The effect of repeated controlled ovarian stimulation in donors

C. Caligara1, J. Navarro1, G. Vargas1, C. Simón1,2, A. Pellicer1,2 and J. Remohí1,2,3

1 Instituto Valenciano de Infertilidad, and 2 Department of Paediatrics, Obstetrics and Gynaecology, Valencia University School of Medicine, Valencia, Spain


    Abstract
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Acknowledgements
 References:
 
BACKGROUND: The effect of repeated controlled ovarian stimulation in young, healthy women who consented to donate their oocytes was assessed through a retrospective study. METHODS AND RESULTS: Between March 1994 and February 2000, 284 donors consented to at least two controlled ovarian stimulation cycles, with four donors undergoing up to nine cycles. Through an intra-group analysis we studied the oocyte retrieval rate, controlled by the duration and doses of stimulation, as well as the interval between cycles, finding that the number of retrieved oocytes was maintained during repeated controlled ovarian stimulation cycles. The oocyte quality, assessed as fertilization, implantation and pregnancy rates, was similar through five repeated controlled ovarian stimulation cycles. The same result was obtained when the interval in days was analysed as a continuous variable or categorized in <90 and 90 days. CONCLUSIONS: The analysis of the data suggests that successive stimulation cycles do not impair ovarian response in terms of quantity and quality of the oocytes obtained in this group of women. Furthermore, the interval between cycles does not influence the result of the stimulations.

Key words: controlled ovarian stimulation/IVF/oocyte donation


    Introduction
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Acknowledgements
 References:
 
Oocyte donation has become a common treatment modality for a large spectrum of infertility conditions, such as premature ovarian failure, low response to ovarian stimulations, high maternal age, recurrent pregnancy loss, cases with heritable maternal genetic abnormalities, or women who have previously undergone multiple unsuccessful IVF attempts (Remohí et al., 1993Go, 1996Go). The number of clinical indications of this reproductive treatment is growing at an exponential rate (Kalfoglou and Gittelsohn, 2000Go).

At our institution, most donors are young healthy women who consent to an anonymous and gratuitous donation. They undergo controlled ovarian stimulation (COS) with fertility drugs in order to promote multiple follicular development. Some of these donors are treated with consecutive COS cycles, with different intervals of time between stimulations. Concern has been raised that consecutive COS cycles might impair the ovarian response, requiring higher doses of gonadotrophins to achieve the previous response, and/or alter the oocyte quality (Diamond et al, 1987Go). Ovarian response in consecutive cycles has been analysed in different studies but with inherent bias, including small numbers of patients and cycles, different ovarian stimulation protocols and women with various ovulation disorders (Ron-El et al., 1990Go; Silverberg et al., 1992Go). It is unclear if repeated COS in donors is a completely risk-free procedure in terms of premature ovarian failure, breast or ovarian cancer induced by these pharmacological therapies (Fauser et al., 1999Go).

Human follicular development from the pre-antral stage to the pre-ovulatory follicle ready to undergo ovulation is estimated to take three menstrual cycles (Gougeon et al, 1986). No published data are available about the influence of the interval between cycles on COS and the outcome.

Our aim was to assess the effect of multiple COS on oocyte retrieval rate, as well as the oocyte's quality, reflected by the recipient's outcome. Another objective of our study was to determine the influence of the length of interval between cycles on oocyte retrieval rate, fertilization, implantation and pregnancy rates.


    Materials and methods
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Acknowledgements
 References:
 
We retrospectively analysed data from donors who underwent multiple COS and oocyte retrievals between March 1994 and February 2000 at Instituto Valenciano de Infertilidad. A total of 284 young, healthy donors underwent at least two cycles of COS. Mean age was 24.9 years (standard deviation 4.2). Of the women, 136 were nulliparous (47.9%), while 148 had had at least one pregnancy. Of them, 114 had deliveries (40.1%), with a range of 1–4 live borns (mean ± SD, 1.6 ± 0.7), while in 28 the pregnancy was voluntarily interrupted, and six had a spontaneous abortion.

Donor's screening
This study included donors aged 18–35 years, with no personal or familial history of congenital malformations or hereditary diseases, who had undergone testing for Human Immunodeficiency Virus, Cytomegalovirus, Syphilis, Neisseria gonorrhoea, Hepatitis B and C, Chlamydia trachomatis, Herpes virus, Rubella, and Toxoplasmosis. All donors had a normal karyotype, normal physical and gynaecological examinations, and none of them had endocrine abnormalities. They were informed of the risks associated with COS and oocyte retrieval procedures, and signed a written agreement. The local ethics committee regulated the number of stimulations in accordance with the number of live births allowed by Spanish law, though the number and interval of COS cycles underwent by each donor was decided by herself.

Ovarian stimulation protocol
All donors underwent our protocol for COS as previously described (Remohí et al., 1995Go). Briefly, a long protocol for pituitary desensitization was carried out, with daily administration of 1 mg s.c. leuprolide acetate (Procrin; Abbott Laboratories, Madrid, Spain) starting in the luteal phase of the previous cycle. After the menstrual period, the dose of leuprolide acetate was decreased to 0.5 mg s.c. daily until HCG administration. Gonadotrophin stimulation was initiated on day 3 after menses, if the ultrasound showed no follicular activity. Human menopausal gonadotrophin (HMG Lepori, Farma-lepori Laboratories, Barcelona, Spain) and ultra purified urinary FSH (Neo-Fertinorm, Serono Laboratories, Madrid, Spain) were used, at a dose of 225 IU of FSH and 75 IU of HMG for 6 days. Monitoring of follicular development commenced on day 7, and adjustments in gonadotrophin dosage were individually tailored according to each patient's follicular response, based on ultrasound monitoring and serum oestradiol determinations. HCG (Profasi, Serono Laboratories, Madrid, Spain) 10 000 IU i.m. was administered when >=2 follicles reached 21 mm in maximum diameter. Ultrasound-guided vaginal oocyte retrievals were scheduled 36 h after HCG administration, using propofol (Propofol Abbott, Abbott Laboratories, Madrid, Spain) for sedation. The mean interval between cycles was 174 ± 139 days, with a minimum interval of 45 days, a maximum of 1166, and a median of 119 days.

Recipients
A total of 1684 oocyte donation cycles in 1017 recipients was completed. Of these, 1466 (87.1%) led to an embryo transfer. The reasons for the 218 (12.9%) no embryo transfers were: in 145 (66.5%) cases embryo quality was not suitable to perform transfer; in 51 (23.4%) there was fertilization failure (48 with IVF, 3 with ICSI); and in 22 (10.1%) embryos were frozen and transferred in a posterior cycle due to inadequate endometrium at the time of transfer. Mean age ± SD of recipients was 38.5 ± 5.2 years (range: 24–55 years).

Indications of suitability for the donation procedure were ovarian failure in 810 cycles (48.1%), low responders in 470 cycles (27.9%), previous assisted reproductive treatment failure in 337 cycles (20.0%), genetic in 41 cycles (2.4%), recurrent abortion in 18 cycles (1.0%), and others in 8 cases (0.5%).

Analysed variables
Donor's age, day of stimulation, dose of gonadotrophins (HMG, FSH), day of GnRH analogue used, and interval between cycles in days were analysed in 922 stimulation cycles. Analysed outcomes included retrieved oocytes, fertilization, implantation and pregnancy rates.

Statistical analysis
To determine the effect of the repetition of COS on oocyte retrieval rate, each donor was compared with herself through consecutive cycles of COS, using a repeated measure analysis of variance controlled by varying covariates.

To assess oocyte quality through repeated cycles of COS, we chose to study donors who had five consecutive cycles (n = 43), in order to preserve the statistical power of the analysis (minimum number required = 30) through an intra-group analysis. Our strategy of studying oocyte quality from the same donor through consecutive cycles was to obtain means of the evaluated parameters of the recipients of each cycle. The distribution of semen characteristics in the recipients group was assessed by a {chi}2 with Monte Carlo method. In all the analysis, the assumed sphericity was verified by the Mauchly test, that is, to assure that no data overlapping is present in the statistical analysis. The eta2 measures the magnitude of the effect (COS repetition) on the evaluated parameters.

To study the effect of the interval between cycles on oocyte quantity and quality, a controlled by varying co-variates ANOVA test was used, with the interval either as a continuous variable, or as categorized into two groups (<90 days, and >=90 days).

The Statistical Package for Social Science (SPSS Software Inc., Chicago, IL, USA) for Windows, version 6.3 and 8.0, was used for the statistical analysis.


    Results
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Acknowledgements
 References:
 
Quantity
Oocyte retrieval rate was maintained through the consecutive cycles. No significant difference was observed in the number of retrieved oocytes up to nine consecutive cycles of COS in the same donor, even after statistically annulling the differences in days of stimulation, gonadotrophin dose, days of GnRH agonist analogue use, and interval in days between cycles. The mean number of oocytes retrieved through consecutive cycles is shown in Table IGo. Median and interquartile ranges of retrieved oocytes in the consecutive cycles are shown in Figure 1Go.


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Table I. Oocytes retrieved per cycle (mean ± SD)
 


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Figure 1. Median and interquartile ranges of retrieved oocytes through consecutive cycles of COS in donors.

 
Quality
After an intra-group analysis of the recipients' cycles of the 43 donors with five stimulations, no differences were found in fertilization, implantation and pregnancy rates according to the oocyte's cycle origin. Results for all the variables are shown in Table IIGo. Mean pregnancy rate obtained from each cohort of oocytes through the cycles is represented in Figure 2Go.


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Table II. Outcomes in recipients from oocytes donated through five consecutive COS cycles (mean ± SD)
 


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Figure 2. Pregnancy rate of donated oocytes (mean) through five consecutive cycles.Krustal–Wallis analysis of variance not significant, P = 0.325.

 
Semen characteristics were distributed equally in the study group (both in the total group and in the recipients of the five-cycle donors) (P > 0.62), and were out of 1684 cycles: normozoospermia 354 cases (21.2%), oligoasthenoteratospermia 1054 cases (62.5%), donor's spermatozoa 175 cases (10.4%), azoospermia 55 cases (3.3%), others 46 cases (2.7%).

Interval
A group of 35 donors who had their second COS cycle <90 days from the first, and the third COS cycle >=90 days from the second was analysed to assess the influence of the interval on the quantity and quality of the retrieved oocytes. No differences were found between the cycles in terms of oocytes retrieved, fertilization, implantation or pregnancy rate. Results are shown in Table IIIGo.


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Table III. Retrieved oocytes and recipients' outcome of 35 donors who had their second COS cycle <90 days from the first, and the third COS cycles >=90 days from the second (mean ± SD)
 

    Discussion
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Acknowledgements
 References:
 
This study was designed to test the effect of repeated COS on ovarian response, with the advantage of analysing young, healthy women from a large ovum donation programme. No adverse effects neither in the quantity nor the quality of the retrieved oocytes were found. We are conscious of the inherent bias associated with the retrospective analysis of these data, but the special characteristics of this type of analysis made the retrospective study a useful tool to achieve our objective.

The artificially high gonadotrophin milieu induced in COS does not probably create a larger cohort of follicles than the one already pre-selected by the ovary. Rather, it provides an overriding mechanism by which many follicles destined to become atretic escape from this end (Schipper et al., 1998Go). Therefore, the finding of this study that oocyte retrieval rate is maintained during repeated COS cycles suggests that each woman has a recruited `ovarian quota' of follicles, which is partly rescued from atresia by the administration of high doses of gonadotrophins.

The impact of repeated COS was recently studied in women with ovarian endometriosis undergoing IVF and compared with women with tubal factor infertility. A progressive decrease of ovarian response in women with ovarian endometriosis was found, whereas, in women with tubal infertility, repeated COS had no influence (Al-Azemi et al., 2000Go). Also, it has been demonstrated that women with normal ovulatory function, undergoing intra-uterine inseminations, are likely to have similar ovarian response in consecutive COS cycles (Ahmed Ebbiary et al., 1995).

Our data show that the pregnancy rate was maintained throughout the five consecutive cycles, and that IVF results were not influenced by the interval between cycles. However, we cannot apply these conclusions to the wide spectrum of infertile patients, and further studies are needed.

The donor–recipient model is useful in evaluating oocyte quality. The implantation capacity of the uterus is not affected by the oocyte donation clinical indication, recipient's age, endometrial thickness, or serum oestradiol level under hormonal replacement therapy (Balmaceda et al, 1994Go; Lydic et al., 1996Go; Abdalla et al, 1997; Remohí et al., 1997Go). If male aetiologies are equally distributed in the population, thus obviating their influence, oocyte quality can be isolated to be analysed through the IVF outcomes.

According to Gougeon's hypothesis, follicle growth from the small pre-antral to the pre-ovulatory stage requires 80–90 days (Gougeon, 1986Go). Follicle development up to the antral stage is gonadotrophin independent, and only the last two weeks of this long trajectory are gonadotrophin dependent (Fauser and Van Heusden, 1997Go). Our study reflects that the quantity and quality of oocytes retrieved are not affected by the interval of time between COS cycles.

In the literature there is not enough coverage about the influence of the interval between consecutive cycles of COS. Studies on consecutive versus alternating cycles of ovarian stimulation using HMG for intercourse or intra-uterine inseminations found more pregnancies in women stimulated in consecutive than in alternating cycles (Silverberg et al, 1992Go). Our results show that the length of the interval has no effect on the outcome of COS.

Each donor programme must develop strategies in order to follow these women, after their donation, for a long time. We suggest the establishment of a world register for oocyte donors in order to detect any adverse effects associated with this assisted reproduction technique. At the same time, this will offer an optimal way to assess any long-term risks associated with the procedure such as an increased incidence of premature ovarian failure, ovarian or breast cancer. At the moment, there are no data published to answer this question.

In summary, the number of oocytes retrieved in young, healthy voluntary donors is maintained during repeated COS cycles, controlled by the duration and doses of stimulation, as well as the interval between cycles.

The oocyte quality, based on fertilization, implantation and pregnancy rates, is similar through five repeated COS cycles. This suggests that successive stimulation cycles do not impair ovarian response in terms of quantity and quality in this selected group of women.

The length of the interval between cycles had no effect on the oocyte retrieval rate, fertilization, implantation and pregnancy rate.


    Acknowledgements
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Acknowledgements
 References:
 
This study was supported by a grant from Agencia Española de Cooperación Internacional to C.C.and supported by FIS 00–0478 to A.P.


    Notes
 
3 To whom correspondence should be addressed at: Guardia Civil, 23 46020 Valencia, Spain. E-mail: ivi{at}ivi.es Back


    References:
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 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 Acknowledgements
 References:
 
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Submitted on April 18, 2001; accepted on August 7, 2001.