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582: Early prediction of fetal gender by using free fetal DNA from maternal plasma
SMFM Abstracts
www.AJOG.org 581
ESTRADIOL INHIBITS HIF-1 EXPRESSION IN FIRST TRIMESTER VILLOUS EXPLANT CULTURES GEUM JOON CHO1, MIN JEONG OH1, YU CHIN PAEK1, OYE SUN SEOK1, HYE MIN YEO1, HYUN-JOO SEOL1, SOON CHEOL HONG1, HAI-JOONG KIM1, 1Department of Obstetrics and Gynecology, College of Medicine, Korea University, Seoul, South Korea OBJECTIVE: Regulation of trophoblast differentiation toward invasive extravillous trophoblasts (EVTs) is critical for establishing successful pregnancy. During the most time of the first trimester, the placenta develops in condition of physiological hypoxia. Expression of Hypoxia inducible factor-1 (HIF-1) in placenta is high in early gestation between 5 to 8 weeks and then falls precipitously around 10 to 12 weeks of gestation. HIF-1 is known to inhibit trophoblast differentiation toward an invasive EVTs. Estradiol begins to rise in 6-8 weeks of gestation when placental function becomes apparent. Recently, it has been reported that estradiol inhibits hypoxia induction of HIF-1 in Hep3B Cells. In this study, we investigated the effects of estradiol on expression of HIF-1 and trophoblast differentiation in human first trimester villous explant cultures. STUDY DESIGN: Villous explant cultures were established from first trimester human placentas (6-8 weeks of gestation, n⫽3) obtained from elective terminations of pregnancies. Normal villous tissues were explanted on matrigel and incubated under 3% O2 tension for 5 days. In the experiments evaluating the effect of estradiol, 1ng/mL of estradiol was added to the culture medium. Morphological integrity and viability of villous explants were monitored. Expression of HIF-1 in villous explant cultures was evaluated by Western blotting. RESULTS: EVTs emerging from first trimester villous explant cultures formed outgrowth of cells from the distal end and invaded into surrounding matrigel. As compared with control villous explants, exposure of villous explants to estradiol showed a decreased outgrowth of cells from the distal end. However, estradiol treatment increased invasion into the surrounding matrigel. On western blots, the expression of HIF-1 decreased after treatment with estradiol under 3% O2 oxygen tension. CONCLUSION: These findings suggest a possible role for estradiol to mediate trophoblast differentiation toward an invasive EVTs by interfering with increases in HIF-1 levels.
583
0002-9378/$ - see front matter doi:10.1016/j.ajog.2008.09.613
0002-9378/$ - see front matter doi:10.1016/j.ajog.2008.09.611 584 582
EARLY PREDICTION OF FETAL GENDER BY USING FREE FETAL DNA FROM MATERNAL PLASMA GIAN CARLO DI RENZO1, ELENA PICCHIASSI1, ALESSIA FANETTI1, MICHELA CENTRA1, GRAZIANO CLERICI1, GIULIANA COATA1, 1University of Perugia, Department of Obstetrics and Gynecology, Perugia, Italy, Italy OBJECTIVE: Free fetal DNAA (ffDNA) is present in maternal plasma and serum during pregnancy. As the use of ffDNA in non invasive prenatal diagnosis is possible only for fetal DNA sequences absent in the mother, the potential clinical applications are concerned with the prediction of RhD genotype, Mendelian diseases and fetal gender. It has been demonstrated the possibility of detecting Y-specific sequences in maternal blood during gestation using real-time quantitative polymerase chain reaction (PCR) and different types of primers and probes specific for chromosome Y,such as the multicopy DYS14 sequence located within the TSPY gene. STUDY DESIGN: We have analyzed 395 samples from healthy pregnant women at 10-12 weeks gestation, using an original DYS14 real-time PCR protocol, to establish the sensitivity, specificity, and predictability for the diagnosis of fetal gender. RESULTS: We compared the diagnostic performance in fetal gender prediction of two tests based on the same DYS14 real time PCR protocol but using different interpretation criteria of the results. In column A the results of our previous work (Picchiassi et al., Prenat Diagn 2008; 28: 525-530) are shown in the table; in column B the results of a work in progress. The difference between the two works is given by the interpretation criteria of the results. In the previous work we used the interpretation criteria present in literature while in the current work we use new criteria elaborated by our lab. CONCLUSION: DYS14 assay with new interpretation criteria is the best approach for early fetal gender assessment because it is sensitive, accurate and efficient.
Total number of patients Gestational age (weeks) Sensitivity (%) Specificity (%) PPV (%) NPV (%) Efficiency (%)
VASCULAR REACTIVITY PATTERNS IN THE CAROTID ARTERY OF OFFSPRING IN THE APOE MOUSE MODEL OF DEVELOPMENTAL PROGRAMMING OF ATHEROSCLEROSIS AT 4 MONTHS OF AGE NIMA GOHARKHAY1, FANGXIAN LU1, EGLE BYTAUTIENE1, PHYLLIS GAMBLE1, ESTHER TAMAYO1, HUAIZHI YIN1, GARY D HANKINS1, MONICA LONGO1, GEORGE SAADE1, 1University of Texas Medical Branch at Galveston, Department of Obstetrics and Gynecology, Galveston, Texas OBJECTIVE: We have previously described a model of developmental programming of atherosclerosis characterized by high cholesterol levels, atherosclerotic plaques, and renal and hepatic lesions in heterozygous mouse offspring born to apoE-knockout mothers. We recently demonstrated the presence of hypercholesterolemia in this model during young adulthood. In the present study we aimed to determine the effect of this atherogenic state on vascular reactivity patterns in the young adult offspring in this model. STUDY DESIGN: We crossbred apoE knockout (apoE⫺/⫺) and wild-type C57BL/6J (apoE⫹/⫹) mice to obtain heterozygous offspring born to hypercholesterolemic apoE⫺/⫺ mothers (apoE⫹/⫺mat), heterozygous offspring born to wild-type mothers (apoE⫹/⫺pat), homozygous knockout offspring (apoE⫺/⫺), and homozygous wild-type offspring (apoE⫹/⫹). The resulting pups were followed until 4 months of age when they were sacrificed. The right carotid was removed and 2 mm segments were mounted in a wire myograph for isometric tension recording. We determined concentration-response curves to phenylephrine (PE) with and without L-NAME (10⫺10⫺10⫺5 M), acetylcholine (Ach, 10⫺10⫺10-⫺5 M), sodium nitroprusside (SNP, 10⫺10⫺10⫺5 M), and isoprotenerol (ISO, 10⫺10⫺10⫺5 M). RESULTS: At 4 months of age, no significant differences were found in contractile and relaxatory responses to the endothelium-dependent and endothelium-independent agents tested among the study groups. CONCLUSION: The atherogenic state programmed by the hypercholesterolemic fetal environment does not result in altered function of the carotid artery in the young adult offspring in the apoE mouse model. Whether an effect can be demonstrated in smaller vessels, or at a later age, is currently under study. Supported by NICHD grant R03HD056087
A
B
145 10-12 98.78 96.82 97.59 98.39 97.93
250 10-12 100.00 100.00 100.00 100.00 100.00
THE TRUE RISK OF ADVERSE PREGNANCY OUTCOME FOLLOWING GENETIC AMNIOCENTESIS ANDRZEJ LYSIKIEWICZ1, ADJUA YEBOA1, LOIS BRUSTMAN1, RICHARD JAFFE1, ODED LANGER1, 1St. Luke’s Roosevelt Hospital Center, New York, New York OBJECTIVE: To estimate the error of ascertainment in the assessment of amniocentesis outcome. STUDY DESIGN: 3112 consecutive amniocenteses were performed by MFM specialists in a single large urban institute over a 5-year period. All patients were followed up to 24 weeks gestation. Patients were stratified into Group I with complete pregnancy outcome data in hospital, genetic, laboratory and ultrasound records. Group II was comprised of patients with no follow up data. A targeted search for patient pregnancy outcome information was sought from private physician records, industry records and direct interviews with patients. For patients who relocated to another geographic area, ”government record search,” ”people search,” ”people finder” and other public search facilities were used to locate them. Pregnancy loss rates following amniocentesis were compared between the two groups. RESULTS: 3118 patients underwent genetic amniocentesis between January 2003 and Dec 2007. In 2944 (98%) data regarding pregnancy outcome were obtained. In only 72 (2%) no patient information was obtained. 2584/2944 (83%) of subjects with complete records readily available, 13 (0.4%) had spontaneous pregnancy losses following amniocentesis. In 462/2944 (15% ) of patients with initially no follow up records of pregnancy outcome, the targeted data search identified 19 subjects (4%) with spontaneous pregnancy losses following amniocentesis. In the group requiring targeted data search, the voluntary termination of pregnancy rate was 53/462 (11%) in comparison to the group with complete records 28/2584 (1%). Finally, every single case missed to follow up changes the pregnancy loss rate by 0.03%. CONCLUSION: Our data suggests that the large number of patients lost to follow up after amniocentesis may cloud the rate of pregnancy loss. 0002-9378/$ - see front matter doi:10.1016/j.ajog.2008.09.614
0002-9378/$ - see front matter doi:10.1016/j.ajog.2008.09.612
Supplement to DECEMBER 2008 American Journal of Obstetrics & Gynecology
S169
www.AJOG.org 581
ESTRADIOL INHIBITS HIF-1 EXPRESSION IN FIRST TRIMESTER VILLOUS EXPLANT CULTURES GEUM JOON CHO1, MIN JEONG OH1, YU CHIN PAEK1, OYE SUN SEOK1, HYE MIN YEO1, HYUN-JOO SEOL1, SOON CHEOL HONG1, HAI-JOONG KIM1, 1Department of Obstetrics and Gynecology, College of Medicine, Korea University, Seoul, South Korea OBJECTIVE: Regulation of trophoblast differentiation toward invasive extravillous trophoblasts (EVTs) is critical for establishing successful pregnancy. During the most time of the first trimester, the placenta develops in condition of physiological hypoxia. Expression of Hypoxia inducible factor-1 (HIF-1) in placenta is high in early gestation between 5 to 8 weeks and then falls precipitously around 10 to 12 weeks of gestation. HIF-1 is known to inhibit trophoblast differentiation toward an invasive EVTs. Estradiol begins to rise in 6-8 weeks of gestation when placental function becomes apparent. Recently, it has been reported that estradiol inhibits hypoxia induction of HIF-1 in Hep3B Cells. In this study, we investigated the effects of estradiol on expression of HIF-1 and trophoblast differentiation in human first trimester villous explant cultures. STUDY DESIGN: Villous explant cultures were established from first trimester human placentas (6-8 weeks of gestation, n⫽3) obtained from elective terminations of pregnancies. Normal villous tissues were explanted on matrigel and incubated under 3% O2 tension for 5 days. In the experiments evaluating the effect of estradiol, 1ng/mL of estradiol was added to the culture medium. Morphological integrity and viability of villous explants were monitored. Expression of HIF-1 in villous explant cultures was evaluated by Western blotting. RESULTS: EVTs emerging from first trimester villous explant cultures formed outgrowth of cells from the distal end and invaded into surrounding matrigel. As compared with control villous explants, exposure of villous explants to estradiol showed a decreased outgrowth of cells from the distal end. However, estradiol treatment increased invasion into the surrounding matrigel. On western blots, the expression of HIF-1 decreased after treatment with estradiol under 3% O2 oxygen tension. CONCLUSION: These findings suggest a possible role for estradiol to mediate trophoblast differentiation toward an invasive EVTs by interfering with increases in HIF-1 levels.
583
0002-9378/$ - see front matter doi:10.1016/j.ajog.2008.09.613
0002-9378/$ - see front matter doi:10.1016/j.ajog.2008.09.611 584 582
EARLY PREDICTION OF FETAL GENDER BY USING FREE FETAL DNA FROM MATERNAL PLASMA GIAN CARLO DI RENZO1, ELENA PICCHIASSI1, ALESSIA FANETTI1, MICHELA CENTRA1, GRAZIANO CLERICI1, GIULIANA COATA1, 1University of Perugia, Department of Obstetrics and Gynecology, Perugia, Italy, Italy OBJECTIVE: Free fetal DNAA (ffDNA) is present in maternal plasma and serum during pregnancy. As the use of ffDNA in non invasive prenatal diagnosis is possible only for fetal DNA sequences absent in the mother, the potential clinical applications are concerned with the prediction of RhD genotype, Mendelian diseases and fetal gender. It has been demonstrated the possibility of detecting Y-specific sequences in maternal blood during gestation using real-time quantitative polymerase chain reaction (PCR) and different types of primers and probes specific for chromosome Y,such as the multicopy DYS14 sequence located within the TSPY gene. STUDY DESIGN: We have analyzed 395 samples from healthy pregnant women at 10-12 weeks gestation, using an original DYS14 real-time PCR protocol, to establish the sensitivity, specificity, and predictability for the diagnosis of fetal gender. RESULTS: We compared the diagnostic performance in fetal gender prediction of two tests based on the same DYS14 real time PCR protocol but using different interpretation criteria of the results. In column A the results of our previous work (Picchiassi et al., Prenat Diagn 2008; 28: 525-530) are shown in the table; in column B the results of a work in progress. The difference between the two works is given by the interpretation criteria of the results. In the previous work we used the interpretation criteria present in literature while in the current work we use new criteria elaborated by our lab. CONCLUSION: DYS14 assay with new interpretation criteria is the best approach for early fetal gender assessment because it is sensitive, accurate and efficient.
Total number of patients Gestational age (weeks) Sensitivity (%) Specificity (%) PPV (%) NPV (%) Efficiency (%)
VASCULAR REACTIVITY PATTERNS IN THE CAROTID ARTERY OF OFFSPRING IN THE APOE MOUSE MODEL OF DEVELOPMENTAL PROGRAMMING OF ATHEROSCLEROSIS AT 4 MONTHS OF AGE NIMA GOHARKHAY1, FANGXIAN LU1, EGLE BYTAUTIENE1, PHYLLIS GAMBLE1, ESTHER TAMAYO1, HUAIZHI YIN1, GARY D HANKINS1, MONICA LONGO1, GEORGE SAADE1, 1University of Texas Medical Branch at Galveston, Department of Obstetrics and Gynecology, Galveston, Texas OBJECTIVE: We have previously described a model of developmental programming of atherosclerosis characterized by high cholesterol levels, atherosclerotic plaques, and renal and hepatic lesions in heterozygous mouse offspring born to apoE-knockout mothers. We recently demonstrated the presence of hypercholesterolemia in this model during young adulthood. In the present study we aimed to determine the effect of this atherogenic state on vascular reactivity patterns in the young adult offspring in this model. STUDY DESIGN: We crossbred apoE knockout (apoE⫺/⫺) and wild-type C57BL/6J (apoE⫹/⫹) mice to obtain heterozygous offspring born to hypercholesterolemic apoE⫺/⫺ mothers (apoE⫹/⫺mat), heterozygous offspring born to wild-type mothers (apoE⫹/⫺pat), homozygous knockout offspring (apoE⫺/⫺), and homozygous wild-type offspring (apoE⫹/⫹). The resulting pups were followed until 4 months of age when they were sacrificed. The right carotid was removed and 2 mm segments were mounted in a wire myograph for isometric tension recording. We determined concentration-response curves to phenylephrine (PE) with and without L-NAME (10⫺10⫺10⫺5 M), acetylcholine (Ach, 10⫺10⫺10-⫺5 M), sodium nitroprusside (SNP, 10⫺10⫺10⫺5 M), and isoprotenerol (ISO, 10⫺10⫺10⫺5 M). RESULTS: At 4 months of age, no significant differences were found in contractile and relaxatory responses to the endothelium-dependent and endothelium-independent agents tested among the study groups. CONCLUSION: The atherogenic state programmed by the hypercholesterolemic fetal environment does not result in altered function of the carotid artery in the young adult offspring in the apoE mouse model. Whether an effect can be demonstrated in smaller vessels, or at a later age, is currently under study. Supported by NICHD grant R03HD056087
A
B
145 10-12 98.78 96.82 97.59 98.39 97.93
250 10-12 100.00 100.00 100.00 100.00 100.00
THE TRUE RISK OF ADVERSE PREGNANCY OUTCOME FOLLOWING GENETIC AMNIOCENTESIS ANDRZEJ LYSIKIEWICZ1, ADJUA YEBOA1, LOIS BRUSTMAN1, RICHARD JAFFE1, ODED LANGER1, 1St. Luke’s Roosevelt Hospital Center, New York, New York OBJECTIVE: To estimate the error of ascertainment in the assessment of amniocentesis outcome. STUDY DESIGN: 3112 consecutive amniocenteses were performed by MFM specialists in a single large urban institute over a 5-year period. All patients were followed up to 24 weeks gestation. Patients were stratified into Group I with complete pregnancy outcome data in hospital, genetic, laboratory and ultrasound records. Group II was comprised of patients with no follow up data. A targeted search for patient pregnancy outcome information was sought from private physician records, industry records and direct interviews with patients. For patients who relocated to another geographic area, ”government record search,” ”people search,” ”people finder” and other public search facilities were used to locate them. Pregnancy loss rates following amniocentesis were compared between the two groups. RESULTS: 3118 patients underwent genetic amniocentesis between January 2003 and Dec 2007. In 2944 (98%) data regarding pregnancy outcome were obtained. In only 72 (2%) no patient information was obtained. 2584/2944 (83%) of subjects with complete records readily available, 13 (0.4%) had spontaneous pregnancy losses following amniocentesis. In 462/2944 (15% ) of patients with initially no follow up records of pregnancy outcome, the targeted data search identified 19 subjects (4%) with spontaneous pregnancy losses following amniocentesis. In the group requiring targeted data search, the voluntary termination of pregnancy rate was 53/462 (11%) in comparison to the group with complete records 28/2584 (1%). Finally, every single case missed to follow up changes the pregnancy loss rate by 0.03%. CONCLUSION: Our data suggests that the large number of patients lost to follow up after amniocentesis may cloud the rate of pregnancy loss. 0002-9378/$ - see front matter doi:10.1016/j.ajog.2008.09.614
0002-9378/$ - see front matter doi:10.1016/j.ajog.2008.09.612
Supplement to DECEMBER 2008 American Journal of Obstetrics & Gynecology
S169