PoSteRS
eARly PRegNANCy
P-117 Homozygocity for an insertion in the HlA-g gene is associated with recurrent miscarriage and low birthweight in children born prior to the diagnosis
O. Christiansen1, M. Dahl2, H.S. Nielsen1, A.M. Kolte3, E.C. Larsen1,
T.V. Hviid4
1Rigshospitalet, Fertility Clinic 4071, Copenhagen, Denmark
2Roskilde University Hospital, Department of Clinical Biochemistry, Roskilde,
Denmark
3Aalborg University Hospital, Department of Obstetrics and Gynaecology,
Aalborg, Denmark
4Roskilde University Hospital, Department of Clinic Biochemistry, Roskilde,
Denmark
Introduction: HLA-G is thought to play an important role in the immuological
recognition of pregnancy since it is the main HLA molecule expressed on the trophoblast and it seems to display immunosuppressive properties during nor-mal implantation and pregnancy. A 14 basepair (bp) long insertion in exon 8 of the HLA-G gene seems to decrease stability of the HLA-G transcript.
We have previously found that plasma concentrations of soluble HLA-G are lower in homozygous carriers of this HLA-G14bp insertion. In theory, homozy-gous carriers of the HLA-G14bp insertion are at an increased risk of impaired trophoblast invasion and growth since immune protection is defect.
Material and Methods: We investigated the HLA-G14bp insertion/deletion
polymorphism in 301 Caucasian patients with three or more consecutive mis-carriages that remained unexplained after routine anatomical, chromosomal, endocrine and autoimmune screening and in 168 Caucasian women with two or more children and no miscarriages. The HLA-G14bp polymorphisms were investigated after amplification of the HLA-G exon 8 using genomic DNA and primers GE14HLAG and RHG4.
Results: In all recurrent miscarriage patients 56 (18.6 %) were homozygous for
the HLA-G14bp insertion compared with 19 (11.3%) of the controls (p < 0.05). HLA-G14bp insertion homozygocity was found in 19.2% of patients with sec-ondary recurrent miscarriage (p < 0.05 compared with controls) and 17.7% of patients with primary recurrent miscarriage (not significant compared with controls).
Among the children born prior to a secondary recurrent miscarriage diag-nosis, the mean birth weight was 2766.3 g in 35 children born to HLA-G14bp homozygous women, which was significantly lower than the mean birth weight of 3082.7g in the 119 children born to women who were HLA-G14bp insertion/ deletion heterozygous or HLA-G14bp deletion homozygous (p < 0.01).
Conclusions: Homozygocity for the HLA-G14bp insertion seems to be
associ-ated with recurrent miscarriage and especially secondary recurrent miscarriage. In first pregnancies of patients with secondary recurrent miscarriage, maternal carriage of this HLA-G14bp genotype seems to predispose to birth of children with significantly reduced birth weight compared with children born to similar patients with other HLA-G14 bp genotypes. HLA-G14bp insertion homozy-gocity may affect maternal immune tolerance to the trophoblast thereby causing miscarriage or intrauterine growth restriction.
P-118 Accuracy of new clearblue digital pregnancy test with conception indicator
J. Pike1, J. Ellis2, S. Johnson3, R. Shaw4, P. Parkinson5, P. Perry6
1SPD, Medical Marketing, Bedford, United Kingdom 2SPD, External Affairs, Bedford, United Kingdom
3SPD, Clinical & Medical Affairs, Bedford, United Kingdom 4SPD, Data and Statistics, Bedford, United Kingdom 5SPD, Clinical Management, Bedford, United Kingdom 6SPD, Research & Development, Bedford, United Kingdom
objective: The Clearblue Digital Pregnancy Test with Conception Indicator
is the latest advance in home pregnancy testing and is available on the market in Europe. It is the very first pregnancy test that is capable of not only telling
2Burnley General Hospital, Obsterics and Gynaecology, Burnley, United
Kingdom
Introduction: World Health Organization predicts there will be 2.3 billion
overweight adults in the world by 2015 and more than 700 million of them will be obese.
In 2006, 24% of adults (aged 16 or over) in England were classified as obese. This represents an overall increase from 15% in 1993. Among all other problems women who are obese have higher rates of amenorrhea and infertil-ity. Obese women have a higher risk of complications during pregnancy such as hypertensive disorders and gestational diabetes. They are at a higher risk of operative delivery and prolonged labour.
Aims of the study:
1. To evaluate the awareness of complications posed by obesity among women with raised BMI (>30) attending fertility clinic in a United Kingdom NHS Hospital.
2. To educate women about the risks of obesity and provide adequate support.
Methodology:
A prospective survey using a predesigned health questionnaire Study period: 6 months.
Subjects: All Women attending reproductive medicine clinic with history of primary or secondary infertility.
Results: The study is still ongoing but preliminary results show 75% of obese
women attending the reproductive clinic is unaware of their Body Mass Index or current weight. 2/3 of these women are unaware of effects of obesity on ovulation or effects of obesity on pregnancy and child birth. Only half of these women have tried some tested means to achieve weight loss before seeking help for their fertility issues.
Conclusion: Overweight and obesity have negative effects on the health of
both mother and child. Obesity costs will rise enormously as more women be-come overweight and obese.
Prevention of obesity at an early age is ideal but difficult to achieve. Obese women wishing pregnancy should be informed about the potential complica-tions of obesity in pregnancy. However for obese women with infertility prob-lems, weight loss programmes should be the first choice of treatment to achieve conception.
P-116 A comparative analysis of assisted reproductive technology cycles in Australia and New Zealand 2004-2007
C. Farquhar1, Y.A. Wang2, A.E. Sullivan2
1University of Auckland, Univ of Auckland - Dept. of Ob/Gyn, Auckland,
New Zealand
2University of New South Wales, 1Perinatal and Reproductive Epidemiology
Research Unit, Sydney, Australia
Introduction: There are marked differences in the funding arrangements for
fertility investigations and treatments between New Zealand (NZ) and Aus-tralia. In NZ public funding of fertility treatments is limited to women under 40 years old, with a BMI less than 32 kg/m2, do not smoke and who meet certain clinical critera. In Australia there are no restriction on access to fertil-ity treatments.
Methods: A comparative analysis of data extracted from the Australia and New
Zealand Assisted Reproduction Database, which includes autologous ART treatment cycles was undertaken between 2004 and 2007. A total of 116111 autologous fresh cycles were included.
Results: Women undergoing treatment in Australia women were more likely
older (>40 years; 23.5 % vs 16.0%, p < 0.01) compared to women having treat-ment in NZ. There were higher rates of blastocyst transfers and double embryo transfers in all age groups in Australia than NZ. In women aged less than 35 years the crude rates of clinical pregnancy (31.2% vs 37.5%, p < 0.01) and live delivery (26% vs 31.6%, p < 0.01) following fresh ART cycles were signifi-cantly higher in New Zealand than Australia. These differences in the type of treatment and the outcomes persisted in older age groups.
Conclusions: There are major differences in the funding arrangements for
fer-tility services in Australia and NZ which in turn has lead to a healthier popula-tion undergoing treatment and fewer embryos being transferred in NZ. These differences may explain the improved pregnancy outcomes in NZ.
compared with sera from group B or group C. This molecule was also signifi-cantly higher in the sera from group D, compared with the sera from group E.
The spermidine level was significantly higher in sera obtained from mice at 24 and 48 hours after mating. Spermidine was not detected in sera obtained from pseudopregnant mice with or without viable embryos.
Conclusions: Although not highly specific for pregnancy, hypotaurine might
be a biomarker of early pregnancy during the pre-implantation period. Sper-midine, which is thought to derive from sperm, remains in the blood for up to 48 hours after mating and might be useful as a marker of mating.
P-120 Chlamydia trachomatis elevates prokineticin receptor 2 in fallopian tube: a possible explanation for the link between chlamydia and ectopic pregnancy
J.L.V. Shaw1, G.S. Wills2, M.O. McClure2, P.J. Horner3, A. Colgan2, H.N.
Jabbour1, H.O.D. Critchley1, G. Entrican4, A.W. Horne1
1University of Edinburgh, Centre for Reproductive Biology, Edinburgh, United
Kingdom
2Imperial College London, Jefferiss Trust Laboratories, London, United
Kingdom
3University of Bristol, Department of Social Medicine, Bristol,
United Kingdom
4Moredun Research Institute, Immunology, Edinburgh, United Kingdom
Introduction: Tubal ectopic pregnancy is a significant cause of maternal
mor-bidity and mortality (1). Current evidence supports the hypothesis that tubal ectopic pregnancy is caused by a combination of retention of the embryo within the Fallopian tube due to impaired embryo-tubal transport and alterations in the tubal environment allowing early implantation to occur (2). The main risk factor for tubal ectopic pregnancy is Chlamydia trachomatis infection, how-ever the mechanism underlying cause of this association is not well understood. We have previously demonstrated altered expression of prokineticin receptor 1 and 2 (PROKR1 and PROKR2) in Fallopian tube from women with tubal ectopic pregnancy (3). PROKR1 and PROKR2 are G-protein coupled receptors for prokineticins 1 and 2 (PROK1 and PROK2). PROKs are known for their pro-angiogenic functions, the regulation of the expression of genes important for embryo implantation and the control of smooth muscle contractility. C.
tra-chomatis has been shown to initiate signaling from members of the toll-like
receptor (TLR) family of pattern recognition proteins, specifically TLR2 and TLR4. We hypothesized that infection of the Fallopian tube by C. trachomatis would result in activation of TLR signaling and altered PROKR expression that predisposes the tubal environment to implantation.
Materials and Methods: Sera and Fallopian tube biopsies were obtained from
fertile women undergoing hysterectomy for benign conditions at time of sur-gery (n = 21). A new ELISA (4), which measures the levels of Pgp3 anti-bodies in patient serum was used to confirm past chlamydial infection. PgP is a protein secreted by C. trachomatis into the cytosol of host cells. Immunohis-tochemistry was used to localize the expression of TLR2 and TLR4 in human Fallopian tube. Fallopian tube explants and immortalized oviductual epithelial cells (OE-E6/E7) were exposed to C. trachomatis and the TLR2 ligand, Pam3-Cys-SerLys (Pam3CSK). Taqman RT-PCR and Western blotting were used to quantify PROKR, TLR2 and TLR4 levels.
Results: PROKR2 levels were higher in Fallopian tube from women with
sero-logical evidence of previous C. trachomatis infection (as measured by ELISA). PROKR2 levels were significantly increased in Fallopian explants (p < 0.01) and OE-E6/E7 cells (p < 0.01) exposed to either UV-killed C. trachomatis (equivalent multiplicity of infection [MOI] of 1.0) or live C. trachomatis (MOI 1.0) for 8 hours, This suggests a sensory mechanism on the cell surface since infection was not required. TLR2 and TLR4 were localized to the epithelium of the Fallopian tube by immunohistochemistry.TLR2 levels were significantly increased in OE-E6/E7 cells following exposure to UV-killed C. trachomatis (MOI 1.0) and live C. trachomatis (MOI 1.0) (p < 0.05) (8 hour exposure). PROKR2 levels were also increased in OE-E6/E7 cells treated with the TLR2 ligand, Pam3CSK (p < 0.05), and the effect was dose dependent.
Conclusions: We provide evidence that TLR2 ligation causes increased
expres-sion of PROKR2 and demonstrate a link between C. trachomatis exposure and PROKR2 expression in the Fallopian tube epithelium. We have previously dem-onstrated increased expression of pro-angiogenic factors and pro-inflammato-ry cytokines involved in implantation following PROK treatment of OE-E6/ E7 cells. We propose that TLR2 signaling, activated by C. trachomatis in the a woman if she is pregnant, but if so, indicating when she conceived. This
is achieved with a Dual Hormone Sensor and digital display that further cat-egorises ‘Pregnant’ results into one of 3 groups; 1-2, 2-3 or 3 + weeks since conception.
The performance of the pregnancy test in providing a ‘Pregnant’ result and correctly categorising the time since conception was assessed using urine sam-ples from pregnant women.
Materials and Methods: Daily urine samples from 84 fertile women seeking
conception, with cycle lengths 21-42 days, were collected until 42 days post-conception. LH surge was identified in the laboratory using home ovulation tests, and samples from day -4 to day 28 (relative to day of expected period) were tested. The success of the test in providing a ‘Pregnant’ result for each day of pregnancy was calculated. The accuracy of the Conception Indicator result was also determined.
Results: The test was over 99% accurate for pregnancy detection when used on
the day the period was due. The detection rates for early testing were: 98% one day before the period was due; 97% two days before; 86% three days before and 55% four days before. The Conception Indicator feature of the test was found to be 92% accurate.
Conclusion: The new Clearblue Digital Pregnancy Test with Conception
In-dicator shows excellent performance characteristics with regards to accuracy, and is a valuable innovation in providing the answers to the two most important questions a woman has when using a pregnancy test: ‘Am I pregnant?’ and ’If so, when did I conceive?’.
Support: Study investigators all employees of, and study funded by, SPD
De-velopment Company Limited, Bedford, UK.
P-119 Metabolomic analysis of sera samples from pregnant mice during pre-implantation period
N. Kuji1, N. Okumura1, M. Takano1, S. Ogawa1, M. Yamada1, T. Hamatani1, Y.
Yoshimura1, M. Kawakami2, A. Hirayama2, M. Tomita2
1Keio University School of Medicine, Ob/Gyn Dept., Tokyo, Japan 2The Institute for Advanced Biosciences Keio University, Systems Biology,
Yamagata, Japan
Introduction: The maternal system for recognizing a fetus, which is
immuno-logically alien to the mother, from the moment of fertilization has been studied, with Morton et al. reporting that early pregnancy factor (EPF) appears in ma-ternal serum within 6 – 24 h after conception. However, the precise molecu-lar structure of EPF has not yet been determined, and no other molecule has been identified as an early pregnant marker prior to the appearance of human chorionic gonadotropin (hCG) in the blood. Recently, metabolomic analysis has been introduced as a high-throughput analytical procedure for identifying biomarkers of ionic low-molecular weight molecules. In this study, we inves-tigated sera samples obtained from pregnant mice during the pre-implantation period in an attempt to identify a biomarker of early pregnancy that could be used to detect pregnancy before implantation or before the detection limit of hCG has been reached.
Materials and Methods: In the first experiment, pseudopregnant ICR mice
were subjected to the transfer of 1) pronuclear-stage embryos (about 10 embry-os per mouse; group A), 2) unfertilized oocytes (about 10 oocytes per mouse; group B), or 3) culture media alone (group C). Forty-eight hours later, up to 1.0 mL of blood was collected from the cardiac cavity of each mouse after con-firming the existence of 8 cell embryos inside the fallopian tube-uterine horn. Blood samples were equilibrated at 4°C overnight, and sera samples were then deep-frozen at -80°C until use.
In the second experiment, superovulated ICR mice were 1) mated with re-tired male mice (group D) or 2) not mated (group E). The mice were then sacri-ficed, and blood samples were collected from the cardiac cavity of each mouse after 24, 48, 72, or 96 hours after mating. The existence of embryos inside the fallopian tube–uterine horn was confirmed for the group D mice. The serum samples were prepared as described above and frozen at -80°C until use.
Ionic low molecular weight compounds in the serum were analyzed and quantified using capillary electrophoresis time-of-flight mass spectrometry (CE-TOFMS). The raw data were calculated using Master Hands software, which we developed. A panel of 303 cationic and anionic metabolites was mea-sured.
Results: Among the 303 low-molecular weight chemicals that were
comprehensive diagnostic follow-up (median number of abortions 3 (range 2-6), > 4 in 43%, > 5 miscarriages in 19%). To evaluate different complement pathways, blood was taken for determination of MBL using a) a quantitative enzyme-linked immunosorbent assay (ELISA) (MBL A), b) a functional lectin pathway assay (MBL B), and in parallel, for CH 50 determination (hemolytic titration) and c) in aliquots of the same serum samples. 136 obstetrically normal women served as a control. Results were statistically analyzed (e.g. Spearman-rank correlation, Wilcoxon Spearman-rank-sum test, Chi-square analysis) with regard to patients´ history, outcome of a broad microbial screening in the lower genital tract and multiple relevant laboratory determinants.
Results: A significant correlation between MBL A and MBL B was found (r
0.8) (p < 0.0001). There was no marked correlation of MBL (A or B) with CH50. Median levels of CH 50, but not of MBL (A and B) were significantly lower in the study group with repeated abortions compared to the obstetrically normal controls (p < 0.0001). MBL deficiency (as defined as MBL A < 50 ng/ ml) was not significantly more prevalent in patients with recurrent miscarriages compared to the controls (5.8%/ 4.4%). However, there was some association of MBL and CH 50 with variables influencing the coagulation cascade (eg. protein S) (p < 0.05). MBL levels were not related to anti-nucleic Ab (ANA), but were significantly associated with thyroid autoimmunity (eg. TPO Ab) (p < 0.04). Results of MBL A and B, as well as of CH50 examination, in the study group were not significantly related to patients´ age, the duration of infertility, the time frame of fetal loss and the number of abortions. The three determinants of complement activity (MBL A, B, CH 50) were not significantly associated with a history of previous genital infections, or with same-day microbial findings in the lower genital tract (mycoplasmas and other pot. path. bacteria, yeasts), and there was no relationship with chlamydial serology [Chlamydia trachomatis Ig G and Ig A antibodies] in same-day serum samples.
Conclusions: The complement system plays an important part in the body´s
immune system. The prevalence of marked MBL deficiency in women suffer-ing from recurrent miscarriages is low. However, results suggest an association of MBL and complement with some autoimmune factors influencing patients´ endocrine status, and also a potential impact on the course of pregnancy via in-terference with fibrinolytic activity. Therefore the investigation of complement in these patients deserves further attention.
P-123 bloodborn angiogenic factors and multiple implantation, a comparison of singleton and twin pregnancies
M.J. Lambers1, E. Groeneveld1, D.A. Hoozemans1, R. Schats1, P.G.A.
Hompes1, C.B. Lambalk1
1VUMC Hospital, Gynaecology Obstetrics and Reproductive Medicine,
Am-sterdam, The Netherlands
Introduction: Recent in vitro fertilisation (IVF) studies have shown that
preg-nancies with double implantations have a lower chance of pregnancy loss than pregnancies with single implantations, reflected by a lower incidence of com-plete pregnancy loss and a lower pregnancy loss per gestational sac. While the chance of multiple implantation at 6 weeks of gestation is mainly determined by (morphological) embryo quality, the chance of continuation of multiple pregnancies beyond 6 weeks of gestation becomes more dependent on mater-nal factors, affecting the uterine milieu. Among these are factors involved in angiogenesis, a process which is essential for embryo implantation, and thus probably also for the maintenance of implanted embryos. The aim of this study was to analyse the potential role of a number of angiogenic factors in sustained multiple pregnancy by measuring their serum concentrations in IVF treatments with double embryo transfer (DET) and comparing these concentrations be-tween ongoing singleton and twin pregnancies.
Material and Methods: In this prospective study we studied sixteen ongoing
singleton and nine ongoing twin pregnancies after DET. We determined longitu-dinal serum concentrations of Vascular Endothelial Growth Factor-A (VEGF-A), Inhibin A, Glycodelin A, Insulin-like growth factor-I (IGF-I), Insulin-like growth factor-II (IGF-II), Insulin-like growth factor binding protein-1 (IGFBP-1) and Insulin-like growth factor binding protein-3 (IGFBP-3) at baseline before any intervention, during the IVF treatment and in early pregnancy.
Results: Baseline VEGF-A levels and Body Mass Index (BMI) are significantly
higher in women who achieve an ongoing twin pregnancy compared to women who achieve an ongoing singleton pregnancy (p = 0.04). Levels of Inhibin A in women with twin pregnancies were significantly higher from early implanta-tion onwards (14-15 days after OPU, p = 0.02), but not before pregnancy. We Fallopian tube epithelium, results in increased levels of PROKR2 and
attenu-ated PROK signaling, resulting in a tubal environment predisposed to implan-tation. These findings may explain the link between C. trachomatis infection ectopic pregnancy.
References:
1 Farquhar (2005) The Lancet, 366: 583-591.
2 Shaw et al. (2010) Human Reproduction Update, Epub ahead of print. 3 Shaw et al. (2010) Fertility and Sterility, Epub ahead of print. 4 Wills et al. (2009) Clinical and Vaccine Immunology, 16: 835-843.
P-121 early spontaneous miscarriage after single and double blastocyst transfer - An analysis of 1020 blastocyst transfers
M. Dosen1, V. Vlaisavljevic1, B. Kovacic1
1University Clinical Centre Maribor, Reproductive Medicine and
Gynecologi-cal Endocrinology, Maribor, Slovenia
Introduction: Early spontaneous miscarriage (SM) in pregnancies achieved by
assisted reproduction procedures was evaluated almost exclusively in cycles in which pre-blastocyst stage embryos were transferred. The primary objective of this retrospective study was to evaluate the association between several impor-tant cycle parameters and SM after blastocyst transfer.
Materials and Methods: Using multivariable logistic regression we analyzed
1020 consecutive IVF/ICSI cycles with positive β-hCG after single or dou-ble blastocyst transfer. Age of the patient, fertilization method (IVF or ICSI), number of retrieved oocytes, number of developed blastocysts, number of blastocysts suitable for freezing, number of transferred blastocysts and quality of transferred blastocysts were analyzed as possible predictors for spontane-ous miscarriage. Majority of patients were stimulated according to a standard gonadotrophin-releasing hormone agonist (GnRH-a) protocols (almost exclu-sively the long protocol); the flexible gonadotrophin-releasing hormone an-tagonist (GnRH-ant) protocol was used in the rest of the cases. The quality of the transferred blastocysts was assessed according to our own grading system, which was a modification of the blastocyst grading system introduced by Gard-ner and Schoolcraft. Data for this analysis were extracted from the electronic database of our department.
Results: The SM rate after transfer of one or two blastocysts was 19.9%
(203/1020): 24.4% (62/254) after transfer of one and 18.4% (141/766) after transfer of two blastocysts. After adjusting for all relevant variables, age of the woman (OR = 1.10; 95% CI 1.06-1.14), quality of transferred blastocysts (OR = 0.67; 95% CI 0.47-0.95), number of transferred blastocysts (OR = 0.59; 95% CI 0.41-0.85) and number of frozen blastocysts (OR = 0.91; 95% CI 0.83-0.99) were significantly associated with SM.
Conclusion: Age of the treated woman, quality and number of transferred
blas-tocysts and number of frozen blasblas-tocysts are predictors of SM after blasto-cyst transfer. Extended embryo culture, transfer of at least one optimal quality blastocyst and double blastocyst transfer are measures which could be used to reduce SM rate in ART pregnancies. We think that more precise evaluation of relationship between number of transferred blastocysts and spontaneous mis-carriage should be conducted in the future studies.
P-122 Clinical relevance of mannan-binding lectin (Mbl) and complement in patients with recurrent miscarriages
S. Horlbeck1, M. Kirschfink2, A. Ziegler1, T. Strowitzki1, W. Eggert-Kruse1
1Women´s Hospital University of Heidelberg, Department of Gynecological
Endocrinology & Reproductive Medicine, Heidelberg, Germany
2Institute of Immunology, University of Heidelberg, Heidelberg, Germany
Introduction: Mannan-binding lectin (MBL) is a C-type lectin which
partici-pates in the innate immune defense by acting as an opsonin and by activating complement as a key mediator of inflammation. Complement defects increase the susceptibility to infection and may be associated with autoimmune diseases. Complement is activated by three pathways: the classical and the alternative pathway and the recently discovered lectin pathway all of which lead to the formation of the cytolytic membrane attack complex. A higher prevalence of MBL deficiency in women suffering from recurrent miscarriages has been sug-gested.
Materials and Methods: Therefore the relevance of MBL and complement
was evaluated in a prospective study, which included 120 randomly cho-sen patients with a history of recurrent miscarriages, who precho-sented for a
P-125 Patients with repeated ARt failures should be tested for an abnormal X chromosome karyotype
T. Seki1, N. Sugawara1, M. Maeda1, T. Manome1, Y. Araki2
1Iwaki Women’s Clinic, Gynecology, Iwaki Fukushima, Japan 2The Institute for ARMT, Laboratory of IVF, Maebashi Gunma, Japan
Introduction: X chromosome mosaicism, including mosaic 45, X, is
occasion-ally encountered; however, it usuoccasion-ally is not considered to be an important infer-tility factor. In contrast to this viewpoint, we recently recognized that patients with X chromosome mosaicism tend to have repeated assisted reproductive technology (ART) failures; thus, they are categorized as severely infertile. For this reason, we evaluated whether X chromosome mosaicism is related to repro-ductive failure in infertile patients.
Materials and Methods: Subjects comprised 108 couples who underwent ART
at Iwaki Women’s Clinic between April 1996 and December 2008. Subjects were divided in three groups: Group A (n = 45), frequent implantation failure despite the transfer of good-quality embryos; Group B (n = 47), failure to obtain good-quality embryos; and Group C (n = 16), poor responders ( ≤ 3 retrieved oocytes). We examined the karyotype via peripheral lymphocyte analysis using G-banding and fluorescent in situ hybridization (FISH). The incidence of mo-saicism, including 45, X karyotype, pregnancy rate, implantation rate and mis-carriage rate, was compared among the three groups. All subjects underwent in
vitro fertilization (IVF).
Results: All semen analyses were in the normal range (WHO criteria). The
average age was 36.6 years for the women. The basal FSH level was 8.0 mIU/ ml. The average number of retrieved oocytes was 3.7 and the fertilization rate was 310/430 (72.1%).
The incidence of mosaicism, including 45, X karyotype, was 21/108 (19.4%); this included 7/45 (15.6%) in Group A, 8/47 (17.0%) in Group B, and 6/16 (37.5%) in Group C. The mosaicism percentage in Group C (poor respond-ers) was higher than that in the other two groups.
The incidence of 46, XX/45, X mosaicism was 14/108 (13.0%). The inci-dence of mosaicism, including (46, XX/47, XXX), (46, XX/45, X/47, XXX), (46, XX/45, X/48, XXXX), and (46, XX/45, X/47, XXX/48, XXXX/49, XXXXX), was 7/108 (6.5%). The pregnancy rate of subjects with mosaicism was lower (12.9%) than that of those without mosaicism (20.3%). In addition, the pregnancy rate per subject was lower (52.4%) in the subjects with mosa-icism than that in those without mosamosa-icism (62.1%). Furthermore, the implanta-tion rate of the subjects with mosaicism was lower (8.6%) than that of subjects without mosaicism (11.3%). The spontaneous abortion rate was not significant-ly different between subjects with and without mosaicism.
Conclusion: This study revealed that infertile patients who had experienced
repeated IVF failures had a higher rate of abnormal X chromosomes with mo-saicism. It is possible that an X chromosome abnormality may negatively affect oocytogenesis. However, these patients may eventually be able to achieve a successful pregnancy outcome via repeated IVF cycles. It is necessary to evalu-ate the relationship between the X chromosome (including mosaicism) and the clinical aspects of infertile patients.
P-126 transabdominal cerclage : preconceptual or first trimester insertion?
R. Farquharson1, F. Dawood1
1Liverpool Women’s Hospital, Department of OB/GYN, Liverpool, United
Kingdom
background: Transabdominal cerclage (TAC) is a recognized treatment for
cervical weakness with a history of recurrent mid-trimester loss (MTL) and a failed vaginal suture.
Methodology: A comparative study was performed of 40 women who
under-went TAC in the first trimester (T1) between 1993 to 2005 with a group of 38 women who underwent pre-conceptual (PC) TAC between 2006 and 2008. All patients underwent a standardized investigation protocol and TAC insertion based at the Recurrent Miscarriage Clinic of Liverpool Women’s Hospital. All patients had identical inclusion criteria with 1 or > MTL and 1 or > failed elec-tive vaginal suture.
Results: Preconceptual investigation showed a similar prevalence of
associ-ated pathology with antiphospholipid syndrome (APS) or bacterial vaginosis (BV) present in 45 % (18/40) and 50 % (19/38) in the T1 and PC groups re-spectively.
did not find significant differences in secretion profiles of Glycodelin A and the various members of the IGF-family between women who conceived of twins and singletons at any moment during the IVF treatment.
Conclusions: Our data indicate that maternal VEGF-A may be involved in
sus-tained (multiple) implantation and that a high BMI is associated with double implantation. Our observation highlights the need for more research on angio-genic factors which may be involved in optimizing the endometrial conditions for implantation. Future studies should also investigate whether the predictive value of VEGF-A for ongoing singleton or twin pregnancies observed here is due to chance and if not, is a useful clinical test.
P-124 High H-y antibody responses in secondary recurrent miscarriage patients correlate with low male birth in surviving pregnancies
H.S. Nielsen1, F. Wu2, Z. Aghai3, R. Steffensen4, A.G.S. van Halteren3,
E. Spierings3, O.B. Christiansen1, D. Miklos2, E. Goulmy5
1University Hospital Copenhagen Rigshospitalet, The Fertility Clinic,
Copenhagen, Denmark
2Stanford University Medical Center, Department of medicine, Stanford CA,
U.S.A.
3Leiden University Medical Center, Department of Immunohematology and
Blood Transfusion, Leiden, The Netherlands
4Aalborg Hospital, Department of Clinical Immunology, Aalborg, Denmark 5Leiden University Medical Center, Department Immunohematology and
Blood Transfusion, Leiden, The Netherlands
Introduction: Three or more consecutive miscarriages after the birth of a live or
stillborn child is defined as secondary recurrent miscarriage (SRM). Among SRM patients, the birth of a boy prior to the miscarriages is significantly more com-mon than a girl and is associated with a poorer chance of a subsequent live birth. We consequently questioned the role of immune responses against male specific, Y-chromosome encoded histocompatibility, (H-Y) antigens initiated in a prior male-fetus pregnancy in these patients. The strong potential of H-Y antibodies has been shown in studies aiming at female sex selection of pre-implantation cattle embryos to increase profitability of dairy and beef cattle production. Between 80%-87% of male bovine embryos that are cultured in high-titer rat H-Y antisera, at the morula stage, stop their development in contrast to female embryos. In this study we explored if H-Y antibodies are distinguishable in women with SRM from women with primary recurrent miscarriage (PRM), and control women with prior uncomplicated male births. We also, correlated the presence of H-Y antibodies in these patients in early pregnancy to the sex of surviving pregnancies.
Material and Methods: Patients were recruited from the Danish Recurrent
Miscarriage Clinic from July 2004 to March 2008. Controls were recruited from July 2004 to July 2006 by advertisements. Serum samples from patients with unexplained SRM (n = 84), unexplained PRM (n = 12), and healthy women (n = 37) were obtained. Serum samples were obtained in the first preg-nancy after referral at gestational week 4-5 for 77 (80%) patients while samples from the remaining patients and the controls were obtained when they were not pregnant but no later than two years after their last pregnancy. Samples were analysed in Dutch and US laboratories specialised in H-Y immunity. All samples were tested by Enzyme-Linked Immunosorbent Assay (ELISA) for the presence of IgG antibodies that specifically recognised any of 5 H-Y recom-binant proteins (EIF1AY, RPS4Y1, ZFY, DDX3Y and UTY) and their 5 H-X homologs or the HIVp24 proteinas (negative control). Random selection of 5 H-Y negative, 5 H-Y specific and 2 H-Y nonspecific samples were subsequently confirmed by Western blot technique.
Results: H-Y antibodies were more frequent in SRM patients 46% compared
to female controls 19% (p = 0.004) and PRM patients 8% (p = 0.01). The pres-ence of H-Y antibodies in early pregnancy correlated with low male birth in surviving pregnancies, as only 12% of children born by H-Y antibody positive patients were boys compared to 42% boys born by H-Y antibody negative pa-tients (p = 0.05) and 51% among Danish newborns (p = 0.002).
Conclusion: The high frequency of H-Y antibodies in SRM patients and the
association between the presence of these antibodies in early pregnancy and the low frequency of surviving male pregnancies, indicate that H-Y antibodies may specifically kill male embryos in SRM patients. These clinical findings suggest that maternal immune responses against paternally inherited genes can cause immune-mediated pregnancy losses. Further exploring the mechanisms may answer some hitherto unanswered questions regarding maternal-fetal interac-tions in normal and pathological pregnancies.
of spontaneous tubal pregnancy was significantly higher for high sperm DNA fragmentation. Sperm DNA integrity may compromise migration of embryo and progression of pregnancy, resulting in tubal pregnancy following spontane-ous conceptions like the other early gestational wastages. Accordingly it is im-portant to identify strategies that may reduce sperm DNA damage which would help reducing the risk of tubal pregnancy. Antioxidant or antibiotic therapy might be a good option prior to planning pregnancy. The results of this study provide the elements necessary to plan such studies which will drive solid con-clusions on tubal pregnancy etiology.
P-128 Acupuncture on the day of embryo transfer does not improve ongoing pregnancy and live birth rates after IVf/ICSI
D. Andersen1, K. Lossl1, A.N. Andersen1, J. Fürbringer1, H. Bach1,
J. Simonsen1, E.C. Larsen1
1The Fertility Clinic - Section 4071, Copenhagen O, Denmark
Introduction: In the last decade several randomized controlled studies have
been published showing increased pregnancy rates when acupuncture is given in relation to embryo transfer. Two meta-analyses have confirmed these results. However, recently a third meta-analysis including eight studies could not dem-onstrate an increased pregnancy rate. The purpose of the present study was to assess further – through a large prospective, randomized, controlled, and double-blinded trial whether acupuncture on the day of embryo transfer could increase both the ongoing pregnancy and live birth rate.
Matrial and Methods: A total of 635 patients undergoing IVF or ICSI were
included in this multi-center study. In 314 patients embryo transfer was ac-companied by acupuncture according to the principles of traditional Chinese medicine. In the control group 321 patients received placebo acupuncture using a validated placebo acupuncture needle using the same acupuncture points as in the acupuncture group but without penetrating the skin. Indeed the procedure was performed blinded to both the patients and to the clinician who performed the embryo transfer.
Results: In the acupuncture group and the placebo group the ongoing
preg-nancy rates were 27% (95%CI: 22-32) and 32% (95%CI: 27-37), respectively. Live birth rates were 25% (95%CI: 20-30) in the acupuncture group and 30% (95%CI: 25–30) in the placebo group. The differences were not statistically significant. The absolute difference in live birth rate was 4.8%; 95% CI –2.5 to 12.0, P = 0.208 (5.04%, 95% CI: -1.93 to 12.02, P = 0.16, when adjusting for centre).
Conclusion: In this large randomized double-blinded placebo controlled trial,
we could not demonstrate any beneficial effect of acupuncture on the day of embryo transfer on either pregnancy or live birth rates.
P-129 the effect of body mass index on the treatment of pregnancy of unknown location with methotrexate
J. McCartney1, P. Dutton2, B.M. Brady2
1University of Edinburgh School of Medicine, Obstetrics and Gynaecology,
Edinburgh, United Kingdom
2Simpsons Centre for Reproductive Health Royal Infirmary of Edinburgh,
Obstetrics and Gynaecology, Edinburgh, United Kingdom
Introduction: Ectopic pregnancy remains the most common cause of maternal
mortality in the first trimester. Extra- uterine implantation can occur anywhere along the reproductive tract with nearly 98% implanting in the fallopian tube (1). The mainstay of treatment is for a pregnancy of unknown location (PUL) is expectant, medical and surgical management. A single dose of intramuscular methotrexate calculated from the patient body surface area (50 mg/m) is cur-rently the most widely used medical treatment. Previous studies have shown about 14 % of women will require a second dose of methotrexate with less than 10% of women who are treated medically, requiring surgical intervention (2). Obesity is rising throughout the United Kingdom and we aimed to assess the effect of BMI on the treatment of PUL with methotrexate. Furthermore, we aimed to assess the incidence of requiring a second dose of methotrexate in comparison to current guidelines (2). We hypothesised that patients with raised BMI would have a higher incidence of treatment failure with methotrexate.
Material and Methods: Data was collected from 182 female subjects
present-ing to Pregnancy Support Units (PSU) throughout Scotland in 2006, with a PUL. The majority of subjects presented to the PSU at the Royal Infirmary Obstetric outcome (BJOG, 2005, 112, 1424-6) demonstrated a 90% success
rate for T1 insertion compared to 95% in PC group. A higher preterm delivery rate is consistently associated with associated pathology (APS and/or BV) es-pecially in the T1 group. (Table 1)
Gestation at delivery first trimester tAC (t1) Preconceptual tAC (PC)
n = 40 n = 20 >34 /40 62% (25) 89% (17) 30-34/40 20% (8 ) 11% (2) 24-30/40 7% (3 ) 0 <24/40 10% (4 ) 5% (1) Overall >24/40 90% (40) 95% (20)
Conclusions: Preconceptual insertion of transabdominal cerclage provides
equivalent efficacy to first trimester insertion and avoids significant surgical complications of T1 insertion such as haemorrhage, bowel and bladder dam-age. Treatment intervention for associated pathology is mandatory for this high risk group.
P-127 Is ectopic tubal pregnancy occurrence related with DNA fragmented spermatozoa?
O. Leylek1, O. Uner2, M.D. Ozcil1, V. Baltaci3
1Suzan Health Care Center, Women’s Clinic, Gaziantep, Turkey 2Genart, IVF Center, Anakara, Turkey
3Ufuk University School of Medicine, Medical Genetics, Ankara, Turkey
Introduction: It is evident that embryo movement in tuba is far more
compli-cated process than initially thought. Impaired tubal function including surgical-ly visualized tubal pathology originating from pelvic infection, endometriosis or previous surgery is associated with tubal ectopic pregnancy. Tubal ectopic risk in women undergoing assisted reproduction technology (ART) increases when the embryo quality is low. Sperm DNA fragmentation (increased DNA strand breaks) was previously shown to be an important factor affecting em-bryo quality. Previous studies also reported the effect of sperm DNA integrity on spontaneous abortion rates and implantation. If the embryo has fragmented DNA originated from the sperm genome, its tubal transport can be affected negatively. Thus the aim of the study is to investigate the relationship between sperm DNA fragmentation and the tubal pregnancy occurrence.
Materials and Methods: A total of 42 spontaneous tubal ectopic pregnancies
(group I), 14 tubal ectopic following ART (group II) and 28 controls (spontane-ous par(spontane-ous woman, group III) were included in the study. Group I patients had not experienced any laparatomic or laparoscopic operations. However when the questionnaire was introduced to groups, all the ART women (group II) had at least one laparoscopic or laparotomic operation. Women who were directed to ART cycles had patent tubes and sufficient peritoneal radioopaque spillage on their hysterosalpingography. They didn’t have a preexisting hydrosalpinx on their HSG. They were all stimulated with standard long protocol and in-tracytoplasmic sperm injection (ICSI) was applied to each oocyte. Three em-bryos were transferred under direct ultrasound guidance on the third day after ICSI. The control group had at least two healthy children and didn’t have any spontaneous abortion. Tubal pregnancy was diagnosed by imaging the adnexal ectopic mass with lacking endometrial gestational sac on transvaginal pelvic sonography and the consecutive beta hCG levels. After diagnosis, sperm DNA fragmentation analysis was made by using SCSA (sperm chromatin structure assay). Student t-test or chi-square test was used where appropriate and a spear-man correlation analysis was done for statistical analysis.
Results: The distribution of the mean fragmentation rate was %19.14 ± 4.19 in group I, %16.7 ± 2.62 in group II and %15.67 ± 2.81 in the control group. Group I patients had a higher DNA fragmentation rate with respect to group II and controls (p < 0.01, p < 0.05 respectively). No significant relationship was found between the mean fragmentation rates of group II and controls. In case of a threshold of % 19, DNA fragmentation rate revealed an odd ratio of 2.20 (95% (CI) 1.49-3.25, p < 0.01) for spontaneous ectopic pregnancy. There was a positive correlation existed between sperm DNA fragmentation and spontane-ous tubal pregnancy (r = 0.39, p = 0.001). But the fragmentation rate was not related to tubal ectopics after ICSI.
Conclusions: Current knowledge on reproduction cannot explain the cellular
and molecular mechanisms leading to tubal ectopic pregnancy. Our findings suggest that there is a positive relationship between spontaneous tubal preg-nancy and sperm DNA fragmentation. Considering the threshold value, the risk
certain controlled ovarian stimulation protocols may contribute to an ectopic implantation. According to our results, a long protocol with GnRH-analogs should be the first choice for women with risk factors for EP. In our series, the recurrence of EP is not increased after ART, and oocyte donation decreases the recurrence rate 3-fold.
P-131 the Vegf and the its receptors Vegf-R1 and Vegf-R2 are over expressed in term placenta of women with Recurrent miscarriage treated with g-CSf
F. Scarpellini1, M. Sbracia1, G. Rossi2
1Hungaria, Private Center, Rome, Italy
2Istologia e Embriologia, Università Tor Vergata, Rome, Italy
Introduction: We have showed previously that women with recurrent
miscar-riage can be treated successfully with G-CSF, a cytokine with Th2 effects on immunosystem and with anti apoptotic role on trophoblast in in vitro studies. In order to investigate the G-CSF positive role on trophoblast growth and devel-opment we havestudied in our patients affected by Recurrent miscarriage that delivered at term after the treatment with G-CSF the expression of VEGF and its receptors VEGF-R1 and VEGF-R2.
Material and Methods: We collected placental tissue from 15 placentas at
term after deliver from women treated with G-CSF for Recurrent Miscarriage. As control we used 15 placental tissues of normal pregnancies at term in normal women no affected by any diseases. All the samples were fixed in formalin and paraffin embedded; sections of 5mm were dewaxed, rehydratated and incubated overnight with anti-VEGF, anti VEGF-R1 and anti VEGF-R2 antibodies (Santa Cruz Biotechnology, Santa Cruz, California). The indirect avidin-biotin com-plex (Vectastin ABC-peroxidase kit) immunoperoxidase assay was used, and staining was obtained with DAB. In order to perform statistical analysis of the immunostaining for each protein, semi-quantitative analysis of specific staining was performed using HSCORE system Statistical analysis was performed using Fisher exact test and Student’t test.
Results: Both cytotrophoblast and syncytiumtrophoblast of villi in normal
pregnancy at term showed immunostaining for VEGF. Also endothelial cells of villi vasa showed a positive staining for this growth factors. The cyto and the syncytium trophoblast of placental villi of the women treated with G-CSF showed a strong increase of specific immunostaing for VEGF in all 15 cases. (P < 0.001).
The cytotrophoblast and the syncytiumtrophoblast of villi in normal preg-nancy at term showed immunostaining for VEGF-R1. Also endothelial cells of villi vasa showed a positive staining for this receptor. The cyto and the syncytium trophoblast of placental villi of the women treated with G-CSF showed a strong increase of specific immunostaing for VEGF-R1 in all 15 cases.(P < 0.001). Both cytotrophoblast and syncytiumtrophoblast of villi of normal pregnancy at term showed immunostaining for VEGF-R2. Also endothelial cells of villi vasa showed a positive staining for this receptor. The cyto and the syncytium trophoblast of placental villi of the women treated with G-CSF showed a strong increase of specific immunostaing for VEGF-R2 in all 15 cases.(P < 0.001).
Conclusions: The treatment with G-CSF during the pregnancy in case of
women with Recurrent Miscarriage increases the expression of VEGF and its receptors in the trophoblast, both in syncytiumtrophoblast that in the cytotro-phoblast, throughout the pregnancy, enhancing the growth and development of placental tissue, with an increase in the blood flow of placenta and consequently of the foetus.
P-132 glycodelin-a suppressed trophoblast invasion by down-regulating urokinase plasminogen activator and extracellular signal regulated kinases activities
K.K.W. Lam1, P.C.N. Chiu1, C.L. Lee1, W.S.B. Yeung1, P.C. Ho1
1The University of Hong Kong, Obstetrics and Gynaecology, Hong Kong,
Hong Kong
Introduction: Invasion of the extravillous cytotrophoblast into the maternal
endometrium is crucial to placenta formation and is strictly regulated by para-crine and autopara-crine interactions. Dysregulation of the process results in a wide spectrum of abnormal pregnancies, like preeclampsia, choriocarcinoma and intrauterine growth restriction. One of the invasive phenotypes of extravillous cytotrophoblast is production of matrix degrading enzyme, including urokinase of Edinburgh. Data was then extracted into an Excel spreadsheet for further
analysis using descriptive statistics (SPSS, version 2007). T-tests were used to determine whether the observed data varied significantly from expectation using the chi squared test with acceptable significance set at P < 0.05.
Results: The incidence of patients requiring a second dose of methotrexate was
13.2%, which is comparable to recent figures of 14% (2). Failure of treatment occurred in 26 patients out of 182 subjects (14.3%). Treatment with methotrex-ate failed in only 7.5% of patients with a BMI of gremethotrex-ater than 30, with the great-est rate of failure seen in patients with a BMI below 20 (35%) (P < 0.05).
Conclusion: The incidence of failure of medical management of PUL with
methotrexate is not increased in women with raised BMI, refuting our hypoth-esis that the greater the BMI, the higher the failure rate. However, our results suggest a trend towards greater risk of treatment failure with methotrexate in women with a lower BMI. Our results support the use of methotrexate as an al-ternative to surgical management in patients with suspected ectopic pregnancy and the associated co-morbidity of a raised BMI.
P-130 Is ectopic pregnancy really increased in ARt?
B. Oriol1, J. Giles2, F. Bronet3, A. Ruiz2, A. Pellicer2, J.A. Garcia-Velasco4
1Klinikum rechts der Isar der TUM, Frauenklinik, Munich, Germany 2Instituto Valenciano de Infertilidad (IVI), Reproductive Medicine, Valencia,
Spain
3Instituto Valenciano de Infertilidad (IVI), Laboratory for Preimplantation
Genetic Diagnosis, Madrid, Spain
4Instituto Valenciano de Infertilidad (IVI), Reproductive Medicine, Madrid,
Spain
Introduction: Assisted reproductive techniques (ART) have traditionally been
considered as a risk factor for ectopic pregnancy (EP). We have studied the in-cidence of EP in 76 077 ART cycles, and analysed which techniques and factors contribute to an ectopic implantation. Furthermore, the recurrence of EP has been estimated after a subsequent spontaneous pregnancy in 15%; we analysed if ART increases the risk of recurrence in this subgroup of patients.
Material and Methods: Between the year 2000 and 2008, 633 EPs were
diag-nosed after ART at our center. A retrospective analysis was performed, calculat-ing the incidence of EP in relation to the total number of gestations after ART. Subgroup analysis was performed according to each ART procedure. 327 EPs occurred after in vitro fertilisation (IVF) and controlled ovarian stimulation; for this subgroup we analysed possible risk factors related to the stimulation protocol. For patients with a history of EP, we calculated the recurrence rate after ART.
Results: The general incidence of EP was 2.2% per pregnancy after ART. For
different techniques the incidence was: 2.6% with general IVF (3.2% after IVF only, and 2.5% after ICSI), and 2.1% after thawed-embryo transfer which is not significantly different from the incidence after fresh embryo transfer. Pre-implantation genetic diagnosis (PGD) did not significantly alter the incidence (2.3%) as compared to ICSI without PGD (2.6%). In oocyte donation cycles, EP-incidence per pregnancy (1.7%) was significantly lower (p = 0.0002) than after IVF with own oocytes (2.6%). This observation may be explained by un-known oocyte-related factors or differences in the stimulation protocols. Age at stimulation onset did not predict the risk of EP. With different stimulation protocols, EP-incidence was higher if clomifen citrate (p = 0.006) or an an-tagonist protocol (p = 0.008) were used, as compared to the long protocol with GnRH-analogs. The number of days with stimulation, total gonadotropin dosis, estradiol peak and number of oocytes retrieved did not significantly differ in women with EP as compared to intrauterine gestations after IVF. The differ-ence in EP-inciddiffer-ence after intrauterine insemination (IUI) with partner-sperm (2.5%) or after sperm-donation (1.8%) did not reach statistical significance. Of all 633 EPs, 28 were heterotopic (4.4%) which makes an incidence of 0.1% per pregnancy. From all patients who recurred to ART, 3.1% had had at least one previous EP. In this subgroup, the recurrence rate per pregnancy after ART was 8.2%. According to the treatment, the recurrence rate per gestation was: 11.5% after IVF; 4.3% after oocyte donation; 4.9% after thawed-embryo transfer; 17.5% after IUI; and 14.3% after timed intercourse.
Conclusions: The incidence of EP after ART was not increased in our series
when compared to the 2% demonstrated for the general population. The inci-dence was not altered by performing PGD which speaks against aneuploidies as an independent risk factor for EP. Nevertheless, oocyte donation significantly lowererd the incidence of EP compared to IVF. Therefore, it can be argued that
interspersed with any other pregnancy. Univariable and multivariable logistic regression analyses were performed.
Results: 2055 women were analyzed for recurrent miscarriage during the study
pe-riod. 373 women were excluded from further analyses since they were diagnosed with other underlying causes for recurrent miscarriage. Of 1682 women included, 301(18%) had APS and 1381 (82%) had unexplained recurrent miscarriage.
The mean age at the time of presentation was 32.6 years, which did not differ between women with and without APS (Odds Ratio (OR) 1.00, 95% Con-fidence Interval (CI) 0.98-1.02). The median number of miscarriages was 3 in both groups (OR 1.02, CI 0.94-1.10). 518 (31%) women had a history of 2 miscarriages, 1164 (69%) women had experienced at least 3 miscarriages (OR 0.97, 0.74-1.27). No difference was observed between women with APS and unexplained recurrent miscarriage in number of first trimester miscarriages or second trimester miscarriages (OR 1.01, CI 0.94-1.10 and OR 1.02, CI 0.84-1.23 respectively). The mean number of children was 0.67 in women with APS and 0.72 in women with unexplained recurrent miscarriage. (OR1.06, CI 0.92-1.23). 845 (50%) women had a history of at least one live birth after 24 weeks gestational age, prior to their first visit, with no difference between the two groups (OR 0.87, CI 0.68-1.11).
The sequence of pregnancies was known in 1499 women. 250 of 258 (97%) APS positive women and 1213 of 1241 (98%) women with unexplained recur-rent miscarriage had a history of consecutive miscarriages (OR 1.39, CI 0.62-3.08). Of the women with a history of at least 3 miscarriages, those miscarriages were consecutive in 136 of 155 (88%) APS positive women and in 672 of 750 (83%) women with unexplained recurrent miscarriage (OR 1.20, CI 0.71-2.05). Multivariable analysis showed no significant differences between women with and without APS for any of the variables.
Conclusions: Neither parameters of obstetric history -the number of
miscar-riages and the sequence of pregnancies- nor maternal age were associated with a diagnosis of APS in women with recurrent miscarriage. Thus, these cannot be used to increase the yield of testing women with recurrent miscarriage for APS. It could therefore be advised so far to offer testing for APS to all women with a history of 2 or more miscarriages.
References:
1 Wilson WA, et al. International consensus statement on preliminary classification cri-teria for definite antiphospholipid syndrome: Arthritis Rheum 1999;42:1309–11.
P-134 Recombinant lH as luteal supplementation method after agonist triggering in IVf. A proof of concept study
E. Papanikolaou1, W. Werpoest2, H. Fatemi2, N. Polyzos3, P. Humaidan4, B.
Tarlatzis5, P. Devroey2, F. Tournaye2
1BIOGENESIS Thessaloniki Greece, Assisted Reproduction Unit, Thessaloniki,
Greece
2Vrije Universiteit Brussels, Centre for Reproductive Medicine, Brussels,
Belgium
3Univesity of Thessalia, Centre for Reproductive Medicine, Larisa, Greece 4Skive Regional Hospital, Centre for Reproductive Medicine, Viborg, Denmark 5Aristotle University of Thessaloniki, Assisted Reproduction Unit,
Thessaloniki, Greece
Introduction: In GnRH-antagonist protocols for IVF, GnRH-agonist
trigger-ing of ovulation carries a lower risk of OHSS. Nevertheless, the significantly reduced reproductive outcome previously reported has cast a swadow over the agonist triggering protocol implicating a luteal phase defect. We hypothesized that the low endogenous LH in the early luteal phase, as a result of steroid downregulation, cannot compensate for appropriate implantation after agonist triggering, whereas in HCG triggering the prerequisite role of LH in implanta-tion is played by the circulating HCG. Our proposal was to supplement the luteal phase with recombinant LH (rec-LH) in conjunction with progesterone in order to overcome the defective luteal endocrine/endometrial environment and to restore implantation capacity after agonist triggering. This study aimed at investigating the role of luteal supplementation by rec-LH.
Material and Methods: 39 patients seeking IVF treatment were recruited to
participate in this randomised controlled trial, however, four patients refrained from further treatment (two due to personal problems, one became pregnant and one due to poor response). Ovarian stimulation was performed with a fixed dose of 187.5 IU recombinant FSH in a fixed day-6 antagonist protocol.The study was approved by the ethical committee of the University Hospital at the Vrije Universiteit Brussel as a proof of concept (EC ref 2005/74). Inclusion criteria plasminogen activator (uPA). Extracellular signal regulated kinase (ERK), a
member of the mitogen activated protein kinase (MAPK) family is known to control trophoblast invasion and migration. Glycodelin-A (GdA), a glycopro-tein abundantly synthesized by the decidua but not the trophoblast, is upregu-lated during first trimester coinciding with the time of trophoblast invasion. We hypothesize that decidual-derived glycodelin-A is a paracrine regulator fine-tuning the invasion of human trophoblast.
Material and Methods: Trans-well invasion assay was used to examine the
ef-fect of GdA and MEK (MAPK kinase) inhibitors (PD98059 & U0126) on the in-vasiveness of an immortalized first trimester extravillous cytotrophoblast (TEV-1) and a choricarcinoma (JEG-3) cell line. The RNA expression, proteinase activity and secretion of uPA were determined by real time quantitative PCR, quantitative urokinase activity assay and ELISA, respectively. Western blotting analysis was employed to examine the effect of GdA on ERK activities.
Results: Transwell invasion assay showed that GdA, but not deglycosylated
GdA, significantly reduced the invasiveness of the cells. At 30 pmol/ml, it in-hibited 47.7 ± 8.7% of TEV-1 and 27.1 ± 4.7% of JEG-3 invasion. The suppres-sive effect of GdA on invasion was associated with a suppression of uPA pro-teinase activity. The transcription and secretion of uPA were also significantly reduced by 72.0 ± 4.3% and 36.13 ± 8.8% respectively after GdA (30 pmol/ml) treatment. Western blotting analysis showed that GdA suppressed the phospho-rylated ERK level in a dose dependent manner. The involvement of ERK in the trophoblast invasion was supported by the observation that MEK inhibitors PD98059 and U0126 that reduced phosphorylation of ERK also significantly suppressed the invasiveness of trophoblast. A dose-dependent down-regulation of uPA mRNA was also noted by the real time quantitative PCR after treatment with these inhibitors at concentrations that did not affect cell viability.
Conclusions: This is the first report demonstrating a suppression of
tropho-blast invasion and uPA transcription through down regulating ERK activities in trophoblast cell lines. Thus, GdA is a potential paracrine regulator at the fetal-maternal interface limiting the extent of trophoblast invasion. The results may have implication in pregnancy complications such as preeclampsia, which is characterized by shallow invasion. Consistently, trophoblast cells from preec-lamptic women have reduced uPA proteolytic activities.
P-133 Risk factors for Antiphospholipid Syndrome (APS) in women with recurrent miscarriage
E. van den Boogaard1, D.M. Cohn2, J.C. Korevaar3, F. Dawood4,
S. Middeldorp5, M. Goddijn1, R.G. Farquharson4
1Academic Medical Center, Department of Obstetrics and Gynaecology Centre
for Reproductive Medicine, Amsterdam, The Netherlands
2Academic Medical Center, Department of Vascular Medicine, Amsterdam,
The Netherlands
3Academic Medical Center, Department of Clinical Epidemiology, Amsterdam,
The Netherlands
4Liverpool Women’s Hospital, Department of Obstetrics and Gynaecology,
Liverpool, United Kingdom
5Leiden University Medical Center, Department of Clinical Epidemiology and
Department of General Internal Medicine, Leiden, The Netherlands
Introduction: Antiphospholipid Syndrome (APS) is an acquired trombophilia
and an established cause of recurrent miscarriage. It is advised to offer testing for lupus anticoagulant (LAC) and anticardiolipin antibodies (aCL) in women with recurrent miscarriage. International definitions of recurrent miscarriage vary in their number of miscarriage and in the outcome and sequence of pre-ceding pregnancies. At present, it is unknown whether the diagnostic yield of testing for APS can be increased by clinical determinants derived from the ob-stetric history. The aim of this study was to investigate the relationship between parameters of obstetric history and a diagnosis of APS, in order to define a risk profile to determine in whom testing for APS is most efficient.
Methods: A cohort study was performed including all women with 2 or more
miscarriages who attended the recurrent miscarriage clinic of Liverpool Wom-en’s Hospital, Liverpool, UK between 1988 and 2006. All women underwent a systematic diagnostic work up including testing for APS. Women with estab-lished causes for recurrent miscarriage other than APS were excluded. Char-acteristics of women with proven APS according to the SAPPORO criteria [1] were compared to women with unexplained recurrent miscarriage. Data on preceding pregnancies were collected; number of previous pregnancies, their outcome and sequence. Miscarriages were considered consecutive when not
