Prognostic factors for the success rate of embryo freezing

To find some prognostic factors for the outcome of frozen-thawed cycles, we have retrospectively analysed all frozen pre-embryos that were thawed during 1993 and 1994 at two in-vitro fertilization (IVF) units in Sweden. Supernumerary pre-embryos were frozen from 551 oocyte retrievals and these resulted in 660 frozen-thawed cycles which lead to 623 thawed embryo transfers. The outcome of these transfers was 137 clinical pregnancies with a pregnancy rate of 22% per frozen-thawed embryo transfers. Women <40 years of age had a higher birth rate than those > or =40 years, 19 and 5% respectively (P < 0.01). Transfers with two and three pre-embryos resulted in pregnancy rates of 23 and 27%, respectively, compared with 14% for transfer of one embryo. A pregnancy resulting from the initial embryo transfers had a predictive value for results of the subsequent frozen-thawed cycle. Embryo grade and cleavage stage at the time of freezing was important for the survival of the frozen-thawed pre-embryos. The pregnancy rate was not influenced by the cleavage stage, but a tendency toward a lower pregnancy rate was seen for the embryos with lower grading. To conclude, cryopreservation seems to be beneficial in women <40 years of age, who have supernumerary pre-embryos of good quality for freezing and of which at least two can be transferred.      

Elective transfer of one embryo results in an acceptable pregnancy rate and eliminates the risk of multiple birth.

To avoid multiple pregnancies without compromising pregnancy rates (PR) is a challenge in assisted reproduction. We have compared pregnancy results among 74 elective one-embryo transfers (group 2) and 94 transfers where only one embryo was available (group 1). All the fresh embryo cycles during 1997 in two clinics in Helsinki were analysed, and cumulative PR among these couples after frozen-thawed embryo transfers up to June 1998 were counted. In group 2, where at least two embryos were available for transfer, and only one was transferred on day 2 or 3, the PR per embryo transfer was 29.7%. In group 1, the PR per embryo transfer was 20.2%. In group 2, the cumulative PR after frozen-thawed embryo transfers was 47.3% per oocyte retrieval. Over the same time, 742 two-embryo transfers were carried out. The PR per embryo transfer was 29.4% in these subjects, but 23.9% of these pregnancies were twins. The implantation rates, as well as the PR, were highest when the embryos were at the four- to five-cell stage on day 2 (35.8 versus 9.7% compared with the two- to three-cell stage, P < 0.001) or at the six- to eight-cell stage on day 3 (45.5%). The PR per embryo transfer was higher when a grade 1 or 2 embryo was transferred compared with a grade three embryo (34. 0 and 26.7% versus 8.8% respectively, P < 0.05). In women 35 years or younger, the PR per elective one-embryo transfer was 32.8%. The corresponding figure in women older than 35 years was 18.8%. On the basis of these results, elective one-embryo transfer can be highly recommended, at least in subjects who are younger than 35 years of age, and who have grade one or grade two embryos available for transfer.     

Turner's syndrome and pregnancies after oocyte donation

A total of 20 clinical pregnancies was achieved among 18 women with Turner's syndrome who were treated in an oocyte donation programme. The oocytes were donated by voluntary unpaid donors. A mean of 1.8 embryos per transfer was given to each recipient by way of 28 fresh and 25 frozen embryo transfers. With fresh and frozen embryos, 13 and seven pregnancies respectively were achieved. The clinical pregnancy rate per fresh embryo transfer was 46%, and the implantation rate 30%, being similar to the corresponding rates among our oocyte recipients with primary ovarian failure in general. The corresponding rates with frozen embryos were 28 and 19%. Of these pregnancies, 40% ended in miscarriage. This high rate may be explained by uterine factors. Six women were hypertensive during pregnancy, a rate comparable with that in other oocyte donation pregnancies. All these women delivered by Caesarean section. Pregnancy and implantation rates after oocyte donation were high in women with Turner's syndrome, but the risk of cardiovascular and other complications is high. Careful assessment before and during follow-up of pregnancy are important. Transfer of only one embryo at a time to avoid the additional complications caused by twin pregnancy is recommended.     

The contribution of embryo cryopreservation to in-vitro fertilization/gamete intra-Fallopian transfer: 8 years experience

In this paper, the authors summarized their experience withembryo cryopreservation over an 8-year period. The results,therefore, reflect the long-term benefit of embryo cryo-preservationto the overall in-vitro fertilization/gamete intra-Fallopiantransfer (IVF/GIFT) programme and to the women who had embryoscryopreserved. The stable survival rate of thawed embryos andpregnancy rate, especially over the past 4 years, suggests thatthe results can reliably be used to evaluate the efficacy ofthe embryo cryopreservation programme. The ongoing pregnancyrate of frozen/thawed embryo transfer is 10.9%, comparable withthe ongoing pregnancy rate of fresh IVF/embryo transfer in ourunit over the same period. In addition to those factors knownto affect the pregnancy rate in fresh IVF/GIFT cycles, suchas age of the recipients and number of embryos transferred,the major factor affecting the efficacy of the cryopreservationprogramme is the number of oocytes retrieved in the initialstimulation cycle, and the number of embryos available for cryopreservation.The storage time of cryopreserved embryos will also have a significanteffect on the realization of the total potential of embryo cryopreservation.Overall the contribution of cryopreservation to our IVF/GIFTprogramme is substantial, increasing pregnancy rate by 4%, whilethe greater net benefit, of course, is for the women who hadembryos cryopreserved (pregnancy rate increased by 7%), especiallyfor those who returned for frozen/thawed embryo transfer cycles(pregnancy rate increased by 11%).     

Pregnancies achieved after frozen-thawed pronuclear oocytes obtained by intracytoplasmic sperm injection with spermatozoa extracted from frozen-thawed testicular tissues from non-obstructive azoospermic men.

The use of frozen-thawed testicular tissue as a source of spermatozoa for intracytoplasmic sperm injection (ICSI) in non-obstructive azoospermia yields favourable fertilization and pregnancy rates while avoiding both repetitive biopsies and unexpected cycle cancellations. Spermatozoa were obtained from frozen-thawed testicular biopsy specimens from 67 non-obstructive azoospermic men. Following fertilization, supernumerary two pronuclear (2PN) oocytes were frozen. After thawing, 17 cycles of embryo transfer were carried out with a mean number of 2.7 embryos and a mean cumulative embryo score (CES) of 18.3 per transfer. The clinical pregnancy and implantation rates per transfer in these cycles (23.5 and 8.3% respectively) were comparable to those of fresh embryo transfers (35.7 and 12.7% respectively) with a mean number of 2.7 embryos and a mean CES of 28.7 per transfer. Abortion rates, although higher with cryopreserved 2PN oocytes were not significantly different. With this approach, cryopreservation of supernumerary 2PN oocytes can be used to improve the cumulative pregnancy rates in a severely defective spermatogenetic population. To our knowledge, these are the first pregnancies reported which have been obtained by the transfer of cryopreserved pronuclear oocytes obtained from ICSI using cryopreserved testicular spermatozoa.     

Cryopreservation of supernumerary oocytes in IVF/ICSI cycles

BACKGROUND: The aim of the present study is to investigate cryopreservation of oocytes in patients refusing embryo cryopreservation for ethical reasons, patients from whom no sperm could be retrieved and patients with enough oocytes to yield a number of fresh and cryopreserved embryos to transfer. METHODS: A total of 2900 oocytes out of 6216 retrieved were cryopreserved in 286 patients undergoing 303 cycles. The reasons for cryopreservation were because no sperm was found in 16 cycles, for ethical or personal reasons in 80, and in 207 only supernumerary oocytes were frozen. In 159 cycles, the oocytes were thawed and the surviving metaphase II oocytes microinjected. RESULTS: A total of 1087 oocytes were thawed, 760 (69.9%) survived and 687 were microinjected. We obtained 368 (53.5%) normally cleaved embryos, 331 were transferred and 37 were cryopreserved. One hundred and forty-five transfers (range 1-3 embryos/patient) were performed and 18 (12.4%) pregnancies were obtained. Twelve patients delivered 13 healthy children, and six first trimester abortions were observed (33.3%). CONCLUSION: Although a low implantation rate was observed and a higher abortion rate than in fresh cycles, our results show that in sibling oocytes, the process of cryopreservation apparently does not affect the fertilization and cleavage rate. In this group of patients, producing a large number of mature gametes, oocyte cryopreservation gives the couple extra chances to achieve a pregnancy within a single retrieval and is a good effort towards reducing the number of embryos cryopreserved and enhancing our experience in this new technology.     

Impact of patients' choice for single embryo transfer of a top quality embryo versus double embryo transfer in the first IVF/ICSI cycle

BACKGROUND: The aim of this study was to evaluate the impact of transferring a single top quality embryo in the first IVF/ICSI cycle of patients <38 years old who chose to have one or two embryos transferred. METHODS: A total of 262 patients participated in the study, and 243 transfers were performed: 156 (64%) patients chose the transfer of a single top quality embryo, if available, and two non-top quality embryos if not available; 87 (36%) patients chose to have a double embryo transfer regardless of embryo quality. RESULTS: In the first group an ongoing pregnancy rate of 40% (63/156) with a twin pregnancy rate of 2% (1/63) was achieved. In the second group the ongoing pregnancy rate was 44% (38/87) with 26% (10/38) twin pregnancies. In the patient group with only one embryo transferred, irrespective of the patient's choice, the ongoing pregnancy rate was 43% (54/127) with no twin pregnancies. For the study population as a whole, the ongoing pregnancy rate was 42% (101/243) with 11% (11/101) twins. CONCLUSION: We conclude that the introduction of single embryo transfer in the first IVF/ICSI cycle is highly acceptable in women <38 years old.     

血LH峰与尿LH峰测定判断冻融胚胎移植时机的方法学比较

目的研究利用电化学发光免疫分析技术监测血LH板或利用尿LH板监测尿LH峰判断冻融胚胎移植时机的方法学比较。方法本中心由1999年5月至今共完成冻融胚胎移植67例,排除冻融胚胎移植周期中所移植的胚胎全部为部分卵裂球受损胚胎的周期后,将其中61例移植周期随机分成A、B两组,A组共12周期,通过使用尿LH板测定尿LH峰来判断冻融胚胎移植时机,B组共49个周期,通过电化学发光免疫分析技术测定血LH峰来判断冻融胚胎移植时机,对两组资料进行回顾性分析。结果两组患者平均冻融胚胎移植数目、冷冻前胚胎形态学评分及解冻后胚胎卵裂球存活程度等均无显著差异(P>0.05),冻融胚胎移植周期的子宫内膜厚度均大于8mm。利用尿LH峰测定判断冻融胚胎移植时机的12周期中,移植冻融胚胎37个,平均每周期移植胚胎3.1±0.4个,周期临床妊娠率33.3%(4/12)。利用血LH峰测定判断冻融胚胎移植时机行冻融胚胎移植的46例49个周期中,移植冻融胚胎137个,平均每周期移植胚胎2.8±0.1个,周期临床妊娠率63.3%(31/49),两组资料临床妊娠率有非常显著性差异(P<0.01)。结论利用电化学发光免疫分析技术监测血LH峰判断冻融胚胎移植时机比利用尿LH板监测尿LH峰判断冻融胚胎移植时机更为准确和灵敏,显著提高临床妊娠率。     

Day 2 elective single embryo transfer in clinical practice: better outcome in ICSI cycles

BACKGROUND: Single embryo transfer (SET) after IVF/ICSI has been shown to result in an acceptable pregnancy rate in selected subjects. In our unit, SET is routinely carried out among women under the age of 36 in the first or second treatment cycle when a top-quality embryo is available. In order to define further the selection criteria for SET, we have analysed the outcome of elective SET (eSET), including the cumulative pregnancy rate after frozen embryo transfers, performed in the years 2000-2002 in the Oulu Fertility Center. METHODS: During the study period, a total of 1271 transfers were performed, and in 468 cycles SET (39% of all transfers) was carried out. Of the SET cycles, in 308 cases a top-quality embryo was transferred on day 2 and extra embryos were frozen. Of these eSET cycles, ICSI was carried out in 87 cycles (28%). RESULTS: The overall clinical pregnancy rate per transfer was 34.7% in the eSET cycles. In the eSET ICSI cycles, the clinical pregnancy rate was significantly higher than in the corresponding IVF cycles (50.6 versus 28.5%, P < 0.001). The cumulative pregnancy rate per patient after fresh and frozen embryo transfers was also significantly higher after ICSI (71.2 versus 53.4%, P < 0.01). CONCLUSIONS: A high cumulative pregnancy rate per oocyte retrieval can be achieved after eSET in daily clinical practice. The implantation rate of fresh top-quality embryos in the ICSI cycles was significantly higher than in the IVF cycles, possibly due to more successful selection of the embryo for embryo transfer on day 2 after ICSI. In addition, our data suggest that embryo quality is a more important determinant of outcome than the age of the woman.     

Should diagnostic testicular sperm retrieval followed by cryopreservation for later ICSI be the procedure of choice for all patients with non-obstructive azoospermia?

BACKGROUND: This was a retrospective study to determine if diagnostic testicular biopsy followed by cryopreservation should be the procedure of choice for all patients with testicular failure. METHODS: The first part of the study analysed 97 ICSI cycles scheduled with frozen-thawed testicular sperm for 69 non-obstructive azoospermia (NOA) patients. The second part focused on a subgroup of 32 patients who underwent 42 ICSI cycles with frozen and 44 cycles with fresh testicular sperm. Sperm characteristics, fertilization, embryo quality, pregnancy and implantation rates were evaluated. RESULTS: Part I: The average time needed to find sperm was 113 min per cycle and 17 min per individual sperm. Fertilization rate, embryo transfer rate, ongoing pregnancy and implantation rates were 58.4%, 83%, 20.8% and 11.3%, respectively. Part II: The search time per sperm was higher (P=0.016) in frozen (18 min) than in fresh suspensions (13 min). A higher embryo transfer rate was observed in fresh cycles than in frozen cycles (93.2% vs 76.2%, P=0.028). Fertilization, ongoing pregnancy and implantation rates were comparable for the two groups. CONCLUSIONS: Even in a programme with low-restrictive criteria for patient allocation and for sperm cryopreservation, diagnostic testicular biopsy followed by cryopreservation can be the procedure of choice for patients with testicular failure.     

Impact of elective single embryo transfer on the twin pregnancy rate

BACKGROUND: It is unclear how the implementation of elective single embryo transfer in clinical practice would affect clinical pregnancy and delivery rates and multiple birth rates. METHODS: This retrospective study analysed 1871 IVF/ICSI cycles carried out from 1997 to 2001 in the IVF programme of a single university infertility clinic. RESULTS: The number of elective single embryo transfers increased from 11 to 56%. At the same time the clinical pregnancy rate was relatively stable; mean 34.0% (range 28-42). The number of embryos per embryo transfer decreased from 1.8 to 1.3. The multiple pregnancy and delivery rates dropped markedly from 25 to 7.5% and from 25 to 5% respectively. CONCLUSIONS: An elective single embryo transfer programme can be adopted in daily practice that decreases the twinning rate to <10% and does not affect the overall pregnancy rate.     

Endocrinology: Long-term evaluation of implantation of fresh and cryopreserved human embryos following ovarian stimulation with buserelin acetate-human menopausal gonadotrophin (HMG) or clomiphene citrate-HMG

This study is a long-term evaluation of the total pregnancypotential of cohorts of fresh and cryopreserved sibling embryosfrom in-vitro fertilization (TVF) cycles stimulated with eitherthe gonadotrophin-releasing hormone analogue buserelin (BUS)(long protocol) or clomiphene citrate (CC) both in combinationwith human menopausal gonadotrophin (HMG). Therefore a retrospectiveanalysis was performed on patients who entered the FVT programmebetween January 1986 and July 1987 and who had triple embryotransfer in the collection cycle. Significantly more fertilizedoocytes developed to good-quality embryos in the CC-HMG group(86.1%) than in the BUS-HMG group (80.8%). Transfer of the threemorphologically best-looking embryos was performed on day 2post-insemination in 106 CC-HMG and 80 BUS-HMG cycles. Supernumeraryembryos were cultured for a further 24 h and multicellular embryoswith up to 20% of fragments were frozen slowly with 1.5 M dimethylsulphoxideon day 3 post-insemination (162 embryos in CC-HMG cycles, 102embryos in BUS-HMG cycles). Outcome was measured by embryo survivalrate, embryo implantation rate and delivery rate in fresh andfrozen embryo transfers. Delivery rates were 31.3 and 21.7%per fresh embryo transfer in BUS-HMG and CC-HMG cycles respectively.Fresh embryo implantation rates were significantly higher incollection cycles stimulated with BUS-HMG (17.9%) than in cyclesstimulated with CC-HMG (113%). Implantation rates were significantlyenhanced in embryos transferred in excess of one in cycles leadingto pregnancy, perhaps indicative of higher embryo quality inBUS-HMG cycles. Almost all cryopreserved embryos have by nowbeen thawed, so the contribution of frozen embryos to overallpregnancy rates can be evaluated. Overall morphological survivalrates of frozen-thawed embryos were similar for 140 embryosfrom CC-HMG cycles (50%) and 100 embryos from BUS-HMG cycles(46%). The percentage of fully intact embryos was, however,significantly lower in the BUS-HMG group (19%) than in the CC-HMGgroup (39.5%). Delivery rates were significantly lower following30 transfers of frozen-thawed embryos from BUS-HMG-stimulatedcycles (3.3%) than following 42 transfers of frozen-thawed embryosfrom CC-HMG cycles (19.1%). Embryo implantation rates were lowerfor frozen-thawed embryos from BUS-HMG cycles (23%) than fromCC-HMG cycles (12.7%). Here we demonstrate that ovarian stimulationwith the long protocol BUS-HMG instead of the CC-HMG protocolled to higher embryo implantation rates in collection cyclesbut to lower intact embryo survival rates and to lower embryoimplantation rates for frozen sibling embryos. Despite the lowerimplantation rates with frozen embryos originating from theBUS-HMG protocol, there was no significant difference betweentotal delivery rate per transfer from cycles stimulated withCC-HMG (30.2%) compared with BUS-HMG (33.8%).     

Elective single day 3 embryo transfer halves the twinning rate without decrease in the ongoing pregnancy rate of an IVF/ICSI programme

BACKGROUND: Data on the effect of elective single embryo transfer (eSET) on the total and multiple pregnancy rates of an IVF/ICSI programme are reported. METHODS AND RESULTS: A retrospective cohort analysis of eSET was carried out over a 4 year period. A total of 1559 cycles resulted in 1464 transfers; 299 transfers of one top quality embryo (20.4%) and 86 of one non-top quality embryo (5.9%) yielded 149 conceptions (49.8%) with 105 ongoing pregnancies (35.1%) and 26 conceptions (30.2%) with 19 ongoing implantations (22.1%) respectively; 1079 transfers of two (n = 853; 58.3%) or more than two (n = 226; 15.4%) embryos yielded 366 ongoing pregnancies (33.9%). The ongoing pregnancy rates for the years between 1998 and 2001 were 35.9, 27.9, 31.9 and 31.0% per oocyte retrieval and 38.5, 29.4, 34.1 and 33.2% per transfer. There were no differences in pregnancy rates between any of the years. The average ongoing pregnancy rate (>12 weeks) over the 4 years was 31.5% per started cycle and 33.5% per transfer; the average number of embryos transferred decreased from 2.26 (1998) to 1.79 (2001); the multiple pregnancy and twinning rates dropped from 33.6 and 29.5% (1998) to 18.6 and 16.3% (2001) respectively. CONCLUSIONS: Judicious application of eSET can halve the twinning rate while maintaining the overall pregnancy rate.     

Frozen embryo transfers: implications of clinical and embryological factors on the pregnancy outcome

BACKGROUND: Frozen embryo transfers are characterized by impaired pregnancy outcome and increased incidence of pregnancy loss as compared with fresh IVF/ICSI embryo transfers. In this study, we performed a retrospective analysis of clinical and embryological factors that potentially influence the outcome of frozen embryo transfer. METHODS: We reviewed the outcome of 1242 frozen embryo transfers with respect to the age of the woman, the method of fertilization, embryo quality before and after freezing and the number of embryos transferred. RESULTS AND CONCLUSIONS: The pregnancy (positive hCG) and clinical pregnancy rates were 25.8 and 21.1%, respectively. A total of 107 (33.3%) of the 321 pregnancies identified by a positive hCG test miscarried either before (18.4%) or after (15%) the clinical recognition of gestational sac(s). The delivery rate for the frozen embryo transfers analysed was 17.2%. Our data revealed that the delivery rate after frozen embryo transfer was dependent on both the woman's age and the quality of embryos transferred, at the same time being unaffected by IVF/ICSI treatment. In addition, the increased woman's age at IVF/ICSI treatment was identified as the only parameter elevating the biochemical pregnancy rate, whereas the clinical abortion rate was found to be unrelated to the clinical or embryological parameters studied.     

Effect of developmental stage of embryo at freezing on pregnancy outcome of frozen-thawed embryo transfer

BACKGROUND: The study aim was to investigate the impact of the developmental stage of embryos on pregnancy outcome of frozen embryo transfer (FET). METHODS: The survival rates of embryos after thawing and pregnancy outcome following FET were compared retrospectively between three cryopreservation strategies utilizing either zygote, day 2 or day 3 embryo freezing. RESULTS: A total of 4006 embryos was analysed in 1657 thaw cycles. The highest (P < 0.0001) survival rate (all cells survived) was observed for zygotes (86.5%), followed by day 2 (61.7%) and day 3 (43.1%) embryos. FET was performed in 1586 (95.7%) of all thaw cycles, resulting in overall clinical pregnancy and implantation rates of 20.7 and 14.2% respectively. The delivery rate per transfer was 16.5%, and live birth rate per transferred embryo 11%. There were no significant differences in clinical pregnancy, implantation, delivery and birth rates between frozen zygote, day 2 and 3 embryo transfers. However, an elevated miscarriage rate was observed in the day 3 group (45%) compared with zygotes (21.3%; P = 0.049) and day 2 embryos (18.3%; P = 0.004). The overall efficacy of FET (birth rate per thawed embryo) was 7.3%. The efficacy was lower in day 3 group (4.2%) than in the zygote (7.1%; P = 0.082) and day 2 (7.6%; P = 0.027) groups. CONCLUSIONS: The developmental stage of embryos at freezing has a profound effect on their post-thaw survival, but seems to have little effect on rates of clinical pregnancy, implantation, delivery and birth after FET. The elevated miscarriage rate for day 3 frozen embryo transfers may be caused by damage during freeze-thaw procedures. The low survival rate and elevated miscarriage rate were both responsible for a reduced overall efficacy for day 3 FET when compared with zygotes and day 2 embryos.     

One year's experience with elective transfer of two good quality embryos in the human in-vitro fertilization and intracytoplasmic sperm injection programmes

High incidences of multiple pregnancies, after transferringa maximum of three embryos, were observed after in-vitro fertilization(IVF) treatment. In a randomized study, it was demonstratedthat, after taking into account embryo quality and other positivelyinterfering parameters, an elective transfer of two good qualityembryos does not significantly influence the pregnancy rate.The intracytoplasmic sperm injection (ICSI) technique was successfullydeveloped in the meantime and high incidences of multiple pregnancieswere also obtained after ICSI. The question arose whether afterICSI there was also room for elective double embryo transferin a well-defined patient group. This report covers 1 year of IVF and ICSI treatment and theresults are presented in relation to the number of embryos transferred.The embryo development is similar for zygotes obtained afterIVF and ICSI; for both techniques 63% of the zygotes developto type A-B embryos and 13% to type C embryos. There is alsono difference in the pregnancy rate after ICSI or IVF. Globally,after IVF, 307 out of the 766 double and triple transfers (40.1%)and 317 out of 774 double and triple transfers (40.9%) afterICSI resulted in a positive HCG. After IVF, 73.9% (227) andafter ICSI 76.3% (242) of the pregnancies were evolutive. Neitherwas there any difference between the two techniques as regardsthe implantation rate per transferred embryo. After IVF, 22.8%of the transferred embryos implanted compared with 21.8% afterICSI. When the elective double embryo transfers were compared,no difference was found between IVF and ICSI. After IVF, 102of the 211 elective double transfers (48.1%) resulted in a pregnancyversus 93 out of 225 (41.3%) after ICSI [not significant (NS)].A high implantation rate per transferred embryo (IVF: 33.2%;ICSI: 26.9%, NS) was obtained in this elective double transfercategory, as was also reported in the randomized study. Thesedata confirm the results obtained in our randomized study andthe effectiveness of the elective double embryo transfer forIVF as well as for ICSI.     

A prospective randomized study to assess the benefit of partial zona pellucida digestion before frozen-thawed embryo transfers

BACKGROUND: Assisted hatching (AH) in fresh embryo transfer (ET) could be associated with increased implantation rates. However, very few prospective randomized studies have specifically addressed the issue of AH during frozen-thawed embryo transfers (FET) cycles, those that have reported controversial results. The aim of this study was to evaluate the benefit of an enzymatic zona pellucida treatment of frozen-thawed embryos before transfer. METHODS: This was a prospective study including 125 non-donor FET cycles from 125 infertile couples. FETs were randomly allocated into AH group (n = 61, embryos pretreated with pronase 5 IU/ml for 1 min at 37 degrees C) or control group (n = 64, untreated embryos). Zona pellucida thickness was measured for each transferred embryo. The main outcome parameters were clinical pregnancy and implantation rates. RESULTS: The two groups were comparable regarding mean women's age, duration and indications of infertility, IVF outcome after fresh ETs, numbers and quality of fresh and frozen embryos, frozen-thawed embryo survival rates and blastomeres survival indexes. Despite a statistically significant decrease of zona pellucida thickness after pronase treatment [(mean +/- SD) 18.5 +/- 2.25 versus 14.5 +/- 2.75 microm; P < 0.0001], implantation (9.6 versus 9.2%) and clinical pregnancy rates (18.0 versus 17.2%) were not statistically different after FETs, with a similar mean number of embryos transferred between AH and control groups, respectively. CONCLUSION: Within the constraints of our protocol, partial enzymatic digestion of zona pellucida by pronase was not related with any benefit of the FET outcome especially concerning the implantation ability of frozen-thawed embryos.     

Resumption of mitosis during post-thaw culture: a key parameter in selecting the right embryos for transfer

This retrospective study of 701 thaw cycles analysed the clinical importance of whether or not embryos resumed mitosis during 24 h of post-thaw culture. A total of 3360 frozen embryos were thawed; 1922 embryos survived the freeze-thaw procedure with at least one intact blastomere and were then cultured for 24 h before transfer. All transfers were registered into either the 'cleaved embryo group' (n = 459), which was defined as transfers where at least one of the transferred embryos cleaved during the post-thaw culture period, or the 'non-cleaved embryo group' (n = 153), where none of the transferred embryos cleaved during the post-thaw culture period. A total of 1408 thawed embryos were transferred in 612 cycles; 459 embryo transfers were in the cleaved embryo group, resulting in an implantation rate of 10%, significantly higher than the 4% in the non-cleaved embryo group (P = 0.0003). A total of 130 pregnancies (28% per transfer) were obtained in the cleaved embryo group which was significantly higher than the 17 pregnancies (11% per transfer) obtained in the non-cleaved embryo group (P = 0.0001). However, the average number of transferred embryos was significantly higher in the cleaved embryo group (2.46 +/- 0.03) compared to the non-cleaved embryo group (1.82 +/- 0.07). No difference was found in the age of the women between the two groups. When analysing transfers where all transferred embryos had cleaved during the post-thaw culture period the clinical pregnancy rate increased significantly from 13% transferring two embryos to 36% transferring three embryos (P = 0.0136). In this latter subgroup an implantation rate as high as 17% was obtained. The overall multiple pregnancy rate was 16%. The multiple pregnancy rate was 19% in the cleaved embryo group. In conclusion, 24 h post-thaw culture may allow a better selection of the embryos and thereby we may be able to increase the implantation and pregnancy rates. This may enable us further to reduce the number of embryos transferred.      

Transfers of frozen-thawed human embryos in cycles stimulated by HMG

A total of 130 transfers of frozen-thawed (F-T) human embryos was carried out after moderate ovarian stimulation with human menopausal gonadotrophin (HMG). Embryos were replaced 3 days after the spontaneous luteinizing hormone (LH) surge or 4 days if ovulation was induced by human chorionic gonadotrophin (HCG). Embryos were thawed a few hours prior to transfer. One-hundred-and-twenty-three transfers were effective and 23 pregnancies were achieved. The rate of ongoing pregnancies per transfer was 17.9% (22/123). The survival rate of embryos originating from cycles stimulated by a combination of an LHRH analogue and HMG in a long protocol (LA-HMG protocol) was significantly lower when compared with the rate of embryos retrieved from clomiphene citrate-HMG (CC-HMG protocol) stimulated cycles (52 versus 67%, P less than 0.05). When fresh embryos originated from cycles stimulated with an LHRH analogue and HMG in a short protocol (SA-HMG protocol), the survival rate was not affected (59 versus 67%, NS). Although the difference was not significant, the ongoing pregnancy rate per transfer according to the three protocols from which the embryos originated seemed to be better with the SA-HMG protocol: 16% with the CC-HMG protocol, 14.5% with the LA-HMG protocol versus 27.6% with the SA-HMG protocol. The success rate was independent of the number of F-T transferred embryos if at least one embryo with 100% intact blastomeres was replaced.     

Human preimplantation embryo cryopreservation: selected aspects

This report describes the results of cryopreserving human preimplantation zygotes and cleaved embryos (2-4 cells) in our in-vitro fertilization programme. Cryopreserved zygotes and cleaved embryos resulted in similar post-thaw survival rates (74.8 versus 70.9%). Pregnancy rates per retrieval cycle (RC) and embryos transferred per pregnancy for frozen-thawed zygotes versus frozen-thawed cleaved embryos were 21.8 versus 11.5% (P less than 0.2) and 12.6 versus 17.5 (P less than 0.2), respectively. Pregnancy rates increased significantly for both fresh (P less than 0.0005) and frozen-thawed (P less than 0.05) embryos as the number of embryos replaced per transfer increased from one to three or more. Frozen-thawed embryos resulted in multiple implantation rates per transfer of 25 compared to 6.4% (P less than 0.1) for fresh embryos when two embryos were replaced. Pregnancy rates were reduced for fresh (P less than 0.05) and frozen-thawed (P less than 0.1) embryos obtained from patient retrieval cycle numbers greater than 3. The method of follicular stimulation during the retrieval cycle did not affect frozen-thawed embryo survival rates. There was no difference in pregnancy rates from frozen-thawed embryos replaced during natural or clomiphene citrate transfer cycles. Patients with cryopreserved embryos had cumulative pregnancy rates of 37.1% (66/178) compared to 23.5% (110/468) (P less than 0.01) for patients with no embryos cryopreserved; cryopreservation of preimplantation embryos is a reliable therapeutic procedure that enhances achievement of pregnancy through in-vitro fertilization.