Early pregnancy termination in rats immunized with denatured chicken riboflavin carrier protein

Immunoneutralization of the maternal riboflavin carrier protein in the pregnant rat with antibodies to chicken egg vitamin carrier has earlier been shown to terminate their pregnancies. In order to understand the nature of the epitopic conformations capable of eliciting antibodies bioneutralizing the endogenous riboflavin carrier protein in the pregnant rat, we compared pregnancy progression in the fertile rodents following active immunization with either the native, SDS-denatured, reduced-carboxymethylated or SDS-treated reduced carboxymethylated avian egg white riboflavin carrier protein. The data revealed that despite the total antibody titers being higher in the animals immunized with the native protein, the antibodies elicited against the denatured avian vitamin carrier exhibited relatively better potencies to bioneutralize the endogenous maternal protein as evidenced by higher rates of early fetal resorption.

Substance misuse problems during pregnancy with special emphasis on buprenorphine

Maternal drug abuse during pregnancy endangers the future health and wellbeing of the infant and growing child. On the other hand, via maternal abstinence, these problems would never occur; so the problems would be totally preventable. Buprenorphine is widely used in opioid maintenance treatment as a substitute medication. In Finland, during 2000 s buprenorphine misuse has steadily increased. In 2009 almost one third of clientele of substance treatment units were in treatment because of buprenorphine dependence. At Helsinki Women s Clinic the first child with prenatal buprenorphine exposure was born in 2001. During 1992-2001 in the three capital area maternity hospitals (Women s clinic, Maternity hospital, Jorvi hospital) 524 women were followed at special antenatal clinics due to substance abuse problems. Three control women were drawn from birth register to each case woman and matched for parity and same place and date of the index birth. According to register data mortality rate was 38-fold higher among cases than controls within 6-15 years after index birth. Especially, the risk for violent or accidental death was increased. The women with substance misuse problems had also elevated risk for viral hepatitis and psychiatric morbidity. They were more often reimbursed for psychopharmaceuticals. Disability pensions and rehabilitation allowances were more often granted to cases than controls. In total 626 children were born from these pregnancies. According to register data 38% of these children were placed in out-of-home care as part of child protection services by the age of two years, and half of them by the age of 12 years, the median follow-up time was 5.8 years. The risk for out-of-home care was associated with factors identifiable during the pre- and perinatal period. In 2002-2005 67 pregnant women with buprenorphine dependence were followed up at the Helsinki University Hospital, Department of Obstetrics and Gynecology. Their pregnancies were uneventful. The prematurity rate was similar and there were no more major anomalies compared to the national statistics. The neonates were lighter compared to the national statistics. They were also born in good condition, with no perinatal hypoxia as defined by standard clinical parameters or certain biochemical markers in the cord blood: erythropoietin, S100 and cardiac troponin-t. Almost 80% of newborns developed neonatal abstinence syndrome (NAS) and two third of them needed morphine medication for it. Maternal smoking over ten cigarettes per day aggravated and benzodiazepine use attenuated NAS. An infant s highest urinary norbuprenorphine concentration during their first 3 days of life correlated with the duration of morphine treatment. The average length of infant s hospital stay was 25 days.

Luteal-phase defect induced by deprivation of FSH at a specific period of the follicular phase prevents pregnancy in the bonnet monkey (Macaca radiata)

Female bonnet monkeys were injected i.v. with 25 µl antiserum to FSH on Days 5, 6 or 7 of the cycle: the length of the luteal phase was shortened but there was no alteration in cycle length. Proven fertile females (N = 6) were caged throughout the period of the experiment (6 cycles) with proven fertile males and treated with 25 µl FSH antiserum on Day 7 of each of 3 successive cycles. Out of 18 cycle exposures during the treatment phase, 17 were ovulatory, but no pregnancies occurred. In the post-treatment phase, 5 monkeys became pregnant within 3 cycle exposures. These results show that it is possible to render female monkeys infertile by creating luteal insufficiency and this can be achieved repeatedly in a reproducible manner by depriving the cyclic females of FSH support on Day 7 of consecutive cycles.

Effect of a specific estrogen antibody on pregnancy establishment in the bonnet monkey (Macaca radiata)

The requirement for estrogen for pregnancy establishment has not been conclusively demonstrated in primates. Selective neutralization of estrogens was achieved in mated female monkeys during preimplantation and postimplantation periods by injecting characterized estrogen antiserum from either day 14 to 18 or day 28 to 32 of cycle. While estrogen deprivation during preimplantation period in 5 animals exposed to 14 ovulatory cycles resulted in only one pregnancy, only 3 of 13 monkeys treated during postimplantation period continued pregnancy to term. In comparison with controls (4 of 5 monkeys becoming pregnant), the percent protection against pregnancy in animals treated during preimplantation period was 93. The pregnancy termination in 10 of 13 monkeys treated during postimplantation period when compared with normal postimplantation pregnancy wastage in our colony (2%) is also highly significant (P less than 0.01). The present study demonstrates a critical need for estrogen during the peri-implantation period for a successful pregnancy establishment in primates.

Antigenic determinants at the carboxy terminus of chicken egg white riboflavin carrier protein (RCP): Epitope mapping and antibody-mediated pregnancy curtailment in rodents

The epitopic core sequences recognized by three monoclonal antibodies raised to chicken riboflavin carrier protein (RCP) were mapped to the C-terminal tail-end of the protein using the pepscan method A 21-residue synthetic peptide corresponding to residues 200-219 of the protein and comprising the regions corresponding to the antibodies was synthesized. Administration of polyclonal antibodies specific to this peptide led to termination of early pregnancy in mice. Also, active immunization of rats with the peptide-purified protein derivative conjugate inhibited establishment of pregnancy. These results demonstrate the functional importance of the C-terminal 200-219 region of chicken RCP. Copyright (C) 1996 Elsevier Science Ltd.

Determinants of self-reported smoking and misclassification during pregnancy, and analysis of optimal cut-off points for urinary cotinine: a cross-sectional study

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Changes of urinary steroid conjugates and gonadotropin excretion in the menstrual cycle and pregnancy in the Yunnan snub-nosed monkey (Rhinopithecus bieti)

The Yunnan snub-nosed monkey (Rhinopithecus bieti) is one of the most endangered species in the world, and it is endemic to China. According to our knowledge, there was no information on reproduction for this species. The present study was designed to und

Prediction and prevention of venous thrombosis in pregnancy

Venous thromboembolism (VTE) remains the leading cause of maternal mortality. Reports identified further research is required in obese and women post caesarean section (CS). Risk factors for VTE during pregnancy are periodically absent indicating the need for a simple and effective screening tool for pregnancy. Perturbation of the uteroplacental haemostasis has been implicated in placenta mediated pregnancy complications. This thesis had 4 main aims: 1) To investigate anticoagulant effects following a fixed thromboprophylaxis dose in healthy women post elective CS. 2) To evaluate the calibrated automated thrombogram (CAT) assay as a potential predictive tool for thrombosis in pregnancy. 3) To compare the anticoagulant effects of fixed versus weight adjusted thromboprophylaxis dose in morbidly obese pregnant women. 4) To investigate the LMWH effects on human haemostatic gene and antigen expression in placentae and plasma from the uteroplacental , maternal and fetal circulation. Tissue factor pathway inhibitor (TFPI), thrombin antithrombin (TAT), CAT and anti-Xa levels were analysed. Real-time PCR and ELISA were used to quantify mRNA and protein expression of TFPI and TF in placental tissue. In women post CS, anti-Xa levels do not reflect the full anticoagulant effects of LMWH. LMWH thromboprophylaxis in this healthy cohort of patients appears to have a sustained effect in reducing excess thrombin production post elective CS. The results of this study suggest that predicting VTE in pregnant women using CAT assay is not possible at present time. The prothrombotic state in pregnant morbidly obese women was substantially attenuated by weight adjusted but not at fixed LMWH doses. LMWH may be effective in reducing in- vivo thrombin production in the uteroplacental circulation of thrombophilic women. All these results collectively suggest that at appropriate dosage, LMWH is effective in attenuating excess thrombin generation, in low risk pregnant women post caesarean section or moderate to high risk pregnant women who are morbidly obese or tested positive for thrombophilia. The results of the studies provided data to inform evidence-based practice to improve the outcome for pregnant women at risk of thrombosis.

The expression and regulation of pregnancy-specific glycoproteins in the mouse

Pregnancy-Specific Glycoproteins (PSG) are the most abundant fetally expressed proteins in the maternal bloodstream at term. This multigene family are immunoglobulin superfamily members and are predominantly expressed in the syncytiotrophoblast of human placenta and in giant cells and spongiotrophoblast of rodent placenta. PSGs are encoded by seventeen genes in the mouse and ten genes in the human. Little is known about the function of this gene family, although they have been implicated in immune modulation and angiogenesis through the induction of cytokines such as IL-10 and TGFβ1 in monocytes, and more recently, have been shown to inhibit the platelet-fibrinogen interaction. I provide new information concerning the evolution of the murine Psg genomic locus structure and organisation, through the discovery of a recent gene inversion event of Psg22 within the major murine Psg cluster. In addition to this, I have performed an examination of the expression patterns of individual Psg genes in placental and non-placental tissues. This study centres on Psg22, which is the most abundant murine Psg transcript detected in the first half of pregnancy. A novel alternative splice variant transcript of Psg22 lacking the protein N1-domain was discovered, and similar to the full length isoform induces TGFβ1 in macrophage and monocytic cell lines. The identification of a bidirectional antisense long non-coding RNA transcript directly adjacent to Psg22 and its associated active local chromatin conformation, suggests an interesting epigenetic gene-specific regulatory mechanism that may be responsible for the high level of Psg22 expression relative to the other Psg family members upon trophoblast giant cell differentiation

Vitamin D status and requirements during pregnancy and lactation

Vitamin D deficiency during pregnancy, lactation, and early infancy has been widely reported. Current understanding of vitamin D metabolism during pregnancy and lactation is incomplete, and to date, experimental data to support vitamin D requirements for these life stages are scarce. There is a shortage of nationally representative data and appropriate reference ranges for serum 25-hydroxyvitamin D (25OHD) during pregnancy, lactation and infancy, including in umbilical cord blood. This thesis described concentrations of total 25OHD and individual metabolites including 25OHD3, 25OHD2, and 3-epi-25OHD3 at 15 weeks’ gestation in a large seasonally balanced pregnancy cohort study (n 1768), carried out in Cork, Ireland (52oN). The prevalence of low 25OHD concentrations in pregnant women was higher than published reports in other Caucasian women, and was highest among non-users of vitamin D-containing supplements during winter. A longitudinal pregnancy study was included which suggested gestational stages had an impact on the total serum 25OHD concentration. This thesis incorporated a randomized controlled trial carried out among 100 women across 3 intervention groups using 20 μg/day of vitamin D3 with or without 500 mg calcium, or placebo, over 12-weeks of lactation to investigate the vitamin D requirement for lactating mothers and the vitamin D content of human milk. A daily intake of 25 μg/day was suggested to meet the requirement of lactating women to maintain a 25OHD levels above 50 nmol/L in 97.5% of the population at 52oN all year around. However, vitamin D content in human milk did not increase in response to supplementation. Serum 25OHD concentration has been used as a predictor of a number of health outcomes. This thesis reported large differences in serum 25OHD concentrations using different methods in 86 umbilical cord samples. The need for international standardization of serum 25OHD measurements was re-emphasized in this thesis.

Pregnancy-specific glycoprotein function, conservation and receptor investigation

Pregnancy-specific glycoproteins (PSGs) are highly glycosylated secreted proteins encoded by multi-gene families in some placental mammals. They are carcinoembryonic antigen (CEA) family and immunoglobulin (Ig) superfamily members. PSGs are immunomodulatory, and have been demonstrated to possess antiplatelet and pro-angiogenic properties. Low serum levels of these proteins have been correlated with adverse pregnancy outcomes. Objectives: Main research goals of this thesis were: 1). To attempt to replicate previously reported cytokine responses to PSG-treatment of immune cells and subsequently to investigate functionally important amino acids within PSG1. 2). To determine whether candidate receptor, integrin αvβ3, was a binding partner for PSG1 and to investigate whether PSG1 possessed functionality in a leukocyte-endothelial interaction assay. 3). To determine whether proteins generated from recently identified putative PSG genes in the horse shared functional properties with PSGs from other species. Outcomes: 1). Sequential domain deletion of PSG1 as well as mutation of conserved residues within the PSG1 Ndomain did not affect PSG1-induced TGF-β1. The investigated response was subsequently found to be the result of latent TGF-β1 contaminating the recombinant protein. Protein further purified by SEC to remove this showed no induction of TGF-β1. The most N-terminal glycosylation site was demonstrated to have an important role in PSG N domain secretion. PSG1 attenuated LPS-induced IL-6 and TNF-α. Investigations into signalling underpinning this proved inconclusive. 2). Integrin αvβ3 was identified as a novel PSG1 receptor mediating an as yet unknown function. Preliminary investigations into a role for PSGs as inhibitors of leukocyte endothelial interactions showed no effect by PSG1. 3). Horse PSG protein, CEACAM49, was shown to be similarly contaminated by latent TGF-β1 particle and once removed did not demonstrate TGF-β1 release. Interestingly horse PSG did show anti-platelet properties through inhibition of the plateletfibrinogen interaction as previously published for mouse and human PSGs.

Caesarean section and subsequent pregnancy outcome: a Danish register-based cohort study

Background and Aims: Caesarean section rates have increased in recent decades and the effects on subsequent pregnancy outcome are largely unknown. Prior research has hypothesised that Caesarean section delivery may lead to an increased risk of subsequent stillbirth, miscarriage, ectopic pregnancy and sub-fertility. Structure and Methods: Papers 1-3 are systematic reviews with meta-analyses. Papers 4-6 are findings from this thesis on the rate of subsequent stillbirth, miscarriage, ectopic pregnancy and live birth by mode of delivery. Results Systematic reviews and meta-analyses: A 23% increased odds of subsequent stillbirth; no increase in odds of subsequent ectopic pregnancy and a 10% reduction in the odds of subsequent live birth among women with a previous Caesarean section were found in the various meta-analyses. Danish cohorts: Results from the Danish Civil Registration System (CRS) cohort revealed a small increased rate of subsequent stillbirth and ectopic pregnancy among women with a primary Caesarean section, which remained in the analyses by type of Caesarean. No increased rate of miscarriage was found among women with a primary Caesarean section. In the CRS data, women with a primary Caesarean section had a significantly reduced rate of subsequent live birth particularly among women with primary elective and maternal-requested Caesarean sections. In the Aarhus Birth Cohort, overall the effect of mode of delivery on the rate and time to next live birth was minimal. Conclusions: Primary Caesarean section was associated with a small increased rate of stillbirth and ectopic pregnancy, which may be in part due to underlying medical conditions. No increased rate of miscarriage was found. A reduced rate of subsequent live birth was found among Caesarean section in the CRS data. In the smaller ABC cohort, a small reduction in rate of subsequent live birth was found among women with a primary Caesarean section and is most likely due to maternal choice rather than any ill effects of the Caesarean. The findings of this study, the largest and most comprehensive to date will be of significant interest to health care providers and women globally.