Background: Pregnant women are at increased risk of venous thrombosis compared to non-pregnant women. Epidemiological and laboratory data suggest that hypercoagulability begins in the first trimester but it is unknown exactly how early in pregnancy this develops. The mechanisms that result in a prothrombotic state may involve oestrogens and progestogens. Methods: Plasma samples were taken prior to conception and five times in early pregnancy, up to Day 59 gestation, from 22 women undergoing natural cycle in vitro fertilization, who subsequently gave birth at term following a normal pregnancy. Thrombin generation, free Protein S, Ddimer, Fibrinogen, factor VIII, estradiol and progesterone were measured. To counter inter-individual variability, the change in laboratory measurements between the pre-pregnant and pregnant state were measured over time. Results: Peak thrombin, Endogenous Thrombin Potential, Velocity Index and fibrinogen significantly increased, and free Protein S significantly decreased, from pre-pregnancy levels, by 32 days gestation. Ddimer and VIII significantly increased from pre-pregnancy levels by 59 days gestation. Estradiol significantly increased by Day 32 gestation with a non-significant increase of 67% by Day 24 gestation. Progesterone significantly increased by Day 32 gestation. Almost all laboratory markers of thrombosis correlated significantly with estradiol and progesterone. Conclusion: Our work is the first to demonstrate that the prothrombotic state develops very early in the first trimester. Laboratory markers of hypercoagulability correlate significantly with estradiol and progesterone suggesting these are linked to the prothrombotic state of pregnancy. Clinicians should consider commencing thromboprophylaxis early in the first trimester in women at high thrombotic risk.

The environment for embryo implantation and fetal growth and development is affected by maternal nutritional, metabolic and health status. The aim of this prospective, cohort study was to test whether plasma metabolic and inflammatory biomarkers can predict pregnancy resulting from in vitro fertilisation (IVF). Women with a natural menstrual cycle undergoing frozen embryo transfer (FET) were recruited and fasting baseline blood samples were collected a mean of 3.4 days prior to the luteinising hormone (LH) surge and a non-fasting blood sample was taken on the day of FET. Ongoing pregnancy was defined by positive fetal heartbeat on ultrasound scan at day 45 post LH surge. Thirty-six pregnancies resulted from FET in 143 women. In an overall stepwise multivariable analysis, erythrocyte saturated to unsaturated fatty acid ratio was positively associated with ongoing pregnancy. A similar model incorporating day of FET covariates found that erythrocyte saturated to unsaturated fatty acid ratio, erythrocyte fatty acid average chain length and plasma log-triglycerides predicted ongoing pregnancy. In conclusion, a higher peri-conceptional saturated to unsaturated fatty acid ratio predicted ongoing pregnancy after natural cycle frozen embryo transfer and may reflect a maternal nutritional status that facilitates pregnancy success in this assisted conception scenario.

Background: Pregnancy is a hypercoagulable state associated with an increased risk of venous thrombosis, which begins during the first trimester, but the exact time of onset is unknown. Thrombin generation, a laboratory marker of thrombosis risk, increases during normal pregnancy but it is unclear exactly how early this increase occurs. Methods: We assessed thrombin generation by Calibrated Automated Thrombography in women undergoing natural cycle in vitro fertilization, who subsequently gave birth at term following a normal pregnancy (n = 22). Blood samples were taken just prior to conception and repeated five times during very early pregnancy, up to Day 59 estimated gestation. Results: Mean Endogenous Thrombin Potential (ETP), peak thrombin generation and Velocity Index (VI) increased significantly from pre-pregnancy to Day 43 gestation (p = 0.024-0.0004). This change persisted to Day 59 gestation. The mean of the percentage change from baseline, accounting for inter-individual variation, in ETP, peak thrombin and VI increased significantly from pre-pregnancy to Day 32 gestation (p = 0.0351-< 0.0001) with the mean increase from baseline persisting to Day 59 gestation. Conclusion: Thrombin generation increases significantly during the very early stages of normal pregnancy when compared to the pre-pregnancy state. The increased risk of venous thrombosis therefore likely begins very early in a woman's pregnancy, suggesting that women considered clinically to be at high thrombotic risk should start thromboprophylaxis as early as possible after a positive pregnancy test. (C) 2017 Elsevier Ltd. All rights reserved.

Obesity increases pre-eclampsia (PE) risk. Adipose tissue inflammation may contribute to the clinical syndrome of PE. We compared adipose tissue macrophage infiltration and release of pro-inflammatory adipokines in PE and healthy pregnancy. Subcutaneous and visceral adipose tissue biopsies were collected from healthy (n=13) and PE (n=13) mothers. Basal and lipopolysaccharide (LPS) stimulated adipocyte TNFα, IL-6, CCL-2, and CRP release was measured. Adipose tissue cell densities of activated (cfms+) and total (CD68+) macrophages were determined. In PE only, visceral adipose tissue TNFα release was increased after LPS stimulation (57 [76] versus 81 [97] pg/ml/µg DNA, P=0.030). Basal TNFα release was negatively correlated insulin sensitivity of visceral adipocytes (r = −0.61, P=0.030) in PE. Visceral adipocyte IL-6 release was increased after LPS stimulation in PE only (566 [696] versus 852 [914] pg/ml/µg DNA, P=0.019). Visceral adipocyte CCL-2 basal (67 [61] versus 187 [219] pg/ml/µgDNA, P=0.049) and stimulated (46 [46] versus 224 [271] pg/ml/µg DNA, P=0.003) release was greater than in subcutaneous adipocytes in PE only. In PE, median TNF mRNA expression in visceral adipose tissue was higher than controls (1.94 [1.13–4.14] versus 0.8 [0.00–1.27] TNF/PPIA ratio, P=0.006). In visceral adipose tissue, CSF1R (a marker of activated macrophages) mRNA expression (24.8[11.0] versus 51.0[29.9] CSF1R/PPIA ratio, P=0.011) and activated (cfms+) macrophage count (6.7[2.6] versus 15.2[8.8] % cfms+/adipocyte, P=0.031) were higher in PE than in controls. In conclusion, our study demonstrates dysregulation of inflammatory pathways predominantly in visceral adipose tissue in PE. Inflammation of visceral adipose tissue may mediate many of the adverse metabolic effects associated with PE.

Free fatty acid transfer across the placenta facilitated by lipoprotein lipase and its relationship with fetal adiposity

CONTEXT:Docosahexaenoic acid (DHA) is an important fatty acid required for neurological development but its importance during early fetal neurological organogenesis is unknown.OBJECTIVE:This study aimed to assess plasma fatty acid changes in early pregnancy in women undergoing natural cycle-frozen embryo transfer as a means of achieving accurately timed periconceptual sampling.DESIGN:Women undergoing frozen embryo transfer were recruited and serial fasting blood samples were taken pre-luteinizing hormone (LH) surge, and at 18, 29, and 45 d post-LH surge and fatty acids were analyzed using gas chromatography.SETTING:This study took place at the Assisted Conception Unit, Glasgow Royal Infirmary, Scotland.MAIN OUTCOME MEASURES:Plasma fatty acid concentrations and influence of twin pregnancies on DHA plasma concentration were measured.RESULTS:In pregnant women, there was a rapid, early increase in the maternal rate of change of plasma DHA concentration observed by 29 d post-LH surge (mean ± SD, from 0.1 ± 1.3 to 1.6 ± 2.9 nmol DHA per mL plasma per day). This early pressure to increase plasma DHA concentration was further emphasized in twin pregnancies where the increase in DHA concentration over 45 d was 2-fold higher than in singleton pregnancies (mean ± SD increase, 74 ± 39 nmol/mL vs 36 ± 40 nmol/mL). An index of delta-6 desaturase activity increased 30% and positively correlated with the rate of change of DHA concentration between 18 and 29 d post-LH surge (R2 adjusted = 41%; P = .0002). DHA was the only fatty acid with a continual accelerated increase in plasma concentration and a positive incremental area under the curve (mean ± SD, 632 ± 911 nmol/mL × d) during the first 45 d of gestation.CONCLUSIONS:An increase in maternal plasma DHA concentration is initiated in human pregnancy prior to neural tube closure which occurs at 28 d gestation.

Obesity increases preeclampsia risk, and maternal dyslipidemia may result from exaggerated adipocyte lipolysis. We compared adipocyte function in preeclampsia with healthy pregnancy to establish whether there is increased lipolysis. Subcutaneous and visceral adipose tissue biopsies were collected at caesarean section from healthy (n=31) and preeclampsia (n=13) mothers. Lipolysis in response to isoproterenol (200 nmol/L) and insulin (10 nmol/L) was assessed. In healthy pregnancy, subcutaneous adipocytes had higher diameter than visceral adipocytes (P<0.001). Subcutaneous and visceral adipocyte mean diameter in preeclampsia was similar to that in healthy pregnant controls, but cell distribution was shifted toward smaller cell diameter in preeclampsia. Total lipolysis rates under all conditions were lower in healthy visceral than subcutaneous adipocytes but did not differ after normalization for cell diameter. Visceral adipocyte insulin sensitivity was lower than subcutaneous in healthy pregnancy and inversely correlated with plasma triglyceride (r=-0.50; P=0.004). Visceral adipose tissue had lower ADRB3, LPL, and leptin and higher insulin receptor messenger RNA expression than subcutaneous adipose tissue. There was no difference in subcutaneous adipocyte lipolysis rates between preeclampsia and healthy controls, but subcutaneous adipocytes had lower sensitivity to insulin in preeclampsia, independent of cell diameter (P<0.05). In preeclampsia, visceral adipose tissue had higher LPL messenger RNA expression than subcutaneous. In conclusion, in healthy pregnancy, the larger total mass of subcutaneous adipose tissue may release more fatty acids into the circulation than visceral adipose tissue. Reduced insulin suppression of subcutaneous adipocyte lipolysis may increase the burden of plasma fatty acids that the mother has to process in preeclampsia.

Obesity and excessive lipolysis are implicated in preeclampsia (PE). Intrauterine growth restriction is associated with low maternal body mass index and decreased lipolysis. Our aim was to assess how maternal and offspring fatty acid metabolism is altered in mothers in the third trimester of pregnancy with PE (n=62) or intrauterine growth restriction (n=23) compared with healthy pregnancies (n=164). Markers of lipid metabolism and erythrocyte fatty acid concentrations were measured. Maternal adipose tissue fatty acid composition and mRNA expression of adipose tissue fatty acid–metabolizing enzymes and placental fatty acid transporters were compared. Mothers with PE had higher plasma triglyceride (21%, P <0.001) and nonesterified fatty acid (50%, P <0.001) concentrations than controls. Concentrations of major n−6 and n−3 long-chain polyunsaturated fatty acids in erythrocytes were 23% to 60% lower (all P <0.005) in PE and intrauterine growth restriction mothers and offspring compared with controls. Subcutaneous adipose tissue Δ−5 and Δ−6 desaturase and very long-chain fatty acid elongase mRNA expression was lower in PE than controls (respectively, mean [SD] control 3.38 [2.96] versus PE 1.83 [1.91], P =0.030; 3.33 [2.25] versus 1.03 [0.96], P <0.001; 0.40 [0.81] versus 0.00 [0.00], P =0.038 expression relative to control gene [square root]). Low maternal and fetal long-chain polyunsaturated fatty acid concentrations in PE may be the result of decreased maternal synthesis.

Introduction Placental cytokine expression differs between normal and complicated pregnancy (Williams et al, 2009). Interleukin-33 (IL-33) and its receptor ST2 are involved in lipid delivery in maternal adipose tissue. Our study hypothesised that IL-33 is an important pregnancy cytokine which influences early placental development and/or regulates fetal lipid supply. Methods Maternal plasma, adipose tissue and placentas were sampled at caesarean section. Non-pregnant plasma was also analysed. IL-33 and ST2 messenger RNA (mRNA) expression was quantified by real-time PCR. IL-33 and ST2 protein was localised in healthy and complicated placentas by immunohistochemistry and plasma soluble ST2 (sST2) measured by ELISA. Results Third trimester pregnant women had significantly higher plasma sST2 than non-pregnant women (p<0.001). Placental IL-33 and ST2 expression increases across gestation. In early pregnancy, IL-33 and ST2 are localised to syncytiotrophoblasts and later to the syncytium. Placental ST2 expression correlated with plasma sST2 levels in healthy pregnancy (r=0.36; p=0.02). IL-33 mRNA expression was 62% lower in IUGR placentas (p=0.003) and ST2 mRNA expression was 207% higher in pre-eclamptic placentas (p=0.002). In pre-eclamptic pregnancies, visceral adipose tissue ST2 expression was also 80% higher (p=0.02). In complicated pregnancy, IL-33 and ST2 staining appeared generally reduced. Conclusions The IL-33/ST2 signalling pathway plays a role healthy pregnancy. The placenta is responsible for increased circulating sST2 in pregnancy. In IUGR, placental IL-33 is down-regulated, which may reduce circulating maternal triglycerides. In pre-eclampsia, placental ST2 expression increases and may promote increased lipolytic activity. Third trimester plasma sST2 levels did not differ between healthy and complicated pregnancy, indicating ST2 retention for IL-33 signalling.

Objective: We aimed to investigate the effects of progesterone on gene expression and function of both myometrium and circulating leukocytes. Methods: We recruited women participating in a randomized clinical trial of progesterone to prevent preterm delivery. These participants had a twin pregnancy and were managed in 1 of 2 tertiary referral centers. Participants were treated with progesterone (90 mg vaginally) or placebo from 24 to 34 weeks of pregnancy. The outcome measures were myometrial and leukocyte gene expression and expression of cell surface markers in circulating leukocytes, all quantified ex vivo. Results: Prolonged in vivo administration of progesterone inhibited myometrial expression of connexins 26 and 43, endothelial nitric acid synthase (eNOS), and the prostaglandin receptor EP2 ex vivo. Administration of progesterone also increased numbers of circulating neutrophils while decreasing lymphocyte proportions and decreasing neutrophil CD11b expression. Conclusion: The observed effects of prolonged in vivo administration of progesterone will minimize the ability of the uterus to contract as a synctium and the ability of peripheral blood leukocytes to migrate into the myometrium during parturition. We suggest that these are putative mechanisms by which progesterone might prevent preterm birth in women at high risk.

Objective. To define the role of Toll-like receptors (TLR-2, TLR-4) and triggering receptor expressed on myeloid cells-1 (TREM-1) in pregnant human myometrium. Study Design. Expression of TLR-2 and -4 mRNA and protein and TREM-1 mRNA was quantified in human myometrial samples. Subsequently, we investigated the effect of medroxyprogesterone acetate (MPA) as a functional inhibitor of the TLR agonist lipopolysaccharide (LPS). Results. Messenger RNA expression of TLR-2 and -4 was significantly higher in myometrium at term compared with preterm (P = .009 and .016, respectively). Toll-like receptor-2 protein expression was significantly higher during labor (P = .002) compared with nonlaboring samples. Triggering receptor expressed on myeloid cells-1 mRNA expression was increased in both myometrium and cervix after labor at term (P < .05). Medroxyprogesterone acetate significantly suppressed LPS-induced production of interleukin 1b (IL-1b), IL-6, and IL-8 in vitro (P < .05). Conclusion. Toll-like receptors 2and 4 and TREM-1 are expressed in human myometrium and may play a role in the mechanism of labor at term, and their functional effects are inhibited by MPA.

We hypothesized that the priming and activation of maternal leukocytes in peripheral blood is a key component of parturition, and that inappropriate preterm priming of leukocytes might initiate preterm labour and delivery. The purpose of this study was to characterize peripheral blood leukocyte activation during human term and preterm labour. We obtained blood samples from pregnant women at term and preterm, both in labour and not in labour. Leukocyteswere characterized according to cell subtype and cell surface marker expression. Additionally, we quantified leukocyte cytokine mRNA production, migratory ability and reactive oxygen species production of neutrophils and macrophages. We found that both term and preterm labour were associated with an increase in monocyte and neutrophil proportion or number-neutrophil migratory ability and cell surface marker expression indicating activation. Messenger RNA expression of IL-1 beta and IL-8, MCP-1 and TLR-2 was also increased. We conclude that leukocytes in peripheral blood are primed in preparation for activation during term and preterm labour, and that this may contribute to the pathophysiological events of parturition. These data may lead to novel therapies and diagnostic tools for the prevention and/or diagnosis of preterm birth.