1 Research Institute of Growth & Development (GROW), University of Maastricht and IVF-Laboratory and 2 Department of Obstetrics & Gynaecology, Academic Hospital Maastricht, Maastricht, The Netherlands
3 To whom correspondence should be addressed at: Research Institute of Growth & Development (GROW), University of Maastricht, Maastricht, The Netherlands. Email: avmn{at}sgyn.azm.nl
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Abstract |
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Key words: assisted reproductive technology/multiple pregnancy/single embryo transfer
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Introduction |
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The high twin pregnancy rate after IVF is the result of the current standard practice of transferring more than one embryo. The elective transfer of only a single embryo (eSET) has been shown to be an effective method to reduce the incidence of twin pregnancies in twin-prone IVF/ICSI patients without compromising the overall ongoing pregnancy rate (Vilska et al., 1999; ESHRE Campus Course Report, 2001
; Gerris et al., 2001
, 2002
; Martikainen et al., 2001
; Tiitinen et al., 2001
, 2003
; De Neubourg and Gerris, 2003
).
It was recently recommended by the ESHRE consensus meeting on risks and complications in ART that eSET should be proposed in the first and second treatment cycles (Land and Evers, 2003). Since 2003 in Belgium, SET in the first, and eSET in the second treatment cycle is mandatory in patients younger than 36 years of age, to obtain maximal reimbursement of the costs of IVF treatment (Ombelet, 2004
,). However, little information is available on eSET results to substantiate this proposal to limit eSET to the first two treatment cycles. In the present study we evaluate the results per treatment cycle number in a cohort of patients in whom eSET was offered as the standard transfer policy during the first three treatment cycles.
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Materials and methods |
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In the study period the costs of the first three IVF/ICSI treatment cycles were fully reimbursed in The Netherlands. For this reason the study has been limited to the first three treatment cycles, including the transfer of frozen embryos, if any.
Ovarian stimulation protocol
Patients were down-regulated with daily s.c. injections of triptorelin (Decapeptyl; Ferring B.V., Hoofddorp, The Netherlands) or nafarelin intranasally (Synarel; Searle BV, Maarssen, The Netherlands) according to the long protocol. To stimulate multiple follicular development, recombinant FSH (Puregon, Organon, Oss, The Netherlands) was used. Follicle growth was monitored by ultrasound and 5000 IU of hCG (Pregnyl, Organon, Oss, The Netherlands) was given as soon as at least three follicles were 18 mm. Ultrasound-guided oocyte retrieval was performed 36 h after hCG administration. The luteal phase was supported by progesterone (Progestan, Organon, Oss, The Netherlands) 600 mg daily intravaginally, starting at the day of ovum pick-up and continued for 1416 days. In the case of pregnancy 600 mg Progestan was continued for three more weeks.
Laboratory and embryo transfer procedures
IVF and ICSI procedures used have been described in detail earlier (Dumoulin et al., 2001). Oocytes and embryos were cultured individually in 5 µl droplets covered by mineral oil in an atmosphere of 5% O2, 5% CO2 and 90% N2 in sequential culture media either from Vitrolife (Göteborg, Sweden) or from Cook (Eight Mile Plains, Queensland, Australia). Embryo transfer was routinely performed on day 2 or day 3 after ovum retrieval. Embryos were evaluated at 2326 h post-injection (in the case of ICSI) or 2528 h post-insemination (in the case of IVF), 4145 h post-injection/insemination, and in the case of day 3 transfer, also at 6569 h post-injection/insemination. For each embryo originating from a normally fertilized oocyte, an embryo score was calculated on the basis of morphological grade (1 to 4, with grade 4 being the best grade, using the grading system of Bolton et al., 1989
), number of blastomeres and presence or absence of multinucleated blastomeres (MNBs). Embryos that had reached the 4- or 5-cell stage on day 2 or the 8-cell stage on day 3, in combination with having the best morphological grade (regular, even sized blastomeres with <20% fragmentation) and an absence of MNBs were classified as good quality embryos (Van Royen et al., 1999
). When patients were eligible for standard eSET policy and if at least one good quality embryo was available, only a single embryo was transferred (eSET). In all other cases, two embryos, if available, were transferred (double embryo transfer, DET). Cryopreservation of supernumerary embryos was performed on the third day after ovum retrieval. Only embryos which had reached the 8-cell stage and which were considered to be of sufficient morphological quality (grades 3 or 4) were cryopreserved. After thawing, two embryos, if available, were transferred, also in cases in which in the fresh transfer eSET was applied.
Pregnancy
An hCG pregnancy test with a detection limit of 50 IU/l in urine was performed 1416 days after embryo transfer and patients with a positive test had an ultrasound examination 3 weeks later. Patients were asked to report an abortion occuring after this ultrasound examination immediately. In this study an ongoing pregnancy was defined as the presence of at least one intrauterine gestational sac with fetal heartbeat on ultrasound at 7/8 weeks gestation and no report of pregnancy loss before 12 weeks gestation.
Statistics
The 2 test with Bonferroni correction was used to compare the pregnancy rates. Differences were considered significant at P<0.05.
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Results |
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The results of the first treatment cycles (performed in the period July 2000 until December 2001) and of all eventual subsequent second or third treatment cycles or frozen embryo replacement cycles (performed in the period July 2000 until May 2004) of the 326 patients who accepted our standard eSET policy can be found in Table I. In these patients, after transfer of fresh embryos, 159 ongoing pregnancies were achieved (49%), of which 40 (25%) were twin pregnancies. In 175 patients (54%), an ongoing pregnancy was achieved, either after transfer of fresh embryos in the first, second or third treatment cycle, or after transfer of cryopreserved embryos.
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In 112 cycles at least one surplus embryo of good morphology was available after transfer and subsequently frozen. After eSET significantly more often embryos could be cryopreserved as compared to after DET (53% and 13% of the treatment cycles, respectively). In Table I, pregnancy results are shown for 65 thaw cycles performed in patients who did not get pregnant in any of their transfers of fresh embryos. This resulted in 16 ongoing pregnancies (25%). In 12 patients, thawing has not yet been performed because of the patients' wishes. The cumulative ongoing pregnancy rates after fresh and frozen transfers were significantly different between eSET and cSET and between eSET and DET (13% cSET, 41% eSET and 30% DET). An additional 35 patients who became pregnant from fresh embryos, had embryos cryopreserved. In seven of these patients, embryos have been thawed, resulting in two pregnancies.
After the first cycle, 48 patients (30% of the patients not getting pregnant in the first cycle) dropped out because of medical reasons or patient wishes. After the second cycle the drop-out rate was 36% (27 patients). The Dutch health insurance system only reimburses three IVF/ICSI cycles. Therefore the drop-out rate after the third cycle increased to 75%.
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Discussion |
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The pregnancy rates obtained in the first two treatment cycles in our study are comparable to those reported by other studies in which eSET was performed in either only the first (Gerris et al., 1999; Vilska et al., 1999
) or in the first two treatment cycles (Martikainen et al., 2001
; De Neubourg and Gerris, 2003
). It is concluded in several publications that eSET should be limited to the first two cycles (Martikainen et al., 2001
; De Neubourg and Gerris, 2003
; Land and Evers, 2003
). However, our results show that pregnancy rates of eSET compared to DET are not significantly different up to and including the third treatment cycle, although it must be noted that the group of patients that received a third treatment cycle is relatively small (four pregnancies in 20 patients receiving only a single embryo). Yet, the pregnancy rates in the third treatment cycle in both the eSET and the DET groups are decreased compared with the previous treatment cycles. These decreasing pregnancy rates in successive IVF cycles were already reported by others (Tan et al., 1992
; Templeton et al., 1996
).
Our results confirm earlier publications showing that cryopreservation clearly improves the cumulative pregnancy rate after eSET (Martikainen et al., 2001; Tiitinen et al., 2001
). In our study, in significantly more eSET cycles could embryos be cryopreserved compared to DET cycles (53 and 13%, respectively). After thawing, two embryos, if available, were transferred. The reason for this was that cryopreserved embryos have a lower potential for implantation and therefore were not considered to be good quality embryos, which is a necessary condition for eSET (Levran et al., 1990
; Edgar et al., 2000a
,b
). At present, almost all embryos from patients who did not became pregnant from fresh embryos have been thawed. A significantly higher cumulative pregnancy rate in the eSET group can be seen, caused by the additive effect of frozen embryo transfers.
In order to improve the results after eSET, effort is taken to improve the embryo selection criteria. In addition, to further reduce the risk of twins, the proportion of eSET transfers needs to be enlarged. Tiitinen et al. (2003) reported an increase over the years in the proportion of eSET in their IVF/ICSI programme from 10% to 56%, while maintaining comparable pregnancy rates. The same applies for Gerris et al., who reported an increase of eSET from 9.8% in 1998 to 23.6% in the year 2001 (Gerris et al., 2002
). This increase was achieved by liberalization of the criteria for application of eSET. However, the twin pregnancy rate in both clinics (ranging from 7.5% to 13.5%) (Tiitinen et al., 2003
; Gerris et al., 2004
) remained above the twin pregnancy rate after a spontaneous pregnancy (ranging from 1% to 1.5%) (Ghai and Vidyasagar, 1988
; Dhont, 2001
). In the present study, 19% of the patients eligible for the eSET policy received one embryo. In the DET group however, a high twin pregnancy rate (23%) was found in spite of suboptimal quality embryos. Apparently our embryo selection criteria for eSET are still too strict. Instead of gradually liberating the criteria for eSET, we decided to start a prospective randomized study in which consenting patients will be allocated by lot for the transfer of either one or two embryos, irrespective of the presence or absence of a top quality embryo. This study will provide more insight into whether we should perform SET in every patient, or how the selection of the embryos and the patients suitable for SET can be improved.
In conclusion, eSET in a selected group of patients (younger than 38 years, with at least one good quality embryo) can be performed not only in the first two treatment cycles, but also in the third cycle while maintaining a pregnancy rate comparable to DET. In this way the proportion of eSET in the total IVF programme can be increased, which will result in a further decline in the twin pregnancy rate.
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Acknowledgements |
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References |
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Submitted on October 20, 2003; resubmitted on September 2, 2004; accepted on October 22, 2004.