOC cell proliferation, migration, and invasion; whereas its inhibition enhanced TGF-ß1-induced phospho-Smad2 and suppressed EOC cell proliferation, migration, and invasion. CONCLUSIONS: Our data unveil a novel insight into the molecular mechanism of crosstalk between the STAT1 and TGF-ß signaling pathways, which affected the cancer cell behavior. Suppression of STAT1 may be a potential therapeutic strategy for targeting ovarian cancer.

Progesterone Via its Type-A Receptor Promotes Myometrial Gap Junction Coupling.

Effective labour contractions require synchronization of myometrial cells through gap junctions (GJs). Clasically, progesterone (P4) is known to inhibit the expression of connexin-43 (Cx43, major component of GJs) and GJ formation in myometrium. Our current study is based on a striking observation that challenges this dogma. We observed conspicuous differences in the intracellular localization of Cx43 protein in PRA versus PRB expressing myocytes. Thus in P4 stimulated PRA cells Cx43 protein forms GJs, whereas in PRB cells the forward trafficking of Cx43 and GJ formation is inhibited even when Cx43 is overexpressed. We found that P4, via PRA/B, differentially regulates Cx43 translation to generate a Cx43-20 K isoform, which facilitates the transport of full length Cx43 to plasma membrane. The P4 mediated regulation of Cx43 trafficking and GJ formation occurs via non-genomic pathway and involves the regulation of mTOR signaling since inhibition of this pathway restored the Cx43 trafficking defect in PRB cells. We propose that PRA is a master regulator of Cx43 expression, GJ formation and myocyte connectivity/synchronization for labour.