Maternal obesity modulates intracellular lipid turnover in the human term placenta.

BACKGROUND: Obesity before pregnancy is associated with impaired metabolic status of the mother and the offspring later in life. These adverse effects have been attributed to epigenetic changes in utero, but little is known about the role of placental metabolism and its contribution to fetal development. OBJECTIVES: We examined the impact of maternal pre-pregnancy obesity on the expression of genes involved in placental lipid metabolism in lean and obese women. SUBJECTS/METHODS: Seventy-three lean and obese women with healthy pregnancy were recruited at term elective cesarean delivery. Metabolic parameters were measured on maternal venous blood samples. Expression of 88 genes involved in lipid metabolism was measured in whole placenta tissue. Proteins of genes differently expressed in response to maternal obesity were quantified, correlated with maternal parameters and immunolocalized in placenta sections. Isolated primary trophoblasts were used for in vitro assays. RESULTS: Triglyceride (TG) content was increased in placental tissue of obese (1.10, CI 1.04-1.24 mg g-1, P<0.05) vs lean (0.84, CI 0.72-1.02 mg g-1) women. Among target genes examined, six showed positive correlation (P<0.05) with maternal pre-pregnancy BMI, namely ATGL (PNPLA2), FATP1 (SLC27A1), FATP3 (SLC27A3), PLIN2, PPARG and CGI-58 (ABHD5). CGI-58 protein abundance was twofold higher (P<0.001) in placentas of obese vs lean women. CGI-58 protein levels correlated positively with maternal insulin levels and pre-pregnancy body mass index (R=0.63, P<0.001 and R=0.64, P<0.001, respectively). CGI-58 and PLIN2 were primarily located in the syncytiotrophoblast and, were upregulated (1.38- and 500-fold, respectively) upon oleic acid and insulin treatment of cultured trophoblast cells. CONCLUSION: Pre-gravid obesity significantly modifies the expression of placental genes related to transport and storage of neutral lipids. We propose that the upregulation of CGI-58, a master regulator of TG hydrolysis, contributes to the turnover of intracellular lipids in placenta of obese women, and is tightly regulated by metabolic factors of the mother.

The placental exposome: placental determinants of fetal adiposity and postnatal body composition.

Offspring of obese and diabetic mothers are at increased risk of being born with excess adiposity as a consequence of their intrauterine environment. Excessive fetal fat accretion reflects additional placental nutrient transfer, suggesting an effect of the maternal environment on placental function. High plasma levels of particular nutrients in obese and diabetic mothers are likely to be the important drivers of nutrient transfer to the fetus, resulting in excess fat accretion. However, not all offspring of obese and diabetic mothers are born large for gestational age and the explanation may involve the regulation of placental nutrient transfer required for fetal growth. The placenta integrates maternal and fetal signals across gestation in order to determine nutrient transfer rate. Understanding the nature of these signals and placental responses to them is key to understanding the pathology of both fetal growth restriction and macrosomia. The overall effects of the maternal environment on the placenta are the product of its exposures throughout gestation, the 'placental exposome'. Understanding these environmental influences is important as exposures early in gestation, for instance causing changes in the function of genes involved in nutrient transfer, may determine how the placenta will respond to exposures later in gestation, such as to raised maternal plasma glucose or lipid concentrations. Longitudinal studies are required which allow investigation of the influences on the placenta across gestation. These studies need to make full use of developing technologies characterising placental function, fetal growth and body composition. Understanding these processes will assist in the development of preventive strategies and treatments to optimise prenatal growth in those pregnancies at risk of either excess or insufficient nutrient supply and could also reduce the risk of chronic disease in later life.

Thrombin weakens the amnion extracellular matrix (ECM) directly rather than through protease activated receptors.

INTRODUCTION: Preterm premature rupture of fetal membranes (pPROM) is a major cause of preterm birth. Abruption associated thrombin production, and infection-inflammation associated cytokine production reportedly play major roles in pPROM. Utilizing an in vitro model-system we have confirmed that both thrombin and inflammatory cytokines remodel and biomechanically weaken amnion, the load-bearing component of FM. Also, we have shown thrombin directly weakens isolated amnion but cytokines weaken amnion only indirectly by initially interacting with choriodecidua and releasing unidentified soluble activator(s). This study's purpose was to determine whether thrombin weakens the isolated amnion through thrombin receptor-protease activated receptors (PARs 1,2,3,4), activation of previously secreted extracellular matrix (ECM) enzymes, or by direct action on the ECM. METHODS: Primary amnion cells and isolated amnion were tested for PARs by immunohistochemistry, Western Blot and rtPCR. Cell-free amnion ECM was produced by devitalizing isolated amnion by exposure to UV light and subsequent freeze-thaw cycles. Devitalized amnion membrane explants were incubated with thrombin and biomechanically tested. RESULTS: PARs were not found in amnion or amnion cells. Thrombin induced dose-dependent weakening of devitalized amnion explants. Preincubation with the thrombin inhibitor hirudin prevented thrombin-induced weakening. Thrombin converted pro-MMP2 embedded in the devitalized amnion ECM to multiple active forms. Thrombin also directly digested gelatin gels in zymograms suggesting the possibility of direct degradation of amnion ECM components. DISCUSSION: Thrombin appears to directly weaken the amnion ECM and additionally activates stored pro-MMP2 to active forms that may further enhance amnion ECM degradation. CONCLUSION: Thrombin weakens amnion directly rather than through PARs.

High molecular mass multimer complexes and vascular expression contribute to high adiponectin in the fetus.

CONTEXT: High plasma adiponectin concentrations in human fetuses and neonates are unique features of early developmental stages. Yet, the origins of the high adiponectin concentrations in the perinatal period remain elusive. OBJECTIVE: This study was undertaken to identify the sources and functional properties of adiponectin in utero. DESIGN AND METHODS: Tissue specimens were obtained at autopsy from 21- to 39-wk-old stillborn human fetuses. Adipose tissue and placenta were obtained at term elective cesarean section. Adiponectin complexes and expression were measured by immunodetection and real-time PCR. RESULTS: Adiponectin mRNA transcripts were detected in fetal sc and omental adipose depots at lower concentrations than in maternal adipose tissue. Immunoreactive adiponectin was also observed in vascular endothelial cells of fetal organs, including skeletal muscle, kidney, and brain. The absence of adiponectin in all placental cell types and lack of correlation between maternal and umbilical adiponectin indicate that umbilical adiponectin reflects its exclusive production by fetal tissues. The most prominent forms of adiponectin in fetal plasma were high and low molecular mass (HMW and LMW) multimers of 340 and 160 kDa, respectively. The proportion of the HMW complexes was 5-fold (P < 0.001) higher in umbilical plasma than in adult. The high HMW and total adiponectin levels were associated with lower insulin concentration and lower homeostasis model of assessment of insulin resistance indices in umbilical plasma, reflecting higher insulin sensitivity of the fetus compared with adult. CONCLUSIONS: The abundance of HMW adiponectin and its vascular expression are characteristics of human fetal adiponectin. Combined with high insulin sensitivity, fetal adiponectin may be a critical determinant of in utero growth.

Obesity in pregnancy stimulates macrophage accumulation and inflammation in the placenta.

Obesity and pregnancy are associated with a combination of insulin resistance and inflammatory changes which exacerbate in combination. Based on the similarity between the inflammatory transcriptomes of adipose tissue and placenta, we hypothesized that the placenta develops exaggerated inflammation in response to obesity. The aim of this study was to characterize placental inflammatory mediators and macrophage accumulation in relation to peripheral inflammation in obesity. Placental macrophages and maternal peripheral blood mononuclear cells (PBMC) from 20 obese and 15 lean women were functionally and phenotypically characterized using immunohistochemistry, flow cytometry and expression for macrophage markers and inflammatory cytokines. The number of resident CD68+ and CD14+ cells was increased 2-3 fold in the placenta of obese as compared to lean women. The macrophage population was characterized by a marked phenotypic heterogeneity with complex subsets of CD14+, CD68+ and CD11b+ (mac-1) cells and by an increased expression of the pro-inflammatory cytokines IL-1, TNF-alpha, IL-6. Placental inflammation was associated with an activation of PBMC gene expression with an increase in the monocyte differentiation and maturation markers CD14 and CD68 in maternal but not fetal PBMC. The inflammatory changes were associated with higher plasma concentrations of C-reactive protein and IL-6 in obese compared to lean women. In conclusion, the chronic inflammation state of pre-gravid obesity is extending to in utero life with accumulation of a heterogeneous macrophage population and pro-inflammatory mediators in the placenta. The resulting inflammatory milieu in which the fetus develops may have critical consequences for short and long term programming of obesity.

[Leptin in pregnancy: facts, questions and future]. / Leptine et grossesse: dogmes, questions et perspectives.

Leptin has been primarily considered as a protein secreted by the adipocyte and a regulator of satiety and energy homeostasis. A role for leptin in pregnancy was later suggested as circulating levels of leptin are high in the pregnant woman and leptin is synthetized within the placenta. Placental leptin production is increased in various obstetrical conditions associated with alterations of fetal growth (diabetes, preeclampsia). Furthermore, umbilical leptin can be viewed as a biomarker of fetal adiposity. Our aim is to review the putative roles of leptin in pregnancy.

Adiponectin in human pregnancy: implications for regulation of glucose and lipid metabolism.

AIMS/HYPOTHESIS: Adiponectin is upregulated during adipogenesis and downregulated in insulin-resistant states. The mechanism(s) governing the re-arrangements from adipogenesis to facilitated lipolysis during pregnancy are unknown. Our purpose was to analyse the role of adiponectin relative to the metabolic changes in human pregnancy. SUBJECTS, MATERIALS AND METHODS: Lean women (BMI <25 kg/m(2)) were evaluated longitudinally before conception, and in early (12-14 weeks) and late (34-36 weeks) pregnancy. Insulin sensitivity was measured using the glucose clamp technique. Venous blood and subcutaneous adipose tissue biopsies were obtained at each time point. RESULTS: Adiponectin concentrations were lower in the third trimester than in the pregravid condition (9.9+/-1.4 vs 13.5+/-1.8 microg/ml). The hypoadiponectinaemia was reflected by a 2.5-fold decrease in white adipose tissue adiponectin mRNA. These changes were associated with a 25% increase in fat mass (23.7+/-2.9 vs 18.9+/-2.9 kg). Insulin infusion decreased high molecular weight adiponectin complexes in pregravid women (9.9+/-0.6 vs 6.2+/-0.06) and the suppressive effect of insulin was lost during pregnancy. The pregnancy-mediated changes in adiponectin were strongly correlated with basal insulin levels and insulin sensitivity (p<0.0001). The relationship between adiponectin and insulin sensitivity was related to the decreased insulin regulation of glucose utilisation (r=0.55, p<0.001) but not of endogenous hepatic glucose production. CONCLUSIONS/INTERPRETATION: These data demonstrate that pregnancy is associated with adiponectin changes in lean women. Hypoadiponectinaemia is reflected by a lower amount of high molecular weight adiponectin and by the ratio of high to low molecular weight multimers. The adiponectin changes relate to decreased insulin sensitivity of glucose disposal rather than alterations of lipid metabolism.

The placenta cytokine network and inflammatory signals.

Throughout its entire lifespan, the placenta is able to produce as well as respond to a variety of inflammatory stimuli. Many signaling molecules and concurrent pathways responsible for the propagation of an inflammatory response have been identified in placental cells. From early developmental stages onward, the secretory activity of placenta cells clearly contributes to increase local as well as systemic levels of cytokines and inflammatory molecules. Two aspects of the progression of an immune response have been particularly investigated: the highly regulated process of invasion and implantation and, the induction of preterm labor associated with infections. With the progression of pregnancy, the physiological role of most placental cytokines is more uncertain. Many placental cytokines are similar to adipose tissue derived cytokines. One possibility is that they contribute to the low grade systemic inflammation developing during the third trimester of pregnancy. The prevalent hypothesis is that activation of some inflammatory pathways is necessary to induce maternal insulin resistance which is required for the progression of normal gestation. As an integrative organ, the placenta may relay or enhance the contribution made by the cells of the adipose tissue and immune system in non-pregnant individuals. In pregnancy complicated with obesity or diabetes mellitus, continuous adverse stimulus is associated with dysregulation of metabolic, vascular and inflammatory pathways supported by increased circulating concentration of inflammatory molecules. It is believed that maternal adipose tissue and placental cells both contribute to the inflammatory situation by releasing common molecules. For example, the accumulation of leptin and TNF-alpha is associated with an increased production for markers of inflammation, fibrotic response, vascular remodeling and proteins facilitating lipid storage within the placenta.

Fetal growth restriction: a workshop report.

Intrauterine growth restriction (IUGR) is associated with significantly increased perinatal morbidity and mortality as well as cardiovascular disease and glucose intolerance in adult life. A number of disorders from genetic to metabolic, vascular, coagulative, autoimmune, as well as infectious, can influence fetal growth by damaging the placenta, leading to IUGR as a result of many possible fetal, placental and maternal disorders. Strict definitions of IUGR and of its severity are needed in order to eventually distinguish among different phenotypes, such as gestational age at onset, degree of growth restriction and presence of hypoxia. This report explores and reviews some of the most recent developments in both clinical and basic research on intrauterine growth restriction, by seeking mechanisms that involve genetic factors, utero-placental nutrient availability and vascular growth factors. New exciting findings on the genomic imprinting defects potentially associated with IUGR, and the placental anomalies associated with the decreased nutrient transport are summarized. Moreover, recent data on angiogenic growth factors as well as new information arising from application of gene chip technologies are discussed.

Transduction of leptin growth signals in placental cells is independent of JAK-STAT activation.

We have characterized the transduction pathways induced by leptin in the placenta, using human BeWo cells that express endogenous leptin receptors and synthesize leptin in a regulated manner. We first examined if the JAK-STAT phosphorylation cascade was functional in these cells. Phosphorylated JAK2 was primarily bound to a short 106kDa leptin receptor isoform and to a lesser extent to a 210kDa molecule. Leptin neither enhanced JAK2 phosphorylation nor activated STAT3 and STAT1 proteins indicating that JAK2 is constitutively activated and that the JAK-STAT transduction pathway is not recruited by leptin in BeWo cells. By contrast, leptin stimulated the transcription of the c-fos gene (3-fold) and cell proliferation (2-fold) as measured by DNA synthesis. Both effects were dependent on the rapid phosphorylation of p42-44 MAPK but not p38 MAPK. We conclude that a functional JAK-STAT pathway is not required for leptin to transduce proliferative signals in human placental cells. These findings extend the physiological action of leptin beyond its central effects, to the control of placental gene transcription and cell proliferation.

Placental leptin receptor isoforms in normal and pathological pregnancies.

Alternate mRNA splicing of human leptin receptor generates four membrane isoforms with different C-terminal sequences. They differ by the length of their intracellular domain which include specific motifs crucial for the specificity of leptin signalling. As a step towards functional studies, we have characterized leptin receptors in human placenta from normal pregnancies and pregnancies associated with diabetes and pre-eclampsia. Leptin and leptin receptors were visualized by immunohistochemistry of placentas obtained from first and third trimester pregnancies. Antibodies against N and C-terminal epitopes showed signals in the apical membrane of the syncytiotrophoblast in early and term placental villi as well as in JAr and BeWo derived trophoblast cells. In addition, a distinct isoform recognized by its extracellular juxtamembrane epitope was exclusively localized in cytotrophoblast cells and likely stains the soluble receptor. At contrast with the transmembrane receptors, the expression of this isoform is increased in placentas of pre-eclamptic and diabetic women which synthesize more leptin than placenta from uncomplicated pregnancy. These data demonstrate that short and long transmembrane leptin receptors are expressed in the trophoblast and indicate that leptin synthetized within the placenta can act locally through both receptor isoforms. Being also accessible to leptin from maternal origin, these transmembrane receptors may signal differently in pregnancy with normal and increased leptin production. The co-localization of leptin and the soluble receptor isoform suggests that this isoform serves for modulating maternal free leptin levels through modification of leptin binding capacities.

Fetal macrosomia and maternal weight gain during pregnancy.

BACKGROUND: To calculate an adjusted maternal weight gain during pregnancy including infant and placental weights to the actual weight gain and secondarily examine its influence on the occurrence of fetal macrosomia. METHODS: The corrected weight gain was calculated as follows: maternal body weight at the last prenatal visit - (pregravid body weight + birth weight + placental weight) in 259 women (65 with type 1 diabetes, 69 with gestational diabetes mellitus, and 125 controls). RESULTS: Although weight gain was greater in non obese normal glucose tolerant and type 1 diabetic mothers of macrosomic infants, the difference was no longer significant when using corrected maternal weight gain. The correlation between birth weight and maternal weight gain decreased when using this index. CONCLUSION: The corrected weight gain is a better estimate of true accretion of maternal weight. Our results suggest that recommendations for weight gain during pregnancy should take this index into account.

Transcriptional effect of hypoxia on placental leptin.

We observed recentlyl that placental leptin is markedly increased in preeclampsia. Since this disorder is associated with vascular disorders, we have tested the hypothesis that hypoxia regulates leptin expression. We show that hypoxia increased leptin mRNA and secretion in trophoblast-derived BeWo cells. This effect was mediated through leptin promoter activation. 5' deletion analysis allowed us to delineate two regions containing 1.87 kb and 1.20 kb of the promoter which conferred respectively high and low responsiveness to hypoxia. These data indicate that leptin is up-regulated by hypoxia through a transcriptional mechanism likely to involve distinct hypoxia-responsive cis-acting sequences on the promoter.

[Placental leptin and pregnancy pathologies]. / Leptine placentaire et pathologies de la grossesse.

Leptin, the protein encoded by the Ob gene, is produced by the white adipose tissue and by the placenta during pregnancy. Placental leptin production makes a substantial contribution to maternal circulating levels during pregnancy which rapidly decrease and return to normal after delivery. Leptin has been detected in fetal plasma as early as week 18 of gestation, and umbilical leptin concentrations are closely related to birth weight. This has led to the hypothesis that fetal fat mass mainly determines fetal circulating leptin. Placental leptin production is increased in choriocarcinoma, preeclampsia and type 1 diabetes. Estrogens, hypoxia and insulin have been suggested as positive regulators of placental leptin production. Maternal leptinemia might act as a sensor of energy balance during pregnancy. The presence of both leptin and leptin receptors in the placenta suggests that leptin can act by autocrine or endocrine pathways in the human placenta. The roles of fetal leptin and consequences of increased placental leptin production in pathological pregnancies have yet to be elucidated.

Dissociation between insulin-mediated signaling pathways and biological effects in placental cells: role of protein kinase B and MAPK phosphorylation.

Beyond the presence of insulin receptors, little is known of the mechanisms underlying the biological effects of insulin in the placenta. We show that phosphorylation of MAPK and protein kinase B were enhanced 286 +/- 23% and 393 +/- 17% upon insulin stimulation of JAr placental cells. MAPK activation was prevented by pretreatment with PD98059 but was unaffected by wortmannin. Insulin stimulation of protein kinase B phosphorylation was abolished by pretreatment with wortmannin, suggesting that it is dependent on phosphatidylinositol 3- kinase activation. Despite protein kinase B phosphorylation, GLUT4 translocation, glucose uptake, and glycogen synthesis were not stimulated by insulin. By contrast, glycogen synthesis was stimulated 20-fold in cells incubated with 11 mM glucose. Mitogenesis assessed by incorporation of [(3)H]thymidine into DNA was enhanced 1.9-fold in response to insulin. Stimulation of DNA synthesis was inhibited by pretreatment with PD98059 but was insensitive to wortmannin. These results indicate that stimulation of mitogenesis is one major biological effect of insulin in placenta cells that implicates the MAPK signaling pathway. Phosphatidylinositol 3-kinase- dependent protein kinase B activation is not sufficient to stimulate glucose transport and glycogen synthesis, highlighting the placenta as a nonclassic target of insulin for the regulation of glucose metabolism.

Heterogeneity of fetal growth in type 1 diabetic pregnancy.

OBJECTIVE: To investigate the frequency of macrosomia in an homogeneous cohort of type 1 diabetic mothers and to analyze the influence of maternal factors and glycemic control on the incidence of fetal macrosomia. MATERIAL AND METHODS: Fifty-five consecutive type 1 diabetic first-pregnancies were prospectively studied. Macrosomia was defined by a ponderal index above the 90(th) percentile. Venous cord blood levels of insulin, C peptide and leptin were measured at delivery. The influence of HbA1c levels and other maternal variables on the occurrence of macrosomia and on the ponderal index was assessed using a stepwise regression logistic model. RESULTS: The mean (+/- SD) birth weight was 3482 (+/- 497) g at 37.4 +/- 1.0 weeks gestation. Macrosomia occurred in 29 cases (53.7%). Fetal insulin, C peptide and leptin levels were significantly higher in macrosomic than in non macrosomic infants. Maternal age, duration of diabetes, pregravid body mass index, parity, weight gain during pregnancy, presence of a microangiopathy, nephropathy, smoking habits, gestational hypertension or preeclampsia, and HbA1c levels throughout pregnancy did not differed between mothers of macrosomic and non macrosomic infants. In the stepwise analysis none of these covariates was explanatory of the ponderal index. CONCLUSIONS: The frequency of macrosomia remains very high in infants of type 1 diabetic mothers despite a reasonable degree of glycemic control. The variability of the fetal growth response to mild hyperglycemia prompts for the identification of other factors involved in the modulation of fetal growth.