[Show abstract][Hide abstract] ABSTRACT: SERPINA3 (Serpin peptidase inhibitor clade A member 3), also known as a1-antichymotrypsin, is a serine protease inhibitor involved in a wide range of biological processes. Recently, it has been shown to be up-regulated in human placental diseases in association with a hypomethylation of the 5' region of the gene. In the present study, we show that the promoter of SERPINA3 is transcriptionally activated by three transcription factors (TFs) (SP1, MZF1 and ZBTB7B), the level of induction being dependent on the rs1884082 single nucleotide polymorphism (SNP) located inside the promoter, the T allele being consistently induced to a higher level than the G, with or without added TFs. When the promoter was methylated, the response to ZBTB7B was allele specific (the G allele was strongly induced, while the T allele was strongly down-regulated). We propose an adaptive model to explain the interest of such a regulation for placental function and homeostasis. Overexpression of SERPINA3 in JEG-3 cells, a trophoblast cell model, decreased cell adhesion to the extracellular matrix and to neighboring cells, but protects them from apoptosis, suggesting a way by which this factor could be deleterious at high doses. In addition, we show in different human populations that the T allele appears to predispose to Intra Uterine Growth Restriction (IUGR), while a G allele at a second SNP located in the second exon (rs4634) increases the risk of preeclampsia. Our results provide mechanistic views inside the involvement of SERPINA3 in placental diseases, through its regulation by a combination of epigenetic, genetic and TF-mediated regulations.
Full-text · Article · Jan 2012 · Human Molecular Genetics
[Show abstract][Hide abstract] ABSTRACT: Preeclampsia (PE) and vascular intra-uterine growth restriction (vIUGR) are two pathological obstetrical conditions originating from placental dysfunction. Recently, methylation changes at the placental level have been shown to be indicative of these diseases. The alteration of such epigenetic marks is therefore a novel pathway that might be critical for these pathologies. Here, we identified a region located in the distal promoter of the T-box-containing transcription factor TBX15 that is differentially methylated in pathological placentas. The level of methylation correlated significantly with the weight and stature of the newborn. The promoter was found to be hypomethylated in vIUGR coinciding with the down-regulation of its expression. PDX1, a transcription factor important for the regulation of insulin metabolism regulation was able to repress the TBX15 promoter in a methylation-dependent manner, which might, at least partially, explain the specific mRNA decrease of TBX15 observed in vIUGR placentas. Overall, the data presented herein suggest that TBX15 might be involved in the pathophysiology of placental diseases.
Full-text · Article · Feb 2011 · Epigenetics: official journal of the DNA Methylation Society
[Show abstract][Hide abstract] ABSTRACT: Intra-uterine growth restriction (IUGR) is defined by a restriction of fetal growth during gestation. It is a prevalent significant public health problem that jeopardizes neonatal health but also that can have deleterious consequences later in adult life. Cullins constitute a family of seven proteins involved in cell scaffold and in selective proteolysis via the ubiquitin-proteasome system. Most Cullins are critical for early embryonic development and mutations in some Cullin genes have been identified in human syndromes including growth retardation. Our work hypothesis is that Cullins, particularly CUL4B and CUL7, are involved in placental diseases and especially in IUGR. Thus, expression of Cullins and their cofactors was analyzed in normal and pathological placentas. We show that they present a constant significant over-expression in IUGR placentas, whose extent is dependent on the position of the interrogated fragment along the cDNAs, suggesting the existence of different isoforms of the genes. Particularly, the CUL7 gene is up-regulated up to 10 times in IUGR and 15 times in preeclampsia associated with IUGR. The expression of cofactors of Cullins participating to functional complexes has also been evaluated and showed a similar significant increase in IUGR. Promoters of Cullin genes appeared to be under the control of the SP1 transcription factor. Finally, methylation levels of the CUL7 promoter in placental tissues are modulated according to the pathological conditions, with a significant hypomethylation in IUGR. These results concur to pinpoint the Cullin family as a new set of markers of IUGR.
[Show abstract][Hide abstract] ABSTRACT: Preeclampsia is a common disease of pregnancy, characterized by high blood pressure and proteinuria appearing from the second trimester of gestation. Preeclampsia has been shown to have a strong genetic component. In 2005 a positional cloning project led to the discovery of the STOX1 transcription factor, and mutations of this gene were proposed as causal for preeclampsia in Dutch families. Despite the publication of three contradictory studies, we have shown by analyzing the functional effects of STOX1 that its overexpression in choriocarcinoma cells recapitulates several transcriptomic aspects of preeclampsia. In this review, the current literature is analyzed to evaluate the possible involvement of STOX1 in the pathogenesis of this disease. While preeclampsia obviously cannot be considered as a disease caused by mutation in a single gene, we argue that STOX1 may be at the center of common pathways leading to preeclampsia.
No preview · Article · Aug 2009 · Journal of Reproductive Immunology
[Show abstract][Hide abstract] ABSTRACT: Gene expression is controlled by two types of mechanisms: Epigenetic chromatin alterations affecting histones (methylations, acetylations, other post-translational modifications) or DNA (methylations/demethylations). In this article, we show, after a systematic analysis of 18 Serine Protease Inhibitors (Serpin) expressed in the placenta, that SERPINA3 is transcriptionnally activated in human placental diseases. This over-expression is correlated with the demethylation of specific sites in the proximal promoter of the gene, making it accessible to new transcription factors. The use of "reverse epigenetics" approaches has also enabled us to identify other methylation alterations in the vicinity of two genes involved in the Notch pathway, TLE3 and SOX1. These epigenetic alterations are associated with a increase of TLE3 gene expression.
No preview · Article · Jun 2008 · Reproduction Humaine et Hormones
[Show abstract][Hide abstract] ABSTRACT: Mutations in STOX1 were proposed to be causal for predisposing to preeclampsia, a hypertensive disorder originating from placental defects, affecting up to 10% of human pregnancies. However, after the first study published in 2005 three other groups have dismissed the polymorphism described in the first paper as a causal mutation.
In the present study, we have produced a choriocarcinoma cell line overexpressing STOX1. This overexpression results in transcriptional modification of 12.5% of the genes, some of them being direct targets as shown by chromatin immunoprecipitation. STOX1 overexpression correlates strongly and specifically with transcriptomic alterations in preeclamptic placentas (r = 0.30, p = 9.10(-7)). Numerous known key modulators of preeclampsia (such as Endoglin, Syncytin, human chorionic gonadotrophin -hCG-, and Glial Cell Missing Homolog -GCM1-) were modified in these transformed choriocarcinoma cells.
Our results contribute to reconcile contradictory data concerning the involvement of STOX1 in preeclampsia. In addition, they strongly suggest that anomalies in STOX1 expression are associated with the onset of preeclampsia, thus indicating that this gene should be the target of future studies. Our cellular model could constitute an invaluable resource for studying specific aspects of this human disease.
[Show abstract][Hide abstract] ABSTRACT: Intra-uterine growth restriction (IUGR) is a frequent disease, affecting up to 10% of human pregnancies and responsible for increased perinatal morbidity and mortality. Moreover, low birth weight is an important cause of the metabolic syndrome in the adult. Protein depletion during the gestation of rat females has been widely used as a model for human IUGR. By transcriptome analysis of control and protein-deprived rat placentas, we were able to identify 2543 transcripts modified more than 2.5 fold (1347 induced and 1196 repressed). Automatic functional classification enabled us to identify clusters of induced genes affecting chromosome structure, transcription, intracellular transport, protein modifications and apoptosis. In particular, we suggest the existence of a complex balance regulating apoptosis. Among repressed genes, we noted several groups of genes involved in immunity, signalling and degradation of noxious chemicals. These observations suggest that IUGR placentas have a decreased resistance to external aggression. The promoters of the most induced and most repressed genes were contrasted for their composition in putative transcription factor binding sites. There was an over-representation of Zn finger (ZNF) proteins and Pdx1 (pancreatic and duodenal homeobox protein 1) putative binding sites. Consistently, Pdx1 and a high proportion of ZNF genes were induced at the transcriptional level. A similar analysis of ZNF promoters showed an increased presence of putative binding sites for the Tata box binding protein (Tbp). Consistently again, we showed that the Tbp and TBP-associated factors (Tafs) were up-regulated in IUGR placentas. Also, samples of human IUGR and control placentas showed that human orthologous ZNFs and PDX1 were transcriptionally induced, especially in non-vascular IUGR. Immunohistochemistry revealed increased expression of PDX1 in IUGR human placentas. In conclusion, our approach permitted the proposition of hypotheses on a hierarchy of gene inductions/repressions leading to massive transcriptional alterations in the IUGR placenta, in humans and in rodents.
Full-text · Article · Nov 2007 · The Journal of Pathology
[Show abstract][Hide abstract] ABSTRACT: Preeclampsia is the major pregnancy-induced hypertensive disorder. It modifies the expression profile of placental genes, including several serine protease inhibitors (SERPINs). The objective of this study was to perform a systematic expression analysis of these genes in normal and pathological placentas and to pinpoint epigenetic alterations inside their promoter regions. Expression of 18 placental SERPINs was analyzed by quantitative RT-PCR on placentas from pregnancies complicated by preeclampsia, intrauterine growth restriction, or both and was compared with normal controls. SERPINA3, A5, A8, B2, B5, and B7 presented significant differences in expression in >or=1 pathological situation. In parallel, the methylation status of the CpG islands located in their promoter regions was studied on a sample of control and preeclamptic placentas. Ten SERPIN promoters were either totally methylated or totally unmethylated, whereas SERPINA3, A5, and A8 presented complex methylation profiles. For SERPINA3, the analysis was extended to 81 samples and performed by pyrosequencing. For the SERPINA3 CpG island, the average methylation level was significantly diminished in preeclampsia and growth restriction. The hypomethylated CpGs were situated at putative binding sites for developmental and stress response (hypoxia and inflammation) factors. Our results provide one of the first observations of a specific epigenetic alteration in human placental diseases and provide new potential markers for an early diagnosis.
[Show abstract][Hide abstract] ABSTRACT: Pre-eclampsia (PE) is a multiple disorder of human pregnancy, occuring from mid-gestation or from the third trimester of pregnancy, albeit most specialists estimate that the driving causes are to be found early (as early as at implantation). The principal clinical characteristics of PE are a maternal hypertension accompanied by proteinuria and sometimes oedemas. In industrialized countries, PE occurs in about 5% of the pregnancies, with an increased frequency in primiparous women (up to 10%). The aetiology of the disease is complex, with a strong familial component (maternal and paternal) as well as environemental component. A plausible genetic model to date postulates that maternal genes dictate a woman's susceptibility for the expression of the preeclampsia phenotype. Recently, one maternally imprinted gene, STOX1, contained five different missense mutations, was found cosegregating with the preeclamptic phenotype and following matrilineal inheritance. However, in the non familial preeclampsia cases, the phenotype of preeclampsia remains paradoxical. A large number of studies is focused on the transcriptome profiling, specifically implicated in the PE physiopathology and on the determination of genetic association to susceptibility of this disease.
No preview · Article · Oct 2006 · Reproduction Humaine et Hormones
[Show abstract][Hide abstract] ABSTRACT: Proinflammatory cytokines produced at the fetomaternal interface, such as IL-1beta, have been implicated in preterm and term labor. The present study was performed to evaluate the influence of IL-1beta on the endothelin (ET)/ET receptor system in human myometrial cells. We report that myometrial cells under basal conditions not only respond to but also secrete ET-1, one of the main regulators of uterine contractions. Prolonged exposure of the cells to IL-1beta led to a decrease in prepro-ET-1 and ET-3 mRNA correlated with a decrease in immunoreactive ET-1 and ET-3 levels in the culture medium. Whereas ETA receptor expression at both protein and mRNA levels was not affected by IL-1beta treatment, we demonstrated an unexpected predominance of the ETB receptor subtype under this inflammatory condition. Whereas the physiological function of ETB remains unclear, we confirmed that only ETA receptors mediate ET-1-induced myometrial cell contractions under basal conditions. By contrast, prolonged exposure of the cells to IL-1beta abolished the contractile effect induced by ET-1. Such a regulation of IL-1beta on the ET release and the balance of ETA to ETB receptors leading to a loss of ET-1-induced myometrial cell contractions suggest that complex regulatory mechanisms take place to constraint the onset of infection-induced premature contractions.
[Show abstract][Hide abstract] ABSTRACT: Villi from first-trimester human placenta were exposed to oxygen concentrations of either 2 or 20% during 3 h to construct two reciprocally subtracted libraries using the suppressive subtractive hybridization (SSH) methodology. After cloning, sequencing, and gene identification, the genes (1,071 clones corresponding to 822 different sequences) were classified according to 1) the subtracted library from which they originated and 2) within 58 groups of gene functions. We then developed a logarithm of the odds (LOD) test to identify a possible excess of genes in each group. We show that genes involved in angiogenesis are significantly overrepresented in the "hypoxic" condition (2% O2), whereas apoptotic genes are overrepresented in the "normoxic" condition (20% O2). Furthermore, we observed an excess of kinases relative to phosphatases and an excess of genes involved in proliferation over genes involved in cell growth in the hypoxic condition. To validate our results, we used quantitative RT-PCR to analyze the set of eight genes involved in angiogenesis on six independent placentas. Finally, we studied the distribution of gene clusters on human chromosomes to check whether their chromosomal distribution was random or not. We observed on human chromosome 11 a clear clustering of genes regulated similarly by O2 tension, and we also discovered indications that such clustering exists on chromosomes 6 and 12.
[Show abstract][Hide abstract] ABSTRACT: As a first step to explore the possible relationships existing between the effects of low oxygen pressure in the first trimester placenta and placental pathologies developing from mid-gestation, two subtracted libraries totaling 2304 cDNA clones were constructed. For achieving this, two reciprocal suppressive/subtractive hybridization procedures (SSH) were applied to early (11 weeks) human placental villi after incubation either in normoxic or in hypoxic conditions. The clones from both libraries (1440 hypoxia-specific and 864 normoxia-specific) were spotted on nylon macroarrays. Complex cDNAs probes prepared from placental villi (either from early pregnancy, after hypoxic or normoxic culture conditions, or near term for controls or pathological placentas) were hybridized to the membranes.
Three hundred and fifty nine clones presenting a hybridization signal above the background were sequenced and shown to correspond to 276 different genes. Nine of these genes are mitochondrial, while 267 are nuclear. Specific expression profiles characteristic of preeclampsia (PE) could be identified, as well as profiles specific of Intra-Uterine Growth Retardation (IUGR). Focusing on the chromosomal distribution of the fraction of genes that responded in at least one hybridization experiment, we could observe a highly significant chromosomal clustering of 54 genes into 8 chromosomal regions, four of which containing imprinted genes. Comparative mapping data indicate that these imprinted clusters are maintained in synteny in mice, and apparently in cattle and pigs, suggesting that the maintenance of such syntenies is requested for achieving a normal placental physiology in eutherian mammals.
We could demonstrate that genes induced in PE were also genes highly expressed under hypoxic conditions (P = 5 x 10(-5)), which was not the case for isolated IUGR. Highly expressed placental genes may be in syntenies conserved interspecifically, suggesting that the maintenance of such clusters is requested for achieving a normal placental physiology in eutherian mammals.
[Show abstract][Hide abstract] ABSTRACT: The endothelin (ET) system is involved in the control of several cellular functions such as proliferation, differentiation and migration processes. Considering the great flexibility of the trophoblastic phenotype, the expression and the production of the three ET isoforms have been investigated in the early and term trophoblasts as well as in the choriocarcinoma cell lines BeWo, JEG-3 and JAR which retain the properties of proliferation and invasion of the first trimester trophoblast. While ET-1 is produced in an ubiquitarian way, ET-2 is specifically synthetized by the early cytotrophoblasts and the choriocarcinoma cell lines. A weak expression of ET-3 is only noticed in the normal trophoblast: cytotrophoblasts and syncytiotrophoblast. In the BeWo cell line the dimethylsulfoxide (DMSO) inhibits the cellular growth without inducing the syncytial differentiation or the apotosis. In a parallel way to the DMSO anti-proliferative action, we notice a significant decrease of ET-2 expression as well as for ETA and ETB receptors. In contrast, the ET-1 remains inchanged. All these DMSO effects are reversible. The ET-2 high rates have been also observed in some tumors and developmental tissues. Although the molecular mechanisms regulating the ET-2 production are still unclear, these findings suggest that in the trophoblast, ET-2 expression is associated with a proliferative phenotype.
No preview · Article · Mar 2004 · Reproduction Humaine et Hormones
[Show abstract][Hide abstract] ABSTRACT: Endothelins exhibit growth regulating properties in many cell types, and there is now considerable evidence that they play a critical pathophysiological role in human diseases such as carcinogenesis. In the choriocarcinoma cell lines JEG-3, JAR and BeWO, we demonstrate by RT-PCR that prepro endothelin (ET)-1 and prepro ET-2 mRNA were expressed, whereas prepro ET-3 was never expressed. Only ET-1 and ET-2 peptides were identified by HPLC/RIA analyses in culture media from these three choriocarcinoma cell lines. In the BeWO line, the cellular growth measured as the cell count and DNA content decreased with increasing concentrations of dimethyl sulfoxide (DMSO; range 0.5-3%). The expression of prepro ET-2 was also suppressed by DMSO, whereas no change was noticed inprepro ET-1 mRNA. All these effects were reversible when DMSO was replaced by 15% foetal calf serum. These effects of DMSO which are correlated to ET-2 down regulation in dividing BeWO cells suggest a role for this endothelin isoform in trophoblast proliferation.
No preview · Article · Feb 2004 · Cellular and molecular biology
[Show abstract][Hide abstract] ABSTRACT: The study of the physiology of fetoplacental circulation is essential to our understanding of severe pathologies in pregnancy, including preeclampsia and the intrauterine growth retardation which often accompanies it. Moreover, for low-birth weight infants, epidemiological data reveal an increased risk of metabolic and cardiovascular problems in the course of their adult life. Though the etiology of preeclampsia remains to be elucidated, it is now recognized that this pathology arises from a precocious lesion concomitant with hemochorial placentation, manifested by hypoxia at the fetomaternal interface and a reduction in placental perfusion. Its effects upon fetal growth depend on the placenta's capacity to respond, via compensatory mechanisms, to the deficit in oxygen and nutrient intake. A number of signalling factors are synthesized by the different tissues of the fetomaternal interface: steroid hormones, eicosanoids, vasoactive peptides, cytokines/growth factors, etc. Disruptions in the balance between some of these signals have been associated with physiopathological mechanisms of pre-eclampsia, and especially with the high production of vasoconstrictors and mitogens to the detriment of vasodilators and antiproliferative agents. Thus, endothelin-1 (ET-1), a powerful vasoactive agent with growth factor properties, and the gene of which can be regulated by hypoxia, has been implicated. Vessels of the chorionic villi (major sites of placenta vascular resistance) are a favorite target of ET-1, whose vasoconstrictive effects upon smooth muscle occur preferentially via activation of ET-A-type receptors. During normal pregnancy the endothelium of these vessels plays a crucial role in production of relaxing factors such as NO which, due to its effect upon the media, ensures the maintenance of blood flow adapted to the needs of the fetus. In contrast, in the presence of fetoplacental hypoxia and alterations in the endothelium, these control mechanisms are disrupted and the smooth muscle cells of the media are directly confronted with abnormally elevated levels of ET-1 and vasoconstrictive and mitogenic eicosanoids. An activating effect of ET-1 and proinflammatory cytokines IL-1β and TNFα upon gene expression and the activity of cyclooxygenase 2, manifested by an increase in production of PGE2 and PGI2, has been demonstrated in these cells.
No preview · Article · Jan 2002 · Reproduction Humaine et Hormones
[Show abstract][Hide abstract] ABSTRACT: The status of the corticosteroid-binding globulin (CBG) at the fetomaternal interface, especially in the maternal intervillous blood space (I), was investigated and compared to that of CBG in the maternal (M) and fetal (umbilical arteries [A] and vein [V]) peripheral circulations at term. Immunoquantitation of plasma CBG showed that the CBG concentration in I was 30% less than that in M (P < 0.001) and threefold higher than that in umbilical cord blood (P < 0.001). The microheterogeneity of CBG studied by immunoaffinoelectrophoresis in the presence of concanavalin A and Western blotting indicated that the CBG in I was mainly of maternal origin and different from fetal CBG. A CBG mRNA, but no classic 50- to 59-kDa CBG, was found in isolated term trophoblastic cells. The steroid environment of the CBG in I differed greatly from that in the peripheral maternal and fetal circulations, because the progesterone:cortisol molar ratio in I was 75-fold higher than that in M and 7- to 10-fold higher than that in the fetal circulation. Binding studies revealed that the affinity constants of CBG for cortisol in I, A, and V were significantly lower than that in M plasma (P < 0.02) in their respective hormonal contexts. The binding parameters for I-CBG stripped of endogenous steroids and lipids were close to those for M-CBG but different from those of fetal CBG (P < 0.001). These data reflect the physiological relevance of the CBG-steroid interaction, especially with very CBG-loaded progesterone at the fetomaternal interface during late pregnancy.
Preview · Article · Mar 2001 · Biology of Reproduction
[Show abstract][Hide abstract] ABSTRACT: We have investigated the distribution of immunoreactive endothelins (irET) in fetal fluids and expression of ET precursor genes in villous tissue during the first trimester. Samples of maternal plasma (n = 6), coelomic fluid (n = 28), amniotic fluid (n = 23) and villous tissue (n = 3) were obtained from 30 pregnancies immediately before surgical termination at 7-12 weeks gestation. irET concentration was measured in plasma and fluids using two different radioimmunoassay kits, i.e. RPA 545 and RPA 555 and high performance liquid chromatography (HPLC). Total RNA was extracted and purified from villous tissue, reverse transcription and polymerase chain reaction (RT-PCR) were performed to evaluate the expression of ET-related genes. The irET concentration as evaluated by both kits was significantly higher (P<0.005) in maternal plasma than in coelomic or amniotic fluid and significantly higher (P<0.005) in coelomic fluid than in amniotic fluid using the RPA 555 kit. The profile of ET obtained by the HPLC- radioimmunoassay (RPA 555 kit) method confirmed significantly (P<0.005) higher ET concentration in coelomic than in amniotic fluid, although a similar distribution pattern for the three ET was observed in both embryonic fliud cavities. ET-3 was the predominant isoform in both fluids, reaching 19.4+/-2.0 pg/ml and 6.3+/-1.6 pg/ml in coelomic and amniotic fluid, respectively. Coelomic or amniotic fluid irET concentration did not change with gestational age irrespective of the kit used. RT-PCR demonstrated that first trimester placenta expresses the genes encoding for prepro-ET-1, -ET-2 and -ET-3. The similar ET distribution pattern in both fluid cavities could reflect their origin from the villous tissue and suggests that ET may play a role in the development of placenta and other fetal organs during organogenesis.
Full-text · Article · Sep 2000 · Molecular Human Reproduction
[Show abstract][Hide abstract] ABSTRACT: The presence of progesterone receptors (PR) throughout the human term fetoplacental vascular tree was investigated. By reverse transcription-polymerase chain reaction (RT-PCR), we showed expression of PR mRNAs in stem villi vessels, chorionic arteries and veins, and umbilical arteries and veins. Binding studies and Scatchard analysis revealed a single class of high-affinity binding sites for (3)H-R5020 (promegestone) in cytosolic extracts of all placental vessels, with K(d) values in the range of 2.5-4 nM. High levels of PR were detected in placental vessels compared to other vascular tissues. Thus, maximum binding capacities of stem villi vessels, chorionic arteries and veins, and umbilical arteries and veins were 247 +/- 25, 377 +/- 58, 295 +/- 40, 371 +/- 118, and 672 +/- 144 fmol/mg protein, respectively. Endothelial cell elimination in chorionic arteries did not significantly modify the number of PR. RT-PCR and binding studies also assessed PR expression in cultured placental vascular smooth muscle cells isolated from stem villi vessels. All these data suggested that most of the PR of fetoplacental vessels were from the media. In conclusion, we report here the first evidence of the presence of PR in the muscular layer of human term fetoplacental vessels. This finding, together with the high progesterone concentrations in cord blood, suggests that the interactions between the PR and its ligand may play a role in the physiology and physiopathology of human fetoplacental vascularization.
No preview · Article · Apr 2000 · Biology of Reproduction
[Show abstract][Hide abstract] ABSTRACT: In human myometrium, the modulation of intracellular cAMP content resulting from agonist-mediated stimulation of the receptor-adenylyl cyclase complex is largely influenced by the rate of cAMP hydrolysis by phosphodiesterase (PDE) isoenzymes. We have previously shown that the PDE4 family contributes to the predominant cAMP-hydrolyzing activity in human myometrium and that elevation of the PDE4B2 messenger RNA steady state level occurs in pregnant myometrial tissue. In the present study, we used a model of human myometrial cells in culture to determine whether an elevated cAMP concentration could influence PDE expression. As in myometrial tissue, high levels of PDE4 activity were detected in these smooth muscle cells. Long term treatment with 8-bromo-cAMP or forskolin resulted in a selective induction of PDE4B and of PDE4D short form messenger RNA variants. Concurrently, an increased immunoreactive signal for the PDE4B- and PDE4D-related isoenzymes was detected. This induction was consistent with an observed significant up-regulation of PDE4 activity. Accordingly, our results demonstrate that in human cultured myometrial cells, cAMP-elevating agents manipulate PDE4 activity through selective induction of synthesis of PDE4B and PDE4D short forms. Such a mechanism might have physiological importance during pregnancy by dampening hormonal stimulation and could thereby be involved in tolerance to the tocolytic effect of beta-adrenoceptor agonists.
[Show abstract][Hide abstract] ABSTRACT: The syncytiotrophoblast, which is delineated by two polar membranes (the microvillous and the basal plasma membranes), is the main placental structural element controlling maternal-fetal exchanges. These studies of the full-term placenta were undertaken in order to determine whether the microvillous membranes, which are bathed by the maternal intervillous circulation, and basal plasma membrane, which lines the fetal blood capillaries, have binding sites for insulin-like growth factor (IGF)-II. The microvillous and basal plasma membranes were purified and found to bind 125I-IGF-II with significantly different (p < 0.0001) Kd (0.51 and 1.02 nM, respectively). There were more available binding sites in the microvillous (4.4+/-0.3 pmol/mg protein) than in the basal (2.7+/-0.4 pmol/mg protein) plasma membranes (p < 0.0001). Both membranes contained three major (250, 135, and 130 kDa) 125I-IGF-II/binding-site protein complexes as determined by affinity cross-linking and PAGE. The 250-kDa band (type 2 IGF receptor) was the main band in the basal plasma membranes (46% total bound 125I-IGF-II). The 135-kDa band (insulin-receptor alpha subunit) was the main one in the microvillous membranes (48% total bound 125I-IGF-II). The amounts of 130-kDa band (type 1 IGF-receptor alpha subunit) in the two types of membranes were similar (30% total bound 125I-IGF-II). Only IGF-II displaced 125I-IGF-II from the 250-kDa band, while 125I-IGF-II bound to the 135-kDa band was displaced by insulin, and ligand bound to the 130-kDa band was displaced by IGF-I. Thus there are IGF receptors in both types of membranes of syncytiotrophoblast in the human full-term placenta, and the distributions of the IGF and insulin receptors are asymmetrical. This may reflect the fact that they face and interact with two independent, different media. Maternal IGF may influence the syncytiotrophoblast by binding to receptors on the microvillous membranes, while fetal IGF may also influence syncytiotrophoblast functions by activating receptors in the basal plasma membranes.
Full-text · Article · Jan 1998 · Biology of Reproduction