Study question Is there a relationship between the chromosomal status of biopsied embryos and the presence of polymorphic variants in the female karyotype in IVF cycles? Summary answer The chromosomal status of biopsied embryos is prejudiced because of the presence of specific maternal polymorphic variants (qh-, qh+ and ps+) or combinations of them. What is known already Due to the increasing average age of patients who undergo IVF cycles with their own oocytes, more cycles are performed with embryo biopsy for preimplantation genetic test of aneuploidies (PGT-A). Thus, advanced maternal age has been defined as the main factor causing embryonic chromosomal abnormalities. However, few studies have established direct relationships with other factors. The prevalence of chromosomal polymorphic variants has a higher incidence in the infertile population, which is why there is a need to know the role they play in the genetic inheritance of embryos. Study design, size, duration Retrospective evaluation of a cohort of women who underwent autologous IVF cycles and karyotyping. The sample included 162 IVF cycles with embryo biopsy on D5 and/or D6 for PGT-A, performed between July 2017-December 2021: control group (CG) with normal karyotype (86) and study group (SG) with presence of polymorphisms (76). We studied the connection between maternal karyotype polymorphisms and molecular genetic outcomes in terms of euploid, mosaic, and aneuploid blastocyst rates. Participants/materials, setting, methods The analysis of the karyotype of women, prior to the IVF cycle, was performed using the guidelines of the International System for Human Cytogenetic Nomenclature (ISCN). The trophoectoderm biopsies on D5 and/or D6 blastocysts were analysed by NGS using the Illumina platform (VeriSeq Illumina®, San Diego, CA, USA). The association between the different polymorphic variants in the female karyotype and chromosomal status of embryos was analysed using R (v. 4.2.0) statistical software. Main results and the role of chance We analyzed the results of 483 embryos from 162 IVF-PGT-A cycles. Mean female age was 39.05±2.72 (CG) and 38.97±2.90 (SG). Mean number of mature oocytes (MII) was 7.16±3.94 (CG) and 8.01±5.33 (SG). Mean number of biopsied embryos was 2.97±2.15 (CG) and 2.93±1.88 (SG). No statistically significant differences were found between the CG and SG groups in terms of the rate of euploid embryos: 26.99% vs 21.50% (p = 0.3), transferable mosaic embryos: 8.90% vs 6.76% (p = 0.5), aneuploid embryos: 61.98% vs 69.96% (p = 0.12) and non-informative embryos: 2.13% vs 1.78% (p = 0.2), respectively. However, an increase in the rate of chromosomally abnormal embryos was observed when analysing each polymorphism individually. In females carrying the qh+ variant, we found a statistically significant decrease in the euploid embryo rate: 10.97% vs 26.00% found in CG (p = 0.036). Moreover, the variants qh+ and ps+ increased the rate of aneuploid embryos: 84.17% vs 63.48% in CG (p = 0.013) and 73.52% vs 61.30% in CG (p = 0.021), respectively. Furthermore, it is found that the qh- variant increased the rate of transferable mosaic embryos: 20.00% vs 8.90% in CG (p = 0.025). The results were corrected for confounding variables such as maternal age, oocyte origin and male factor variables, including polymorphic variants in the male karyotype. Limitations, reasons for caution Presence of unmeasured variables that may influence the results. Larger prospective studies including homogeneous cohorts are needed in order to corroborate our initial results. Wider implications of the findings The results show that polymorphic variants in the female karyotype may have a direct effect on the embryonic ploidy and mosaicism status. Therefore, it would be advisable for patients who undergo IVF-PGT-A cycles to have a karyotype performed and be informed of the implications that it may entail. Trial registration number Not applicable
Study question Do polymorphic variants in the karyotype of women undergoing an IVF cycle directly affect oocyte and embryo laboratory parameters? Summary answer The presence of certain polymorphic variants negatively affects the oocyte survival rate and the blastocyst quality in an IVF cycle. What is known already In the recent years, there has been a growing interest in the study of polymorphic variants in infertile patients because their incidence, compared to the fertile population, is increased. However, most research has focused on the male patient study. Several studies have reported information about clinical outcomes, but the influence they may have on IVF laboratory procedures and embryo development has rarely been studied up to the blastocyst stage. In addition, it is very rare to find publications that show the study of polymorphisms according to their type or combination. Study design, size, duration Retrospective evaluation of a cohort of women who underwent autologous IVF cycles and karyotyping. The sample included 424 IVF cycles performed between July 2017-December 2021: control group (CG) with normal karyotype (211) and study group (SG) with polymorphisms (213). We studied the correlation between karyotype polymorphisms and laboratory outcomes in terms of: Number of Oocytes Retrieved, Fresh Oocyte Maturity (MII), Oocyte Survival after Thawing (TS), Fertilization (FZ), Oocyte Degeneration (OD) and Non-viable embryo rates. Participants/materials, setting, methods Analysis of the women karyotype, prior to the IVF cycle, was performed using the guidelines of the International System for Human Cytogenetic Nomenclature (ISCN). Differences between the different study groups (presence or absence of different chromosomal polymorphisms) were assessed with the appropriate statistical test according to the normality or non-normality of the variable distribution. Statistical analysis was performed using R statistical software (v.4.2.0) and SPSS (v.23.0, Chicago, IL, USA). Main results and the role of chance Average female age was 37.72 ± 3.98 (CG) and 36.73 ± 3.59 (SG). Average number of mature oocytes (MII) was: 6.45 ± 4.96(CG) and 7.59 ± 5.10 (SG). Statistically significant differences were found regarding the number of oocytes retrieved between the CG (8.14 ± 5.90) and the SG (9.53 ± 6.63) (p = 0.036), increasing when the ps+ variant was present (9.75 ± 7.21) (p = 0.045). However, no statistically significant differences were found between the presence of polymorphism (SG) and the CG in terms of: number of MII: 79.85% vs. 80.48% (p = 0.447), FZ: 73.76% vs. 70.49% (p = 0.352) and OD: 8.76% vs. 8.36% (p = 0.563) rates. In addition, the TS rate was statistically significant when there was the ps+ variant (82.95%) (p = 0.010) and/or combinations of more than one polymorphic variant (87.80%) (p = 0.044), compared to the CG (93.51%). Finally, according to embryo quality there was an increase in the non-viable embryos rate, on day 5 and/or day 6 of development, between the CG (34.17%) and the SG (43.02%) (p < 0.001), increasing when the ps+ variable was present (45.57%) (p < 0.001) and/or combinations of more than one polymorphic variant (p = 0.045). The results were corrected for confounding variables such as maternal age, oocyte origin and male factor variables, including polymorphic variants in the male karyotype. Limitations, reasons for caution Other variables that have not been analyzed may influence the outcomes. Larger prospective studies including homogeneous cohorts are needed in order to corroborate our initial results. Wider implications of the findings The polymorphic variants in the female karyotype, especially the “ps+” variant and the combination of multiple variants, could influence certain parameters in the laboratory. Therefore, it is important to request a karyotype to all patients before starting IVF treatment. Trial registration number NOT APPLICABLE
Study question Are there any advantages in using High security tubes rather than High Security straws for conventional slow sperm freezing? Summary answer Freezing sperm in High Security tubes (HST) improved post-thaw recovery rate and motility, and also reduced processing and handling compared to High Security straws (HSS). What is known already The use of High Security freezing consumables (HSFC) in an IVF setting is a safe and effective way of eliminating concerns related to viral cross-contamination during storage. The lower diameter of HSS does make them susceptible to warming during handling. The HSFC used in this study is the only CE marked products that are made of resin, leak-proof and shatter-proof in all cryogenic temperatures even in LN2. No previous studies have compared the use of HST with HSS for conventional human sperm freezing. This study sets out to investigate the performance of HST compared to HSS. Study design, size, duration The study was designed as a controlled split-sample study with blind post-thaw analysis. Following the routine WHO analysis of 20 semen samples, the remainder of each of the samples was evenly divided and cryopreserved by conventional slow freezing in each of the two different HSFC. The freeze was conducted simultaneously by the same practitioner, employing the same freezing protocol and cryoprotectant. The pre-freeze and post-thaw concentration, total and progressive sperm motility were recorded. Participants/materials, setting, methods At one IVF clinic, semen samples with sperm density ≥15million/ml, ≥40% motility, ≥1.5ml were included. Cryoprotectant (SpermFreeze, Fertipro) was added dropwise to unprepared semen and kept at room temperature for 10 minutes before loading into HSFC (0.5ml CBS™HSS; CBS™HST). HSFC were heat-sealed (SYMS; SYMSIII sealers) and placed in vapour for 30 minutes before plunging into LN2. Samples were thawed by immersion in a 37Cº water bath for 5 minutes and analysed using WHO methods. Main results and the role of chance Paired-t test was used to compare the percentage motility between the different HSFC. All analysis was considered statistically significant when p < 0.01. We demonstrated that the sperm recovery rate (Percentage total motility post-thaw/ Percentage total motility pre-freeze) in HST was 66.63 ± 14.94 (mean ± standard deviation) compared to 40.80 ± 14.69 in HSS. In the HSS, the percentage post-thaw total motility was 19.99 ± 7.21 and the percentage post-thaw progressive motility was 12.26 ± 2.59. In the HST, the percentage post-thaw total motility was 32.57 ± 8.33 and the percentage post-thaw progressive motility was 23.08 ± 5.53. The overall improvement when using HST against HSS was 12.53 ± 5.69, 10.44 ± 5.29 for the total motility and the progressive motility respectively. Comments were recorded regarding the handling and the condition of the HSS and HST for each freeze event. Neither device displayed any leakage of LN2 or any explosion during the warming. The freezing process was easier and faster using HST rather than HSS. It was also noted that the entire sample can be recovered from the HST, unlike the HSS. Limitations, reasons for caution The study looked at sperm recovery in terms of motility only. DNA damage was not considered as a parameter of sperm quality. Also, fertilization, pregnancy rates, live birth rates and the use of poorer quality sperm samples have not been investigated. Wider implications of the findings: For conventional sperm freezing, the use of HST resulted in improved sperm motility and progression post-thaw, when compared to HSS. This finding supports the use of HST to improve the post thaw quality of sperm, benefitting patients with own frozen samples, recipients of donor sperm and donor sperm banks. Trial registration number Not applicable.
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