Article Case report: Partial hydatidiform mole following the transfer of single frozen thawed embryo subsequent to ICSI

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1 RBMOnline - Vol 9. No Reproductive BioMedicine Online; on web 4 August 2004 Article Case report: Partial hydatidiform mole following the transfer of single frozen thawed embryo subsequent to ICSI Dr Ulun Ulug was born in He received his MD degree from Hacettepe University, Ankara, Turkey and completed the Obstetrics and Gynecology residency in Marmara University, Istanbul. He worked as a research fellow at GBMC, Baltimore, USA and Haemek Medical Centre, Israel. He joined the Bahceci Women Health Care Centre in He is married and already has a newborn boy. Dr Ulun Ulug Ulun Ulug 1, Nadir H Ciray 1, Pinar Tuzlali 2, Mustafa Bahçeci 1,3,4 1 Bahçeci Women Health Care Centre and German Hospital at Istanbul, Istanbul, Turkey; 2 Department of Pathology, Taksim State Hospital, Istanbul, Turkey; 3 Yeditepe University School of Medicine, Istanbul, Turkey 4 Correspondence: Azer Is Merkezi 44/17 Kat 5, Abdi Ipekci Cad, Nisantasi 80200, Istanbul, Turkey. Fax: ; mbahceci@superonline.com Abstract Hydatiform mole is a gestational trophoblastic disease characterized by the dominance of dispermic fertilization. Micromanipulation techniques in assisted reproduction technologies have enabled direct evaluation of the zygotes and the formation of pronuclei in the zygote. Intracytoplasmic sperm injection (ICSI) of oocytes ensures that only a single spermatozoon enters the ooplasma. This study reports a case of partial hydatiform mole following the transfer of day 3 frozen thawed embryo. ICSI was used as the assisted fertilization method because there was male factor infertility due to severe oligoasthenoteratozoospermia. Possible predisposing factors for partial hydatidiform mole after ICSI are also discussed. Keywords: frozen thawed embryo, ICSI, molar pregnancy, partial hydatidiform mole 442 Introduction The introduction of assisted reproduction technologies has given infertile patients the ability to conceive. Among the assisted reproduction techniques available to patients, intracytoplasmic sperm injection (ICSI) has the advantage of enabling the fertilization of retrieved oocytes despite the presence of coexisting severe male factor. Micromanipulation techniques have enabled direct evaluation of the zygotes and early stage embryos, including the formation of pronuclei in the zygote and the nuclear morphology of cleaved blastomeres. Hydatidiform mole is a trophoblastic disease associated with increased maternal age, and is more commonly seen in Far Eastern populations. Hydatidiform mole is subdivided into partial hydatidiform mole, which is characterized by the presence of fetal components, and complete hydatidiform mole, in which no fetal tissue is present. Both complete and partial hydatidiform mole are associated with the presence of two paternally derived genomes. Because many women now delay conception until later in life and subsequently require assisted reproduction techniques such as IVF to achieve a successful pregnancy, hydatidiform mole may become more common. However reports of hydatidiform mole following assisted reproduction remain scarce (Ibrahim et al., 1989; Woods et al., 2002). In contrast to classical insemination during IVF, ICSI theoretically prevents the insertion of multiple spermatozoa into the oocyte. Thus, use of ICSI may be a therapeutic option to minimize the incidence of triploidy (Pal et al., 1996). Of the reports of hydatidiform mole following IVF, the majority have described complete molar pregnancies, since occurrence of

2 partial molar pregnancy is reduced by mechanisms that decrease the chances of producing or transferring a triploid embryo. This study describes a case of partial hydatidiform mole following the transfer of a single frozen thawed ICSI embryo, and discusses the possible predisposing factors for partial hydatidiform mole after ICSI. Case report A 36-year-old woman was admitted to an assisted reproduction centre 6 years ago for primary infertility due to male factor. Her spouse had severe oligoasthenoteratozoospermia (OAT), with 100,000 spermatozoa/ml, 10% motility, and severe head defect. She had regular menses, and her baseline hormonal concentrations were within normal limits. Neither her nor her spouse s peripheral lymphocytes showed karyotype abnormalities. In 1999 and 2000, she underwent two cycles of ovarian stimulation, ICSI and embryo transfer in the centre (Table 1). Although the centre generally uses its own needles in performing ICSI, commercially available ICSI needles (Cook, Brisbane, Australia) were used during both cycles because of the severe head defect of her spouse s spermatozoa. During her first cycle she conceived a singleton pregnancy, but it aborted at 11 weeks. In both assisted fertilization cycles pronuclear morphologies were evaluated 17 h after ICSI. Three years after the second cycle, she underwent a frozen thawed embryo transfer cycle. An oestradiol valerate transdermal patch (8 mg; Estraderm, Novartis, Istanbul, Turkey) was utilized for endometrial preparation for embryo transfer. During the treatment period, the couple was reminded that they must completely abstain from intercourse. Among the four frozen embryos, one survived; it was scored as 7-cell, grade II (between 10 and 20% fragmentation) after thawing. Embryo transfer was performed under ultrasound guidance. The patient s serum β-human chorionic gonadotrophin (HCG) concentrations following embryo transfer are shown in Figure 1. Her first transvaginal ultrasonogram (TVUS) (Aplio 80; Toshiba, Japan), performed 30 days after embryo transfer, revealed a large gestational sac of transverse diameter 20 mm and without a yolk sac (Figure 2A). There were no enlarged cystic areas in either ovary. A second TVUS, performed 4 days later, revealed a gestational sac filling one-third of the intrauterine cavity, and a yolk sac was identified. At the posterior aspect of the gestational sac, a large hyperechogenic mass was protruding into the endometrial cavity (Figure 2B). At this time, 6.5 weeks of gestation, neither embryonic pole nor embryo heartbeat was detected. The patient did not have any dominant complaints, including hyperemesis, tremor or palpitation. Her vital signs were all within normal limits. Her last TVUS, performed 7 days later, showed a very large gestational sac with a yolk sac and embryonic pole. The posterior of the gestational sac was covered by an excessively large hyperechogenic area, presumably due to increased chorionic activity. No embryo heartbeat was detected by M-mode Doppler. A missed abortion was diagnosed, and termination of pregnancy was recommended. Dilatation and evacuation were performed under general anaesthesia. Pathological evaluation of the specimen revealed hydropic villi and increased trophoblastic activity, both indicative of partial hydatidiform mole (Figure 3). Karyotyping of the specimen demonstrated XXX triploidy in 50 metaphase plaques (Figure 4). The follow-up of the patient was unremarkable, and her serum β-hcg concentrations steadily decreased (Figure 1). Table 1. Breakdown of previous ICSI cycles. Cycle Protocol Type of Total MII 2PN Cleaved Assisted ET Pregnancy Outcome no. gonado- oocytes hatching trophins used retrieved 1 Long GnRH Highly purified Abortion agonist FSH 2 Short GnRH Highly purified embryos agonist FSH frozen Figure 1. Serum β-hcg concentrations following embryo transfer. 443

3 a b Figure 2. (a) Transvaginal ultrasound scan of gestational sac without yolk sac. (b) Large gestational sac with yolk sac and embryonic pole and excessively large hyperechogenic lesion. Figure 3. Microscopic appearance of the specimen of evacuated gestational product. Scale bar = 0.1 mm. 444

4 Figure 4. Karyotyping of evacuated gestational product. Discussion Complete and partial hydatidiform mole are two abnormal concepti that can be identified by their clinical, ultrasonographic, gross morphological, histological, and genetic characteristics (Szulman and Surti, 1978). Patients with partial moles usually do not present with clinical features characteristic of complete molar pregnancy. Partial hydatidiform mole is generally not diagnosed until after histological review of curettage specimens. Genetically, complete hydatidiform mole is a pregnancy abnormality due to a diploid conception that is generally androgenetic in origin (i.e. all 46 chromosomes are paternally derived) (Kajii and Ohama, 1977; Wake et al., 1978). Complete hydatidiform mole may be monospermic, arising from fertilization of an enucleated egg by a single spermatozoon, which then doubles to provide a diploid chromosome complement, or it may be dispermic, arising from fertilization of an enucleated egg by two spermatozoa (Ohama et al., 1981). In partial hydatidiform mole, there are also two paternal contributions to the nuclear genome; however, in contrast to complete hydatidiform mole, partial hydatidiform mole also has a maternal contribution. Partial hydatidiform moles are usually triploid (Szulman and Surti, 1978) and generally arise by dispermic fertilization of an enucleated egg (Jacobs et al., 1982; Lawler et al., 1982). In both complete and partial hydatidiform mole, trophoblastic hyperplasia, a characteristic of a molar pregnancy, is associated with the presence of two paternal genomes. Thus, trophoblastic hyperplasia involves imprinted genes, that is, genes normally expressed only from the maternally or paternally derived allele (Walsh et al., 1995). Further evidence that trophoblastic hyperplasia of molar pregnancies results from increased expression of paternally derived genes is provided by studies of the development of reconstituted mouse eggs (Kaufman et al., 1989). These studies indicate that hydatidiform mole is an imprinted condition, in that the pathology is dependent on the parental origin of the genome (Wake et al., 1978, 1998). Since molar pregnancies are related to paternal factors, the characteristics of the spermatozoa may be important during ICSI. Diploidy is the most common chromosomal anomaly found in the sperm of patients with meiotic disorders (Egozcue et al., 2000), including oligozoospermic males (Bernardini et al., 1997). Clinically, patients with severe oligoasthenoteratozoospermia have a higher risk of producing triploid embryos after ICSI (Macas et al., 2001). ICSI of oocytes ensures that only a single spermatozoon enters the egg, thus preventing hydatidiform mole, which arises by dispermy. In contrast, an excess of XYY triploid 445

5 446 zygotes, indicating a possible increase of dispermic fertilization related to IVF, has been observed (Rosenbusch et al., 1997). Despite the widespread use of assisted reproduction, reports of hydatidiform mole following assisted reproduction are scarce (Shozu et al., 1998). The practice of checking for dipronuclear fertilization should markedly reduce, although not necessarily eliminate, the possibility of transferring a triploid embryo. However, triploidy arising from diploid spermatozoa is not usually apparent at the pronuclear stage. Furthermore, the pronuclear stage may not be observed in some embryos, or pronuclear number may be incorrectly assessed. A case of partial hydatidiform mole has been reported in frozen thawed blastocyst stage embryos derived from IVF (Fluker and Yuzpe, 2000). So far as is known, however, the case described here is the first reported case of partial hydatidiform mole following transfer of a frozen thawed embryo derived from ICSI. Close follow-up of patients after evacuation of a molar pregnancy is mandatory, although the prevalence of persistent gestational trophoblastic disease after partial hydatidiform mole is <10% (Berkowitz et al., 1985). Some of the clinical factors that lead to IVF treatment for couples, including advanced maternal/paternal age and poor oocyte quality, can also predispose to molar pregnancy. Thus, this complication is not likely to be directly related to the IVF technique but rather to the characteristics of the women and their partners. For patients with repetitive hydatidiform mole (Al Hussaini and Abdel-Alim, 2001), it has been suggested that ICSI followed by preimplantation genetic diagnosis can be used to prevent recurrence (Reubinoff et al., 1997). The future management of these couples should thus include these details of their reproductive history. A sperm chromosome study by fluorescence in-situ hybridization may be required in some cases. 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