European Journal of Obstetrics & Gynecology and Reproductive Biology, 44 (1992) 47-52
47
0 1992 Elsevier Science Publishers B.V. All rights reserved 002%2243/92/$05.00 EUROBS 01273
Prenatal diagnosis by amniocentesis in 82 pregnancies after in vitro fertilization Wolfgang Wiirfel a, Hertraut Haas-Andela b, Gottfried Kriismann a, Maria Rothenaicher ‘, Peter Hirsch a, Hemyk K. Kwapisz b, Joachim Haas b, Inge Hijgemann b and Klaus Fiedler a a Frauenklinik Dr. Kriismann, Munich and b Institut fiir Priinantale Diagnostik, Giessen, FRG Accepted for publication 26 August 1991
Summary We report on the outcome of 82 amniocenteses (AC) carried out during pregnancies after in vitro fertilization (IVF) from 1-1-1985 to 31-12-1989. The main indication for amniocentesis was a maternal age of 2 35 years. In 48 cases, we found an anterior placenta and assumed that this was related to the position in which the uterine embryo transfer was performed. In six pregnancies, we found an abnormal karyotype, including two cases of trisomy 21; the two couples decided for abortion. Four aberrations in the fetal karyotypes were also present in either the mother or the father, the resulting children are healthy. The further course of pregnancies after IVF and AC was characterized by a higher incidence of toxemia, uterine bleeding before the 28th week of gestation, abruptio placentae, and premature deliveries, when compared to the course of pregnancies after spontaneous conception. We believe that these occurrences were not caused by AC, as the incidence was higher in all our pregnancies after IVF (without AC) and has also been reported in pregnancies after ovarian hyperstimulation without IVF. Therefore, we see no reason to renounce AC after IVF. However, the special risks inherent in pregnancies after IVF must always be discussed with the couple. Amniocentesis; Down’s syndrome; Embryo transfer (ET); Intratubal embryo transfer; In vitro fertilization (IVF); Prenatal diagnosis
Introduction In vitro fertilization (IVF) and embryo transfer (ET) have become into widespread use for patients who have incurable tubal blockage [10,12,13]. Besides, IVF is an established form of treatment for other indications, such as unex-
Correspondence:
Wolfgang Wiirfel, M.D., Universitlts Frauenklinik Wiirzburg, Josef-Schneider-Str. 4, 87 Wiir burg, FGR.
plained infertility and male subfertility [20]. In all these patients without tubal occlusion, the embryo (or pronucleus) can also be transferred directly into the fallopian tubes (zygote intrafallopian transfer (ZIFT), pronuclear stage tubal transfer (PROST), intratubal embryo transfer (IT-ET) [11,35,36]). By combining all of these indications, between 1982 and 1989 we counted more than 2000 pregnancies in the Federal Republic of Germany [29], and about ten times as many worldwide [5,6,17]. Therefore, the problems of pregnancy care for IVF patients are becoming
48
more and more important. Because of the higher average age of IVF patients compared to that of women with normal pregnancies [5,6,29], prenatal diagnosis of fetal malformations and/or chromosomal disorders is of particular interest. In this article, we report on our experience with amniocentesis in 82 pregnancies after IVF. Material and Methods Indications
anestthesia and always under ultrasound control. The whole procedure was recorded (Polaroid; Printer, Mitsubishi). We attempted to aspirate a total of 20 ml of amniotic fluid during each puncture. The number of attempts was limited to two per patient. There were five twin pregnancies in which the location of the placentas and of the fetuses did not allow us to perform the amniocentesis during one puncture; thus, two punctures were carried out. In cases of anterior placentas, we tried to perform the puncture without perforating the placenta. In eight patients, however, we were forced to do the puncture through the placenta. After amniocentesis, all patients remained in bed for at least 3 to 4 h. During this time and approximately 24 h later, we checked each patient for possible uterine contractions (by tocography) and signs of infection (elevated leukocyte counts, rise in temperature). Rh-negative patients routinely received IgG (PartubolinTM i.m.> prophylactically.
The most important indication for amniocentesis was a maternal age of 35 years or more (56 patients). In four of these pregnancies, the mother’s age was > 35 years and the father’s age was > 45 years. A paternal age of 45 years or above was not considered to be a strict indication [30]; however, after discussion with such couples, three wanted an amniocentesis to be performed. Two women had had a previous pregnancy in which a chromosomal aberration was detected (trisomy 21, with a subsequent elective abortion). Both of these pregnancies with Down’s_syndrome fetuses were achieved after IVF. In 21 cases, there was no medical indication for prenatal diagnosis by amniocentesis. In all of these cases, however, we respected the couple’s request for the procedure. Ten couples did not consent to amniocentesis in spite of a maternal age of > 35 years. Four of these ten would have opposed an abortion in any case; and six patients had uterine bleeding during the first 12 weeks of gestation, which was considered a relative contraindication for performance of amniocentesis. Thus, the total number of patients who underwent amniocentesis was 80.
From 1-1-1985 to 31-12-1989 we performed 82 amniocenteses in 80 women who were pregnant after IVF/ET or intratubal embryo transfer (IVF/IT-ET). Punctures were carried out in five women pregnant with twins; no puncture was carried out in one patient pregnant with triplets. We found an anterior placenta in 48 pregnancies (Table I). Because of the position of the uterus, 46 of these patients had received an embryo transfer in the knee-elbow position. In the other two patients, the transfer was performed in
Technique All amniocenteses
TABLE I
were carried out between the 16th and 17th week of gestation. The ultrasound apparatus which we used was an array-scan (7000 S, Picker). Before the puncture, the ultrasound probe was cleaned thoroughly with warm water, covered with gel, and wrapped in sterile foil. The abdomen was disinfected with BetaisodonnaTM, which served as the transducer medium as well. The puncture was carried out with spinal needles (diameter: 9 mm), usually without local
Results
Transfer position of patient and occurrence intrauterine embryo transfer (n = 76) Position of uterus
No. of transfers
Position
Anteflexed Mid position Retroflexed
52 13 11
52 13 -
knee-elbow
in all cases of
No. with lithotriptic
11
anter’or placenta 38 8 2
49 TABLE II Abnormal fetal karyotypes found in six amniocenteses. The prenatal diagnosis was confirmed by genetic examination after delivery and by pathologic and genetic examination after abortion Patient no.
Patient age (years)
AFP
Ache activity
Fetal karyotype
Parental karyotypes (lymphocyte culture)
Further course of pregnancy
1
35
normal
negative
47,XX, +21
M *: 46,Xx; P ‘: 46,XY
2
38
normal
negative
46,XY, 15p +
3
36
normal
negative
46,XY, 16qh +
4
28
normal
negative
5
37
normal
negative
46,Xx, inv (7) (P12 q221 47,Xx, + mar
6
35
normal
negative
47,xx,
M: 46,Xx P: 46,XY,15p + M: 46,XY, 16qh + P: 46,XY M: 46,Xx P: 46,XY, inv (7) (~12 q22) M: 47,Xx, + mar P: 46,XY M: 46,Xx P: 46,XY, inv (9)
Abortion, diagnosis of Down’s syndrome confirmed Cesarean section, child healthy Normal delivery, child healthy Normal delivery, child healthy Cesarean section, child healthy Abortion, diagnosis of Down’s syndrome confirmed
+21
* M = maternal karyotype ’ P = paternal karyotype
the lithotriptic position. In all cases of tubal embryo transfer, we found a fundal placenta (6 patients). Contaminated amniotic fluid was aspirated during three punctures; in two cases, blood was removed by centrifugation. In the third case, the amniotic fluid was brownish, and centrifugation led to no change in the color. Subsequent culture showed no growth of cells, and the pregnancy ended in intrauterine death of the fetus 8 days
later. We assume that the pregnancy was already in a critical state when the amniocentesis was performed. In one patient, we registered uterine contractions within the first 24 h after puncture. Another patient developed an abdominal hematoma. No uterine bleeding, leakage, or fever was observed in any case. In six pregnancies (Table II), a numerical or _ structural aberration was found in the fetal karyotype. In two patients (one of them after tubal
TABLE III Further course of single pregnancies after in vitro fertilization spontaneous conception (Bayerische Perinatalerhebung, BPE)
(IVF) and amniocentesis
(AC), after IVF only, and after
Pregnancies with complications
IVF and AC (1985-1989) (% (No.11
IVF only (1985-1989) (%I
Toxemia Uterine bleeding before 28th week of gestation Intra-uterine death of fetus Induced abortion Abruptio placenta Premature rupture of membranes Premature labor ( < 37th week of gestation)
9.1 (7) 5.2 (4) 1.3 (1) 2.6 (2) 3.9 (3) 15.6 (12) 10.4 (8)
9.7 10.4 2.7 0.7 3.8 20.8 10.7
a Includes all children born by 30-6-1990.
BPE 1987 (%I 3.1 3.8 0.5 0.5 19.2 4.2
50
embryo transfer [34]), the fetal karyotype showed trisomy 21 (47,Xx, +21). Both of the couples decided for abortion, after which the prenatal diagnosis of Down’s syndrome was confirmed. In one of these couples, the paternal karyotype showed a pericentric inversion of chromosome 9 in the heterochromatic area. The wife became pregnant again by IVF, and on amniocentesis the fetus now showed a regular karyotype. Of the other four pathologic aberrations of the fetal karyotype, each was found also in the karyotype of either the mother or the father. All of these four children were born and were healthy. The further course of all single pregnancies after IVF and AC is given in Table III and is compared to the outcome of all our single pregnancies after IVF from 1985 to 1989 and to the outcome of single pregnancies after spontaneous conception in Bavaria. Among the five twin pregnancies, we had 2 cases of toxemia, 3 cases with premature rupture of membranes, and 3 premature deliveries. Discussion The lack of published reports on the performance and results of amniocentesis in pregnancies after IVF is probably due to the fact that patients who are pregnant after IVF often do not undergo pregnancy care in the IVF center itself. Amniocentesis is usually carried out elsewhere, and it is difficult to obtain complete data. In our IVF program, amniocentesis was performed at our hospital for all of the pregnancies. If the further care was not provided at our hospital, the patient and her gynecologist were given a questionnaire concerning the further course of the pregnancy. With this system, we obtained a complete survey of all IVF pregnancies achieved at our hospital, including the 82 pregnancies in which amniocentesis was performed. The most frequent indication for amniocentesis was a maternal age of 2 35 years (or a paternal age of 2 45 years, or both). This is not surprising, as the average age of patients undergoing IVF in the F.R.G. is over 30 years [291. The second category was that in which the parents requested amniocentesis. Judging from several
reports on the outcome of pregnancies after IVF [2,5,6,15,18,24,25], it appears unlikely that a pregnancy after IVF poses a higher risk of chromosoma1 aberrations and/or fetal malformations (e.g., neural tube defects [7,9]) than does a normal pregnancy. However, because this has not been proved, we decided to follow the parents’ request for a prenatal diagnosis whenever such a request was made. A less important indication was a history of previous pregnancies in which there were pathologic findings. Only the two women with the earlier Down’s syndrome fetuses are mentioned here, because they underwent amniocentesis once again in the following pregnancy (again after IVF). The fact that this category constitutes only 2.3% of all indications in our patients is not surprising, because a pregnancy after IVF usually is the first pregnancy for most of these patients. The incidence of anterior placentas in cases of intrauterine embryo transfer was remarkably high. We assume that this is related to our technique of transuterine ET: if the uterus is anteflexed or in the mid-position (as determined by ultrasound examination), we prefer to have the patient in the knee-elbow position during the procedure. The ET was performed in the lithotriptic position only if the uterus was retroflexed. It is well known that implantation of the embryo takes place a few days after the transfer [21-231, and in the intervening time the embryo is believed to float in the uterine cavity. However, the coincidence of the transfer position with the location of the placenta leads us to conclude that a pregnancy is more likely to be established if floating of the embryo can be avoided and implantation takes place at the site to which the embryo has been transferred. This confirms other findings indicating that a traumatic performance of the ET leads to expulsion of embryos [281 and that the embryo transfer should therefore be done as atraumatitally as possible [193. Besides, the incidence of 56% of anterior placentas in our pregnancies must be taken into consideration as a risk factor for amniocentesis in these IVF patients. The rate of perioperative complications (2 patients) was low and is in agreement with reports on the outcome of amniocentesis in a larger num-
51
ber of patients with spontaneous pregnancies 127,301.When large number of anterior placentas is taken into consideration, the aspiration of bloody amniotic fluid in two patients is not surprising. As already mentioned, we believe that the intra-uterine death of one fetus was not related to the puncture itself. In six pregnancies, we found a fetal karyotype that was abnormal. The two cases of Down’s syndrome among 56 IVF patients older than 35 years amounted to an incidence of 3.5%. This is only slightly higher than the incidence in spontaneous pregnancies [30]. Based on our small number of patients, we do not believe that this represents a general increase in the incidence of trisomy 21 after IVF treatment. In one of the Down’s syndrome cases, the genetics report did not indicate any connection between the paternal inv(9> and the fetal trisomy 21, and amniocentesis in the subsequent IVF pregnancy meanwhile showed a normal fetal karyotype. The total of five chromosome aberrations of the parents, either father or mother, represents an incidence of 6.0%. Compared to the incidence of all chromosomal aberrations that can be expected in clinical pregnancies (approximately 1.5% [4], including spontaneous abortions) our figure was considerably higher. We have no explanation for this difference. Because all of these aberrations were also found in one of the parents, they are not related to the ovarian hyperstimulation performed on the patients that is often discussed as a cause for chromosomal aberrations in these pregnancies [31-331. The further course of the single pregnancies after IVF and amniocentesis was characterized by a high incidence of toxemia, uterine bleeding before the 28th week of gestation, premature abruptio placentae, and premature labor. However, these findings must be compared with the outcome of IVF pregnancies without amniocentesis. In 1989, we published an analysis of the obstetric outcome of IVF pregnancies achieved at our hospital [14]. The data, which were completed for 1989, now include 347 pregnancies and are shown in Table III. It is clear from this data that the higher percentage of these gestational and obstetric complications was not caused by the
performance of amniocentesis. This conclusion is supported by similar results obtained by other investigators [2,5,6]. As discussed in our previous paper [14], we believe that the main cause for these complications is to be seen not in the performance of IVF, but in the performance of ovarian hyperstimulation with clomifen citrate or gonadotropins, which is usually associated with IVF treatment [1,8,16,26]. Therefore, our experience does not indicate that the performance of amniocentesis introduces any enhanced risk for the further course of pregnancy after IVF. Conclusion
Prenatal diagnosis by amniocentesis in pregnancies after in vitro fertilization and intrauterine or tubal embryo transfer requires no difference in handling compared to that for amniocentesis in spontaneous pregnancies. The higher risk of encountering problems during the course of pregnancy is due to the IVF treatment and/or to ovarian hyperstimulation and must be discussed with each couple. Our data show that the incidence of trisomy 21 was not greater after IVF than in spontaneous conceptions. Acknowledgment
The authors thank Elisabeth Lanzl, The University of Chicago, for her editorial assistance. References 1 Adashi EY, Rock JA, Sapp KC et al. Gestational outcome of clomiphene-related conceptions. Fert Steril 1979;31: 620-626. 2 Australian In Vitro Fertilisation Collaborative Group. High incidence of preterm births and early losses after in vitro fertilization. Br Med J 1985;291:1160-1163. 3 Bayerische Perinatalerhebung 1987. Kassenlrztliche Vereinigung Bayern, Munich, 1987. 4 Beier HM. Die internationale Entwicklung der Reproduktionsmedizin und der Forschung an menschlichen Embryonen 1989. Fertilitat 1990;6:74-84. 5 Cohen J, de Mouzon J. Outcome of IVF pregnancies in Europe. XIII World Congress on Fertility and Sterility, Marrakesch, Abstr No. 698, 1989.
52 6 Cohen J, Mayaux MJ, Guihard-Moscato ML. Pregnancy outcomes after in vitro fertilization: a collaborative study on 2342 pregnancies. Ann NY Acad Sci 1988;541:1-6. 7 Cornel MC, ten Kate LP, Dukes MNG et al. Ovulation induction and neural tube defects. Lancet 1989;i:l386. 8 Correy JF, Marsden DE, Schokman FCM. The outcome of pregnancy resulting from clomiphene-induced ovulation. Aust NZ J Obstet Gynaecol 1982;22:18-25. 9 Czeizel A. Ovulation induction and neural tube defects. Lancet 1989;ii:167. 10 DeCherney AH. In vitro fertilization. Curr Opinion Obstet Gynecol 1989;1:43-44. 11 Devroey P, Broeckman SP, Smitz J et al. Pregnancy after translaparascopic zygote intrafallopian transfer on a patient with sperm antibodies. Lancet 1986;i:1329. 12 Edwards RG. A decade of in vitro fertilization. Res Reprod 1990;22:1-2. 13 Edwards RG, Brinsden P, Elder K et al. Benefits of in vitro fertilisation. Lancet 1989;ii:1328. 14 Fiedler K, Wiirfel W, Kriismann G et al. Zum Schwangerschafts- und Geburtsverlauf nach in-vitro Fertilisation. Z Geburtsh Perinat 1990;194:8-12. 15 Finne PH, Hurum J, Lindemann R. Assisted fertilization. Pediatric aspects. Tidsskr Nor Laegeforen 1989;109:19111912. 16 Hack M, Brish M, Serr DM et al. Outcome ofpregnancies after induced ovulation: follow-up of pregnancies and children born after gonadotrophin therapy. JAMA 1972;220: 1329-1333. 17 In vitro fertilization/embryo transfer in the United States: 1987 results from the National IVF-ET Registry. Fert Steril 1989;51:13-19. 18 Lancaster PA. Congenital malformations after in vitro fertilization. Lancet 1987;ii;1392-1393. 19 Leeton J, Trounson A, Jessup D et al. The technique of embryo transfer. Fertil Steril 1982;38:156-161. 20 London SN, Miller MM, Weitzman GA. In vitro fertilization: an overview. J Arkansas Med Sot 1989;85:523-526. 21 Lopata A, Hay DL. The surplus human embryo: its potential for growth, blastulation, hatching, and human chorionic gonadotropin production in culture. Fertil Steril 1989;51:984-991. 22 Lopata A, Hay DL. The potential of early human embryos to form blastocysts, hatch from their zona and secrete hCG in culture. Human Reprod 1989; suppl4:87-94. 23 Malter HE, Cohen J. Blastocyst formation and hatching in
24
25
26
27
28 29
30
31
32
33
34
35
36
vitro following zona drilling of mouse and human embryos. Gamete Res 1989;24:67-80. Morin NC, Wirth FH, Johnson DH, et al. Congenital malformations and psychosocial development in children conceived by in vitro fertilization. J Pediatr 1989;115:222227. Plachot M. Chromosome analysis of spontaneous abortions after IVF. A European survey. Hum Reprod 1989;4:425-429. Plachot M, Junta AM, Mandelbaum J et al. Les echecs du developpement embryonnaire (Failure of embryonic development). Reprod Nutr Dev 1988;28:1781-1790. Rhoads GG, Jackson LG, Schlesselman SE et al. The safety and efficiency of chorionic villus sampling for early prenatal diagnosis of cytogenetic abnormalities. N Engl J Med 1989;320:609-617. Schulmann JS. Delayed expulsion of transfer fluid after IVF/ET. Lancet 1986;i:44. Siebzehnrilbl ER. Aktueller Stand der IVF- und GIFTTherapie in der Bundesrepublik Deutschland. Fertilitat 1990;6:43-49. Tabor A, Madsen M, Obel EB et al. Randomised controlled trial of genetic amniocentesis in 4606 low-risk women. Lancet 1986;i:1288-293. Vogel R, Hammerstein J, Spielmann H. Chromosomenanomalien in Gameten, Zygoten und Praimplantationsembryonen als Ursache von Fertilitatsstorungen und friihem Embryoverlust beim Menschen. Fertilitat 1988;4: 164-172. Vogel R, Rux H, Lankes T et al. Untersuchungen zur Embryotoxizitiit nach Superovulation durch Gonadotropine bei der Maus. Ferilitat 1989;5:33-39. Wramsby H, Fredga K, Liedholm P. Chromosome analysis of human oocytes recovered from preovulatory follicles in stimulated cycles. N Engl J Med 1987;316:121-124. Wiirfel W, Kriismann G, Hirsch P et al. Down syndrome (trisomy 21) in a pregnancy after intratubal embryo transfer (IVF/IT-ET). J In Vitro Fertil Embryo Transfer 1990;7:63-64. Wiirfel W, Kriismann G, Rothenaicher M et al. Schwangerschaft nach in-vitro-Fertilisation und intratubarem Embryotransfer. Geburtsh Frauenheilk 1988;48:179-181. Yovich JL, Blackledge DG, Richardson PA et al. Pregnancies following pronuclear stage tubal transfer. Fertil Steril 1987;48:851-857.
47
0 1992 Elsevier Science Publishers B.V. All rights reserved 002%2243/92/$05.00 EUROBS 01273
Prenatal diagnosis by amniocentesis in 82 pregnancies after in vitro fertilization Wolfgang Wiirfel a, Hertraut Haas-Andela b, Gottfried Kriismann a, Maria Rothenaicher ‘, Peter Hirsch a, Hemyk K. Kwapisz b, Joachim Haas b, Inge Hijgemann b and Klaus Fiedler a a Frauenklinik Dr. Kriismann, Munich and b Institut fiir Priinantale Diagnostik, Giessen, FRG Accepted for publication 26 August 1991
Summary We report on the outcome of 82 amniocenteses (AC) carried out during pregnancies after in vitro fertilization (IVF) from 1-1-1985 to 31-12-1989. The main indication for amniocentesis was a maternal age of 2 35 years. In 48 cases, we found an anterior placenta and assumed that this was related to the position in which the uterine embryo transfer was performed. In six pregnancies, we found an abnormal karyotype, including two cases of trisomy 21; the two couples decided for abortion. Four aberrations in the fetal karyotypes were also present in either the mother or the father, the resulting children are healthy. The further course of pregnancies after IVF and AC was characterized by a higher incidence of toxemia, uterine bleeding before the 28th week of gestation, abruptio placentae, and premature deliveries, when compared to the course of pregnancies after spontaneous conception. We believe that these occurrences were not caused by AC, as the incidence was higher in all our pregnancies after IVF (without AC) and has also been reported in pregnancies after ovarian hyperstimulation without IVF. Therefore, we see no reason to renounce AC after IVF. However, the special risks inherent in pregnancies after IVF must always be discussed with the couple. Amniocentesis; Down’s syndrome; Embryo transfer (ET); Intratubal embryo transfer; In vitro fertilization (IVF); Prenatal diagnosis
Introduction In vitro fertilization (IVF) and embryo transfer (ET) have become into widespread use for patients who have incurable tubal blockage [10,12,13]. Besides, IVF is an established form of treatment for other indications, such as unex-
Correspondence:
Wolfgang Wiirfel, M.D., Universitlts Frauenklinik Wiirzburg, Josef-Schneider-Str. 4, 87 Wiir burg, FGR.
plained infertility and male subfertility [20]. In all these patients without tubal occlusion, the embryo (or pronucleus) can also be transferred directly into the fallopian tubes (zygote intrafallopian transfer (ZIFT), pronuclear stage tubal transfer (PROST), intratubal embryo transfer (IT-ET) [11,35,36]). By combining all of these indications, between 1982 and 1989 we counted more than 2000 pregnancies in the Federal Republic of Germany [29], and about ten times as many worldwide [5,6,17]. Therefore, the problems of pregnancy care for IVF patients are becoming
48
more and more important. Because of the higher average age of IVF patients compared to that of women with normal pregnancies [5,6,29], prenatal diagnosis of fetal malformations and/or chromosomal disorders is of particular interest. In this article, we report on our experience with amniocentesis in 82 pregnancies after IVF. Material and Methods Indications
anestthesia and always under ultrasound control. The whole procedure was recorded (Polaroid; Printer, Mitsubishi). We attempted to aspirate a total of 20 ml of amniotic fluid during each puncture. The number of attempts was limited to two per patient. There were five twin pregnancies in which the location of the placentas and of the fetuses did not allow us to perform the amniocentesis during one puncture; thus, two punctures were carried out. In cases of anterior placentas, we tried to perform the puncture without perforating the placenta. In eight patients, however, we were forced to do the puncture through the placenta. After amniocentesis, all patients remained in bed for at least 3 to 4 h. During this time and approximately 24 h later, we checked each patient for possible uterine contractions (by tocography) and signs of infection (elevated leukocyte counts, rise in temperature). Rh-negative patients routinely received IgG (PartubolinTM i.m.> prophylactically.
The most important indication for amniocentesis was a maternal age of 35 years or more (56 patients). In four of these pregnancies, the mother’s age was > 35 years and the father’s age was > 45 years. A paternal age of 45 years or above was not considered to be a strict indication [30]; however, after discussion with such couples, three wanted an amniocentesis to be performed. Two women had had a previous pregnancy in which a chromosomal aberration was detected (trisomy 21, with a subsequent elective abortion). Both of these pregnancies with Down’s_syndrome fetuses were achieved after IVF. In 21 cases, there was no medical indication for prenatal diagnosis by amniocentesis. In all of these cases, however, we respected the couple’s request for the procedure. Ten couples did not consent to amniocentesis in spite of a maternal age of > 35 years. Four of these ten would have opposed an abortion in any case; and six patients had uterine bleeding during the first 12 weeks of gestation, which was considered a relative contraindication for performance of amniocentesis. Thus, the total number of patients who underwent amniocentesis was 80.
From 1-1-1985 to 31-12-1989 we performed 82 amniocenteses in 80 women who were pregnant after IVF/ET or intratubal embryo transfer (IVF/IT-ET). Punctures were carried out in five women pregnant with twins; no puncture was carried out in one patient pregnant with triplets. We found an anterior placenta in 48 pregnancies (Table I). Because of the position of the uterus, 46 of these patients had received an embryo transfer in the knee-elbow position. In the other two patients, the transfer was performed in
Technique All amniocenteses
TABLE I
were carried out between the 16th and 17th week of gestation. The ultrasound apparatus which we used was an array-scan (7000 S, Picker). Before the puncture, the ultrasound probe was cleaned thoroughly with warm water, covered with gel, and wrapped in sterile foil. The abdomen was disinfected with BetaisodonnaTM, which served as the transducer medium as well. The puncture was carried out with spinal needles (diameter: 9 mm), usually without local
Results
Transfer position of patient and occurrence intrauterine embryo transfer (n = 76) Position of uterus
No. of transfers
Position
Anteflexed Mid position Retroflexed
52 13 11
52 13 -
knee-elbow
in all cases of
No. with lithotriptic
11
anter’or placenta 38 8 2
49 TABLE II Abnormal fetal karyotypes found in six amniocenteses. The prenatal diagnosis was confirmed by genetic examination after delivery and by pathologic and genetic examination after abortion Patient no.
Patient age (years)
AFP
Ache activity
Fetal karyotype
Parental karyotypes (lymphocyte culture)
Further course of pregnancy
1
35
normal
negative
47,XX, +21
M *: 46,Xx; P ‘: 46,XY
2
38
normal
negative
46,XY, 15p +
3
36
normal
negative
46,XY, 16qh +
4
28
normal
negative
5
37
normal
negative
46,Xx, inv (7) (P12 q221 47,Xx, + mar
6
35
normal
negative
47,xx,
M: 46,Xx P: 46,XY,15p + M: 46,XY, 16qh + P: 46,XY M: 46,Xx P: 46,XY, inv (7) (~12 q22) M: 47,Xx, + mar P: 46,XY M: 46,Xx P: 46,XY, inv (9)
Abortion, diagnosis of Down’s syndrome confirmed Cesarean section, child healthy Normal delivery, child healthy Normal delivery, child healthy Cesarean section, child healthy Abortion, diagnosis of Down’s syndrome confirmed
+21
* M = maternal karyotype ’ P = paternal karyotype
the lithotriptic position. In all cases of tubal embryo transfer, we found a fundal placenta (6 patients). Contaminated amniotic fluid was aspirated during three punctures; in two cases, blood was removed by centrifugation. In the third case, the amniotic fluid was brownish, and centrifugation led to no change in the color. Subsequent culture showed no growth of cells, and the pregnancy ended in intrauterine death of the fetus 8 days
later. We assume that the pregnancy was already in a critical state when the amniocentesis was performed. In one patient, we registered uterine contractions within the first 24 h after puncture. Another patient developed an abdominal hematoma. No uterine bleeding, leakage, or fever was observed in any case. In six pregnancies (Table II), a numerical or _ structural aberration was found in the fetal karyotype. In two patients (one of them after tubal
TABLE III Further course of single pregnancies after in vitro fertilization spontaneous conception (Bayerische Perinatalerhebung, BPE)
(IVF) and amniocentesis
(AC), after IVF only, and after
Pregnancies with complications
IVF and AC (1985-1989) (% (No.11
IVF only (1985-1989) (%I
Toxemia Uterine bleeding before 28th week of gestation Intra-uterine death of fetus Induced abortion Abruptio placenta Premature rupture of membranes Premature labor ( < 37th week of gestation)
9.1 (7) 5.2 (4) 1.3 (1) 2.6 (2) 3.9 (3) 15.6 (12) 10.4 (8)
9.7 10.4 2.7 0.7 3.8 20.8 10.7
a Includes all children born by 30-6-1990.
BPE 1987 (%I 3.1 3.8 0.5 0.5 19.2 4.2
50
embryo transfer [34]), the fetal karyotype showed trisomy 21 (47,Xx, +21). Both of the couples decided for abortion, after which the prenatal diagnosis of Down’s syndrome was confirmed. In one of these couples, the paternal karyotype showed a pericentric inversion of chromosome 9 in the heterochromatic area. The wife became pregnant again by IVF, and on amniocentesis the fetus now showed a regular karyotype. Of the other four pathologic aberrations of the fetal karyotype, each was found also in the karyotype of either the mother or the father. All of these four children were born and were healthy. The further course of all single pregnancies after IVF and AC is given in Table III and is compared to the outcome of all our single pregnancies after IVF from 1985 to 1989 and to the outcome of single pregnancies after spontaneous conception in Bavaria. Among the five twin pregnancies, we had 2 cases of toxemia, 3 cases with premature rupture of membranes, and 3 premature deliveries. Discussion The lack of published reports on the performance and results of amniocentesis in pregnancies after IVF is probably due to the fact that patients who are pregnant after IVF often do not undergo pregnancy care in the IVF center itself. Amniocentesis is usually carried out elsewhere, and it is difficult to obtain complete data. In our IVF program, amniocentesis was performed at our hospital for all of the pregnancies. If the further care was not provided at our hospital, the patient and her gynecologist were given a questionnaire concerning the further course of the pregnancy. With this system, we obtained a complete survey of all IVF pregnancies achieved at our hospital, including the 82 pregnancies in which amniocentesis was performed. The most frequent indication for amniocentesis was a maternal age of 2 35 years (or a paternal age of 2 45 years, or both). This is not surprising, as the average age of patients undergoing IVF in the F.R.G. is over 30 years [291. The second category was that in which the parents requested amniocentesis. Judging from several
reports on the outcome of pregnancies after IVF [2,5,6,15,18,24,25], it appears unlikely that a pregnancy after IVF poses a higher risk of chromosoma1 aberrations and/or fetal malformations (e.g., neural tube defects [7,9]) than does a normal pregnancy. However, because this has not been proved, we decided to follow the parents’ request for a prenatal diagnosis whenever such a request was made. A less important indication was a history of previous pregnancies in which there were pathologic findings. Only the two women with the earlier Down’s syndrome fetuses are mentioned here, because they underwent amniocentesis once again in the following pregnancy (again after IVF). The fact that this category constitutes only 2.3% of all indications in our patients is not surprising, because a pregnancy after IVF usually is the first pregnancy for most of these patients. The incidence of anterior placentas in cases of intrauterine embryo transfer was remarkably high. We assume that this is related to our technique of transuterine ET: if the uterus is anteflexed or in the mid-position (as determined by ultrasound examination), we prefer to have the patient in the knee-elbow position during the procedure. The ET was performed in the lithotriptic position only if the uterus was retroflexed. It is well known that implantation of the embryo takes place a few days after the transfer [21-231, and in the intervening time the embryo is believed to float in the uterine cavity. However, the coincidence of the transfer position with the location of the placenta leads us to conclude that a pregnancy is more likely to be established if floating of the embryo can be avoided and implantation takes place at the site to which the embryo has been transferred. This confirms other findings indicating that a traumatic performance of the ET leads to expulsion of embryos [281 and that the embryo transfer should therefore be done as atraumatitally as possible [193. Besides, the incidence of 56% of anterior placentas in our pregnancies must be taken into consideration as a risk factor for amniocentesis in these IVF patients. The rate of perioperative complications (2 patients) was low and is in agreement with reports on the outcome of amniocentesis in a larger num-
51
ber of patients with spontaneous pregnancies 127,301.When large number of anterior placentas is taken into consideration, the aspiration of bloody amniotic fluid in two patients is not surprising. As already mentioned, we believe that the intra-uterine death of one fetus was not related to the puncture itself. In six pregnancies, we found a fetal karyotype that was abnormal. The two cases of Down’s syndrome among 56 IVF patients older than 35 years amounted to an incidence of 3.5%. This is only slightly higher than the incidence in spontaneous pregnancies [30]. Based on our small number of patients, we do not believe that this represents a general increase in the incidence of trisomy 21 after IVF treatment. In one of the Down’s syndrome cases, the genetics report did not indicate any connection between the paternal inv(9> and the fetal trisomy 21, and amniocentesis in the subsequent IVF pregnancy meanwhile showed a normal fetal karyotype. The total of five chromosome aberrations of the parents, either father or mother, represents an incidence of 6.0%. Compared to the incidence of all chromosomal aberrations that can be expected in clinical pregnancies (approximately 1.5% [4], including spontaneous abortions) our figure was considerably higher. We have no explanation for this difference. Because all of these aberrations were also found in one of the parents, they are not related to the ovarian hyperstimulation performed on the patients that is often discussed as a cause for chromosomal aberrations in these pregnancies [31-331. The further course of the single pregnancies after IVF and amniocentesis was characterized by a high incidence of toxemia, uterine bleeding before the 28th week of gestation, premature abruptio placentae, and premature labor. However, these findings must be compared with the outcome of IVF pregnancies without amniocentesis. In 1989, we published an analysis of the obstetric outcome of IVF pregnancies achieved at our hospital [14]. The data, which were completed for 1989, now include 347 pregnancies and are shown in Table III. It is clear from this data that the higher percentage of these gestational and obstetric complications was not caused by the
performance of amniocentesis. This conclusion is supported by similar results obtained by other investigators [2,5,6]. As discussed in our previous paper [14], we believe that the main cause for these complications is to be seen not in the performance of IVF, but in the performance of ovarian hyperstimulation with clomifen citrate or gonadotropins, which is usually associated with IVF treatment [1,8,16,26]. Therefore, our experience does not indicate that the performance of amniocentesis introduces any enhanced risk for the further course of pregnancy after IVF. Conclusion
Prenatal diagnosis by amniocentesis in pregnancies after in vitro fertilization and intrauterine or tubal embryo transfer requires no difference in handling compared to that for amniocentesis in spontaneous pregnancies. The higher risk of encountering problems during the course of pregnancy is due to the IVF treatment and/or to ovarian hyperstimulation and must be discussed with each couple. Our data show that the incidence of trisomy 21 was not greater after IVF than in spontaneous conceptions. Acknowledgment
The authors thank Elisabeth Lanzl, The University of Chicago, for her editorial assistance. References 1 Adashi EY, Rock JA, Sapp KC et al. Gestational outcome of clomiphene-related conceptions. Fert Steril 1979;31: 620-626. 2 Australian In Vitro Fertilisation Collaborative Group. High incidence of preterm births and early losses after in vitro fertilization. Br Med J 1985;291:1160-1163. 3 Bayerische Perinatalerhebung 1987. Kassenlrztliche Vereinigung Bayern, Munich, 1987. 4 Beier HM. Die internationale Entwicklung der Reproduktionsmedizin und der Forschung an menschlichen Embryonen 1989. Fertilitat 1990;6:74-84. 5 Cohen J, de Mouzon J. Outcome of IVF pregnancies in Europe. XIII World Congress on Fertility and Sterility, Marrakesch, Abstr No. 698, 1989.
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