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Is there an increased risk of malformations after assisted reproductive technologies?
RBMOnline - Vol 10. Suppl. 3 2005 83–89 www.rbmonline.com/Article/1759
Is there an increased risk of malformations after assisted reproductive technologies? Dr Michael Ludwig was born in 1968 in Germany and concluded university in 1994. He then became Gynaecologist and Obstetrician at the University Hospital Lübeck and ran the Division of Reproductive Medicine and Gynaecologic Endocrinology at the Department of Gynaecology and Obstetrics. Since 2003 he has worked at the Endokrinologikum Hamburg, a private centre for hormone and metabolic diseases, gynaecological endocrinology and reproductive medicine and has recently been promoted to Head of the Division of Gynecologic Endocrinology and Reproductive Medicine. Dr Ludwig has published extensively in different fields of reproductive medicine, especially assisted reproductive techniques. His special interests lie in the field of evidence-based reproductive medicine and epidemiological studies in assisted reproductive treatment. Dr Michael Ludwig Michael Ludwig Endokrinologikum Hamburg, Zentrum für Hormon- und Stoffwechselerkrankungen, Reproduktionsmedizin und Gynäkologische Endokrinologie, Lornsenstraße 6, 22767 Hamburg, Germany Correspondence: Tel: +49 40 30628320; Fax: +49 40 30628349; e-mail: [email protected]
Abstract The question as to whether there is an increased risk of major malformations after assisted reproductive technologies has been answered insofar that it is slightly increased (30% relative risk when absolute figures analysed). This means an increased risk of 1:15 to 1:12 pregnancies when naturally conceived and intracytoplasmic sperm injection (ICSI) pregnancies are compared. Furthermore, there seems to be no risk difference between more invasive techniques (ICSI) or less invasive ones (conventional IVF). However, there are as yet no sufficiently robust data to describe the risk of children born either after ovarian stimulation only or intrauterine insemination procedures. With these data, it would be possible to clarify whether the increased risk after IVF and ICSI is due to the in-vitro culture techniques or to the ovarian stimulation procedure, or whether infertility per se is a risk factor. The latter seems to be suggested by retrospective studies. The different theories behind the increased risk of major malformations and the current database worldwide are described in this review. Keywords: ICSI, IVF, malformation, risk
Introduction The birth of a healthy singleton is increasingly becoming the major and most important outcome parameter after assisted reproductive technologies (Wennerholm and Bergh, 2004). Data have been collected in recent years to study whether or not there is an increased risk, especially for pregnancy complications or major malformations in the children born (Ludwig and Diedrich, 2002). Firstly, it is not possible to give an answer regarding this risk or the risk for pregnancy and birth complications for all types of assisted reproduction. Only in IVF and intracytoplasmic sperm injection (ICSI) have enough valid data been collected in recent years to give a true estimation. Currently, for intrauterine insemination cycles there are insufficient data to estimate the risks. Regarding ovarian stimulation cycles using either gonadotrophins or clomiphene citrate, insufficient data can be found in the literature. Some studies, however, have concentrated on particular abnormalities, e.g. hypospadias (Sorensen et al., 2005), but did not look for the overall risk. Furthermore, certain limitations in study design will make it quite impossible, even in the future, to give a true estimation of the
influence of assisted reproduction and the underlying impairment of fertility in treated couples on the health outcome of children (Buck Louis et al., 2005). Regarding pregnancy complications, a recent meta-analysis and systematic review has proved that children born after IVF with or without ICSI have an increased risk (Helmerhorst et al., 2004; Jackson et al., 2004). The most important results are shown in Table 1. The present paper will concentrate on whether major malformations are also increased, and what the cause of such an increase might be.
Is there an increased risk of major malformations? The question as to whether there is an increased risk of major malformations in assisted reproduction can be answered easily, since two recent meta-analyses have already collected all data published on that topic up to March 2003 (Hansen et al., 2005) and September 2003 (Rimm et al., 2004) respectively. Hansen et al. (2005) showed, in the meta-analysis with 25
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Risk of malformations after assisted reproduction - M Ludwig
Table 1. Results of a systematic review and a meta-analysis on the outcome of singleton pregnancies after assisted reproduction treatments. Data according to Helmerhorst et al. (2004) and Jackson et al. (2004). Jackson et al. (2004)
Helmerhorst et al. (2004)
Publication period analysed
1978–2002
1985–2002
Inclusion criteria
– Comparison of IVF to spontaneous conception – More than 50% in the IVF group had received standard IVF (defined as ovulation induction–egg retrieval–IVF–intrauterine embryo transfer), (non-standard: GIFT, ICSI, frozen embryo transfers, use of donor embryos) – More than 50% in the control group were fertile – Control for at least maternal age and parity – Singleton gestations reported and analysed separately from multiple gestations – Outcomes explicitly defined – Risk ratios with 95% confidence intervals provided or sufficient data to enable calculation
– Categorical data present on any of the following outcomes (gestational age and weight at birth, Caesarean section, perinatal death, and admission to neonatal intensive care) – Studies without a control group of natural conceptions or that did not distinguish singleton from multiple pregnancies were excluded
Study cohort
12,283 singletons
5361 singletons
Control cohort
1.9 million
7038
Risk of perinatal mortality
OR 2.2 (95% CI 1.6–3.0)
RR 1.68 (95% CI 1.11–2.55)
Risk of preterm delivery (<37 weeks)
OR 2.0 (95% CI 1.7–2.2)
RR 2.04 (95% CI 1.80–2.32)
Risk of very preterm delivery (<32 weeks)
n/a
RR 3.27 (95% CI 2.03–5.28)
Risk of low birth weight (<2.500 g)
OR 1.8 (95% CI 1.4–2.2)
RR 1.40 (95% CI 1.15–1.71)
Risk of very low birth weight (<1.500 g)
OR 2.7 (95% CI 2.3–3.1) RR 3.00 (95% CI 2.07–4.36)
Risk of small for gestational age
OR 1.6 (95% CI 1.3–2.0)
RR 1.40 (95% CI 1.15–1.71)
Risk of Caesarean section
n/a
RR 1.54 (95% CI 1.44–1.66)
Risk of admission to neonatal intensive care unit
n/a
RR 1.27 (95% CI 1.16–1.40)
GIFT = gamete intra-Fallopian transfer; ICSI = intracytoplasmic sperm injection; OR = odds ratio; RR = relative risk; CI = confidence interval; n/a = not available.
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studies, an overall increase in major malformations of about 30%. Details of sub-analysis are given in Table 2. Interestingly, 18 studies originated from Europe, and half of the studies were published in the last 4 years of the time period analysed. Five of the studies analysed concentrated on ICSI only (Hansen et al., 2005). Lie et al., in a recent metaanalysis, compared data on major malformation risk in children born after ICSI with those born after other IVF techniques. Only four studies were identified that were performed prospectively, did not overlap with other studies and provided reliable and comparable data on birth defects for children either conceived after ICSI or another IVF technique. These studies included data on 5395 children born after ICSI and 13,086 children born after other IVF techniques. The data gave a relative risk for the presence of major malformations after ICSI of 1.2 [95% confidence interval (CI) 0.97–1.28] (Lie et al., 2004). All these findings were confirmed by an independent metaanalysis performed by Rimm et al. that described an overall odds ratio of 1.29 [95% CI 1.01–1.67] for children born after IVF or ICSI as compared with spontaneously conceived children. The risk for singletons born after ICSI [odds ratio 1.33 (95% CI 0.90–1.95)] and IVF [odds ratio 1.51 (95% CI 0.85–2.7)] was increased compared with spontaneously conceived children, but this difference was not significant.
No significant difference was found for the risk of children born after IVF or ICSI, when these groups were compared directly with each other (Rimm et al., 2004). Figure 1 shows in an overview the present status of knowledge based on the current available data. Regarding ovarian stimulation cycles and intrauterine insemination, the question of an increased risk of major malformations cannot be answered yet. There are, of course, many studies that also include data on major malformation rates; however, the data cohorts are almost always small, or there is no clear definition of major malformations, the manner of data collection is not clear, or, most important, the manner of data collection regarding major malformations is not clearly defined (Hack et al., 1970, 1972; Caspi et al., 1976; Adashi et al., 1979; Barrat and Leger, 1979). One of the most important limitations of studies looking for major malformation rates is the missing control cohort, the use of historical control cohorts with unclear definitions, differences in the means of data collection in the control and study cohort, or problems in the definitions of the term ‘major malformation’ (Kurinczuk and Bower, 1997; Ludwig, 2004). However, the question is still open as to the origin of this increased risk.
Table 2. Analysis of sub-cohorts in a meta-analysis on the risk of major malformations after IVF and ICSI. Data according to Hansen et al. (2005).
Major birth defects All infants (singletons and multiples) Singletons only IVF only ICSI only
No. studies
Pooled OR
95% CI
15 17 15 12 5
1.32 1.36 1.31 1.94 1.28
1.20–1.45 1.28–1.45 1.17–1.46 1.50–2.50 1.14–1.43
ICSI = intracytoplasmic sperm injection; OR = odds ratio; CI = confidence interval.
Figure 1. Risk of major malformations after ovarian stimulation and different assisted reproduction treatments as compared with spontaneously conceived children from fertile couples. For most differences no valid data are present, shown by the question mark. Only for the comparison of malformation rates in IVF and intracytoplasmic sperm injection (ICSI) conceived pregnancies to those spontaneously conceived in fertile couples an increased risk is well documented.
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What is the cause of the increased risk of major malformations? At different steps during the treatment cycle for assisted reproduction, factors may be involved that lead to an increased risk in major malformations (Figure 2). Principally, it is difficult to compare couples who become pregnant following IVF or ICSI with those who conceive spontaneously. There are many differences, e.g. patient’s age or genetic factors (Buck Louis et al., 2005). In several studies, an increased risk for chromosomal abnormalities could be shown for those couples who underwent ICSI treatment. This held true not only for the male, but also for the female partner in these relationships (Meschede et al., 1998; Scholtes et al., 1998; Peschka et al., 1999). This, per se, might be a relevant risk factor. Furthermore, it is well known that spermatozoa show a higher rate of chromosomal abnormalities in men suffering from oligoasthenoteratozoospermia (OAT syndrome), even without constitutional chromosomal abnormalities (Egozcue et al., 1997). Other genetic factors may be related to the problem of cystic fibrosis transmembrane regulator mutations (Chillon et al., 1995), an increase in trinucleotide repeats of the androgen receptor gene (Dowsing et al., 1999), or all the unknown genetic factors that might influence male and female fertility. It is important to note that the invasiveness of the assisted reproduction procedure chosen in the individual case of infertility problems does not seem to increase the risk of major malformations. There is no difference between ICSI or IVF (Bonduelle et al., 2002; Hansen et al., 2002, 2004; Lie et al., 2004; Rimm et al., 2004). This is in contrast to all the suggestions that were published some years ago (Ludwig et al., 2001; Schröder et al., 2001), as well as the well documented abnormalities in the fertilization process after ICSI as compared with that in IVF in rhesus monkeys (Hewitson et al., 1999): apparently, the difference in the fertilization process and the ‘choreography of fertilization’ (Hewitson et al., 1999) does not matter for the development of the embryo and the health of children born.
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Theoretically, each manipulation of embryos, e.g. the in-vitro culture process per se, assisted hatching, polar body biopsy or blastomere biopsy, might have an influence on its constitution and influence the health outcome (Ludwig et al., 2001; Schröder et al., 2001; Cetin et al., 2003). The data on these techniques have never shown such influence; however, the numbers are still very small and systematic analysis on children’s health is often missing. It had taken several years of prospective comparative and retrospective controlled studies to evaluate the risk of major malformations in IVF and ICSI treatment. It is therefore important to establish multinational registries to collect data of these children, as has already been done so thoroughly for preimplantation genetic diagnosis (Sermon et al., 2005). Finally, it is possible, that the temperature change during embryo transfer, which cannot be avoided since it is done at room temperature with a very small medium droplet, might influence epigenetic factors in the human embryo, as was shown several decades ago for the Drosophila embryo (Edwards and Ludwig, 2003). Quality management in the IVF laboratory should therefore include, as far as possible, the control of culture conditions and environmental factors to ensure maximal stability for the embryo (Kastrop, 2003; Mayer et al., 2003).
Increase of diseases due to imprinting errors: a special form of malformations Recently, case reports and case–control studies have been published, which show an increased risk of certain syndromes, which are due to imprinting errors. These are cases of Angelman syndrome (Cox et al., 2002; Orstavik et al., 2003; Ludwig et al., 2005) and Beckwith–Wiedeman syndrome (Debaun et al., 2003; Maher et al., 2003; Chang et al., 2005). Initial case reports that have suggested this association are now supported by large case–control studies. A recent study from Australia demonstrated a 17.8-fold increased risk for Beckwith–Wiedeman syndrome (Halliday et al., 2004). However, even with this multifold increase, the absolute risk still remains small, since these disorders are very rare.
Figure 2. Possible influences during different steps of the treatment course in ovarian stimulation and assisted reproduction treatments.
Risk of malformations after assisted reproduction - M Ludwig
Why is assisted reproduction associated with an increased risk of imprinting errors? Up to now, no studies have been published in humans showing a direct relationship between ovarian stimulation procedures and imprinting errors. However, there are several observations on abnormalities that may be related to the ovarian stimulation procedure (Tanbo et al., 1995; Ertzeid and Storeng, 2001; Keizer et al., 2004; Mitwally et al., 2004), the in-vitro culture conditions (Khosla et al., 2001), or infertility per se. Infertility may be associated with a certain genetic disposition to increase the transmittance of imprint errors to the offspring. One group has suggested the association of the severity of male factor subfertility with imprinting errors (Marques et al., 2004). This, however, has still to be confirmed by others.
Subfertility per se as a risk factor for major malformations Common points There is one drawback to all studies done in this field related to complications in pregnancy and children born after assisted reproduction: even with adjustment of study groups regarding age, previous pregnancies and genetic disorders, there remains one major difference: one group is subfertile, the others are not (Buck Louis et al., 2005). Therefore, the optimal control group to study complications in pregnancy and following birth after assisted reproduction would be a group of either spontaneously conceived pregnancies in subfertile couples, especially those who have had IVF or ICSI cycles in the past, or treatment of fertile couples with intrauterine insemination, IVF or ICSI. The latter is for obvious ethical reasons not possible.
Subfertility per se as a risk factor for pregnancy complications Before the coincidence of subfertility and major malformations are discussed, it is useful to highlight the coincidence of subfertility and pregnancy complications. Even if these are two different things, they are both consequences of infertility treatment regimens and subfertility. Some studies have investigated whether subfertile couples who had conceived spontaneously had the same problems in pregnancy and after birth as those who had undergone treatment. In a large cohort of patients with idiopathic infertility, a group from the UK showed a similar increased risk for abruptio placentae, pre-eclampsia and Caesarean section as compared with fertile couples, independently of whether the subfertile couples conceived spontaneously or after any infertility treatment (Pandian et al., 2001). In 1991, a study was published on children born in Boston (USA) to subfertile or fertile women. Subfertility was defined as 1 year or more to conception (Williams et al., 1991). The authors saw an increased risk for a low birth weight and intrauterine growth retardation [relative risk (RR) 2.3, 95% CI 1.2–4.4] (Williams et al., 1991). Similar data regarding premature birth have been published from Denmark in two cohorts with more than 8000 and nearly 4000 singleton pregnancies
(Henriksen et al., 1997). The risk for a premature born singleton was significantly increased by 1.6- to 1.8-fold after more than 12 months time to conception. The risk for preeclampsia was also related to the time to conception, with an increased RR of 1.62 (95% CI 1.14–2.30) after 12 months time to pregnancy (Basso et al., 2003). All these risks were present without any infertility treatment.
Subfertility per se as a risk factor for major malformations Regarding the risk of major malformations in spontaneously conceived pregnancies in subfertile couples, only one study has been published so far (Ghazi et al., 1991). Ghazi et al. analysed a cohort of 379,779 females with 384,589 children in Sweden in 1983–1986. In all, 7.8% showed subfertility, defined by a time to pregnancy of more than 12 months (n = 29,821). In this sub-cohort, 24% had had treatment with clomiphene citrate and the risk for multiple pregnancies was increased [odds ratio 1.2 (95% CI 1.1–1.2)]. No IVF or ICSI treatment and no intrauterine insemination was performed. However, despite that, the risk for premature birth in singleton pregnancies was increased from 5.4 to 7.1% (P < 0.001). The risk of low birth weight showed a strong relationship to the duration of time to pregnancy (Table 3). Similarly, the risk of major malformations was increased depending on the time to pregnancy (Table 3). The risk was increased after at least 5 years time to conception as compared with those with 1, 2, 3, or 4 years [odds ratio 1.18 (95% CI 1.01–1.37)] and after 4 years as compared with those with 1–3 years [odds ratio 1.17 (95% CI 1.03–1.33)] (Ghazi et al., 1991). Therefore, the theory of a link between subfertility itself and the risks in pregnancy and at birth is quite convincing, due to a growing body of evidence from well designed retrospective cohort studies.
Table 3. Risk of premature birth (a) and major malformations (b) depending on the time to pregnancy (Ghazi et al., 1991). Time to pregnancy (years)
Odds ratio
95% Confidence interval
(a) 1 2 3 4 ≥5
1.03 1.13 1.16 1.29 1.60
1.01–1.26 1.01–1.26 1.00–1.35 1.07–1.55 1.44–1.77
(b) 1 2 3 4 ≥5 1–3 ≥4
0.97 1.00 1.01 1.14 1.18 0.99 1.17
0.85–1.10 0.87–1.16 0.83–1.24 0.90–1.45 1.01–1.37 0.90–1.08 1.03–1.33
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Conclusion A lot of questions regarding the risk of major malformations are still unanswered. There is enough evidence for an increased risk of major malformations following conventional IVF as well as after ICSI. There is also some evidence that there is no difference between ICSI and other IVF techniques (Figure 1). However, the question is open for discussion as to why the risk is increased at all, even after matching for important variables. It seems that not only are the in-vitro culture conditions responsible, but that infertility per se contributes to this risk, perhaps through genetic factors. Further studies are needed, especially to investigate the risk of major malformations in children born after ovarian stimulation only, to evaluate the contribution of ovarian stimulation and, in children born after intrauterine insemination, to evaluate the contribution of in-vitro culture conditions. Those couples who conceived spontaneously after more than 12 months of time-to-pregnancy must now be counselled about an increased risk of major malformations.
References
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Scholtes MC Behrend C, Dietzel-Dahmen J et al. 1998 Chromosomal aberrations in couples undergoing intracytoplasmic sperm injection: influence on implantation and ongoing pregnancy rates. Fertility and Sterility 70, 933–937. Schröder AK, Diedrich K, Ludwig M 2001 Fertilization and preimplantation development after intracytoplasmic sperm injection. Reproductive BioMedicine Online 3, 247. Sermon K, Moutou C, Harper J et al. 2005 ESHRE PGD Consortium data collection IV: May–December 2001. Human Reproduction 20, 19–34. Sorensen HT, Pedersen L, Skriver MV et al. 2005 Use of clomifene during early pregnancy and risk of hypospadias: population based case-control study. British Medical Journal 330, 126–127. Tanbo T, Dale PO, Lunde O et al. 1995 Obstetric outcome in singleton pregnancies after assisted reproduction. Obstetrics and Gynecology 86, 188–192. Wennerholm UB, Bergh C 2004 What is the most relevant standard of success in assisted reproduction? Singleton live births should also include preterm births. Human Reproduction 19, 1943–1945. Williams MA, Goldman MB, Mittendorf R, Monson RR 1991 Subfertility and the risk of low birth weight. Fertility and Sterility 56, 668–671.
Received 21 February 2005; refereed 24 March 2005; accepted 18 April 2005.
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Is there an increased risk of malformations after assisted reproductive technologies? Dr Michael Ludwig was born in 1968 in Germany and concluded university in 1994. He then became Gynaecologist and Obstetrician at the University Hospital Lübeck and ran the Division of Reproductive Medicine and Gynaecologic Endocrinology at the Department of Gynaecology and Obstetrics. Since 2003 he has worked at the Endokrinologikum Hamburg, a private centre for hormone and metabolic diseases, gynaecological endocrinology and reproductive medicine and has recently been promoted to Head of the Division of Gynecologic Endocrinology and Reproductive Medicine. Dr Ludwig has published extensively in different fields of reproductive medicine, especially assisted reproductive techniques. His special interests lie in the field of evidence-based reproductive medicine and epidemiological studies in assisted reproductive treatment. Dr Michael Ludwig Michael Ludwig Endokrinologikum Hamburg, Zentrum für Hormon- und Stoffwechselerkrankungen, Reproduktionsmedizin und Gynäkologische Endokrinologie, Lornsenstraße 6, 22767 Hamburg, Germany Correspondence: Tel: +49 40 30628320; Fax: +49 40 30628349; e-mail: [email protected]
Abstract The question as to whether there is an increased risk of major malformations after assisted reproductive technologies has been answered insofar that it is slightly increased (30% relative risk when absolute figures analysed). This means an increased risk of 1:15 to 1:12 pregnancies when naturally conceived and intracytoplasmic sperm injection (ICSI) pregnancies are compared. Furthermore, there seems to be no risk difference between more invasive techniques (ICSI) or less invasive ones (conventional IVF). However, there are as yet no sufficiently robust data to describe the risk of children born either after ovarian stimulation only or intrauterine insemination procedures. With these data, it would be possible to clarify whether the increased risk after IVF and ICSI is due to the in-vitro culture techniques or to the ovarian stimulation procedure, or whether infertility per se is a risk factor. The latter seems to be suggested by retrospective studies. The different theories behind the increased risk of major malformations and the current database worldwide are described in this review. Keywords: ICSI, IVF, malformation, risk
Introduction The birth of a healthy singleton is increasingly becoming the major and most important outcome parameter after assisted reproductive technologies (Wennerholm and Bergh, 2004). Data have been collected in recent years to study whether or not there is an increased risk, especially for pregnancy complications or major malformations in the children born (Ludwig and Diedrich, 2002). Firstly, it is not possible to give an answer regarding this risk or the risk for pregnancy and birth complications for all types of assisted reproduction. Only in IVF and intracytoplasmic sperm injection (ICSI) have enough valid data been collected in recent years to give a true estimation. Currently, for intrauterine insemination cycles there are insufficient data to estimate the risks. Regarding ovarian stimulation cycles using either gonadotrophins or clomiphene citrate, insufficient data can be found in the literature. Some studies, however, have concentrated on particular abnormalities, e.g. hypospadias (Sorensen et al., 2005), but did not look for the overall risk. Furthermore, certain limitations in study design will make it quite impossible, even in the future, to give a true estimation of the
influence of assisted reproduction and the underlying impairment of fertility in treated couples on the health outcome of children (Buck Louis et al., 2005). Regarding pregnancy complications, a recent meta-analysis and systematic review has proved that children born after IVF with or without ICSI have an increased risk (Helmerhorst et al., 2004; Jackson et al., 2004). The most important results are shown in Table 1. The present paper will concentrate on whether major malformations are also increased, and what the cause of such an increase might be.
Is there an increased risk of major malformations? The question as to whether there is an increased risk of major malformations in assisted reproduction can be answered easily, since two recent meta-analyses have already collected all data published on that topic up to March 2003 (Hansen et al., 2005) and September 2003 (Rimm et al., 2004) respectively. Hansen et al. (2005) showed, in the meta-analysis with 25
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Risk of malformations after assisted reproduction - M Ludwig
Table 1. Results of a systematic review and a meta-analysis on the outcome of singleton pregnancies after assisted reproduction treatments. Data according to Helmerhorst et al. (2004) and Jackson et al. (2004). Jackson et al. (2004)
Helmerhorst et al. (2004)
Publication period analysed
1978–2002
1985–2002
Inclusion criteria
– Comparison of IVF to spontaneous conception – More than 50% in the IVF group had received standard IVF (defined as ovulation induction–egg retrieval–IVF–intrauterine embryo transfer), (non-standard: GIFT, ICSI, frozen embryo transfers, use of donor embryos) – More than 50% in the control group were fertile – Control for at least maternal age and parity – Singleton gestations reported and analysed separately from multiple gestations – Outcomes explicitly defined – Risk ratios with 95% confidence intervals provided or sufficient data to enable calculation
– Categorical data present on any of the following outcomes (gestational age and weight at birth, Caesarean section, perinatal death, and admission to neonatal intensive care) – Studies without a control group of natural conceptions or that did not distinguish singleton from multiple pregnancies were excluded
Study cohort
12,283 singletons
5361 singletons
Control cohort
1.9 million
7038
Risk of perinatal mortality
OR 2.2 (95% CI 1.6–3.0)
RR 1.68 (95% CI 1.11–2.55)
Risk of preterm delivery (<37 weeks)
OR 2.0 (95% CI 1.7–2.2)
RR 2.04 (95% CI 1.80–2.32)
Risk of very preterm delivery (<32 weeks)
n/a
RR 3.27 (95% CI 2.03–5.28)
Risk of low birth weight (<2.500 g)
OR 1.8 (95% CI 1.4–2.2)
RR 1.40 (95% CI 1.15–1.71)
Risk of very low birth weight (<1.500 g)
OR 2.7 (95% CI 2.3–3.1) RR 3.00 (95% CI 2.07–4.36)
Risk of small for gestational age
OR 1.6 (95% CI 1.3–2.0)
RR 1.40 (95% CI 1.15–1.71)
Risk of Caesarean section
n/a
RR 1.54 (95% CI 1.44–1.66)
Risk of admission to neonatal intensive care unit
n/a
RR 1.27 (95% CI 1.16–1.40)
GIFT = gamete intra-Fallopian transfer; ICSI = intracytoplasmic sperm injection; OR = odds ratio; RR = relative risk; CI = confidence interval; n/a = not available.
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studies, an overall increase in major malformations of about 30%. Details of sub-analysis are given in Table 2. Interestingly, 18 studies originated from Europe, and half of the studies were published in the last 4 years of the time period analysed. Five of the studies analysed concentrated on ICSI only (Hansen et al., 2005). Lie et al., in a recent metaanalysis, compared data on major malformation risk in children born after ICSI with those born after other IVF techniques. Only four studies were identified that were performed prospectively, did not overlap with other studies and provided reliable and comparable data on birth defects for children either conceived after ICSI or another IVF technique. These studies included data on 5395 children born after ICSI and 13,086 children born after other IVF techniques. The data gave a relative risk for the presence of major malformations after ICSI of 1.2 [95% confidence interval (CI) 0.97–1.28] (Lie et al., 2004). All these findings were confirmed by an independent metaanalysis performed by Rimm et al. that described an overall odds ratio of 1.29 [95% CI 1.01–1.67] for children born after IVF or ICSI as compared with spontaneously conceived children. The risk for singletons born after ICSI [odds ratio 1.33 (95% CI 0.90–1.95)] and IVF [odds ratio 1.51 (95% CI 0.85–2.7)] was increased compared with spontaneously conceived children, but this difference was not significant.
No significant difference was found for the risk of children born after IVF or ICSI, when these groups were compared directly with each other (Rimm et al., 2004). Figure 1 shows in an overview the present status of knowledge based on the current available data. Regarding ovarian stimulation cycles and intrauterine insemination, the question of an increased risk of major malformations cannot be answered yet. There are, of course, many studies that also include data on major malformation rates; however, the data cohorts are almost always small, or there is no clear definition of major malformations, the manner of data collection is not clear, or, most important, the manner of data collection regarding major malformations is not clearly defined (Hack et al., 1970, 1972; Caspi et al., 1976; Adashi et al., 1979; Barrat and Leger, 1979). One of the most important limitations of studies looking for major malformation rates is the missing control cohort, the use of historical control cohorts with unclear definitions, differences in the means of data collection in the control and study cohort, or problems in the definitions of the term ‘major malformation’ (Kurinczuk and Bower, 1997; Ludwig, 2004). However, the question is still open as to the origin of this increased risk.
Table 2. Analysis of sub-cohorts in a meta-analysis on the risk of major malformations after IVF and ICSI. Data according to Hansen et al. (2005).
Major birth defects All infants (singletons and multiples) Singletons only IVF only ICSI only
No. studies
Pooled OR
95% CI
15 17 15 12 5
1.32 1.36 1.31 1.94 1.28
1.20–1.45 1.28–1.45 1.17–1.46 1.50–2.50 1.14–1.43
ICSI = intracytoplasmic sperm injection; OR = odds ratio; CI = confidence interval.
Figure 1. Risk of major malformations after ovarian stimulation and different assisted reproduction treatments as compared with spontaneously conceived children from fertile couples. For most differences no valid data are present, shown by the question mark. Only for the comparison of malformation rates in IVF and intracytoplasmic sperm injection (ICSI) conceived pregnancies to those spontaneously conceived in fertile couples an increased risk is well documented.
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Risk of malformations after assisted reproduction - M Ludwig
What is the cause of the increased risk of major malformations? At different steps during the treatment cycle for assisted reproduction, factors may be involved that lead to an increased risk in major malformations (Figure 2). Principally, it is difficult to compare couples who become pregnant following IVF or ICSI with those who conceive spontaneously. There are many differences, e.g. patient’s age or genetic factors (Buck Louis et al., 2005). In several studies, an increased risk for chromosomal abnormalities could be shown for those couples who underwent ICSI treatment. This held true not only for the male, but also for the female partner in these relationships (Meschede et al., 1998; Scholtes et al., 1998; Peschka et al., 1999). This, per se, might be a relevant risk factor. Furthermore, it is well known that spermatozoa show a higher rate of chromosomal abnormalities in men suffering from oligoasthenoteratozoospermia (OAT syndrome), even without constitutional chromosomal abnormalities (Egozcue et al., 1997). Other genetic factors may be related to the problem of cystic fibrosis transmembrane regulator mutations (Chillon et al., 1995), an increase in trinucleotide repeats of the androgen receptor gene (Dowsing et al., 1999), or all the unknown genetic factors that might influence male and female fertility. It is important to note that the invasiveness of the assisted reproduction procedure chosen in the individual case of infertility problems does not seem to increase the risk of major malformations. There is no difference between ICSI or IVF (Bonduelle et al., 2002; Hansen et al., 2002, 2004; Lie et al., 2004; Rimm et al., 2004). This is in contrast to all the suggestions that were published some years ago (Ludwig et al., 2001; Schröder et al., 2001), as well as the well documented abnormalities in the fertilization process after ICSI as compared with that in IVF in rhesus monkeys (Hewitson et al., 1999): apparently, the difference in the fertilization process and the ‘choreography of fertilization’ (Hewitson et al., 1999) does not matter for the development of the embryo and the health of children born.
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Theoretically, each manipulation of embryos, e.g. the in-vitro culture process per se, assisted hatching, polar body biopsy or blastomere biopsy, might have an influence on its constitution and influence the health outcome (Ludwig et al., 2001; Schröder et al., 2001; Cetin et al., 2003). The data on these techniques have never shown such influence; however, the numbers are still very small and systematic analysis on children’s health is often missing. It had taken several years of prospective comparative and retrospective controlled studies to evaluate the risk of major malformations in IVF and ICSI treatment. It is therefore important to establish multinational registries to collect data of these children, as has already been done so thoroughly for preimplantation genetic diagnosis (Sermon et al., 2005). Finally, it is possible, that the temperature change during embryo transfer, which cannot be avoided since it is done at room temperature with a very small medium droplet, might influence epigenetic factors in the human embryo, as was shown several decades ago for the Drosophila embryo (Edwards and Ludwig, 2003). Quality management in the IVF laboratory should therefore include, as far as possible, the control of culture conditions and environmental factors to ensure maximal stability for the embryo (Kastrop, 2003; Mayer et al., 2003).
Increase of diseases due to imprinting errors: a special form of malformations Recently, case reports and case–control studies have been published, which show an increased risk of certain syndromes, which are due to imprinting errors. These are cases of Angelman syndrome (Cox et al., 2002; Orstavik et al., 2003; Ludwig et al., 2005) and Beckwith–Wiedeman syndrome (Debaun et al., 2003; Maher et al., 2003; Chang et al., 2005). Initial case reports that have suggested this association are now supported by large case–control studies. A recent study from Australia demonstrated a 17.8-fold increased risk for Beckwith–Wiedeman syndrome (Halliday et al., 2004). However, even with this multifold increase, the absolute risk still remains small, since these disorders are very rare.
Figure 2. Possible influences during different steps of the treatment course in ovarian stimulation and assisted reproduction treatments.
Risk of malformations after assisted reproduction - M Ludwig
Why is assisted reproduction associated with an increased risk of imprinting errors? Up to now, no studies have been published in humans showing a direct relationship between ovarian stimulation procedures and imprinting errors. However, there are several observations on abnormalities that may be related to the ovarian stimulation procedure (Tanbo et al., 1995; Ertzeid and Storeng, 2001; Keizer et al., 2004; Mitwally et al., 2004), the in-vitro culture conditions (Khosla et al., 2001), or infertility per se. Infertility may be associated with a certain genetic disposition to increase the transmittance of imprint errors to the offspring. One group has suggested the association of the severity of male factor subfertility with imprinting errors (Marques et al., 2004). This, however, has still to be confirmed by others.
Subfertility per se as a risk factor for major malformations Common points There is one drawback to all studies done in this field related to complications in pregnancy and children born after assisted reproduction: even with adjustment of study groups regarding age, previous pregnancies and genetic disorders, there remains one major difference: one group is subfertile, the others are not (Buck Louis et al., 2005). Therefore, the optimal control group to study complications in pregnancy and following birth after assisted reproduction would be a group of either spontaneously conceived pregnancies in subfertile couples, especially those who have had IVF or ICSI cycles in the past, or treatment of fertile couples with intrauterine insemination, IVF or ICSI. The latter is for obvious ethical reasons not possible.
Subfertility per se as a risk factor for pregnancy complications Before the coincidence of subfertility and major malformations are discussed, it is useful to highlight the coincidence of subfertility and pregnancy complications. Even if these are two different things, they are both consequences of infertility treatment regimens and subfertility. Some studies have investigated whether subfertile couples who had conceived spontaneously had the same problems in pregnancy and after birth as those who had undergone treatment. In a large cohort of patients with idiopathic infertility, a group from the UK showed a similar increased risk for abruptio placentae, pre-eclampsia and Caesarean section as compared with fertile couples, independently of whether the subfertile couples conceived spontaneously or after any infertility treatment (Pandian et al., 2001). In 1991, a study was published on children born in Boston (USA) to subfertile or fertile women. Subfertility was defined as 1 year or more to conception (Williams et al., 1991). The authors saw an increased risk for a low birth weight and intrauterine growth retardation [relative risk (RR) 2.3, 95% CI 1.2–4.4] (Williams et al., 1991). Similar data regarding premature birth have been published from Denmark in two cohorts with more than 8000 and nearly 4000 singleton pregnancies
(Henriksen et al., 1997). The risk for a premature born singleton was significantly increased by 1.6- to 1.8-fold after more than 12 months time to conception. The risk for preeclampsia was also related to the time to conception, with an increased RR of 1.62 (95% CI 1.14–2.30) after 12 months time to pregnancy (Basso et al., 2003). All these risks were present without any infertility treatment.
Subfertility per se as a risk factor for major malformations Regarding the risk of major malformations in spontaneously conceived pregnancies in subfertile couples, only one study has been published so far (Ghazi et al., 1991). Ghazi et al. analysed a cohort of 379,779 females with 384,589 children in Sweden in 1983–1986. In all, 7.8% showed subfertility, defined by a time to pregnancy of more than 12 months (n = 29,821). In this sub-cohort, 24% had had treatment with clomiphene citrate and the risk for multiple pregnancies was increased [odds ratio 1.2 (95% CI 1.1–1.2)]. No IVF or ICSI treatment and no intrauterine insemination was performed. However, despite that, the risk for premature birth in singleton pregnancies was increased from 5.4 to 7.1% (P < 0.001). The risk of low birth weight showed a strong relationship to the duration of time to pregnancy (Table 3). Similarly, the risk of major malformations was increased depending on the time to pregnancy (Table 3). The risk was increased after at least 5 years time to conception as compared with those with 1, 2, 3, or 4 years [odds ratio 1.18 (95% CI 1.01–1.37)] and after 4 years as compared with those with 1–3 years [odds ratio 1.17 (95% CI 1.03–1.33)] (Ghazi et al., 1991). Therefore, the theory of a link between subfertility itself and the risks in pregnancy and at birth is quite convincing, due to a growing body of evidence from well designed retrospective cohort studies.
Table 3. Risk of premature birth (a) and major malformations (b) depending on the time to pregnancy (Ghazi et al., 1991). Time to pregnancy (years)
Odds ratio
95% Confidence interval
(a) 1 2 3 4 ≥5
1.03 1.13 1.16 1.29 1.60
1.01–1.26 1.01–1.26 1.00–1.35 1.07–1.55 1.44–1.77
(b) 1 2 3 4 ≥5 1–3 ≥4
0.97 1.00 1.01 1.14 1.18 0.99 1.17
0.85–1.10 0.87–1.16 0.83–1.24 0.90–1.45 1.01–1.37 0.90–1.08 1.03–1.33
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Conclusion A lot of questions regarding the risk of major malformations are still unanswered. There is enough evidence for an increased risk of major malformations following conventional IVF as well as after ICSI. There is also some evidence that there is no difference between ICSI and other IVF techniques (Figure 1). However, the question is open for discussion as to why the risk is increased at all, even after matching for important variables. It seems that not only are the in-vitro culture conditions responsible, but that infertility per se contributes to this risk, perhaps through genetic factors. Further studies are needed, especially to investigate the risk of major malformations in children born after ovarian stimulation only, to evaluate the contribution of ovarian stimulation and, in children born after intrauterine insemination, to evaluate the contribution of in-vitro culture conditions. Those couples who conceived spontaneously after more than 12 months of time-to-pregnancy must now be counselled about an increased risk of major malformations.
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Received 21 February 2005; refereed 24 March 2005; accepted 18 April 2005.
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