Human Reproduction, Vol. 15, No. 5, 1189-1194,
May 2000
© 2000 European Society of Human Reproduction and Embryology
Obstetric outcome of pregnancies following ICSI, classified according to sperm origin and quality
1 Department of Obstetrics and Gynaecology, Sahlgrenska University Hospital, Göteborg and 2 Fertility Center Scandinavia, Carlanderska Hospital, Göteborg, Sweden
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Abstract |
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The aim of this study was to report the outcome of all clinical pregnancies obtained after intracytoplasmic sperm injection (ICSI) performed during a 5 year period at two fertility clinics, with special reference to delivery outcome associated with different sperm origin and quality and the transfer of fresh or frozen–thawed pre-embryos. A total of 1293 clinical pregnancies was analysed. Deliveries occurred in 75.9% (n = 982) and early spontaneous abortion, late spontaneous abortion and ectopic pregnancy in 21.4, 1.0 and 1.2% respectively. Multiple birth occurred in 21.3% (208 sets of twins and one set of triplets) of the deliveries, with the highest incidence in the epididymal sperm group (30.2%) and lowest in the cryopreserved group (13.7%). A total of 1192 infants was born. Preterm birth occurred in 15.7% of all deliveries. Preterm birth was not related to sperm origin or quality but was related to multiple birth. The prematurity rate was 8.4%, 42.3% and 100% for singletons, twins and triplets respectively. Singleton infants born after cryopreservation as embryos had a significantly higher birthweight than the ejaculated sperm group with fresh embryo transfer. The perinatal mortality rate was 11.7 per 1000 born infants. Eighty-seven of the 1192 infants (7.3%) had a malformation, 40 of which were minor. The perinatal mortality rate and the malformation rate were similar in the different subgroups. Prenatal karyotyping was performed on 149 fetuses (12.5%) and abnormal results were found in four cases (2.7%). In conclusion, obstetric outcome of ICSI pregnancies was similar to that of conventional IVF and was not influenced by sperm origin or quality. The high incidence of multiple births is still the major concern.
Key words: epididymal spermatozoa/ICSI/pregnancy outcome/sperm parameters/testicular spermatozoa
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Introduction |
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The development of intracytoplasmic sperm injection (ICSI) during the 1990s has dramatically improved the outlook for couples with male factor subfertility, often yielding pregnancy rates resembling those of conventional IVF (Tournaye et al., 1992
). Except for cases in which all spermatozoa are immobile, fertilization rates after ICSI have been shown to be independent of sperm quality, even with spermatozoa derived directly from the epididymis or testis (Nagy et al., 1995; Tournaye et al., 1995). Today, however, there is increasing evidence that different types of genetic abnormalities may account for a substantial part of the aetiological background to male subfertility. Sex chromosome aberrations, translocations, microdeletions in the Y chromosome and mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene are examples of male factors which cause azoospermia/oligozoospermia (Assche et al., 1996; Lissens et al., 1996; Girardi et al., 1997). It has also been known for a long time that the frequency of chromosome aberrations increases with decreasing sperm count (Kjessler, 1966); obviously, ICSI may increase the risk of transmitting these abnormalities to the offspring. Data have recently been published indicating an increased risk of chromosomal anomalies, predominantly affecting the sex chromosomes, after ICSI (Bonduelle et al., 1998).
In order to provide a reliable answer to the concerns expressed about the safety of ICSI, special effort must be made to follow up the pregnancies and the short-term and long-term health of children born after ICSI. It is also important to analyse the effects of sperm quality and origin on this outcome. There is some evidence that the use of severely defective spermatozoa in ICSI is an additional risk factor for an adverse perinatal outcome (Aytoz et al., 1998); however, this has yet to be confirmed in further studies.
In this study, we report the outcome of a 5 year cohort of pregnancies obtained after ICSI, classified according to sperm origin and quality. The main endpoint was delivery outcome and the secondary endpoint was early pregnancy loss.
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Material and methods |
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The outcome of all clinical pregnancies obtained after ICSI with embryo transfer between January 1993 and January 1998 at the two IVF clinics in Göteborg, Sahlgrenska University Hospital and Fertility Center Scandinavia, was analysed. Pregnancies were divided into six subgroups according to sperm origin, sperm quality and the replacement of fresh (groups 1–5) or frozen–thawed (group 6) pre-embryos as follows: group 1, ejaculated sperm concentration
5x106/ml or total number 15x106 and infertility due to a male factor or unexplained infertility; group 2, ejaculated sperm concentration 5x106/ml or total number 15x106 and infertility due to a female factor (tubal factor or endometriosis); group 3, ejaculated sperm concentration <5x106/ml or total number <15x106; group 4, epididymal spermatozoa; group 5, testicular spermatozoa; group 6, transfer of frozen–thawed pre-embryos derived from ICSI using ejaculated, epididymal or testicular sperm preparations.
The reasons for this particular group division were the following: group 1 included patients with mild/moderate oligozoospermia as the sole known infertility factor but could have included women with oocyte factors; group 2 resembled group 1 when it came to male factors but included cases with known female factors as well; group 3 included couples with severe oligozoospermia. The values of <15x106 or <5x106/ml spermatozoa are the lower limits for severe oligozoospermia, accepted internationally and by WHO (World Health Organization,1992). An increased incidence of chromosomal aberrations has been detected in men with severe oligozoospermia and non-obstructive azoospermia (Assche et al., 1996).
Standard oocyte retrieval, embryo culture and transfer procedures have been described previously (Wikland et al., 1983, 1994). The sperm preparation and micro-insemination procedures utilized by us have also been reported earlier (Hamberger et al., 1995). Epididymal spermatozoa were obtained by microsurgical epididymal sperm aspiration (MESA) (Temple Smith et al., 1985; Silber et al., 1994) or percutaneous sperm aspiration (PESA) (Craft and Shrivastav, 1994; Shrivastav et al., 1994; Westlander et al., 1999). Testicular spermatozoa were obtained by testicular sperm extraction (TESE) (Schoysman et al., 1993) or by fine-needle testicular sperm aspiration (TESA) (Tsirigotis and Craft, 1995).
For cryopreservation, we used the technique previously described (Lassalle et al., 1985), with embryos frozen at the 2–4-cell stage, corresponding to day 2 after oocyte retrieval.
In the epididymal sperm group (group 4), frozen–thawed spermatozoa were used in 16 pregnancies but due to the small number, this group was analysed together with pregnancies achieved with fresh epididymal spermatozoa having first established that there was no detectable difference between them. Only fresh spermatozoa were used in the other groups.
Details of treatment and fertility history were collected from the clinics' medical records and obstetric and perinatal histories were obtained from records at the obstetric departments where the patients were treated and/or gave birth. Data were obtained by telephone interview in 10 cases. The study was approved at the outset by the Ethics Committee in Göteborg.
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Definitions |
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Gestational age: In the fresh embryo transfer groups (1–5), gestational age was calculated from the day of oocyte retrieval which was converted into menstrual age by adding 14 days. In the cryopreserved group, the women were monitored with ultrasound and by measurements of LH in serum/urine. Frozen–thawed embryos were transferred 2 days after ovulation and 3 days after the LH surge. Gestational age was calculated from the day of embryo transfer, which was converted into menstrual age by adding 16 days.
A pregnancy was defined as clinical if a gestational sac(s) was visualized by ultrasonography in the seventh week of gestation. The diagnosis of an ectopic pregnancy was confirmed by laparoscopy. Early spontaneous abortion: pregnancy loss before 12 completed weeks of gestation. Late spontaneous abortion: pregnancy loss after 12 weeks and before 28 weeks of gestation. Stillborn: intrauterine death after 28 weeks of gestation. Delivery: the birth of a liveborn or stillborn infant. Early neonatal death: the death of a liveborn infant during the first 7 days after birth. Perinatal mortality: the sum of stillborn and early neonatal deaths per 1000 infants born. Preterm birth: delivery before 37 weeks of gestation. Very preterm birth: delivery before 32 weeks of gestation. Low birthweight (LBW): a liveborn or stillborn infant weighing <2500 g at birth. Very low birthweight (VLBW): a liveborn or stillborn infant weighing <1500 g at birth. Small for gestational age (SGA): birthweight <–2 SD, according to the Swedish reference tables for birthweight (Marsál et al., 1996). Malformation: any congenital malformation defined in the International Classification of Diseases (ICD, 1977, 1992; Chapter 14 in ICD-9 and Chapter 17 in ICD-10).
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Statistical methods |
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For comparisons between the groups, analysis of variance (ANOVA) and Kruskal–Wallis were used for continuous variables and
2 for comparisons between proportions. Paired comparisons between two groups were made only when ANOVA or Kruskal–Wallis showed a significant main effect. To control for type I error, the Bonferroni correction was used. Two-tailed tests were used. P < 0.05 was deemed statistically significant. Only statistically significant values are given in the tables. Odds ratio (OR) with 95% confidence interval (CI) was calculated using the approximation of Woolf (In Stat 2.01; GraphPad Software, San Diego, CA, USA). |
Results |
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A total of 1293 pregnancies resulted from ICSI performed at the two institutions in Göteborg during the 5 year period between January 1993 and January 1998. The number of pregnancies, classified by sperm origin and quality and the transfer of fresh of frozen–thawed pre-embryos, is shown in Table I
. Of the 1293 pregnancies, early spontaneous abortion occurred in 21.4%, late spontaneous abortion in 1.0%, ectopic pregnancy in 1.2% and delivery in 75.9% (n = 982). There was no statistically significant difference in the incidence of early or late spontaneous abortion or the incidence of ectopic pregnancy between the different subgroups (Table I). Four pregnancies were interrupted because of the presence of fetal abnormalities, which were detected by second trimester ultrasound scanning. These abnormalities included trisomy 18, polycystic kidney disease and acrania (two cases).
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Multiple birth occurred in 21.3% of the 982 deliveries (208 sets of twins and one set of triplets). A total of 1192 infants was born. Embryo reduction to twins was performed in three women with triplet pregnancies, and a reduction to singleton was performed in three triplet pregnancies. Maternal characteristics (age at delivery and parity), the number of multiple deliveries and the mean number of embryos transferred in the different subgroups are shown in Table II
. The incidence of multiple birth was highest in the epididymal sperm group 4 (30.2%) and lowest in the cryopreserved group 6 (13.7%); however, the difference was not statistically significant.
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Gestational age
Preterm birth and very preterm birth occurred in 15.7% (n = 154) and 3.5% (n = 34) respectively of the 982 deliveries. The median (range) gestational age for 773 singleton deliveries was 280 days (189–300 days) and 262 days (173–288 days) for 208 twin deliveries. Preterm birth and very preterm birth rates for singletons were 8.4% (n = 65) and 1.7% (n = 13) respectively and for twins 42.3% (n = 88) and 10.1% (n = 21) respectively. The only set of triplets was delivered at a gestational age of 232 days. The OR for preterm delivery in twins compared to singletons was 8.0 (95% CI, 5.5–11.6). Gestational age for singletons and twins divided into the different subgroups is presented in Table III
. As expected, the prematurity rate was not influenced by the origin or quality of spermatozoa but was related to multiple birth. Preterm birth occurred less frequently, both in the case of singletons and twins, in the cryopreserved embryo group compared to the fresh embryo transfer groups, but the difference was not statistically significant.
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Birthweight
LBW and VLBW occurred in 18.9% (n = 226) and 3.9% (n = 47) of the 1192 infants. The median (range) birthweight for the 773 singletons was 3470 g (795–5060 g) and 2640 g (370–4010 g) for the 416 twins. LBW and VLBW occurred in 7.6% (n = 59) and 1.4% (n = 11) respectively in singletons and in 39.4% (n = 164) and 8.4% (n = 35) respectively in twins. The OR for LBW in twins compared to singletons was 7.9 (95% CI, 5.7–11.0). The frequency of SGA was 6.5% (n = 50) and 17.1% (n = 71) in singletons and twins respectively. The triplets weighed 1680, 1665 and 1440 g respectively. The incidence of LBW, VLBW and SGA was comparable in the subgroups (Table IV
). The cryopreserved embryo singleton group had a significantly higher median birthweight than the ejaculated sperm groups (1, 2 and 3) with fresh embryo transfer (P = 0.0007, 0.0004 and 0.0026 respectively).
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Perinatal mortality
The perinatal mortality rate is presented in Table V
. Fourteen infants (11.7 per 1000 infants born) died during the perinatal period, including six intrauterine deaths and eight cases of early neonatal deaths. Among singletons, three intrauterine deaths [one fetus with trisomy 13, one SGA (<–3 SD) and one unexplained intrauterine death at 40 weeks of gestation] and three cases of early neonatal death (two infants delivered preterm at 27 and 30 weeks gestation respectively, and one multi-malformed infant, possibly trisomy 13) were reported. Among twins, three intrauterine deaths (two unexplained at week 28 and 32 respectively, and one caused by placental abruption in week 35) and five early neonatal deaths [two infants delivered preterm at 24 and 31 weeks gestation respectively, and three infants with malformations (diaphragmatic hernia, arthrogryphosis and unbalanced translocation)] occurred. The perinatal mortality rate was 7.8 and 19.2 per 1000 infants born, for singletons and twins respectively. All triplets survived. Perinatal mortality was comparable in the different subgroups (Table V).
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Malformations
Eighty-seven of the 1192 infants (7.3%) had a malformation, 40 of which were classified as minor. No difference was seen between the subgroups (Table VI
). The malformations have been described in detail previously (Wennerholm et al., 2000).
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Karyotypes
As a routine all women with singleton pregnancies were offered an early amniocentesis for karyotyping, but it was not compulsory. Because of the small risk associated with the procedure and for ethical reasons, prenatal karyotyping was only performed on 149 fetuses (12.5%) (148 amniocenteses and one cordocentesis). Abnormal results were found in four cases (three singletons and one twin) (2.7%). One singleton was a trisomy 18 and the parents chose to terminate the pregnancy in this case. Two were familial balanced structural anomalies inherited from the father. One twin had an unbalanced translocation with partial monosomy of chromosome 12 with associated multiple malformations and early neonatal death. In addition, three singletons were born with clinical signs of chromosome aberrations. Postnatal karyotypes were obtained in two of the infants and either trisomy 13 or 21 were verified. The third infant was stillborn with clinical signs of trisomy 13 but this was never verified. No sex chromosome aberrations were found.
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Discussion |
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The results of this report analysing pregnancy outcome of children born after ICSI, classified according to sperm origin and quality, are reassuring. The obstetric outcome for singletons and twins were satisfactory and similar in all groups and were also in accordance with earlier studies analysing obstetric outcome after IVF and ICSI (Bonduelle et al., 1995
, 1996; Palermo et al., 1996; Wennerholm et al., 1996; Wisanto et al., 1996; Bergh et al., 1999; Loft et al., 1999). A significantly higher median birthweight was noted for singleton infants born after ICSI with transfer of cryopreserved–thawed pre-embryos, compared to infants born after ICSI with ejaculated spermatozoa and fresh embryo transfer. The outcome in the cryopreserved group was not stratified further according to sperm origin and quality due to the limited number of patients included. However, a similar trend towards a higher birthweight after cryopreservation has previously been observed in connection with conventional IVF (Wennerholm et al., 1997; Bergh et al., 1999). Neither early nor late pregnancy loss reached unexpectedly high levels in the whole material or in any subgroup. In a recent publication (Aytoz et al., 1998), where pregnancy outcome after ICSI was also analysed according to sperm origin and quality, certain differences were found in the rate of very low birthweight and perinatal deaths between the groups, but it was concluded that the major problems were due to the high rate of multiple gestations.
Analysis of the malformation and chromosomal aberration rates after different kinds of assisted reproduction technologies is, no doubt, crucial. Concerns have been raised regarding the possibility of an increased rate of malformations and chromosomal aberrations after ICSI, based on the fact that the ICSI technique itself is invasive and makes it possible for spermatozoa of inferior quality to fertilize oocytes. In addition, it is well known that men with severe oligozoospermia and non-obstructive azoospermia carry a higher rate of chromosomal aberrations that might be transmitted to the offspring (Lundin et al., 1998). In the present study, a total of 87/1192 infants (7.3%) with malformations were recorded, of which 40/1192 (3.4%) were classified as minor and 47/1192 (3.9%) as relatively serious malformations. The malformations were distributed evenly, with no accumulation in any particular subgroup.
The rate of malformations might appear slightly higher than registry data from the general population, in which the reported rate is often 3–4% (Bergh et al., 1999). However, it must be emphasized that the data in this study have been analysed by scrutinizing almost all individual records for the 1192 infants. In fact, in a recent report (Wennerholm et al., 2000) it was found that 4/47 (8.5%) serious malformations were not reported to the Medical Birth Registry or the Registry of Congenital Malformations in Sweden. Therefore, for proper comparison, data have to be collected in the same way and strictly identified control groups are preferable. Even if the malformation rate seems reassuring, one has to remember that the power to detect an increase in malformations in this study is rather low. With 80% power and a significance level of 5%, it would only be possible to detect an almost doubled malformation rate, i.e. an increase from 3.5 to 6.0%, in the total ICSI population of 1192 infants. In the severe oligozoospermia subgroup (220 infants), the power to detect a doubled rate, i.e. an increase from 3 to 6%, is only 33%. In previous studies, available data on major birth defects in children born after ICSI, using mainly ejaculated spermatozoa, has shown that there are no differences between these children and children born after conventional IVF or spontaneous conception (Bonduelle et al., 1995; Govaerts et al., 1998; Tarlatzis and Bili, 1998). Recently, however, data from the French national registry (FIVNAT) were reported showing a significant increase in the malformation rate (including chromosomal anomalies) compared with conventional IVF (Rossin-Amar et al., 1999).
Chromosomal aberrations were found in four of 149 prenatal karyotypes and in two postnatal karyotypes in this study. No sex chromosome aberrations were detected. This might be of interest since earlier studies have indicated a possible increased rate of sex chromosome anomalies in children born after ICSI (In't Veld et al., 1995; Bonduelle et al., 1998).
In summary, the results from this study concerning the obstetric outcome of children born after ICSI, classified according to sperm origin and quality, showed that the outcome was similar to other published series for ICSI and conventional IVF. The major problem seems to be the high rate of twin pregnancies and neither the ICSI technique per se nor sperm origin or quality seemed to cause a substantial increase in adverse outcome.
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Acknowledgments |
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This study was supported by grants from the Göteborg Medical Society, the Regional Authority of West Sweden, the `Ordenssällskapet W:6' Society and the Swedish Medical Research Council (no. 2893). We would like to thank the midwives and technicians at the IVF clinics at the Fertility Center, Scandinavia and Sahlgrenska University Hospital (in alphabetical order) for their help and co-operation: K.Borg, A.-S.Forsberg, H.Holter, E.Nilsson, E.Reismer and E.Samuelsson.
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Notes |
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3 To whom correspondence should be addressed at: Institute for Women's and Children's Health, Department of Obstetrics and Gynaecology, Sahlgrenska University Hospital, Östra, 416 85 Göteborg, Sweden
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References |
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Submitted on November 11, 1999; accepted on February 14, 2000.
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U. Ulug, E. A. Jozwiak, A. Mesut, M. M. Berksoy, and M. Bahceci Survival rates during the first trimester of multiple gestations achieved by ICSI: a report of 1448 consecutive multiples Hum. Reprod., February 1, 2004; 19(2): 360 - 364. [Abstract] [Full Text] [PDF]
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V. Vernaeve, M. Bonduelle, H. Tournaye, M. Camus, A. Van Steirteghem, and P. Devroey Pregnancy outcome and neonatal data of children born after ICSI using testicular sperm in obstructive and non-obstructive azoospermia Hum. Reprod., October 1, 2003; 18(10): 2093 - 2097. [Abstract] [Full Text] [PDF]
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M. Ludwig, A. Katalinic, and for the German ICSI Follow-Up Study Group Pregnancy course and health of children born after ICSI depending on parameters of male factor infertility Hum. Reprod., February 1, 2003; 18(2): 351 - 357. [Abstract] [Full Text] [PDF]
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M. Bonduelle, E. Van Assche, H. Joris, K. Keymolen, P. Devroey, A. Van Steirteghem, and I. Liebaers Prenatal testing in ICSI pregnancies: incidence of chromosomal anomalies in 1586 karyotypes and relation to sperm parameters Hum. Reprod., October 1, 2002; 17(10): 2600 - 2614. [Abstract] [Full Text] [PDF]
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I. Mateizel, G. Verheyen, E. Van Assche, H. Tournaye, I. Liebaers, and A. Van Steirteghem FISH analysis of chromosome X, Y and 18 abnormalities in testicular sperm from azoospermic patients Hum. Reprod., September 1, 2002; 17(9): 2249 - 2257. [Abstract] [Full Text] [PDF]
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M. Bonduelle, I. Liebaers, V. Deketelaere, M.-P. Derde, M. Camus, P. Devroey, and A. Van Steirteghem Neonatal data on a cohort of 2889 infants born after ICSI (1991-1999) and of 2995 infants born after IVF (1983-1999) Hum. Reprod., March 1, 2002; 17(3): 671 - 694. [Abstract] [Full Text] [PDF]
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S. Kelleher, S.M. Wishart, P.Y. Liu, L. Turner, I. Di Pierro, A.J. Conway, and D.J. Handelsman Long-term outcomes of elective human sperm cryostorage Hum. Reprod., December 1, 2001; 16(12): 2632 - 2639. [Abstract] [Full Text] [PDF]
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