Improved implantation rate after chemical removal of the zona pellucida

OBJECTIVE: To evaluate the implantation rate achieved after chemical removal of the zona pellucida from day 5 human in vitro-derived embryos. DESIGN: Prospective, randomized, controlled study. SETTING: A tertiary care infertility clinic. PATIENT(S): Two hundred fifty-seven patients undergoing IVF with transfer of morulas or blastocysts on day 5 after oocyte retrieval. All patients had had at least two previous implantation failures. INTERVENTION(S): Chemical removal of zona pellucida by using acidic Tyrode's solution vs. no removal (controls). MAIN OUTCOME MEASURES: Clinical pregnancy rate and implantation rate per transfer. RESULT(S): Embryos without zona pellucida implanted at nearly twice the rate of control embryos (15.7% vs. 27.5%). The pregnancy rate was also significantly higher in the zona pellucida-free group than the control group (31.0% vs. 46.1%). Removal of zona pellucida was most effective in embryos with delayed development, which reached the morula or early cavitating stage on day 5 of in vitro culture (implantation rate, 12.1% vs. 25.7%). CONCLUSION(S): Chemical removal of zona pellucida from day 5 in vitro cultured human embryos is an effective and safe method of significantly improving the implantation rate, especially of embryos with delayed development.     

Morphology of vitrified/warmed day-5 embryos predicts rates of implantation,pregnancy and live birth

Although some post-thaw morphological predictors of pregnancy have been investigated in slow freezing of blastocysts, no such data have been published for vitrified and warmed blastocysts. Therefore, a prospective four-part score was applied to vitrified/warmed day-5 embryos to evaluate whether certain morphological parameters could serve as predictors of implantation, pregnancy and live birth. All morulae/blastocysts that were considered to be viable after warming were scored according to a previously unpublished grading system based on re-expansion, hatching (out of an artificial gap in the zona pellucida), extensive cytoplasmic granulation and presence of necrotic foci. Overall, 74% (202/273) of the vitrified concepti were found to be viable after warming. Early blastocysts showed better survival versus extended/hatching blastocysts (P < 0.01). Of the morphological parameters analysed, immediate re-expansion (P < 0.05) and hatching (P < 0.001) were positive predictors of the rates of implantation, pregnancy and live birth. The opposite holds for extensive cytoplasmic granulation (P < 0.05), which was negatively related. Accurate scoring of warmed blastocysts (within the first 2 h) allows for prediction of pregnancy outcome, and thus will help to further reduce the number of transferred embryos.     

Zona pellucida removal and vitrified blastocyst transfer outcome: a preliminary study

The objective of this study was to investigate whether a change in assisted hatching technique from partial opening to total removal of the zona pellucida improved the outcome of vitrified blastocyst transfer. This was a preliminary observational study conducted from November 2003 to April 2006. Partial opening using acid Tyrode's solution was performed in 45 cycles, while total removal using a laser and mechanical pipetting was performed in 57 cycles. The clinical pregnancy, implantation, and delivery rates were higher in the total removal group than in the partial opening group (67% versus 42%, P < 0.02; 55% versus 30%, P < 0.01; 56% versus 36%, P < 0.04, respectively). These results suggest that total removal of the zona pellucida is associated with higher pregnancy, implantation and delivery rates compared with partial opening for vitrified blastocyst transfer.     

Comparison of Zona Manipulated and Zona Intact Blastocyst Transfers: A Prospective Randomized Trial

Purpose: In this prospective randomized study the effectsof enzymatic treatment of zona pellucida of blastocysts onimplantation and pregnancy rates were evaluated in a groupof patients who had more than five embryos on day 3. Methods: Forty-six patients with a mean age of 29.8 ± 4.5years and mean duration of infertility of 6.72 ± 0.63 yearshad blastocyst stage transfers, with a mean number of2.9 ± 0.1 embryos replaced per patient. Patients were randomlydivided into two groups. The first group consisted ofpatients (n = 22) who had zona intact blastocyst stagetransfers and the second group consisted of patients (n = 24)who had zona manipulated (enzymatic treatment) blastocyststage transfers. Patient and cycle characteristics were similarin both groups. A commercial cell and serum-free sequentialculture system was used for all embryos. Results: Overall blastocyst formation rate was 50.3%.Transfer could be done in all patients. The positive -humanchorionic gonadotropin rate in the zona intact group was50% (11/22) and in the zona-manipulated group was 70.8%(17/24). Clinical pregnancy, ongoing pregnancy, andimplantation rates in zona intact and manipulated groupswere 45.5%, 27.3%, and 19%, and 62.5%, 45.8%, and 24%respectively. Although implantation and pregnancy rates inthe zona-manipulated group were higher, there were nostatistically significant differences in terms of these variablesbetween two groups. No triplet pregnancy was obtained ineither group, and the twin pregnancy rate was 20% (2/10)in the zona intact group and 13.3% (2/15) in thezonamanipulated group. Conclusions: With further improvements in the embryoculture systems it will become possible in the near future toachieve high implantation rates even with single blastocysttransfers. Enzymatic treatment of the zona pellucida seemsthat it does not alter the pregnancy and implantation rates,but further studies with larger group of patients are neededto clarify the real effect of this zona manipulation onpregnancy outcome.     

Chemical removal of zona pellucida versus laser assisted hatching after repeated failures of assisted reproductive technology

Background and Aims:  To evaluate outcomes after zona pellucida removal by pronase or laser assisted hatching in women with repeated assisted reproduction failures.
Methods:  Of 389 procedures (January 2004 to November 2005), 203 control cycles had an intact zona, 116 cycles had chemical removal of the zona and 70 cycles had laser assisted hatching. Rates of pregnancy, implantation and abortion were compared, and pregnancy rate was secondarily evaluated for fresh or frozen-thawed blastocysts.
Results:  Pregnancy rates were 33.5% (68/203) for controls, 29.3% (34/116) for chemical removal and 30.0% (21/70) for laser. Implantation rates were 24.8% (68/274) for controls, 21.8% (34/156) for chemical removal and 30.0% (21/105) for laser. There were no significant differences among groups. Abortion rates were 15.6% (10/64) for controls, 13.9% (5/36) for chemical removal and 14.3% (3/21) for laser. No difference was observed by blastocyst type for control or laser assisted hatching cycles. In the chemical removal group, both pregnancy and implantation rates were higher for frozen-thawed blastocysts than for fresh blastocysts. (41.5% vs 13.2% and 30.7% vs 11.1%, respectively).
Conclusions:  Assisted hatching did not show a significant benefit. Chemical zona pellucida removal might increase pregnancy rates for frozen-thawed blastocysts. (Reprod Med Biol 2006; 5: 263–267)     

Chemical zona pellucida thinning with acidified Tyrode’s solution: comparison between partial and circumferential techniques

Purpose To compare effectiveness of two different chemical zona thinning techniques. Method We studied 163 patients who had experienced IVF or ICSI failures in two or more cycles. Patients were assigned to one of three groups: zona intact (n = 72), partial thinning (n = 59), or circumferential thinning (n = 73). Before transfer, the zonae pellucidae of embryos were thinned partially or circumferentially using acidified Tyrode’s solution. Results Implantation rates were 8.9% in the intact zona group, 17.6% in the partial thinning group, and 11.9% in the circumferential thinning group: respective clinical pregnancy rates were 16.7% (12/72), 32.2% (19/59), and 27.4% (20/73). Both rates were significantly higher in the partial thinning group than the intact zona group. For circumferential thinning versus zona intact groups, differences fell short of significance. Conclusions Following embryo transfer failure, partial thinning would be recommended over circumferential thinning for successful assisted hatching. In patients with previous failure of embryo transfer, partial zona pellucida thinning for assisted hatching improved implantation and clinical pregnancy rates.     

Chemical removal of the outside of the zona pellucida of day 3 human embryos has no impact on implantation rate

Two hundred eighteen consenting patients entered a randomized study of the application of chemical zona pellucida thinning on their day 3 embryos, prior to uterine transfer. Of those control patients (n =108), whose embryos remained unmanipulated, 40 (37.0%) have ongoing/delivered pregnancies, while in the experimental group (n =110), whose embryos had their zonae pellucidae chemically thinned, there are 49 patients (44.6%) who have ongoing/delivered pregnancies. Although this difference is not significant, clearly the application of this micromanipulative intervention has not been detrimental, and this bodes well for routine application of embryonic micromanipulation procedures in general. Certain patient subgroups were studied including older women, those with elevated basal follicle stimulating hormone levels, patients with embryos of differing zona thickness, and patients with embryos of differing uniformity of zona thickness. No significant influence of chemical removal of the outside of the zona on the implantation rate of embryos in any of these subgroups was observed other than a marginally significant (P =0.095) improvement of implantation of embryos with less than 4.0 µm variation in zona thickness when chemical zona thinning was applied. Failure of chemical zona thinning to enhance human embryo implantation significantly, compared to assisted hatching by complete zona drilling, strongly suggests that the bilayered human zona pellucida needs to be fully breached, unlike that of the mouse.     

Outcomes of blastocysts biopsied and vitrified once versus those cryopreserved twice for euploid blastocyst transfer

Trophectoderm biopsy with comprehensive chromosome screening (CCS) has been shown to increase implantation and pregnancy rates. Some patients desire CCS on previously cryopreserved blastocysts, resulting in blastocysts that are thawed/warmed, biopsied, vitrified and then warmed again. The effect of two cryopreservation procedures and two thawing/warming procedures on outcomes has not been effectively studied. Cycles were divided into two groups: group 1 patients underwent a cryopreserved embryo transfer with euploid blastocysts that were vitrified and warmed once; group 2 patients had a cryopreserved embryo transfer of a euploid blastocyst that was cryopreserved, thawed/warmed, biopsied, vitrified and warmed. Groups 1 and 2 included 85 and 17 women aged 35.6 ± 3.9 and 35.3 ± 4.9 years, respectively (not significantly different). Blastocyst survival in group 1 (114/116, 98.3%) and survival of second warming in group 2 (21/24, 87.5%) was significantly different (P = 0.0354). There was no difference between biochemical (68.2% and 62.5%) and clinical (61.2% and 56.3%) pregnancy rates, implantation rate (58.4% and 52.4%) and live birth/ongoing pregnancy rate (54.0% and 47.6%) between groups 1 and 2, respectively. Although it is unconventional to thaw/warm, biopsy, revitrify and rewarm blastocysts for cryopreserved embryo transfer, the results indicate that outcomes are not compromised.Trophectoderm biopsy and screening the embryos for chromosomal abnormalities has been reported to increase implantation and pregnancy rates. There is a category of patients requesting chromosomal screening on previously cryopreserved blastocysts. This scenario requires blastocysts to be thawed/warmed, biopsied, cryopreserved, and thawed/warmed again. The effect of double cryopreservation procedures and double thawing/warming procedures on pregnancy is unknown. Patients were divided into two groups, group 1 underwent a cryopreserved embryo transfer with a chromosomally normal blastocyst that was vitrified and warmed once and group 2 included patients that had a cryopreserved embryo transfer of a chromosomally normal blastocyst that was cryopreserved, thawed/warmed, biopsied, vitrified, and rewarmed. A total of 85 and 17 women aged 35.6 ± 3.9 and 35.3 ± 4.9 years were included in groups 1 and 2, respectively. The survival rate for group 1 (114 of 116, 98.3%) compared with the second warming for group 2 (21 of 24, 87.5%) was significantly higher. There was no difference between biochemical (68.2% and 62.5%), and clinical pregnancies (61.2% and 56.3%), implantation (58.4% and 52.4%), and live birth/ongoing rates (54.0% and 47.6%) between groups 1 and 2. Although it is unconventional to twice cryopreserve and twice thaw/warm a blastocyst, our results indicate that outcomes are not compromised.     

Zona-intact versus zona-free blastocyst transfer: a prospective,randomized study

OBJECTIVE: To compare the outcome of zona-intact versus zona-free blastocyst transfer. DESIGN: Prospective, randomized study. SETTING: Tertiary care private hospital IVF center. PATIENT(S): A total of 240 patients undergoing blastocyst stage ET (119 zona intact and 121 zona free). INTERVENTION(S): In vitro culture of embryos to the blastocyst stage was followed by random allocation to zona-intact or zona-free transfer. MAIN OUTCOME MEASURE(S): Treatment cycle characteristics, implantation, and pregnancy rates. RESULT(S): Transfer of zona-free blastocysts was associated with a higher implantation rate. Subgroup analysis, however, indicated that better outcome was achieved only in the group that received poor-quality blastocysts. Patients undergoing blastocyst transfer for the first time did not seem to benefit from zona-free blastocyst transfer. CONCLUSION(S): Zona-free blastocyst transfer increases the success of blastocyst-stage transfer in patients with poor-quality blastocysts.     

A comparison of the effects of different degrees of zona pellucida damage followed by cryopreservation on the postthaw development of mouse embryos

Purpose: The totally intact zona pellucida is not essential for the development of embryos. It is still unclear how much effect the degree of damages to the zona pellucida will have on the developmental potential of postthaw embryos after cryopreservation. We compared the developmental potential of cryopreserved mouse embryos after induction of two degrees of mechanical damage to the zonae pellucidae by micromanipulation. Methods: In experiment I, the development of 124 cryopreserved ICR mouse embryos to the blastocyst stage after zona pellucida penetration of two-cell embryos as in the procedures of subzonal sperm insertion (SUZI) was compared with the development of zona-intact cryopreserved embryos. In experiment II, the zonae pellucidae of 93 two-cell mouse embryos were dissected as in the procedures of partial zonal dissection (PZD), following which the embryos were frozen. This postthaw development was also compared with that of zona-intact two-cell cryopreserved embryos. All the embryos were thawed and cultured to the blastocyst stage. Additional controls were provided by culturing zonaintact and zona-penetrated or zona-dissected embryos without cryopreservation. Results: The development of unfrozen mouse embryos was not affected by either zona penetration (P=0.433) or zona dissection (P=0.659). The developmental potential of cryopreserved mouse embryos was significantly affected after zona dissection (blastocyst rate, 31% ZD vs 72%, control; P<0.001) but not after zona penetration (blastocyst rate, 59% ZP vs 64% control; P=0.441). Conclusions: The quality of cryopreserved embryos was affected by a large hole on the zona pellucida created by zona dissection but not by simple zona penetration.     

Transfer of frozen-thawed blastocysts that underwent quarter laser-assisted hatching at the day 3 cleaving stage before freezing

OBJECTIVE: To examine the pregnancy potential of frozen-thawed blastocysts that underwent quarter laser-assisted hatching (AH) at the cleaving stage before freezing and to compare clinical and embryo characteristics between the groups that succeeded in and failed to achieve pregnancy. DESIGN: Prospective observational study. PATIENT(S): Thirty-four of 112 patients with frozen blastocysts requiring transfer of thawed embryos between January 2000 and December 2001. SETTING: Assisted reproductive technology unit in a tertiary medical center. INTERVENTION(S): Embryos of patients undergoing blastocyst transfers routinely underwent quarter AH using a nontouch 1.48-microm diode laser. Blastocysts not transferred were cryopreserved using a six-step freezing protocol with glycerol as the cryoprotectant. MAIN OUTCOME MEASURE(S): Postthaw embryo survival and zona pellucida (ZP) maintenance, implantation rate, and clinical pregnancy rate per transfer of thawed blastocysts. RESULT(S): A total of 118 frozen blastocysts was thawed. Of these, 89 (75.4%) embryos survived and were transferred in 35 cycles. Assisted hatching-manipulated ZP tolerated the freeze-thaw procedures without shape distortion in surviving embryos. Eleven (31.4%) clinical pregnancies with 15 intrauterine gestational sacs occurred, for an implantation rate of 16.9%. Those who succeeded in pregnancy tended to have more embryos available before freezing, more original blastocysts of good quality for freezing, and more surviving blastocysts after thawing for transfer than did those who failed to achieve pregnancy. CONCLUSION(S): An acceptable clinical pregnancy rate was obtained from transfer of thawed blastocysts that underwent laser AH on the ZP at the day 3 cleaving embryo stage in fresh cycles in selected patients. Embryo characteristics before freezing played major roles in determining implantation potential of thawed embryos.     

Influence of Zona Pellucida Thickness of Human Embryos on Clinical Pregnancy Outcome Following In Vitro Fertilization Treatment

Purpose: To evaluate the correlation between the degree ofzona pellucida thickness variation (ZPTV) of transferredembryos with identical morphologies and subsequent clinicalpregnancy rates during 141 intracytoplasmic sperminjection (ICSI) treatment cycles. Methods: A total of 141 women participating in the studygroup comprising mostly male factor infertilities and somerepeat in vitro fertilization (IVF) failures were transferred,2 days after ovum pickup, with two to three embryos withidentical grades created by fertilization with ICSI. Allselected embryos were subjected to zona pellucida thicknessmeasurements immediately prior to transfer using a computerizedembryo measurement program from videocinematographyrecordings. Results: A total of 326 identical-grade transferredpre-embryos resulted in 70 clinical pregnancies with live-bornimplantation rate of 27.6%. A highly significant correlationwas observed between ZPTV of transferred embryos andthe IVF outcome with 77.1% and 83.64% of the clinicalpregnancies resulting from transferred embryos with ZPTVvalues greater than 20 and 25, respectively. The mean ZPTVvalues for 70 conceptual cycles and 62 nonconceptual cycleswere 28 ± 6.43 and 17.85 ± 8.11, respectively. No significantcorrelation between ZP thickness and number of blastomeresin the transferred embryos was evident, though embryoswith better scores had significantly thinner zonae and higherZPTV values. Though average zona thickness of embryosdeclined with age, the mean ZPTV value for women lessthan 30 years old was significantly higher (25.84 ± 8.57)as compared with those from women older than 35 years(20.72 ± 8.45). Conclusions: The degree of ZPTV of the transferred embryosexhibits a strong correlation with clinical pregnancy outcome following IVF treatment. This potentially reliable indicatorof IVF success rate could be used as a criteria forembryo selection during clinical transfers.     

Zona pellucida induced acrosome reaction,sperm morphology,and sperm-zona binding assessments among subfertile men

Purpose : The study aimed to evaluate the relationship between the zona pellucida induced acrosome reaction (ZIAR) and (i) percentage normal spermatozoa as well as (ii) sperm–zona pellucida binding potential among men referred for a routine semen analysis. Methods : Semen samples of 164 consecutive men referred to the andrology laboratory for routine semen analysis were studied. Semen samples were analyzed using the new WHO standards (strict criteria). ZIAR was recorded with a lectin conjugated Pisum sativum agglutinin microassay, while sperm–zona binding was evaluated with a standard hemizona assay (HZA). Results : Andrology patients were divided according to the percentage normal spermatozoa in the ejaculate, namely <4% normal forms (n = 71), 5–14% normal forms (n = 73), and >14% normal forms (n = 20). ZIAR data of the <4%, 5–14%, and >14% groups was (9.6 ± 0.6)%, (13.9 ± 0.5)%, and (15.0 ± 1.1)%, respectively. The ZIAR data of fertile control men was (26.6 ± 1.4)% which differed significantly from the three andrology referrals groups. Likewise significant differences were recorded during the hemizona assay namely, 38.0% (<4% normal forms), 54.5% (5–1% normal forms), and 62.6% (>14% normal forms). Among the group with >14% normal forms, five cases had impaired ZIAR outcome (<15%). Three of these men had normal morphology and HZAs. Conclusions : ZIAR testing should become part of the second level of male fertility investigations, i.e., sperm functional testing, since 15% of andrology referrals revealed an impaired acrosome reaction response to solubilized zona pellucida.     

Effect of pretreatment with a levonorgestrel-releasing intrauterine system on IVF and vitrified–warmed embryo transfer outcomes in women with adenomyosis

Research questionDoes the use of a levonorgestrel-releasing intrauterine system (LNG-IUS) improve the ongoing pregnancy rate of vitrified–warmed embryo transfer in women with adenomyosis undergoing IVF?DesignThis retrospective study included 358 women with adenomyosis undergoing IVF. Of these, 134 women were enrolled in the LNG-IUS group and another 224 women were in the control group. All women were screened for adenomyosis by transvaginal ultrasound and magnetic resonance imaging (MRI). There was no significant difference in the ages of women, FSH, cause of infertility, body mass index, total dose of gonadotrophin used and number of oocytes collected between the two groups. All comparisons performed were between patients undergoing vitrified–warmed embryo transfer.ResultsStatistical differences were found in the ongoing pregnancy rates (41.8% vs 29.5%, P = 0.017) between the LNG-IUS group and control group. Logistic regression analysis showed that the odds ratio (OR) of ongoing pregnancy was significantly increased with LNG-IUS usage (adjusted OR = 1.628, 95% confidence interval 1.011–2.622). Also, differences were found in implantation rates (32.1% vs 22.1%, P = 0.005) and clinical pregnancy rates (44% versus 33.5%, P = 0.045) between the LNG-IUS group and control group.ConclusionsThe results of this study offer some support for evaluating the effect of pretreatment with LNG-IUS in women with adenomyosis in future randomized controlled trials.     

The impact of serum oestradiol concentration prior to progesterone administration on live birth rate in single vitrified–warmed blastocyst transfer cycles

Research questionCan serum oestradiol concentrations on the day of progesterone initiation predict live birth rates in single, autologous vitrified–warmed blastocyst transfers following artificial endometrial preparation?DesignThis retrospective study included the first transfers of 468 patients with unexplained or tubal factor infertility who underwent freeze-all cycles using single, top-quality blastocysts after artificial endometrial preparation from January 2015 to January 2018. Patients were stratified into four groups based on serum oestradiol concentration percentiles on the day of progesterone initiation: Group 1 (<25th percentile), Group 2 (25–50th percentile), Group 3 (51–75th percentile) and Group 4 (>75th percentile). The primary outcome was live birth rate. The secondary outcomes were implantation, clinical pregnancy and multiple pregnancy rates. Receiver operating characteristic (ROC) curves were generated to evaluate serum oestradiol concentrations in predicting implantation, clinical pregnancy and live birth.ResultsSimilar live birth rates of 51.6%, 55.1%, 54.9% and 56.4% for Groups 1, 2, 3 and 4, respectively, were found. The groups also showed similar implantation and clinical pregnancy rates. ROC analysis revealed that serum oestradiol concentrations on the day of progesterone initiation were not predictive for implantation (area under the curve [AUC] 0.490, 95% CI 0.445–0.554), clinical pregnancy (AUC 0.507, 95% CI 0.453–0.561) or live birth (AUC 0.514, 95% CI 0.461–0.566).ConclusionsSerum oestradiol concentration monitoring just prior to progesterone administration does not appear to be predictive for live birth rates in good prognosis patients undergoing single, autologous vitrified–warmed blastocyst transfer after artificial endometrial preparation. Therefore, the current practice of monitoring serum oestradiol concentration is not supported by this study.     

Uterine flushing with supernatant embryo culture medium in vitrified warmed blastocyst transfer cycles: a randomized controlled trial

Purpose

Does transfer of supernatant embryo culture fluid (stimulation of endometrial embryo transfer - SEET) prior to vitrified warmed blastocyst transfer result in better clinical pregnancy and live birth rates than direct vitrified warmed blastocyst transfer?

Methods

This randomized controlled trial compared SEET group and direct transfer group (control) in 60 women undergoing vitrified warmed blastocyst transfers. The duration of the study was 3 years. The patients were undergoing vitrified warmed blastocyst transfer at university level infertility centre. Sixty women were randomized to SEET (n = 30) or control (n = 30).

Results

Data was available for analysis from all the 30 women in the SEET group and 30 women in the control group. There were no drop outs in the trial. The implantation rate was significantly lower in the SEET group compared to the control group (27 vs. 44 %, P = 0.018). The clinical pregnancy rates were similar in both the groups (47 vs. 53 %) but the live birth rate was also significantly lower in SEET group (23 vs. 50 %, P = 0.03).

Limitations

The sample size based on clinical pregnancy rates was small and hence not adequately powered to detect differences in live birth rates. Lack of blinding leading to possible bias cannot be ruled out.

Conclusion

There was no evidence of an improvement in clinical pregnancy rate following SEET in vitrified warmed blastocyst transfer compared to direct transfer.     

Successful Use of a Laser for Human Embryo Biopsy in Preimplantation Genetic Diagnosis: Report of Two Cases

Purpose: The use of Tyrode's acid to drill the zona pellucida for embryo biopsy is the most widely used methodology in preimplantation genetic diagnosis. Instead of this, we propose the use of a 1.48-m diode noncontact laser, which is quicker, simpler, and safer. Methods: The laser beam was tangentially guided to the zona pellucida of the embryo. Depending on zona pellucida measurement, two to four consecutive shots of 8–22 msec were necessary to drill the zona pellucida of the 13 embryos biopsied for two patients (hemophilia carriers). Results: Female embryos were replaced into the uterus of the patients (1.5 embryos/replacement). One single pregnancy was established (33.3% implantation rate). Coculture of untransferable embryos showed a blastocyst rate of 66.7% (4/6) for male embryos and 25% (1/4) for abnormal ones. Conclusions: These results demonstrate the safety and usefulness of laser methodology in preimplantation genetic diagnosis.     

Pregnancy and live birth following the transfer of vitrified–warmed blastocysts derived from zona- and corona-cell-free oocytes

This study reports two clinical pregnancies and one live birth following the transfer of vitrified blastocysts developed from oocytes with neither zona pellucida nor corona cells. Two zona-free oocytes obtained from two patients of advanced maternal age undergoing minimal stimulation were normally fertilized after intracytoplasmic sperm injection. In case 1, all four blastomeres of the zona-free embryo were loosely associated and inserted back into ruptured zona on day 2. Zona-free embryo from case 2 had tight contacts between blastomeres and was cultured without zona. Both embryos derived from zona-free oocytes progressed to blastocyst stage and were cryopreserved by vitrification. When patients came back for a cryopreserved embryo transfer, both vitrified blastocysts survived warming. In case 1, transfer of a warmed blastocyst with reconstructed zona resulted in a clinical pregnancy that ended in a spontaneous abortion at 22 weeks. In case 2, live birth was achieved with a normal healthy baby (male) weighing 2381 g at 40 weeks’ gestation. This report emphasizes the importance of maintenance of normal cell arrangement on the subsequent embryonic development for a zona-free oocyte. Zona-free oocytes may provide a valuable source of embryos for infertility patients, especially for those with a limited number of oocytes.In this study, we report two clinical pregnancies and one live birth following the transfer of vitrified blastocysts developed from oocytes with neither zona pellucida nor corona cells. Two zona-free oocytes obtained from two patients of advanced maternal age undergoing minimal stimulation were normally fertilized after intracytoplasmic sperm injection. In case 1, all four blastomeres of the zona-free embryo were loosely associated and inserted back into ruptured zona on day 2. Zona-free embryo from case 2 had tight contacts between blastomeres and was cultured without zona. Both embryos derived from zona-free oocytes progressed to blastocyst stage and were cryopreserved by vitrification. When patients came back for a cryopreserved embryo transfer, both vitrified blastocysts survived warming. In case 1, transfer of a warmed blastocyst with reconstructed zona resulted in a clinical pregnancy that ended in a spontaneous abortion at 22 weeks. In case 2, live birth was achieved with a normal healthy baby (male) weighing 2381 g at 40 weeks’ gestation. Our report emphasizes the importance of maintenance of normal cell arrangement on the subsequent embryonic development for a zona-free oocyte. Zona-free oocytes may provide a valuable source of embryos for infertility patients, especially for those with a limited number of oocytes.     

A Quantitative and Morphological Analysis of Oocytes Collected During 438 IVF Cycles

Purpose: Our purpose was to determine if the number ofretrieved oocytes, oocyte maturity, morphology, and otherembryological parameters are related to the outcome oftreatment. Methods: This retrospective study on 438 IVF cyclesanalyzes the numbers of retrieved oocytes, fractured zonaoocytes, germinal vesicle-stage oocytes, normally andabnormally fertilized oocytes, pregnancy rate, age of femalepartner, ovarian stimulation protocol, day of hCG injection,and serum estradiol concentration. Results: (1) Pregnancy rate increases with an increase inthe number of retrieved oocytes, (2) a high incidence offractured zona pellucida oocytes has a negative effect onfertilization rate but none on pregnancy rate, (3) a highincidence of immature oocytes is associated with improvedfertilization and pregnancy rates, and (4) an inverserelationship between the presence of immature oocytes and oocyteswith fractured zona pellucida is suggested. Conclusions: Precise oocyte assessment in IVF cyclesprovides informations useful for the analysis and improvementof ovarian stimulation protocols.     

A comparison of the survival and implantation rates of blastocysts that were vitrified on post-fertilization day five,six and seven

The goal of this retrospective cohort study was to compare survival, implantation, clinical and ongoing pregnancy rates between blastocysts that were vitrified on post-fertilization days 5, 6 and 7. Before vitrification, blastocysts were evaluated in terms of morphology and blastocyst expansion, inner cell mass and trophectoderm quality. They were thawed and transfered in a subsequent artificial cycle. Embryo implantation rates were 39%, 25% and 25% for blastocysts that were vitrified on days 5, 6, and 7, respectively (p?=?0.006). Clinical and ongoing pregnancy rates were 19%, 12%, 13% (p?=?0.100) and 9%, 7%, 12% (p?=?0.99) for days 5, 6 and 7 blastocysts, respectively. Day 5 blastocysts had significantly higher full-collapsing score after assisted-hatching compared to days 6 and 7 blastocysts (p?=?0.014). As blastocyst quality increased, implantation and clinical pregnancy rates increased in all groups and both parameters were statistically significantly higher on day 5 blastocysts than on days 6 or 7 (p?=?0.001). It was clearly found that good quality blastocysts obtained on day 5 have higher implantation and clinical pregnancy rates than 6th and 7th day cryopreserved embryos. There were no statistically significant differences between the cryopreserved embryos on days 6 and 7 regarding the implantation, clinic and ongoing pregnancy rates.