Human Reproductive Unit, Department of Obstetrics and Gynecology, País Vasco University, Hospital from Cruces, Baracaldo, Vizcaya, Spain
Author whom correspondence should be addressed at: María Diaz de Haro 7, 6iz. 48013 Bilbao, Spain. Fax: 34 946006476; Email: rmatorras{at}hcru.osakidetza.net
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Abstract |
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Key words: embryo transfer/implantation rate/interval loading-discharging embryos/prognostic factors/time
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Introduction |
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Difficult transfers are associated with an impaired implantation (Englert et al., 1986; Mansour et al., 1990
; Matorras et al., 2002
), presumably because of endometrial damage (Marikiniti et al., 2003
), presence of blood (Goudas et al., 1998
) and the triggering of uterine contractions due to cervical manipulation (Fanchin et al., 1998
; Lesny et al., 1998
).
There is, however, one aspect that, as far as we know, has not been studied: the time employed in the performance of the transfer. It could be speculated that embryos could be vulnerable to exposure to the environmental temperature, considerably lower than the 37°C of the culture medium and of the human body. Additional external factors such as the light or the catheter composition could have a detrimental effect. Thus the time elapsed between embryo loading and embryo deposition could be a prognostic factor of the implantation rate and the pregnancy rate.
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Material and methods |
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Main population characteristics were: woman age, 34.18±3.12 years, male age, 36.27±3.34 years, infertility duration, 5.14±2.56 years, 90% primary infertility. The main IVF-ICSI indications were: male factor (49.5%), tubal factor (26.47%), endometriosis (15.88%), IUI failure (25.84%), idiopathic infertility (28.65%).
Our IVF cycle management has been previously described (Matorras et al., 1998; Matorras et al., 2002
). Briefly, it consisted of down-regulation with gonadotrophin-releasing hormone analogue, triptoreline acetate (Decapeptyl, Laboratorios Lasa, Madrid, Spain) on long protocol, ovarian stimulation with recombinant FSH (Gonal F, Laboratorios Serono, Spain) and highly purified urinary menopausal gonadotropins (Menopur, Ferring, Spain), ovulation being triggered with 250 mcg Ovitrelle (Serono, Spain). Transvaginal ultrasound-guided oocyte retrieval was scheduled 36 h after hCG injection. The luteal phase was supplemented with micronized progesterone (Utrogestan, Laboratorios Seid, Spain), vaginally 200 mg/12 h.
A mock transfer was performed 34 weeks before oocyte pickup. When the study was performed our policy of embryo transfer consisted in transferring three embryos (when available) in cases with a good prognosis, whereas in poor prognosis cases (women older than 35, previous IVF failures, bad quality embryos) four embryos were transferred. ET was carried out at 4872 h post oocyte retrieval. All patients were placed in the lithotomy position. A bivalve speculum was placed to expose the cervix. The exocervix was cleaned with Ham F-10 medium (GIBCO BRL). Cervical mucus was gently aspired. Concurrently, in the adjacent embryo culture laboratory, the morphological appearance of the embryos was evaluated and the best embryos were selected and loaded into the soft Frydman catheter (Laboratoire CCD, Paris, France). Drawing up the embryos into the insertion catheter was done with the aid of a disposable tuberculin syringe using the three-drop procedure in which the embryos are separated by a bubble of air from a preceding and a following drop of medium.
Embryo transfer was always performed under ultrasound guidance, as previously described (Matorras et al., 2002). The embryos were released when the ultrasound scan showed the catheter to be within 11.5 cm of the uterine cavity fundus.
In all cases the Frydman catheter was used. Transfers were classified as easy (no cervical tenaculum needed), of moderate difficulty (cervical tenaculum needed) and difficult (cervical tenaculum and hysterometer) (Matorras et al., 2002).
In cases with an easy mock transfer, transfer technique was as follows. With the embryos already loaded, ET was tried with the soft inner sheath of the catheter, without the outside sheath. If in this way it was not possible to access into the uterine cavity, the catheter was gently pushed by means of dissection forceps. If it was still not possible to access into the uterine cavity, the outer sheath was employed.
In cases with mock transfers classified as moderately difficult or difficult, immediately before the ET we performed another mock ET, firstly only with the inner sheath, and if this failed with the outer sheath. ET was performed afterwards according to these results.
In very rare cases where it was not possible to accede into the uterine cavity as described, cervical tenaculum was used (n=16). In no case was the help of the hysterometer needed.
The catheter was carefully removed after a period of 10 s. The catheter was then checked under the microscope and flushed for embryos retained within the lumen or adherent to the outside of the catheter. After ET, the patient was transferred to the recovery area, and allowed to empty her bladder within 1015 min of embryo transfer and she was discharged 30 min after ET.
All procedures were performed with powder-free surgical gloves. Pregnancy was defined by the visualization of a gestational sac at the 6th week of amenorrhea.
Statistical analysis was performed by means of 2,
2 for trend, Fisher's exact test, MannWhitney and Student's t-tests following the standard criteria of applicability. Statistical significance limit was defined as alpha=0.05.
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Results |
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When only easy transfers were considered, the duration of the ILDE was significantly lower in conceptional than in non conceptional cycles (50.5 s±35.3 vs 60.2±44.7) (P<0.05), the remaining characteristics being similar. There were no differences concerning the duration of ILDE in non-easy transfers regarding conceptional (275.5 s±50.2) and non conceptional cycles (132.7 s±79.4), but there were few cases of such transfers (n=2 and 14).
Pregnancy rates and duration of ILDE (Table II)
Four groups were established on the basis of ILDE duration, with the following cut-offs: 30 s, 60 s and 120 s. There were no differences among the four populations according demographic parameters.
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Concerning implantation rates, the following results were obtained: ILDE <30 s=21.2% (73/345), ILDE 3160=15.4% (103/668), ILDE 61120=15.9% (37/233) and ILDE >120=9.4% (14/149) (2 for a trend=7.56, P=0.006). When only easy transfers were considered, similar results were obtained: 21.4% (72/336), 15.4% (103/668), 16.2% (37/229) and 8.8% (10/114) (
2 for a trend=11, P=0.009).
Statistical analysis was not possible regarding non-easy transfers, due to the small size of the population.
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Discussion |
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On the other hand, a number of studies have shown how some environmental factors can have a detrimental effect on oocytes and embryos such as exposure to light (Smith et al., 1993; Evans et al., 1999
), temperature (Rocha et al., 1998
), polyvinyl chloride (Hunter et al., 1988
), polychlorinated biphenils (Pocar et al., 2001
), and other environmental conditions (Cohen et al., 1997
; Rocha et al., 1998
). The importance of keeping embryo temperature close to 37°C and O2 and CO2 concentrations under specific ranges is well known (Bavister, 1995
; Cohen et al., 1997
), and a number of technological devices have been designed to avoid their fluctuations in oocytes and embryos during their manipulation and storage. However, from the moment of embryo loading, and up to embryo discharge the aforementioned environmental factors are difficult to control. Thus, it could be speculated that a long ILDE could decrease the implantation rates.
In our study we observed an inverse association between the duration of ILDE and the pregnancy and implantation rates. The higher pregnancy and implantation rates were observed with ILDE <30 s, and after that time there was a trend to lower pregnancy rates, that decreased sharply with ILDE >120 s. However, since difficult transfers are associated with lower pregnancy rates (Matorras et al., 2002; Tomás et al., 2002
), an association between long ILDE and the difficulty of the transfer could be expected. Thus, our data were reanalyzed considering only easy transfers, and a similar evolution of the pregnancy rates was observed: 40.0% (ILDE <30 s), 34.3% (3160), 32.0% (61120), 19.4% (>120). The same occurred when implantation rates were considered.
Thus, from our data it can be concluded that the duration of ILDE is a prognostic factor of IVF, independent of transfer difficulty.
It could be speculated that lower pregnancy and implantation rates associated with an increased duration of ILDE can be related with embryo exposure to a suboptimal environment: outside light, room temperature and even the theoretical influence of the composition of transfer catheter. However it can not be ruled out that a relatively prolonged endocervical manipulation could also play a role.
According to our data, the longer the ILDE duration, the lower the pregnancy and implantation rates. With ILDE >120 s there was a remarkable decrease in the pregnancy and implantation rates. Although our study was not a controlled study, we recommend that, when possible ILDE should be shortened, ideally to under 30 s. ILDE duration >120 s should be avoided.
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References |
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Submitted on March 1, 2004; accepted on May 20, 2004.
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