Effect of time post warming to embryo transfer on human blastocyst metabolism and pregnancy outcome.

PURPOSE: This study is aiming to test whether variation in post warming culture time impacts blastocyst metabolism or pregnancy outcome. METHODS: In this single center retrospective cohort study, outcomes of 11,520 single frozen embryo transfer (FET) cycles were analyzed from January 2015 to December 2020. Patient treatments included both natural and programmed cycles. Time categories were determined using the time between blastocyst warming and embryo transfer: 0 (0- <1h), 1 (1-<2h), 2 (2-<3h), 3(3-<4h), 4 (4-<5), 5 (5-<6), 6 (6-<7) and 7 (7-8h). Non-invasive metabolic imaging of discarded human blastocysts for up to 10h was also performed using Fluorescence lifetime imaging microscopy (FLIM) to examine for metabolic perturbations during culture. RESULTS: The mean age of patients across all time categories were comparable (35.6 ± 3.9). Live birth rates (38-52%) and miscarriage rate (5-11%) were not statistically different across post-warming culture time. When assessing pregnancy outcomes based on the use of PGT-A, miscarriage and live birth rates were not statistically different across culture hours in both PGT-A and non-PGT cycles. Further metabolic analysis of blastocysts for the duration of 10h of culture post warming, revealed minimal metabolic changes of embryos in culture. CONCLUSION: Overall, our results show that differences in the time of post warming culture have no significant impact on miscarriage or live birth rate for frozen embryo transfers. This information can be beneficial for clinical practices with either minimal staffing or a high number of patient cases.

Day after rescue ICSI: eliminating total fertilization failure after conventional IVF with high live birth rates following cryopreserved blastocyst transfer.

STUDY QUESTION: What is the impact of day after rescue ICSI (r-ICSI) on success of fresh and frozen embryo transfers? SUMMARY ANSWER: The use of r-ICSI can virtually allay fears of total fertilization failure (TFF) after conventional IVF (C-IVF) and achieve high live birth rates after frozen blastocyst transfer. WHAT IS KNOWN ALREADY: More infertility clinics have resorted to the use of ICSI in place of C-IVF in IVF treatment owing to fear of TFF or a low fertilization rate. r-ICSI has been attempted either on the day of IVF or the day after. Day after r-ICSI has proved unsuccessful in the past. STUDY DESIGN, SIZE, DURATION: A retrospective data analysis was performed of 16 608 qualifying cases between April 2010 and July 2021 conducted at a single private academically affiliated fertility clinic. PARTICIPANTS/MATERIALS, SETTING, METHODS: r-ICSI was performed principally on patients with >4 metaphase II oocytes, showing no signs of fertilization 18 h after C-IVF. C-IVF was performed on patients who had >4 million total motile sperm after preparation. r-ICSI was then performed 18-24 h after insemination, using the sperm sample from the previous day. r-ICSI fertilization rates, cryopreservation of cleavage and blastocysts embryos, and pregnancy rates after fresh or frozen transfer were then assessed. MAIN RESULTS AND THE ROLE OF CHANCE: r-ICSI was performed on 377 patients (2.3% of eligible retrieval cycles) who had a mean (±SD) female and male age of 35.9 ± 4.5 and 38.1 ± 9.1 years, respectively. A total of 5459 oocytes were initially retrieved. Of the oocytes undergoing r-ICSI, 2389 (49.5%) fertilized normally, and 205 (54.4%) patients underwent a fresh embryo transfer. The live birth rates were 23/186 (12.3%) for fresh cleavage and 5/19 (26.3%) for fresh blastocyst stage transfers. In 145 cycles a blastocyst was frozen, and 137 transfers were performed with a 64/137 (46.7%) live birth rate. Of the 377 cycles receiving r-ICSI only, 25 of the qualifying cases failed to have any fertilization, reducing TFF to 25/16 608 (0.15%). LIMITATIONS, REASONS FOR CAUTION: This was a single-center retrospective study on a specific subset of patients, which may limit its generalizability to other clinics. WIDER IMPLICATIONS OF THE FINDINGS: r-ICSI allows a second opportunity to fertilize oocytes despite poor initial outcomes. Patients who had a frozen blastocyst transfer achieved high live birth rates, indicating that a resynchronization of the embryo with the endometrium can optimize r-ICSI cases. r-ICSI allays fears of TFF when using C-IVF, providing evidence that the overuse of ICSI in patients without male factor may not be warranted. STUDY FUNDING/COMPETING INTEREST(S): The study was internally funded by Boston IVF. The authors declare that they have no conflict of interest in relation to the data published in the article. TRIAL REGISTRATION NUMBER: N/A.

The effect of rapid and delayed insemination on reproductive outcome in conventional insemination and intracytoplasmic sperm injection in vitro fertilization cycles.

PURPOSE: The precise timing of insemination after oocyte retrieval is sometimes challenging. In this study, we have assessed the effect of the variation in insemination timing on reproductive outcome for both conventional insemination (CI) and intracytoplasmic sperm injection (ICSI) cycles. METHODS: A single-center retrospective cohort data analysis was performed on 6559 patients (9575 oocyte retrievals) from January 2017 to July 2019. The main outcome measured was live birth rates. Secondary outcomes included fertilization rate per all oocytes retrieved, blastocyst utilization, clinical pregnancy, and miscarriage rates. The time interval between oocyte retrieval and insemination was analyzed in eight categories: 0 (0- < 0.5 h), 1 (0.5- < 1.5 h), 2 (1.5- < 2.5 h), 3 (2.5- < 3.5 h), 4 (3.5- < 4.5), 5 (4.5- < 5.5), 6 (5.5-6.5), and 7 (6.5- < 8 h). The number of retrievals in each group (0-7) was 586, 1594, 1644, 1796, 1836, 1351, 641, and 127 respectively. RESULTS: The mean fertilization rate for CI ranged from 54.1 to 64.9% with a significant difference between time categories 0 and 5 (p < 0.001) and 1 and 5 (p < 0.0.001). The mean fertilization rate for ICSI ranged from 52.8 to 67.3% with no significant difference between time categories. Blastocyst rate for CI and ICSI was not significantly different. Miscarriage and clinical pregnancy rates in CI and ICSI were not significantly different. Live birth rates differed significantly (p < 0.05) in CI with time categories 0 and 7 representing the lowest rates, but not in the ICSI group. CONCLUSION: If performing CI or ICSI before 1.5 h and > 6.5 h, any detrimental effects are moderate on fertilization but do not affect blastocyst usage and birth rates. TRIAL REGISTRATION: Institutional Review Board Approval from the Beth Israel Deaconess Medical Centre [IRB Protocol #: 2015P000122].

Types and frequency of non-conformances in an IVF laboratory.

STUDY QUESTION: How many non-conformances occur in an ART laboratory and how often do they occur? SUMMARY ANSWER: The limited data to date demonstrate that IVF laboratories have a very low non-conformance rate compared with reported non-conformances in other medical laboratories, especially when one considers the high-complexity of procedures performed. WHAT IS KNOWN ALREADY: ART involves a series of very complex patient and laboratory procedures. Although it is assumed that strict measures control ART laboratories, there is very little published data on non-conformances. STUDY DESIGN, SIZE, DURATION: In accordance with the ISO 9001:2008 standard, Boston IVF has created an electronic database to record non-conformances in the IVF laboratory. We reviewed the non-conformances reported between March 2003 and December 2015. The non-conformances were categorized into four grades largely based upon their impact on the outcome or continuation of an IVF treatment cycle: None/Minimal (not measurably decreasing the likelihood of success), Moderate (a negative impact but not loss of a cycle), Significant (loss of a cycle or majority of gametes or embryos) and Major (infrequent errors that have an extreme impact on a patient or patients such as a confirmed pregnancy or birth involving misidentification of sperm, egg or embryo, or an extreme equipment or documentation failure that affects numerous patients). The category of problem or error associated with the Non-conformance Report was also noted. PARTICIPANTS/MATERIALS, SETTING, METHOD: Retrospective analysis of an electronic database registering non-conformances at a large IVF laboratory. MAIN RESULTS AND THE ROLE OF CHANCE: During the study period, a total of 36 654 IVF treatment cycles (fresh and frozen embryo transfer cycles) were conducted which involved a total of 181 899 individual laboratory procedures encompassing egg retrievals, sperm preparations, inseminations, embryo transfers, etc. When combining both moderate and significant non-conformances, 99.96% of procedures and 99.77% of cycles proceeded with no non-conformances. No Major grade non-conformances were reported. LIMITATIONS, REASONS FOR CAUTION: A comparison of non-conformances between IVF clinics is difficult because of different classifications. WIDER IMPLICATIONS OF THE FINDINGS: Errors are inevitable and it is incumbent on all IVF centers to be honest and transparent, both within the organization and with patients when errors occur. Robust systems for identifying, documenting, analyzing and implementing improvements should be established and maintained. STUDY FUNDING/COMPETING INTEREST(S): No external funding was used for this study. The authors have no conflicts of interest.

Predicting personalized multiple birth risks after in vitro fertilization-double embryo transfer.

OBJECTIVE: To report and evaluate the performance and utility of an approach to predicting IVF-double embryo transfer (DET) multiple birth risks that is evidence-based, clinic-specific, and considers each patient's clinical profile. DESIGN: Retrospective prediction modeling. SETTING: An outpatient university-affiliated IVF clinic. PATIENT(S): We used boosted tree methods to analyze 2,413 independent IVF-DET treatment cycles that resulted in live births. The IVF cycles were retrieved from a database that comprised more than 33,000 IVF cycles. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): The performance of this prediction model, MBP-BIVF, was validated by an independent data set, to evaluate predictive power, discrimination, dynamic range, and reclassification. RESULT(S): Multiple birth probabilities ranging from 11.8% to 54.8% were predicted by the model and were significantly different from control predictions in more than half of the patients. The prediction model showed an improvement of 146% in predictive power and 16.0% in discrimination over control. The population standard error was 1.8%. CONCLUSION(S): We showed that IVF patients have inherently different risks of multiple birth, even when DET is specified, and this risk can be predicted before ET. The use of clinic-specific prediction models provides an evidence-based and personalized method to counsel patients.

Influence of embryo sex on development to the blastocyst stage and euploidy.

OBJECTIVE: To compare the prevalence of blastocyst development and euploidy in XX versus XY embryos. DESIGN: Retrospective cohort study. SETTING: Boston IVF, a large university-affiliated reproductive medicine practice. PATIENT(S): All patients who underwent their first preimplantation genetic screening cycle between January 1, 2006, and December 31, 2007. INTERVENTION(S): In vitro fertilization and preimplantation genetic screening. MAIN OUTCOME MEASURE(S): Proportion of embryos that developed to the blastocyst stage by day 5 and prevalence of euploidy for chromosomes 8, 13, 14, 15, 16, 17, 18, 20, 21, and 22 in XX versus XY embryos. RESULT(S): Seven hundred fifty-eight embryos from 138 cycles in 138 patients were analyzed. Three hundred sixty-six (48%) were XX, and 392 (52%) were XY. XX and XY embryos were equally likely to develop to the blastocyst stage by day 5 and were equally likely to be euploid for the analyzed chromosomes. CONCLUSION(S): Our data suggest that extending embryo culture to day 5 does not lead to sex selection and that euploidy and aneuploidy are not sex dependent.

Derivation of pre-X inactivation human embryonic stem cells under physiological oxygen concentrations.

The presence of two active X chromosomes (XaXa) is a hallmark of the ground state of pluripotency specific to murine embryonic stem cells (ESCs). Human ESCs (hESCs) invariably exhibit signs of X chromosome inactivation (XCI) and are considered developmentally more advanced than their murine counterparts. We describe the establishment of XaXa hESCs derived under physiological oxygen concentrations. Using these cell lines, we demonstrate that (1) differentiation of hESCs induces random XCI in a manner similar to murine ESCs, (2) chronic exposure to atmospheric oxygen is sufficient to induce irreversible XCI with minor changes of the transcriptome, (3) the Xa exhibits heavy methylation of the XIST promoter region, and (4) XCI is associated with demethylation and transcriptional activation of XIST along with H3K27-me3 deposition across the Xi. These findings indicate that the human blastocyst contains pre-X-inactivation cells and that this state is preserved in vitro through culture under physiological oxygen.

Influence of patient age on the association between euploidy and day-3 embryo morphology.

A retrospective cohort study conducted in 138 patients undergoing their first preimplantation genetic screening (PGS) cycle between January 1, 2006, and December 31, 2007, demonstrated that embryos with good day-3 morphology were more likely to be euploid for X/Y, 8, 15, and 18 than those with poor morphology. The strength of association between euploidy and day-3 morphology was not influenced by maternal age.

Prognosis for clinical pregnancy and delivery after total fertilization failure during conventional in vitro fertilization or intracytoplasmic sperm injection.

OBJECTIVE: To determine the prognosis for clinical pregnancy and delivery after total fertilization failure. DESIGN: Retrospective analysis of patient treatment cycles. SETTING: Private fertility clinic. PATIENT(S): 555 couples who had total fertilization failure during a cycle of conventional in vitro fertilization (IVF) or intracytoplasmic sperm injection (ICSI). INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Delivery rates, peak estradiol level, number of mature follicles, number of oocytes retrieved, number of mature oocytes, sperm concentration, and sperm motility. RESULT(S): Delivery rates for IVF patients who elected to continue treatment after fertilization failure were 44% per patient, 25% per embryo transfer (ET), and 22% per cycle. Delivery rates for ICSI patients were 36% per patient, 23% per ET, and 18% per cycle. The number of mature oocytes was always statistically significantly lower in the total fertilization failure cycle when compared with fertilization cycles that occurred either before or after, whether ICSI or conventional IVF was involved. CONCLUSION(S): The prognosis for pregnancy is encouraging in subsequent cycles after total fertilization failure. Fertilization failure was a result of suboptimal response to ovarian stimulation.