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In some IVF cycles, no fresh embryo transfer in the stimulated cycle is advisable. The cryopreservation of zygotes and the transfer of blastocysts in a cryo-embryo transfer is an option to circumvent an inadequate uterine environment due to risk of ovarian hyperstimulation syndrome, inappropriate endometrium build up, endometrial polyps or uterine myomas. For this strategy, highly secure and safe cryopreservation protocols are advisable. This study describes a protocol for aseptic vitrification of zygotes that results in high survival rates and minimizes the potential risk of contamination in liquid nitrogen during cooling and long-term storage. In mouse zygotes, there was no difference in efficiency as compared with a conventional open vitrification system. In IVF patients, aseptically vitrified zygotes showed no difference in blastocyst formation rate as compared with sibling zygotes kept in fresh culture. A clinical study comprising 173 cryo-cycles with a transfer of blastocysts originating from vitrified zygotes showed an ongoing pregnancy rate of 40.9%. The live birth rate per patient was 36.8%. A combination of good clinical results and increased safety conditions due to aseptic vitrification encourages the use of cryo-embryo transfer for patients with a suboptimal uterine environment in a fresh cycle. In stimulated IVF cycles, high doses of hormones are given to stimulate multifollicular growth. One drawback of the hormonal substitution is that the uterine environment is not at the same time optimally prepared for embryo implantation. A solution, which is increasingly under discussion, is to cryopreserve the embryos obtained in the stimulated cycle and to transfer them back into the optimal uterine environment in a subsequent cryo-cycle. This procedure requires highly secure and safe cryopreservation protocols in order to ensure benefits for both pregnancy and birth rates. We have established a protocol for the vitrification of zygote-stage embryos in aseptic devices, which minimize the potential risk of contamination during cooling and storage. The vitrified zygotes showed the same blastocyst development as compared with sibling zygotes in fresh culture. A clinical study comprising 173 cryo-cycles with transfer of blastocysts originating from vitrified zygotes shows an ongoing pregnancy rate of 40.9%. The live birth rate per patient was 36.8%. A combination of good clinical results and increased safety conditions due to aseptic vitrification conditions contributes to a change in transfer strategy and encourages us to increase the cryo-embryo transfer rate for an optimal uterine environment., (Copyright © 2012 Reproductive Healthcare Ltd. Published by Elsevier Ltd. All rights reserved.)

To determine the best day for the selection and transfer of a single embryo, a prospective, randomized study was undertaken that compared the ongoing pregnancy rate (PR) after single embryo transfer (SET) on day 3 with that after single blastocyst transfer (SBT) on day 5. Our results show an overall significantly higher PR after SBT (32.8%) compared with SET (23.2%), and a PR of 40.8% after SBT versus 25.6% after excellent-quality embryos became available.

Luteal phases after in vitro fertilization (IVF) and embryo replacement have been studied in 241 cycles. A positive correlation was observed between the follicular estradiol (E2) peak and the progesterone (P) level on day 3 of the luteal phase, but no correlation was found between the E2-peak value and the luteal-phase duration or midluteal P concentration. When the trials were classified in relation to their outcome (i.e., clinical pregnancies, chemical pregnancies, or failures), the mean P level on day 3 of the luteal phase was significantly higher in clinical pregnancies than in chemical pregnancies and in failures. Mean E2 levels on day 3 were not significantly different among the three groups. Values of the E2/P ratio were significantly higher in chemical pregnancies than in the other groups. No significant differences were observed among the three groups on day 8. When comparing trials ending in failure to those leading to clinical pregnancy for the same patients, pregnancies were obtained in cycles in which early luteal P was higher and the early luteal E2/P ratio was lower than in failures cycles. These data suggest that high P levels and a low E2/P ratio in the early luteal phase might have a favorable influence on the implantation process in human IVF.

After ovarian stimulation with clomiphene citrate combined with human menopausal gonadotropin for in vitro fertilization, the appearance of a spontaneous luteinizing hormone (LH) surge before fulfillment of the minimal criteria of follicular maturity (at least one follicle greater than 19 mm and serum estradiol [E2] greater than 400 pg/ml/follicle greater than 17 mm) is associated with reduced pregnancy rates. In these cases, follicles are smaller and serum E2 values are lower at the time of the LH surge. Oocyte recovery rate is reduced, embryonic anucleate fragments are more frequently observed, and the level of luteal progesterone on day 4 after oocyte retrieval is lower. Hyperandrogenic patients are more prone to display such premature spontaneous LH surge. We concluded that in case of an untimely LH rise, laparoscopy for oocyte retrieval should be cancelled.

Estradiol growth rate (EGR) during active follicular development was calculated for 89 stimulated in vitro fertilization cycles by exponential curve fit (r = 0.83). Cycles could be divided into four groups with very low, low, moderate, and high EGR values. Cases without oocyte fertilization and/or embryo replacement did not occur in the moderate EGR range, which also corresponded to a significantly better ratio of replaceable embryos versus oocytes recovered. This group was also endowed with a pregnancy rate amounting to 28.5% per laparoscopy and per replacement, i.e., about three times higher than in the three other groups. Very low or high EGR entailed significantly higher percentages of missed oocyte recovery and/or fertilization failure. The frequency of occurrence of a spontaneous luteinizing hormone peak was negatively correlated with EGR. The data indicate that a better outcome of in vitro fertilization may be expected when the estrogen rise starts early in the cycle and adopts a moderate growth rate (0.3 to 0.4).

One hundred thirty-three couples were classified into four groups according to previous sperm analyses. These couples underwent 237 in vitro fertilization trials that led to 46 clinical pregnancies (19.4%/trial). Clinical pregnancy rate per laparoscopy of the group with male defects was significantly higher than that of the normospermic couples (25 versus 14%, P less than 0.05). The fertilization rate was significantly reduced only if two sperm anomalies were present (P less than 0.05). However, the mean number of embryos transferred did not differ significantly among the four groups, but the proportion of trials without transfer was increased with regard to the severity of sperm defect. Implantation rate per embryo in cases of triple transfer was significantly higher in the group with sperm anomalies as compared with the normospermic cases (18 versus 10%, P less than 0.02). Correction of the male problem through IVF treatment allows higher female fertility to be disclosed in these cases.

Between March 1983 and March 1986, 108 pregnancies were obtained at the IVF clinic of St Pierre Hospital in Brussels. There were 29 chemical pregnancies (26.8%), five ectopic pregnancies (4.6%), 15 abortions (14%) and 59 ongoing pregnancies of over 20 weeks (54%). Patients who had experienced a chemical pregnancy at first trial displayed a significantly higher rate of ongoing pregnancy at their second attempt. Among the 15 cases of abortion, a chromosomal anomaly was detected in two cases and suspected in a third. Four of the five ectopic pregnancies occurred in patients with previously documented tubal pathology. The ongoing pregnancies were distributed as 44 singletons, 13 pairs of twins, one set of triplets and one set of quadruplets. The Caesarian section rates were 21 and 40% for single and multiple pregnancies, respectively. The prematurity rate was low for singletons (4.5%) but reached 46.6% in multiple pregnancies. Two minor malformations were observed and five perinatal deaths occurred; among these latter, four cases were twins. It appears that perinatal pathology is substantially higher among IVF pregnancies than in the normal population. It is clear, however, that most of this difference is accounted for by the considerably higher rate of twin pregnancy displayed by the former group.

STUDY QUESTION What is the effect of a legal limitation of the number of embryos that can be transferred in an assisted reproductive technology (ART) cycle on the multiple delivery rate? SUMMARY ANSWER The Belgian national register shows that the introduction of reimbursement of ART laboratory costs in July 2003, and the imposition of a legal limitation of the number of embryos transferred in the same year, were associated with a >50% reduction of the multiple pregnancy rate from 27 to 11% between 2003 and the last assessment in 2010, without any reduction of the pregnancy rate per cycle. WHAT IS KNOWN ALREADY Individual Belgian IVF centres have published their results since the implementation of the law, and these show a decrease in the multiple pregnancy rate on a centre by centre basis. However, the overall national picture remains unpublished. STUDY DESIGN, SIZE, DURATION Cohort study from 1990 to 2010 of all ART cycles in Belgium (2685 cycles in 1990 evolving to 19 110 cycles in 2010), with a retrospective analysis from 1990 to 2000 and prospective online data collection since 2001. PARTICIPANTS/MATERIALS, SETTING, METHODS Registration evolved from paper written reports per centre to a compulsory online registration of all ART cycles. From 2001 up to mid-2009, data were collected from Excel spread sheets or MS Access files into an MS Access database. Since mid-2009, data collection is done via a remote and secured web-based system (www.belrap.be) where centres can upload their data and get immediate feedback about missing data, errors and inconsistencies. MAIN RESULTS AND THE ROLE OF CHANCE National Belgian registration data show that reimbursement of IVF laboratory costs in July 2003, coupled to a legal limitation in the number of embryos transferred in utero, were associated with a 50% reduction of the multiple pregnancy rate from 27 to 11% without reduction of the pregnancy rate per cycle, and with an increase in the number of fresh and frozen ART cycles due to improved access to treatment. LIMITATIONS, REASONS FOR CAUTION There is potential underreporting of complications of ART treatment, pregnancy outcome and neonatal health. WIDER IMPLICATIONS OF THE FINDINGS Over the 20 years of registration, the pregnancy rate has remained constant, despite the reduction in the number of embryos transferred, optimization of laboratory procedures and stimulation protocols, introduction of quality systems and implementation of the EU Tissue Directive over the period 2004–2010. STUDY FUNDING/COMPETING INTEREST(S) No external funding was sought for this study. None of the authors has any conflict of interest to declare. [ABSTRACT FROM AUTHOR]

Partially or completely hatched blastocysts were obtained from women with at least one previous cycle with failure of implantation. On day 5 after fresh embryo transfers, the remaining vitrification cycles (VC) were divided into three categories according to the types of blastocysts. The first group (20 VC) contained blastocysts (n = 38) with open zona pellucida (ZP), the second group of 34 VC contained blastocysts (n = 100) with both open and intact ZP, while the third group (14 VC) contained blastocysts with intact ZP (n = 39). Blastocysts were exposed to two mixtures of cryoprotectants. At 24 h after warming, survival rates of 82% (31/38), 72% (72/100) and 64% (25/39) were observed in the three groups respectively. Numbers of embryo transfers and blastocysts in the three groups respectively were 20, 31 and 13 transfers, and 31, 60 and 25 blastocysts, resulting in corresponding ongoing pregnancy rates per VC of 35, 26 and 21%. Blastocysts with a larger blastocoelic cavity survived vitrification better when they had partially or completely hatched. Survival rates significantly increased (P <0.01) from 55 to 81% after warming expanded blastocysts with an intact ZP (n = 42) compared with an open ZP (n = 54) respectively. This study shows that partially or completely hatched blastocysts can be cryopreserved by a simple vitrification procedure using the hemi-straw as embryo carrier. [ABSTRACT FROM AUTHOR]

Thirty-eight single and 10 multiple pregnancies obtained after in-vitro fertilization were compared. In the group of multiple gestations, maternal age was lower and the amounts of ovulatory drugs given were significantly smaller than in relation to single pregnancies. All multiple pregnancies arose from triple embryo transfers and the embryos from this group exhibited significantly higher vitality scores. In both groups, plasma levels of oestradiol and progesterone followed the same pattern until day 8 after oocyte retrieval. Following implantation, the secretion of these hormones increased more rapidly in multiple pregnancies pointing at greater luteal activity in this group. HCG levels became significantly higher in multiple gestation on day 25 after oocyte collection. Echographic examination showed that, compared to normal pregnancy, growth in both groups of IVF conceptuses was initially retarded but caught up with normal evolution at approximately 30 days after egg retrieval. The need for adjusting the number of embryos transferred not only to expected success rates but also to the risk of high rank multiples is emphasized. [ABSTRACT FROM AUTHOR]

A total of 740 cycles of intracytoplasmic sperm injection (ICSI) were performed: 625 cycles when < 6 x 10(5) total motile spermatozoa were harvestable from the ejaculate and 115 cycles in cases with a history of previous fertilization failure after classic in-vitro fertilization or subzonal sperm injection. An average of two pronuclei were observed in 63% of the injected oocytes, allowing 725 transfers of a maximum of three embryos (98%). Of 214 pregnancies initiated, 179 were established (25% of ICSI attempts). Because the fertilization rates from our initial 80 ICSI cycles were 2-fold less than those achieved previously, we changed the injection procedure and analysed, in 740 ICSI attempts, the importance of interfering technical factors and how to establish a successful ICSI programme. A remarkable change in the fertilization rate up to 68% (595 cycles) occurred when two steps in the injection procedure were performed well, i.e. immobilization of the spermatozoon and placement of the spermatozoon into the ooplasm after cytoplasmic aspiration into the pipette until oolema rupture. This immobilization, by touching the tail with the pipette, is mandatory for increasing the percentage of fertilization, even with totally non-motile spermatozoa (41%). Because aspiration of the cytoplasm is an invasive part of the ICSI procedure and influences the quality of the embryos, it is essential to reduce the amount of cytoplasm drawn into the pipette. This could be attained by using a spikeless injection pipette with the smallest possible internal diameter. [ABSTRACT FROM AUTHOR]

The aim of this study was to determine whether the presence of a hydrosalpinx influences in-vitro fertilization (IVF) prognosis. Comparisons were made between 69 IVF cycles in 37 patients carrying hydrosalpinges (hydrosalpinx group) and 67 IVF cycles in 41 patients without tubes or surgically sterilized (control group). Twenty-two patients carrying hydrosalpinges underwent salpingectomy or salpingoplasty (operated group); they then underwent 42 IVF trials which were compared with the two former groups. In the hydrosalpinx group, pregnancy rates by oocyte retrieval were 10.1% for clinical and ongoing pregnancies. In the control group, the corresponding pregnancy rates were 23.0 and 21.3% respectively. The implantation rate per embryo was 4.2% for clinical and ongoing pregnancies in the hydrosalpinx group and 11.0 and 10.4% respectively in the control group. The operated group had pregnancy rates of 38.1% for clinical pregnancies and 31.0% for ongoing pregnancies, with implantation rates of 17.4 (clinical) and 14.8% (ongoing) respectively. Pregnancy and implantation rates were statistically lower in the hydrosalpinx group as compared with controls and with the operated group. The differences between control and operated groups were not significant. In conclusion, the presence of a hydrosalpinx is thus associated with poor prognosis in IVF treatment. Surgical correction of such lesions appears to restore high success rates. [ABSTRACT FROM AUTHOR]

Abstract: Vitrification is a cryopreservation strategy where cells are converted into a glass-like amorphous solid which is free of any crystalline structure. Such process is achieved by a combination of high concentration of cryoprotectant and an extremely high cooling rate. In the last years, survival rates of up to 80% after thawing and pregnancy rates of almost 30% could be achieved after transfer of vitrified embryos at the zygote, cleavage, morula and blastocyst stages. Also deliveries of healthy babies have been reported numerous times. To this day, a limited interest in this technique can be noted. The explanation may lye in the apprehension of many ART units regarding exposure of embryos to high concentrations of cryoprotectants and storage in non sterile conditions. The aim of the first part of this article, is to analyse if such fears are justified on the basis that vitrification mimics conditions already in use for many years in slow-cooling procedures where cells are plunged into liquid nitrogen at around –30 °C and secondly since storage of embryos are now possible in high aseptic conditions. In the second part, results on survival after thawing, pregnancy rates and baby take home rates of vitrified embryos will be presented and the problems associated with vitrification of blastocysts will be discussed. [Copyright &y& Elsevier]