Low-dose aspirin does not improve ovarian responsiveness or pregnancy rate in IVF and ICSI patients: a randomized, placebo-controlled double-blind study

M. Päkkilä1,5, J. Räsänen1, S. Heinonen3, H. Tinkanen4, L. Tuomivaara2, K. Mäkikallio1, M. Hippeläinen3, J.S. Tapanainen1 and H. Martikainen1

1 Department of Obstetrics and Gynecology, University of Oulu, 2 Infertility Clinic, Family Federation of Finland, 90220 Oulu, 3 Department of Obstetrics and Gynecology, University of Kuopio, 70211 Kuopio and 4 Department of Obstetrics and Gynecology, University of Tampere, 33521 Tampere, Finland

5 To whom correspondence should be addressed. Email: mervi.pakkila{at}oulu.fi


    Abstract
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Acknowledgements
 References
 
BACKGROUND: Poor ovarian and endometrial responses to gonadotrophin stimulation in assisted reproduction techniques lead to decreased pregnancy rates. The aim of the present study was to test the hypothesis that low-dose aspirin started prior to controlled ovarian stimulation improves ovarian responsiveness, pregnancy rate (PR) and pregnancy outcome. METHODS: A total of 374 women who were to undergo IVF/ICSI were randomized to receive 100 mg of aspirin (n=186) or placebo (n=188) daily. Treatment was started on the first day of controlled ovarian stimulation. It was continued until menstruation or a negative pregnancy test. Pregnant women continued the medication until delivery. The main outcome measures were the number of oocytes, number and quality of embryos, the clinical PR and pregnancy outcome. RESULTS: The mean (±SD) number of oocytes (12.0±7.0 versus 12.7±7.2), the total mean number of embryos (5.82±4.35 versus 5.99±4.66), the mean number of top quality embryos (0.99±1.39 versus 1.18±1.51) and the number of embryos transferred (1.64±0.64 versus 1.63±0.71) did not differ in the aspirin and placebo groups. Between the aspirin and placebo group, there was no statistically significant difference in clinical PR per embryo transfer (25.3%, n=44 out of 174 versus 27.4%, n=48 out of 175) or clinical PR per cycle initiated (23.7% versus 25.5%). Birth rate per embryo transfer did not differ significantly between the aspirin (18.4%) and placebo (21.1%) groups. The incidence of poor responders [12 (6.5%) versus 13 (6.9%)] was similar in both groups. CONCLUSIONS: The present results indicate that low-dose aspirin treatment does not have any beneficial effect on ovarian responsiveness, PR and pregnancy outcome in unselected women undergoing IVF/ICSI.

Key words: ICSI/IVF/low-dose aspirin/ovarian response/pregnancy rate


    Introduction
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Acknowledgements
 References
 
The clinical pregnancy rate among women undergoing fresh embryo transfer (ET) after IVF or ICSI varies from 20 to 35% per transfer (Nygren and Andersen, 2001Go; Harlin et al., 2002Go). The main factors that affect the outcome of IVF and ICSI treatments are age of the woman, number of oocytes retrieved, quality of the embryos, number of embryos transferred, success of embryo transfer and endometrial receptiveness (Weckstein et al., 1997Go; Ebner et al., 2000Go; Terriou et al., 2001Go; Ziebe et al., 2001Go; Tomas et al., 2002Go). When good quality embryos are available, a high pregnancy rate (PR) can be achieved by single ET in selected women (Martikainen et al., 2001Go; de Sutter et al., 2003Go; Tiitinen et al., 2003Go).

Low-dose acetylsalicylic acid (aspirin) irreversibly inhibits the enzyme cyclo-oxygenase in platelets, preventing the synthesis of thromboxane (Vane, 1971Go; Willis, 1974Go), which is the most potent vasoconstrictive agent in the human body. By decreasing platelet aggregation and inhibiting vasoconstriction, low-dose aspirin may enhance uterine and ovarian blood flow and tissue perfusion and thus improve the results of IVF and ICSI treatments (Wada et al., 1994Go; Rubinstein et al., 1999Go). Previous studies have shown that low-dose aspirin started in the second trimester of pregnancy in high-risk populations decreases the incidence of pre-eclampsia and preterm labour, and increases the birth weight of the newborn (Italian Study of Aspirin in Pregnancy, 1993Go; Collaborative Low-dose Aspirin Study in Pregnancy Collaborative Group, 1994Go). Low-dose aspirin has also been found to improve endometrial receptiveness in cases of thin endometrium (Weckstein et al., 1997Go). Furthermore, women with recurrent miscarriages and antiphospholipid syndrome may benefit from low-dose aspirin therapy, especially when combined with low-molecular weight heparin (Sher et al., 1994Go; Rai and Regan, 1997Go; Tulppala et al., 1997Go). However, the results of low-dose aspirin treatment as regards ovarian responsiveness and pregnancy rate in unselected subjects undergoing artificial reproduction techniques have remained controversial (Rubinstein et al., 1999Go; Urman et al., 2000Go).

In the present randomized double-blind prospective study, we hypothesized that low-dose aspirin therapy (100 mg daily) improves ovarian responsiveness and the clinical PR in unselected subjects undergoing IVF/ICSI when started concomitantly with controlled ovarian stimulation.


    Materials and methods
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Acknowledgements
 References
 
This multicentre randomized double-blind prospective study involved 374 women undergoing IVF (n=235), ICSI (n=120) or combined treatment (n=19) in 2000–2003. The main outcome measures were the number of oocytes, number and quality of embryos, the clinical PR and pregnancy outcome. Each patient was included only once in the study. The Ethics Committees in each institution approved the study protocol and all the subjects gave written informed consent. In all four centres (Oulu University Hospital, The Family Federation of Finland, Oulu, Tampere University Hospital and Kuopio University Hospital), the stimulation and treatment protocols were identical (Martikainen et al., 2001Go). Inclusion criteria were: (i) age <40 years; (ii) fewer than four previous ovarian stimulations; and (iii) no contraindications for aspirin. A long GnRH agonist protocol was carried out, and Buserelin (Suprecur®; Hoechst AG, Frankfurt, Germany) or nafarelin (Synarela®; Syntex Nordica AB, Södertälje, Sweden) were started for downregulation on the cycle days 20–22. When ovarian suppression was confirmed by ultrasound, recombinant FSH (rFSH, Gonal-F®, Laboratories Serono; or Puregon®, Organon, Oss, The Netherlands) or HMG (Menogon®; Ferring, The Netherlands) was used for controlled ovarian stimulation [mean dose of gonadotrophin was 2324 IU (±865) in the aspirin group and 2184 IU (±995) in the placebo group]. Each clinic used their routine hormone regimens. Oocyte retrieval was performed 34–36 h after injection of 5000–10 000 IU of HMG (Profasi®, Ares-Serono; or Pregnyl®, Organon).

Randomization was carried out in blocks of four with sealed envelopes by the pharmacist at Oulu University Hospital. The women were treated with identical looking tablets of aspirin or placebo (Bayer AG, Leverkusen, Germany). The patients were randomized on the first day of gonadotrophin stimulation to receive 100 mg of oral aspirin (n=186) or placebo (n=188) daily in one dose until menstruation or a negative pregnancy test result (Figure 1). Pregnant women continued the medication until delivery. Fertilized oocytes were cultured in MediCult Medium® (Medi-Cult A/S, Copenhagen, Denmark), IVF-500 Medium® (Scandinavian IVF Science, Gothenburg, Sweden) or Sydney IVF Medium® (Cook IVF, Queensland, Australia). In two centres, one top quality embryo (n=82 out of 193) was electively transferred into the uterine cavity 46–50 h after oocyte retrieval in subjects under 39 years undergoing their first or second stimulation. In the other two centres, single ET was performed in 21 out of 181 cases. Otherwise, in all centres, two (n=227 out of 374) or three (n=9/374) embryos were transferred. The criteria for a top quality embryo were: normal fertilization (2PN), 4–5 blastomeres on day 2, <20% fragmentation and no multinuclear blastomeres (van Royen et al., 1999Go). Natural progesterone (Lugesterone®, Leiras, Finland) was given transvaginally (200 mg x3) for luteal support for 14 days. Clinical pregnancies were confirmed by transvaginal ultrasonography 5 weeks after ET.



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Figure 1. Flowchart for the randomized study.

 
Sample size and statistical analysis
Sample size was calculated by means of Stata Statistics/Data Analysis 8.0 (Stata Corporation, College Station, TX) so as to be able to reveal a 15% increase in clinical PR per cycle in favour of the aspirin group. To detect such an increase with {alpha}=0.05 and a power >80%, we needed 176 subjects in each group. In comparisons between the groups, {chi}2 tests and two-tailed t-tests were used as intention to treat analysis.


    Results
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Acknowledgements
 References
 
There were no significant differences in maternal age, aetiology of infertility or previous pregnancy history between the aspirin and placebo groups (Table I). rFSH was used for ovarian stimulation in 155 subjects in the aspirin group and in 151 subjects in the placebo group, and HMG was used in 31 subjects in the aspirin group and in 37 subjects in the placebo group. IVF, ICSI and combined techniques were used in 119 (64%), 57 (31%) and 10 (5%) subjects in the aspirin group and in 116 (62%), 63 (34%) and nine (4%) subjects in the placebo group, respectively. The outcome of treatment in both groups is presented in Table II. No significant differences in ovarian responsiveness (number and quality of oocytes), fertilization rate, number of top quality embryos transferred, total number of top quality embryos, PR and pregnancy outcome were observed between the groups.


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Table I. Characteristics of the groups

 

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Table II. The outcome of the treatment, fresh embryo transfer and pregnancy in the study groups

 
ET was possible in 174 (93.5%) subjects in the aspirin group and in 175 (93.1%) subjects in the placebo group. Elective single ET was performed in 40 (23.0%) cases in the aspirin group and in 42 (24.0%) cases in the placebo group, whereas double ET was carried out in 116 (66.7%) and in 111 (63.4%) cases, respectively. Only one embryo was available for transfer in 14 (8.0%) subjects in the aspirin group and in 17 (9.7%) subjects in the placebo group. Three embryos were transferred in four (2.3%) subjects in the aspirin group and in five (2.9%) subjects in the placebo group. In the aspirin group, no embryos were transferred in 12 (6.5%) cases and in the placebo group in 13 (6.9%) cases owing to a poor response to gonadotrophin treatment, unsuccessful fertilization, premature ovulation or a low oocyte cleavage rate. One subject in the placebo group withdrew from the study after signing the informed consent. Clinical PR per ET did not differ statistically in aspirin- (25.3%) and placebo- (27.4%) treated subjects (Table II). When clinical PR per ET was analysed separately in each clinic, the results did not differ significantly between the aspirin (17.6–37.5%) and placebo (20.8–29.6%) groups. When the subjects were divided into two groups according to age (<35 years and ≥35 years) and the cycle number (first versus second–third), no statistically significant difference were found in variables of ovarian responsiveness or in PR between the study groups.

No significant difference was observed in the pregnancy outcome between the aspirin and placebo groups. There were 32 out of 174 (18.4 %) live births per ET in the aspirin group and 37 out of 175 (21.1 %) in the placebo group. The incidence of miscarriage per clinical pregnancy was 18.2% (eight out of 44) in the aspirin group and 16.7% (eight out of 48) in the placebo group, and the incidence of extrauterine pregnancy per clinical pregnancy was 9.0% (four out of 44) and 6.3% (three out of 48), respectively.


    Discussion
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Acknowledgements
 References
 
This randomized, placebo-controlled double-blind trial with 374 subjects revealed that 100 mg of aspirin daily, started concomitantly with gonadotrophin stimulation for IVF/ICSI, did not increase ovarian responsiveness, improve the quality of the oocytes and embryos or improve the PR or birth rate.

The findings are in agreement with those in a study by Urman et al. (2000)Go, who found no difference in the number of oocytes or PR between aspirin-treated (80 mg, n=139) and non-treated (n=136) ICSI subjects, whose main indication for treatment was male factor, in 85% of the cases. Generally the couples with male infertility have a good result in ICSI treatment and, according to the mechanism of action of low-dose aspirin, it is possible that male infertility cases would benefit less from aspirin treatment compared with other infertility etiologies.

Some selected groups of patients, such as poor responders, ageing women or patients with previous implantation failures, could benefit from low-dose aspirin treatment. By altering the balance of prostacyclin and thromboxane, low-dose aspirin may enhance uterine and ovarian blood flow (Wada et al., 1994Go; Rubinstein et al., 1999Go) and improve endometrial receptivity and ovarian responsiveness to hormonal stimulation. However, the randomized and placebo-controlled study by Lok et al. (2004)Go has revealed that low-dose aspirin (80 mg/day) started at the time of commencement of gonadotrophin stimulation does not improve ovarian responsiveness (mature follicles, oocytes retrieved and total number of embryos) or PR in poor responders undergoing IVF (Lok et al., 2004Go). Accordingly, we did not find any statistical significance in ovarian responsiveness or PR between aspirin and placebo groups in patients with previous IVF/ICSI failures or in women aged ≥35 years.

In contrast, Rubinstein et al. (1999)Go reported a remarkable increase in number of oocytes and an improvement in pregnancy rate after controlled ovarian stimulation in aspirin- (100 mg daily) treated IVF subjects with tubal factor, versus placebo. A recent randomized study showed increased birth rate per ET in aspirin-treated patients with a short regimen compared with the no treatment group in a non-selected IVF population with 1380 cycles (1022 patients) (Waldenstrom et al., 2004Go). However, in this study, there are many aspects which make the interpretation of the results difficult. First, in the aspirin group, the number of embryos transferred was significantly higher than in the no-treatment group, which could contribute to the higher PR in the aspirin group. Secondly, the randomization method used did not allow an equal possibility for every patient to receive either aspirin or no treatment randomly. Thirdly, the study was open and not placebo controlled, both of which may increase the risk for study bias.

As regards the dose of aspirin, 80–160 mg daily in healthy volunteers (Cerletti et al., 2003Go) and 0.5–2.0 mg/kg daily in hypertensive pregnant women (Vainio et al., 1999Go) have been shown to increase the prostacyclin–thromboxane ratio. Thus, the dose of 100 mg daily used in the present study should have been sufficient to demonstrate the possible beneficial effects of aspirin.

The sample size of this study is able to demonstrate a 15% increase in PR in the aspirin-treated group. It is obvious that a smaller increase would have been clinically significant. However, to prove a 5% increase in PR in favour of the aspirin group, almost 1200 patients would have been needed in each randomization arm to demonstrate the significance with 80% power ({alpha}=0.05).

In conclusion, the results of this randomized, placebo-controlled, double-blind study showed that low-dose aspirin does not improve ovarian responsiveness, clinical PR or pregnancy outcome in subjects undergoing IVF/ICSI treatments.


    Acknowledgements
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Acknowledgements
 References
 
We thank all the staff of the participating infertility clinics, and Bayer AG, for making this study possible. Financial support was provided by the Academy of Finland, University of Oulu.


    References
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Acknowledgements
 References
 
Cerletti C, Dell'Elba G, Manarini S, Pecce R, Castelnuovo A, Scorpiglione N, Feliziani V and de Gaetano G (2003) Pharmacokinetic and pharmacodynamic differences between two low dosages of aspirin may affect therapeutic outcomes. Clin Pharmacokinet 42, 1059–1070.[ISI][Medline]

Collaborative Low-dose Aspirin Study in Pregnancy Collaborative Group (1994) CLASP: a randomised trial of low-dose aspirin for the prevention and treatment of pre-eclampsia among 9364 pregnant women. Lancet 343, 619–629.[CrossRef][ISI][Medline]

de Sutter P, van der Elst J, Coetsier T and Dhont M (2003) Single embryo transfer and multiple pregnancy rate reduction in IVF/ICSI: a 5-year appraisal. Reprod Biomed Online 6, 464–469.[Medline]

Ebner T, Yaman C, Moser M, Sommergruber M, Feichtinger O and Tews G (2000) Prognostic value of first polar body morphology on fertilization rate and embryo quality in intracytoplasmic sperm injection. Hum Reprod 15, 427–430.[Abstract/Free Full Text]

Harlin J, Aanesen A, Csemiczky G, Wramsby G and Fried G (2002) Delivery rates following IVF treatment, using two recombinant FSH preparations for ovarian stimulation. Hum Reprod 17, 304–309.[Abstract/Free Full Text]

Italian Study of Aspirin in Pregnancy (1993) Low-dose aspirin in prevention and treatment of intrauterine growth retardation and pregnancy-induced hypertension. Lancet 341, 396–400.[CrossRef][ISI][Medline]

Lok IH, Yip SK, Cheung LP, Yin Leung PH and Haines CJ (2004) Adjuvant low-dose aspirin therapy in poor responders undergoing in vitro fertilization: a prospective, randomized, double-blind, placebo-controlled trial. Fertil Steril 81, 556–561.[CrossRef][ISI][Medline]

Martikainen H, Tiitinen A, Tomas C, Tapanainen J, Orava M, Tuomivaara L, Vilska S, Hyden-Granskog C and Hovatta O (2001) One- versus two-embryo transfer after IVF and ICSI: a randomized study. Hum Reprod 16, 1900–1903.[Abstract/Free Full Text]

Nygren KG and Andersen AN (2001) for the European IVF-monitoring programme (EIM) Assisted reproductive technology in Europe, 1998. Results generated from European registers by ESHRE. European Society of Human Reproduction and Embryology. Hum Reprod 16, 2459–2471.[Abstract/Free Full Text]

Rai R and Regan L (1997) Antiphospholipid antibodies, infertility and recurrent miscarriage. Curr Opin Obstet Gynecol 9, 279–282.[ISI][Medline]

Rubinstein M, Marazzi A and Polak DF (1999) Low-dose aspirin treatment improves ovarian responsiveness, uterine and ovarian blood flow velocity, implantation, and pregnancy rates in patients undergoing in vitro fertilization: a prospective, randomized, double-blind placebo-controlled assay. Fertil Steril 71, 825–829.[CrossRef][ISI][Medline]

Sher G, Feinman M, Zouves C, Kuttner G, Maassarani G, Salem R, Matzner W, Ching W and Chong P (1994) High fecundity rates following in-vitro fertilization and embryo transfer in antiphospholipid antibody seropositive women treated with heparin and aspirin. Hum Reprod 9, 2278–2283.[Abstract]

Terriou P, Sapin C, Giorgetti C, Hans E, Spach JL and Roulier R (2001) Embryo score is a better predictor of pregnancy than the number of transferred embryos or female age. Fertil Steril 75, 525–531.[CrossRef][ISI][Medline]

Tiitinen A, Unkila-Kallio L, Halttunen M and Hyden-Granskog C (2003) Impact of elective single embryo transfer on the twin pregnancy rate. Hum Reprod 18, 1449–1453.[Abstract/Free Full Text]

Tomas C, Tikkinen K, Tuomivaara L, Tapanainen JS and Martikainen H (2002) The degree of difficulty of embryo transfer is an independent factor for predicting pregnancy. Hum Reprod 17, 2632–2635.[Abstract/Free Full Text]

Tulppala M, Marttunen M, Soderstrom-Anttila V, Foudila T, Ailus K, Palosuo T and Ylikorkala O (1997) Low-dose aspirin in prevention of miscarriage in women with unexplained or autoimmune related recurrent miscarriage: effect on prostacyclin and thromboxane A2 production. Hum Reprod 12, 1567–1572.[Abstract]

Urman B, Mercan R, Alatas C, Balaban B, Isiklar A and Nuhoglu A (2000) Low-dose aspirin does not increase implantation rates in patients undergoing intracytoplasmic sperm injection: a prospective randomized study. J Assist Reprod Genet 17, 586–590.[CrossRef][ISI][Medline]

Vainio M, Maenpaa J, Riutta A, Ylitalo P, Ala-Fossi SL and Tuimala R (1999) In the dose range of 0.5–2.0 mg/kg, acetylsalicylic acid does not affect prostacyclin production in hypertensive pregnancies. Acta Obstet Gynecol Scand 78, 82–88.[CrossRef][ISI][Medline]

Vane JR (1971) Inhibition of prostaglandin synthesis as a mechanism of action for aspirin-like drugs. Nature 231, 232–235.

Van Royen E, Mangelschots K, De Neubourg D, Valkenburg M, Van de Meerssche M, Ryckaert G, Estermans W and Gerris J (1999) Characterization of a top quality embryo, a step towards single-embryo transfer. Hum Reprod 14, 2345–2349.[Abstract/Free Full Text]

Wada I, Hsu CC, Williams G, Macnamee MC and Brinsden PR (1994) The benefits of low-dose aspirin therapy in women with impaired uterine perfusion during assisted conception. Hum Reprod 9, 1954–1957.[Abstract]

Waldenstrom U, Hellberg D and Nilsson S (2004) Low-dose aspirin in a short regimen as standard treatment in in vitro fertilization: a randomized, prospective study. Fertil Steril 81, 1560–1564.[CrossRef][ISI][Medline]

Weckstein LN, Jacobson A, Galen D, Hampton K and Hammel J (1997) Low-dose aspirin for oocyte donation recipients with a thin endometrium: prospective, randomized study. Fertil Steril 68, 927–930.[CrossRef][ISI][Medline]

Willis AL (1974) An enzymatic mechanism for the antithrombotic and antihemostatic actions of aspirin. Science 183, 325–327.[ISI][Medline]

Ziebe S, Loft A, Petersen JH, Andersen AG, Lindenberg S, Petersen K and Andersen AN (2001) Embryo quality and developmental potential is compromised by age. Acta Obstet Gynecol Scand 80, 169–174.[CrossRef][ISI][Medline]

Submitted on August 9, 2004; resubmitted on March 9, 2005; accepted on March 14, 2005.





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