1 Departments of Obstetrics and Gynaecology, 2 Central Laboratory, 3 Medical Informatics Research Centre in Turku (MIRCIT), University of Turku, Kiinamyllynkatu 48, FIN-20520 Turku, 4 Department of Central Laboratory and Rehabilitation Research Centre of the Social Insurance Institution, Peltolantie 3, FIN-20720 Turku, 5 The Family Federation of Finland, Maariankatu 3A, FIN-20100 Turku, 6 Central Laboratory, Oulu University Hospital, P.O.Box 22, 90221 Oulu, 7 Department of Central Laboratory, Tampere University Hospital, P.O.Box 2000, 33521 Tampere and 8 Department of Obstetrics and Gynaecology, University of Helsinki, Haartmaninkatu 2, P.O.X. 140, HUS, FIN-00029 Helsinki, Finland
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Abstract |
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Key words: AFP/ß-HCG/Down's syndrome/FET/ICSI/IVF
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Introduction |
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The reason behind the marker level alterations in assisted reproduction pregnancies has remained unknown. The fact that the median age of women becoming pregnant with assisted reproduction is higher than in spontaneous pregnancies cannot alone explain the higher FPR (Heinonen et al., 1996; Wald et al., 1999
). Recently, it was suggested that the high FPR in IVF and oocyte donation pregnancies could reflect factors associating with the pregnancy itself and not with the assisted reproduction procedure (Maymon and Shulman, 2001
).
The aim of this study was to evaluate the effects of the number of embryos transferred and the amount of gonadotrophin medication for ovarian stimulation for IVF and ICSI on maternal serum levels of ß-HCG and AFP in the second trimester Down's syndrome screening. We hypothesized that the ovulation stimulation regimens may have an effect both on the ovum and the maturation of the endometrium, and thereby on the implantation of the embryo on the uterus wall. To test this hypothesis, we studied the marker levels in frozen embryo transfer (FET) pregnancies, where ovarian stimulation was not used, but had affected the ovum prior to fertilization.
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Materials and methods |
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Screening for Down's syndrome was performed by analysing maternal serum levels of ß-HCG and AFP between pregnancy weeks 14 and 18, and the risk ratio for Down's syndrome was counted with Wald's formula (Wald et al., 1994). The data were collected from different registers: birth register, hospital registers, malformation register, legal abortion register and laboratory registers in Finland between years 1994 and 1999. The control group consisted of 6548 singleton pregnancies with known normal outcome and no congenital anomalies. The information on the number of embryos transferred and the medication used for ovarian stimulation was collected from the patient documents. The data concerning the number of embryos transferred was missing from four IVF pregnancies and one ICSI pregnancy.
Serum AFP and ß-HCG were analysed with dual-labelled time-resolved immunofluorometric assay (hAFP/free HCG beta Dual kit; Wallac, Turku, Finland).
The differences in the levels of the two analyses between the different study groups and the control group were evaluated with the KruskalWallis and Wilcoxon tests. An analysis of co-variance (ANCOVA) for log-transformed multiples of the median (MoM) values was used to adjust the age differences between the groups. Spearman's correlation coefficient was used to assess whether the number of embryos transferred or the amount of medication used had an effect on serum ß-HCG or AFP levels. SAS 6.12 for Windows was used in all calculations.
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Results |
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The post-hoc comparisons of the serum MoM levels of free ß-HCG were made between the study groups and the controls: a statistically significant difference in the ß-HCG levels was observed only between the FET group and the controls (P = 0.012) (Table II). After exclusion of the four cases on estrogen in the FET group, the ß-HCG level remained the highest among the study groups, but the difference did not reach statistical significance. There were no differences in AFP levels between any of the study groups and the controls (Table II
).
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Discussion |
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To our knowledge, the effect of the number of embryos transferred on the marker levels has not been evaluated previously, although an early implantation of several embryos may interfere with the marker levels and thereby cause the false positive results in Down's syndrome screening in assisted reproduction pregnancies. We found a weak positive correlation between the number of embryos transferred and the serum marker levels only in the IVF group. However, the statistical power of this analysis is diminished by the fact that both one- and four-embryo transfers were rarely performed in Finland during years 19941999, which makes the sizes of the corresponding subgroups small. According to our data, the number of embryos transferred may have an influence on the marker levels and should be re-evaluated with larger data. Although an early ultrasound examination was performed around week 7 of pregnancy to exclude multiple pregnancies, early implantation of several embryos could affect HCG levels even during second trimester screening.
One of our basic hypotheses was that the dose of gonadotrophin used for ovarian stimulation may cause alterations in the marker levels in assisted reproduction pregnancies. However, we failed to find any association between the levels of maternal serum markers and the amount of ovarian stimulation medication. Unfortunately, we were unable to evaluate the effect of actual ovarian response during stimulation on the marker levels. One may speculate that in maternal mid-trimester serum screening an increased ß-HCG level can occur as a result of an excessive ovarian response, where multiple follicles and corpora lutea develop. However, the FET group, which we assumed to be the closest to the controls in marker levels, exhibited the highest serum HCG MoM levels and highest FPR (30.8%). Instead of ovulation stimulation, the mechanism behind elevated serum HCG levels in the FET group may be due to a placentation failure that could result in changes in trophoblast function and thus in HCG production. Among our study population the implantation rate was slightly lower in the FET group (49%) than in the IVF (54%) or the ICSI (51%) groups. This is in line with the higher pregnancy rate after transfers of fresh than cryopreserved embryos. Interestingly, after exclusion of subjects using estrogen, the ß-HCG levels diminished slightly. It has been suggested that some unidentified endocrine/metabolic errors might explain sub/infertility. These factors may affect the serum screening results in assisted reproduction pregnancies and may also be causative for the complications among these pregnancies (McElrath and Wise, 1997; Maymon and Shulman, 2001
).
It has been speculated that the high HCG levels in assisted reproduction pregnancies may be caused by the higher progesterone levels in these pregnancies than in spontaneous pregnancies (Wald et al., 1999). These workers suggested that the reason for high progesterone levels could be multiple follicle development in IVF and ICSI pregnancies and subsequent formation of multiple corpora lutea, which could in turn increase the production of HCG from the placenta. The results obtained with the FET group in the present study and the recent data concerning pregnancies after oocyte donation (Maymon and Shulman, 2001
) are confusing with regard to this theory, because in FET and oocyte donation pregnancies excessive follicles or corpora lutea do not exist. In the FET cases on estrogen replacement therapy no corpus luteum is involved, but progesterone substitution is used up to week 8 of pregnancy, as in all IVF/ICSI/FET protocols of the present study. Since the half-life of natural progesterone is only 4 h, we assumed that progesterone substitution cannot interfere the mid-trimester (week 1418 of pregnancy) marker levels in assisted reproduction pregnancies. It was also recently shown that in IVF pregnancies serum levels of progesterone, but not HCG, are significantly higher than in normal pregnancies up to 8 weeks post-conception (Costea et al., 2000
).
According to the present study, the FPR in Down's syndrome screening in assisted reproduction pregnancies is higher than in spontaneous pregnancies. This results in an increased need for invasive procedures, e.g. amniocentesis, which carries 0.51% risk of abortion. Even this small risk can be too much in these valuable pregnancies where infertility may have lasted for many years.
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Notes |
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References |
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accepted on October 3, 2001.