RESUMO
OBJECTIVE: Cell-free DNA (cfDNA) is released into the spent blastocyst media (spent media) by the embryo. However, optimal timing to determine maximal cfDNA in the case of frozen-thawed blastocysts undergoing non-invasive preimplantation genetic testing for aneuploidy (niPGT-A) remains to be elucidated. In the current study we investigated the ideal time in culture to optimize embryo cell-free DNA (cfDNA) analysis in frozen-thawed blastocysts undergoing niPGT-A. DESIGN: In this prospective observational study, 135 spent media and corresponding whole blastocysts were collected from January 2021 through March 2022. SUBJECTS: Day-5 frozen-thawed blastocysts were cultured for 8 hours (Day-5 Short) or 24 hours (Day-5 Long), while day-6 frozen-thawed blastocysts were cultured for 8 hours (Day-6 Short). The spent media and whole blastocysts were then collected for further analysis. Spent media and whole blastocysts were amplified using whole genome amplification (WGA) and sequenced using Next Generation Sequencing (NGS). MAIN OUTCOME MEASURES: Informativity and concordance rates between cfDNA in spent media and whole blastocysts DNA were compared according to the different time in culture. RESULTS: When comparing time in culture, informativity rates for spent media were significantly higher (p<0.0001) for Day-5 Long and Day-6 Short (>91%) compared to the Day-5 Short group (<60%). A similar trend was observed for cases with and without a previous PGT-A. Regarding blastocyst expansion grade, informativity rates were lower in Day-5 Short, compared to Day-5 Long and Day-6 Short, regardless of expansion degree. This decrease was significant for Gardner grade expansion grade 3 (p=0.0005), 4 (p=0.0366) and 5-6 (p=0.0002). In addition, for a similar time in culture, the grade of expansion did not have an impact on the informativity rates. For concordance rates, no significant differences were observed among the three groups. In all cases concordance rates were 90.5% for Day-5 Short, 93.6% for Day-5 Long and 92.3% for Day-6 Short. No impact of the expansion grade was observed on concordance rates. CONCLUSION: niPGT-A in frozen-thawed blastocysts yields very high concordance rates with whole blastocysts, possibly limiting the need for invasive PGT-A and making it available for a wider range of patients.
RESUMO
Does sperm preparation using the FERTILE PLUS™ Sperm Sorting Chip improve fertilization rates, blastocyst formation, utilization, and euploidy rates in patients undergoing intracytoplasmic sperm injection (ICSI), compared with density gradient centrifugation (DGC)? A single-cohort, retrospective data review including data from 53 couples who underwent ICSI cycles within a 12-month period. For each couple, the two closest, consecutive cycles were identified, where one used the standard technique of sperm preparation (DGC) and the subsequent used FERTILE PLUS™, therefore, couples acted as their own controls. Paired samples t-test was used to compare means for the outcomes (fertilization, blastocyst formation, utilization, and euploidy rates). Binary logistic regression analysis assessed the relationship between female age, the presence of male factor infertility, and euploidy rates. Blastocyst, utilization, and euploidy rates were significantly higher for cycles using FERTILE PLUS™ compared to DGC (76% vs 56%, p = 0.002; 60% vs 41%, p = 0.005, and 40% vs 20%, p = 0.001, respectively). Although there was an increase in fertilization rates for cycles using FERTILE PLUS™, this was not significant (72% vs 68%, p = 0.449). The euploidy rates of females ≤ 35 years were significantly increased when the FERTILE PLUS™ sperm preparation method was used, compared to the older age group (OR 2.31, p = 0.007). No significant association was found between the presence or absence of male factor infertility and euploidy rates between the two cycles. This study provides tentative evidence that the FERTILE PLUS™ microfluidic sorting device for sperm selection can improve blastocyst formation, utilization, and euploidy rates following ICSI in comparison to the DGC method.
RESUMO
The presence of smooth endoplasmic reticulum aggregates (SERa) in the ooplasm is considered as the most severe oocyte dysmorphism due to its serious and potentially lethal outcomes in offspring. In the present case report, a couple underwent their first intracytoplasmic sperm injection (ICSI) cycle using a gonadotrophin releasing hormone (GnRH) antagonist protocol, followed by fetal ultrasound scanning and amniocentesis. SERa were observed in all oocytes retrieved. A singleton pregnancy was established. The second trimester fetal ultrasound scan revealed a female fetus with overlapping fingers in both hands, and amniocentesis was performed for the detection of chromosomal abnormalities. Comprehensive genetic analysis with the combined use of array-comparative genomic hybridization (CGH), fluoresence in situ hybridization (FISH) and conventional cytogenetics revealed a complex chromosome rearrangement (CCR) involving three break points on two chromosomes, resulting in a reciprocal translocation with a cryptic 2q31 deletion. A week following amniocentesis, there was rupture of amniotic membranes and a stillborn was delivered. This is the first case in the literature to report a CCR with concomitant 2q31 deletion resulting in a well-defined and clinically recognizable contiguous gene syndrome with an abnormal phenotype in a fetus arising from a cohort of oocytes affected by SERa. It is suggested that fertilization and transfer of oocytes with SERa should be avoided, until further research establishes whether there is a causal relationship between the presence of SERa and chromosomal abnormalities in the resulting fetus. ABBREVIATIONS: SER: smooth endoplasmic reticulum; ICSI: intracytoplasmic sperm injection; GnRH: gonadotrophin releasing hormone; CGH: comparative genomic hybridization; FISH: fluoresence in situ hybridization; FSH: follicle stimulating hormone; hCG: human chorionic gonadotrophin; OHSS: ovarian hyperstimulation syndrome; IVF: in vitro fertilization; MII: metaphase II; GV: germinal vesicle; CCR: complex chromosome rearrangement.
