ABSTRACT
RESEARCH QUESTION: Does artificial shrinkage before fresh blastocyst transfer improve clinical pregnancy rates in IVF? DESIGN: In this monocentric prospective, randomized, double-blind, controlled pilot study, 150 couples undergoing fresh single-blastocyst transfer were randomized between 20 May 2018 and 22 February 2022. In the artificial shrinkage group (AS group), a single laser pulse was directed to the cellular junction of the trophectoderm on the opposite side of the inner cell mass in each blastocyst. IVF outcomes were clinical pregnancy, multiple pregnancy and live birth rates. Cell-free DNA (cfDNA) concentration was also measured by quantitative real-time PCR in the blastocyst culture medium. RESULTS: In total, 142 couples underwent fresh single-blastocyst transfer: control group, no artificial shrinkage, nâ¯=â¯47; and AS group, artificial shrinkage, nâ¯=â¯95; An intention-to-treat (ITT) analysis was employed. After a reassessment and the exclusion of patients with major protocol deviations, 139 couples underwent fresh single-blastocyst transfer under optimal conditions: control group, nâ¯=â¯47; and AS group, nâ¯=â¯92; a per-protocol analysis was used here. The clinical and laboratory characteristics were not significantly different between the groups. The clinical pregnancy rate was similar in the control and AS groups (ITT: 48.9% versus 49.5%, Pâ¯=â¯0.97; per protocol: 48.94% versus 51.1%, Pâ¯=â¯0.89). The multiple pregnancy rate and the live birth rate were also similar between the groups. No significant differences in gestational age, birthweight or proportion of male/female newborns were observed. The concentration of cfDNA in the blastocyst culture medium was not associated with IVF outcome. CONCLUSIONS: Large-scale randomized controlled trials are required to confirm these preliminary results.
Subject(s)
Fertilization in Vitro , Pregnancy Rate , Humans , Female , Pregnancy , Pilot Projects , Adult , Fertilization in Vitro/methods , Male , Double-Blind Method , Embryo Transfer/methods , Blastocyst , Prospective Studies , Embryo Culture Techniques , Pregnancy Outcome , Birth RateABSTRACT
Progesterone (P4) is essential for pregnancy. A controlled ovarian stimulation (COS) leads to a iatrogenic luteal defect that indicates a luteal phase support (LPS) at least until pregnancy test day. Some clinicians continue the LPS until week 8 or later, when P4 is mainly secreted by syncytiotrophoblast cells.Measuring serum P4 on pregnancy test day after a fresh embryo transfer could help to identify women who might benefit from prolonged LPS. In women with LPS based on P4 administered by the rectal route, P4 concentration on pregnancy test day was significantly higher in patients with ongoing pregnancy than in patients with abnormal pregnancy.This monocentric retrospective study used data on 99 consecutive cycles of COS, triggered with human chorionic gonadotropin, followed by fresh embryo transfer resulting in a positive pregnancy test (>100 IU/L) (from November 2020 to November 2022). Patients undergoing preimplantation genetic screening or with ectopic pregnancy were excluded. All patients received standard luteal phase support (i.e. micronized vaginal progesterone 600 mg per day for 15 days). The primary endpoint was P4 concentration at day 15 after oocyte retrieval (pregnancy test day) in women with ongoing pregnancy for >12 weeks and in patients with miscarriage before week 12 of pregnancy.The median P4 concentration [range] at pregnancy test day was higher in women with ongoing pregnancy than in women with miscarriage (55.9 ng/mL [11.6; 290.6] versus 18.1 ng/mL [8.3; 140.9], p = 0.002). A P4 concentration ≥16.5 ng/mL at pregnancy test day was associated with higher ongoing pregnancy rate (OR = 12.5, 95% CI 3.61 - 43.33, p <0.001). A P4 concentration ≥16.5 ng/mL at pregnancy test day was significantly associated with higher live birth rate (OR = 11.88, 95% CI 3.30-42.71, p <0.001).After COS and fresh embryo transfer, the risk of miscarriage is higher in women who discontinue luteal support after 15 days, as recommended, but with P4 concentration <16.5 ng/mL. The benefit of individualized prolonged luteal phase support should be evaluated.
Subject(s)
Abortion, Spontaneous , Pregnancy Tests , Pregnancy , Humans , Female , Progesterone , Fertilization in Vitro/methods , Retrospective Studies , Lipopolysaccharides , Embryo Transfer/methods , Ovulation Induction/methodsABSTRACT
Introduction: Poor responder patients remain a challenge in assisted reproductive technologies. The "short agonist stop" (SAS) stimulation protocol uses a double stimulation (flare up effect with the gonadotropin-releasing hormone (GnRH) agonist (GnRH-a) then gonadotropins) associated with a less strenuous blockage (discontinuation of GnRH-a) to favor follicular recruitment in order to obtain a better ovarian response. This study aims to compare the number of oocytes obtained after a SAS stimulation protocol with those obtained after the previous stimulation protocol, in the same women, with poor ovarian response (POR) diagnosed according to the POSEIDON criteria. Design: This therapeutic observational retrospective cohort from 2018 to 2022, with a case-control evaluation compared with the same patients' previous performance, included women with POR undergoing IVF with SAS stimulation protocol. The primary outcome was the number of total oocytes recovered and secondary outcomes were the numbers of mature oocytes, total embryos observed at day 2 and usable cleaved embryos and blastocysts (day 5/6). Results: 63 patients with SAS and previous cycles were included. In the SAS group, the mean number of oocytes was significantly higher: 7.3 vs 5.7, p=0.018 in comparison with the previous attempt. So was the number of mature oocytes (5.8 vs 4.1, p=0.032) and the total mean number of embryos obtained at day 2 (4.1 versus 2.7, p=0.016). The SAS stimulation generated 84 usable embryos: 57 cleaved embryos and 27 blastocysts. The mean number of usable embryos was similar in both groups (1.64 vs 1.31, respectively, p=0.178). In total, out of 63 patients, after the SAS protocol, and subsequent embryo transfers (fresh and frozen, n=54), 9 patients had ongoing pregnancies and no miscarriage occurred. The cumulative ongoing pregnancy rate (cOPR) after the SAS protocol was 14.3% (9/63) per oocyte pick-up and 16.7% (9/54) per transfer. Conclusion: SAS stimulation is a short and original protocol strengthening the therapeutic arsenal of poor responders, that may offer promising results for those patients with low prognosis and previous failed IVF. Results must be confirmed with a randomized controlled trial.
Subject(s)
Fertilization in Vitro , Ovulation Induction , Female , Pregnancy , Humans , Pilot Projects , Retrospective Studies , Reproductive Techniques, Assisted , Gonadotropin-Releasing HormoneABSTRACT
Introduction: Low serum progesterone concentration on frozen embryo transfer (FET) day in hormone replacement therapy (HRT) cycles results in lower reproductive outcomes. Recent studies showed the efficiency of a "rescue protocol'' to restore reproductive outcomes in these patients. Here, we compared reproductive outcomes in HRT FET cycles in women with low serum progesterone levels who received individualized luteal phase support (iLPS) and in women with adequate serum progesterone levels who underwent in vitro fertilization for pre-implantation genetic testing for structural rearrangements or monogenic disorders. Design: This retrospective cohort study included women (18-43 years of age) undergoing HRT FET cycles with pre-implantation genetic testing at Montpellier University Hospital between June 2020 and May 2022. A standard HRT was used: vaginal micronized estradiol (6mg/day) followed by vaginal micronized progesterone (VMP; 800 mg/day). Serum progesterone was measured after four doses of VMP: if <11ng/ml, 25mg/day subcutaneous progesterone or 30mg/day oral dydrogesterone was introduced. Results: 125 HRT FET cycles were performed in 111 patients. Oral/subcutaneous progesterone supplementation concerned 39 cycles (n=20 with subcutaneous progesterone and n=19 with oral dydrogesterone). Clinical and laboratory parameters of the cycles were comparable between groups. The ongoing pregnancy rate (OPR) was 41.03% in the supplemented group and 18.60% in the non-supplemented group (p= 0.008). The biochemical pregnancy rate and miscarriages rate tended to be higher in the non-supplemented group versus the supplemented group: 13.95% versus 5.13% and 38.46% versus 15.79% (p=0.147 and 0.182 respectively). Multivariate logistic regression analysis found that progesterone supplementation was significantly associated with higher OPR ââ (adjusted OR = 3.25, 95% CI [1.38 - 7.68], p=0.007). Conclusion: In HRT FET cycles, progesterone supplementation in patients with serum progesterone concentration <11 ng/mL after four doses of VMP significantly increases the OPR.
Subject(s)
Luteal Phase , Progesterone , Pregnancy , Humans , Female , Dydrogesterone , Retrospective Studies , Embryo Transfer/methods , Genetic TestingABSTRACT
Preimplantation genetic testing (PGT) is widely used to select unaffected embryos, increasing the odds of having a healthy baby. During the last few decades, it was accepted that monozygotic dichorionic diamniotic twin pregnancies occurred from the embryo splitting before Day 3 postfertilization according to Corner's dogma. Hence, the occurrence of a dichorionic diamniotic twin pregnancy after a single blastocyst transfer was considered a dizygotic pregnancy resulting from blastocyst transfer and concurrent natural fertilization. In our study, we have provided for the first time molecular proof that a single blastocyst transfer can result in a monozygotic dichorionic diamniotic twin pregnancy, invalidating Corner's dogma. In this case, we recommend systematically assessing the genetic status of dichorionic twins after single blastocyst transfer using prenatal diagnosis to exclude the risk from a potential concurrent spontaneous pregnancy and to ensure that both fetuses are unaffected. To achieve this goal, we have developed here an innovative noninvasive prenatal diagnosis by exclusion of paternal variants with droplet digital PCR, maximizing the reliability of genetic diagnosis. Further multicentric prospective studies using genetic testing are now required to establish the rate of blastocyst splitting leading to dichorionic pregnancy in PGT and to identify the risk factors.
Subject(s)
Pregnancy, Twin , Twins, Monozygotic , Blastocyst , Embryo Transfer , Female , Genetic Testing , Humans , Pregnancy , Pregnancy, Twin/genetics , Prospective Studies , Reproducibility of Results , Retrospective Studies , Twins, Monozygotic/geneticsABSTRACT
Progesterone plays a key role in implantation. Several studies reported that lower luteal progesterone levels might be related to decreased chances of pregnancy. This systematic review was conducted using appropriate key words, on MEDLINE, EMBASE, and the Cochrane Library, from 1990 up to March 2021 to assess if luteal serum progesterone levels are associated with ongoing pregnancy (OP) and live birth (LB) rates (primary outcomes) and miscarriage rate (secondary outcome), according to the number of corpora lutea (CLs). Overall 2,632 non-duplicate records were identified, of which 32 relevant studies were available for quantitative analysis. In artificial cycles with no CL, OP and LB rates were significantly decreased when the luteal progesterone level falls below a certain threshold (risk ratio [RR] 0.72; 95% confidence interval [CI] 0.62-0.84 and 0.73; 95% CI 0.59-0.90, respectively), while the miscarriage rate was increased (RR 1.48; 95% CI 1.17-1.86). In stimulated cycles with several CLs, the mean luteal progesterone level in the no OP and no LB groups was significantly lower than in the OP and LB groups [difference in means 68.8 (95% CI 45.6-92.0) and 272.4 (95% CI 10.8-533.9), ng/ml, respectively]. Monitoring luteal serum progesterone levels could help in individualizing progesterone administration to enhance OP and LB rates, especially in cycles without corpus luteum. Systematic Review Registration: https://www.crd.york.ac.uk/prospero/display_record.php?RecordID=139019, identifier 139019.
Subject(s)
Abortion, Spontaneous , Progesterone , Birth Rate , Corpus Luteum , Female , Humans , Luteal Phase , Pregnancy , Pregnancy RateABSTRACT
RESEARCH QUESTION: Should whole-genome investigations be considered systematically before a complex chromosomal abnormality preimplantation genetic testing for structural chromosomal rearrangements (PGT-SR) management is carried out using conventional cytogenetic techniques? DESIGN: A male carrying a putative rare interchromosomal reciprocal insertion (IRI) 46,XY,ins(14;?)(q11;?).ish der(14)ins(14;22)(q11.2;q11.2q11.2)(xcp14+,xcp22+,N25+,3'TRA/D+),der(22)ins(22;14)(q11.2;q11.2q11.2)(xcp22+,xcp14+,N25-,5'TRA/D+), and his partner were referred to our centre for preimplantation genetic testing analysis after three spontaneous miscarriages. Whole-genome sequencing was used to distinguish between the proposed IRI and an alternative explanation of reciprocal translocation. Fluorescence in-situ hybridization was used to detect all chromosome segments involved in this chromosomal rearrangement, to identify transferable normal and balanced embryos. RESULTS: Whole-genome sequencing allowed the determination of the number of chromosomal breakpoints involved in chromosomal rearrangement between chromosomes 14 and 22. Finally, only two breakpoints were identified instead of four in IRI rearrangements, which suggests a reciprocal translocation rearrangement. A probe strategy was established to highlight all chromosomal imbalances, whether IRI or reciprocal translocation, and preimplantation genetic testing cycles were achieved. CONCLUSION: Conventional cytogenetic techniques are not capable of identifying all complex chromosomal rearrangements, especially those involving centromeric regions and short arms of acrocentric chromosomes. The advent of new sequencing technologies has allowed for a better appreciation of genome complexity. In this study, whole-genome analysis provided additional information to explain the occurrence of genomic events and confirmed that the initial diagnosis of IRI identified by conventional cytogenetic techniques was, in fact, a simple reciprocal translocation. A reliable PGT-SR strategy was proposed for this couple to achieve their parental project.
Subject(s)
Preimplantation Diagnosis , Chromosome Aberrations , Female , Genetic Testing/methods , Humans , In Situ Hybridization, Fluorescence , Male , Pregnancy , Preimplantation Diagnosis/methods , Translocation, GeneticABSTRACT
Oxygen (O2) concentration is approximately 5% in the fallopian tube and 2% in the uterus in humans. A "back to nature" approach could increase in vitro fertilization (IVF) outcomes. This hypothesis was tested in this monocentric observational retrospective study that included 120 couples who underwent two IVF cycles between 2014 and 2019. Embryos were cultured at 5% from day 0 (D0) to D5/6 (monophasic O2 concentration strategy) in the first IVF cycle, and at 5% O2 from D0 to D3 and 2% O2 from D3 to D5/6 (biphasic O2 concentration strategy) in the second IVF cycle. The total and usable blastocyst rates (44.4% vs. 54.8%, p = 0.049 and 21.8% vs. 32.8%, p = 0.002, respectively) and the cumulative live birth rate (17.9% vs. 44.1%, p = 0.027) were significantly higher with the biphasic (5%-2%) O2 concentration strategy. Whole transcriptome analysis of blastocysts donated for research identified 707 RNAs that were differentially expressed in function of the O2 strategy (fold-change > 2, p value < 0.05). These genes are mainly involved in embryo development, DNA repair, embryonic stem cell pluripotency, and implantation potential. The biphasic (5-2%) O2 concentration strategy for preimplantation embryo culture could increase the "take home baby rate", thus improving IVF cost-effectiveness and infertility management.
Subject(s)
Birth Rate , Blastocyst/metabolism , Embryo Culture Techniques/methods , Fertilization in Vitro/methods , Infertility/therapy , Live Birth , Oxygen/metabolism , Adult , Cost-Benefit Analysis , Embryo Implantation/genetics , Embryo Transfer/methods , Embryonic Development/genetics , Female , Fertilization in Vitro/economics , Gene Expression Regulation, Developmental , Humans , Male , Retrospective Studies , Transcriptome/genetics , Treatment OutcomeABSTRACT
PURPOSE: To assess if the ovarian response of FMR1 premutated women undergoing preimplantation genetic testing (PGT) for Fragile X syndrome is lower compared with fully mutated patients, due to their frequent premature ovarian failure. METHODS: In a retrospective cohort study from January 2009 to March 2019, we compared PGT outcomes in 18 FMR1 premutated women and 12 fully mutated women and aimed to identify predictive factors of stimulation outcomes. RESULTS: Eighty-six IVF/PGT-M cycles for FMR1 PGT were analyzed. Premutation and full mutation patients were comparable in terms of age, body mass index (BMI), basal FSH, antral follicular count, and cycle length. However, premutation carriers had significantly lower AMH (1.9 versus 4.0 ng/mL, p = 0.0167). Premutated patients required higher doses of FSH (2740 versus 1944 IU, p = 0.0069) but had similar numbers of metaphase II oocytes (7.1 versus 6.6, p = 0.871) and embryos (5.6 versus 4.9, p = 0. 554). Pregnancy rates (37.1% versus 13.3%, p = 0.1076) were not statistically different in both groups. CONCLUSION: In spite of lower ovarian reserve and thanks to an increased total dose of FSH, FMR1 premutated selected patients seem to have similar ovarian response as fully mutated patients. Neither the number of CGG repeats in FMR1 gene nor FMR1 mutation status was good predictors of the number of retrieved oocytes.
Subject(s)
Fragile X Mental Retardation Protein/genetics , Fragile X Syndrome/genetics , Mutation , Ovarian Reserve/genetics , Pregnancy Rate , Anti-Mullerian Hormone/blood , Anti-Mullerian Hormone/genetics , Female , Fertilization in Vitro , Heterozygote , Humans , Ovarian Reserve/physiology , Pregnancy , Preimplantation Diagnosis , Retrospective StudiesABSTRACT
RESEARCH QUESTION: What is the real-world effectiveness of Fertistartkit® in women undergoing assisted reproductive technology (ART)? DESIGN: Retrospective cohort study including anonymized data of women undergoing ovarian stimulation for ART with Fertistartkit between April 2016 and November 2017 and follow-up of clinical outcomes up to February 2018. Data were collected from the electronic patient databases of 12 French ART centres. The main outcome was number of oocytes retrieved. All data were categorized according to female age (<25, 25-29, 30-34, 35-37, 38-39 and >39 years). RESULTS: A total of 1006 cycles from 914 women treated with Fertistartkit were included. At the time of first ovarian stimulation in the study, women were 34.9 ± 5.0 years old, with a median body mass index of 22.7 kg/m². Couples had been infertile for more than 4 years, with all patterns of causes of infertility. Ovarian stimulation was started with a median dose of 300 IU (interquartile range [IQR]: 150-300 IU) of Fertistartkit for 10 days (IQR: 9-11 days), so a median total dose of 2700 IU (IQR: 1800-3300 IU). The mean number of oocytes retrieved per cycle was 9.5 ± 6.8, and the mean number of mature oocytes per cycle was 7.4 ± 5.5. The obtained ongoing pregnancy per started cycle was 26.0% (95% confidence interval [CI]: 24.1-27.9) and the obtained ongoing pregnancy per puncture was 27.0% (95% CI: 25.0-29.0). CONCLUSIONS: This is the first cohort to describe Fertistartkit treatment management in real-life conditions. The real-world data show that Fertistartkit is an effective option for ovarian stimulation.
Subject(s)
Fertilization in Vitro/methods , Oocyte Retrieval , Ovulation Induction/methods , Reproductive Techniques, Assisted , Adult , Female , Humans , Pregnancy , Pregnancy Rate , Retrospective Studies , Treatment Outcome , Young AdultABSTRACT
Human endometrium has a key role in implantation process. The measurement of endometrial thickness is the most commonly used in clinical practice. Managing patients with thin endometrium still represents a major challenge for clinicians. The objective of this systematic review was to investigate all available interventions to improve endometrial thickness (EMT) in women with history of thin endometrium undergoing fresh or frozen-thawed embryo transfers (ET). We performed a comprehensive search of relevant studies from January 1978 to February 2018. The different strategies were categorized as hormonal, vascular, and growth factor approaches and specifically analyzed according to the type of ET. Thirty-one studies were included. Overall, quality of the evidence ranged from very low to moderate, with only few randomized controlled trials that support the use of either GnRH analogues in fresh ET or sildenafil in frozen ET for enhancing endometrial growth. Besides, intensified estradiol administration is a common approach that might improve EMT in frozen ET. The present review evidences the paucity of reliable data regarding the efficiency of different interventions aiming at increasing EMT before fresh or frozen-thawed ET. Robust and high-quality randomized controlled trials are still needed before guidelines can be established.
Subject(s)
Embryo Transfer/methods , Endometrium/physiopathology , Infertility/therapy , Embryo Implantation/physiology , Female , Humans , Pregnancy , Pregnancy RateABSTRACT
RESEARCH QUESTION: To analyse why unbalanced viable offspring are derived mainly from the 3:1 segregation mode in t(11;22)(q23;q11.2) reciprocal translocation. DESIGN: Retrospective analysis of 24 pre-implantation genetic testing for chromosomal structural re-arrangements (PGT-SR) cycles was performed on seven male and five female carriers of t(11;22) translocation. Sperm analysis was performed on each male carrier. These patients were directed to the study centre after several years of miscarriages and/or abortions, primary infertility for male carriers or birth of an affected child. RESULTS: Twenty-four PGT-SR cycles were performed to exclude imbalances in both male and female carriers. The unbalanced embryos derived from the adjacent-1 segregation mode were the most represented in both male and female carriers (68.4% and 50%, respectively). These results were positively related with meiotic segregation analysis of reciprocal translocation in spermatozoa. A thorough analysis of the unbalanced embryo karyotypes determined that the expected viable +der22 karyotype resulting from 3:1 malsegregation was less represented at 5.3%. CONCLUSIONS: These findings highlight the divergence that may exist between meiotic segregation and post-zygotic selection. Post-zygotic selection would be responsible for the elimination of unbalanced embryos derived from the adjacent-1 segregation mode. The combined action of several factors occurs at the beginning of post-zygotic selection. Genetic counselling must consider the risk of a birth related to the adjacent-1 segregation mode, irrespective of the sex of the translocation carrier. These results will allow deeper understanding of the PGT results of t(11;22) carriers, which often include a high number of aneuploid embryos.
Subject(s)
Chromosomes, Human, Pair 11/genetics , Chromosomes, Human, Pair 22/genetics , Inheritance Patterns/genetics , Preimplantation Diagnosis/methods , Translocation, Genetic , Adult , Chromosome Mapping/methods , Chromosome Mapping/statistics & numerical data , Female , Gene Frequency , Genetic Carrier Screening/methods , Humans , In Situ Hybridization, Fluorescence/methods , In Situ Hybridization, Fluorescence/statistics & numerical data , Karyotyping , Male , Pregnancy , Preimplantation Diagnosis/statistics & numerical data , Retrospective Studies , Semen Analysis/methods , Semen Analysis/statistics & numerical data , Translocation, Genetic/geneticsABSTRACT
This study provides an overview of preimplantation genetic diagnosis (PGD) for single gene diseases and the management of expanding indications in the context of a fully financially covered service at Montpellier's regional hospital centre. Within the framework of a restrictive law ruling PGD in France, only the parental genetic risk can be studied in embryos (concurrent aneuploidy screening is not allowed). PCR-based techniques were developed combining mutation detection and closely linked short tandem repeat markers within or flanking the affected genes, and set up more than 100 different robust fluorescent multiplex assays for 61 monogenic disorders. This strategy was used to analyse blastomeres from cleavage-stage embryos. Overall, 893 cycles were initiated in 384 couples; 727 cycles proceeded to oocyte retrieval and 608 cycles to embryo transfer, resulting in 184 deliveries. Clinical pregnancy rate per transfer, implantation and miscarriage rates were 33.6%, 25.1% and 8.8%, respectively. Our PGD programme resulted in the birth of 214 healthy babies for 162 out of 358 couples (45.3%), constituting a relevant achievement within an organizational framework that does not allow aneuploidy screening but provides equal access to PGD, both geographically and socioeconomically. This is a rare example of a fully free-of-charge PGD service.
Subject(s)
Preimplantation Diagnosis/statistics & numerical data , Female , France , Genetic Diseases, Inborn/diagnosis , Hospitals, Public/statistics & numerical data , Humans , Male , National Health Programs , Pregnancy , Retrospective StudiesABSTRACT
In human eggs, aneuploidy increases with age and can result in infertility and genetic diseases. Studies in mouse oocytes suggest that reduced centromere cohesion and spindle assembly checkpoint (SAC) activity could be at the origin of chromosome missegregation. Little is known about these two features in humans. Here, we show that in human eggs, inter-kinetochore distances of bivalent chromosomes strongly increase with age. This results in the formation of univalent chromosomes during metaphase I (MI) and of single chromatids in metaphase II (MII). We also investigated SAC activity by checking the localization of BUB1 and BUBR1. We found that they localize at the kinetochore with a similar temporal timing than in mitotic cells and in a MPS1-dependent manner, suggesting that the SAC signalling pathway is active in human oocytes. Moreover, our data also suggest that this checkpoint is inactivated when centromere cohesion is lost in MI and consequently cannot inhibit premature sister chromatid separation. Finally, we show that the kinetochore localization of BUB1 and BUBR1 decreases with the age of the oocyte donors. This could contribute to oocyte aneuploidy.
Subject(s)
Aneuploidy , Kinetochores/metabolism , Oocytes/metabolism , Protein Serine-Threonine Kinases/metabolism , Humans , Oocytes/cytology , Protein TransportABSTRACT
This manuscript presents a molecularly demonstrated gonadal mosaicism from paternal origin for X-linked dominant chondrodysplasia punctata by single sperm typing. A couple who had experienced two medical terminations of pregnancy of female fetuses was referred to our pre-implantation genetic diagnosis (PGD) centre with the diagnosis of maternally derived gonadal mosaicism. Indeed, genetic analyses of different DNA samples - including semen - from the healthy parents failed to detect the variant found in the fetuses. Six embryos, all male, were obtained during the PGD cycle. The causative variant was not detected in any embryo, whereas five embryos had inherited the 'at-risk' maternal haplotype. The assumption of a maternal gonadal mosaicism was still possible, but this finding allowed us to consider the possibility of a paternal rather than maternal gonadal mosaicism. It prompted us to perform extensive single sperm analyses, demonstrating a low-frequency paternal germline mosaicism, which led to completely different haplotype phasing and PGD counselling. In conclusion, this case further exemplifies that germline mosaicism is a pitfall in PGD where diagnosis largely relies on linkage analysis and suggests that tracing the parental inheritance through polar body analysis and/or single sperm typing experiments is of major importance for adequate genetic counselling and accurate PGD. © 2016 John Wiley & Sons, Ltd.
Subject(s)
Chondrodysplasia Punctata/diagnosis , Genetic Diseases, X-Linked/diagnosis , Mosaicism , Paternal Inheritance/genetics , Preimplantation Diagnosis , Single-Cell Analysis/methods , Spermatozoa/metabolism , Adult , Chondrodysplasia Punctata/genetics , Diagnostic Errors , Female , Genetic Diseases, X-Linked/genetics , Genetic Testing/methods , Germ Cells , Humans , Male , Maternal Inheritance/genetics , Pedigree , Pregnancy , Preimplantation Diagnosis/methods , Preimplantation Diagnosis/standards , Recurrence , Spermatozoa/cytologyABSTRACT
OBJECTIVE: To examine if a balanced female embryo with X-autosome translocation could, during its subsequent development, express an abnormal phenotype. DESIGN: Preimplantation genetic diagnosis (PGD) analysis on two female carriers with maternal inherited X-autosome translocations. SETTING: Infertility center and genetic laboratory in a public hospital. PATIENT(S): Two female patients carriers undergoing PGD for a balanced X-autosome translocations: patient 1 with 46,X,t(X;2)(q27;p15) and patient 2 with 46,X,t(X;22)(q28;q12.3). INTERVENTION(S): PGD for balanced X-autosome translocations. MAIN OUTCOME MEASURE(S): PGD outcomes, fluorescence in situ hybridization in biopsied embryos and meiotic segregation patterns analysis of embryos providing from X-autosome translocation carriers. RESULT(S): Controlled ovarian stimulation facilitated retrieval of a correct number of oocytes. One balanced embryo per patient was transferred and one developed, but the patient miscarried after 6 weeks of amenorrhea. In X-autosome translocation carriers, balanced Y-bearing embryos are most often phenotypically normal and viable. An ambiguous phenotype exists in balanced X-bearing embryos owing to the X inactivation mechanism. In 46,XX embryos issued from an alternate segregation, der(X) may be inactivated and partially spread transcriptional silencing into a translocated autosomal segment. Thus, the structural unbalanced genotype could be turned into a viable functional balanced one. It is relevant that a discontinuous silencing is observed with a partial and unpredictable inactivation of autosomal regions. Consequently, the resulting phenotype remains a mystery and is considered to be at risk of being an abnormal phenotype in the field of PGD. CONCLUSION(S): It is necessary to be cautious regarding to PGD management for this type of translocation, particularly in transferred female embryos.
Subject(s)
Chromosome Disorders/genetics , Embryo Transfer/methods , Phenotype , Preimplantation Diagnosis/methods , Translocation, Genetic/genetics , X Chromosome Inactivation/genetics , Adult , Chromosome Disorders/diagnosis , Female , Humans , Male , Ovulation Induction/methods , PregnancyABSTRACT
Cell-free DNA (cfDNA) fragments, detected in blood and in other biological fluids, are released from apoptotic and/or necrotic cells. CfDNA is currently used as biomarker for the detection of many diseases such as some cancers and gynecological and obstetrics disorders. In this study, we investigated if cfDNA levels in follicular fluid (FF) samples from in vitro fertilization (IVF) patients, could be related to their ovarian reserve status, controlled ovarian stimulation (COS) protocols and IVF outcomes. Therefore, 117 FF samples were collected from women (n = 117) undergoing IVF/Intra-cytoplasmic sperm injection (ICSI) procedure and cfDNA concentration was quantified by ALU-quantitative PCR. We found that cfDNA level was significantly higher in FF samples from patients with ovarian reserve disorders (low functional ovarian reserve or polycystic ovary syndrome) than from patients with normal ovarian reserve (2.7 ± 2.7 ng/µl versus 1.7 ± 2.3 ng/µl, respectively, p = 0.03). Likewise, FF cfDNA levels were significant more elevated in women who received long ovarian stimulation (> 10 days) or high total dose of gonadotropins (≥ 3000 IU/l) than in women who received short stimulation duration (7-10 days) or total dose of gonadotropins < 3000 IU/l (2.4 ± 2.8 ng/µl versus 1.5 ± 1.9 ng/µl, p = 0.008; 2.2 ± 2.3 ng/µl versus 1.5 ± 2.1 ng/µl, p = 0.01, respectively). Finally, FF cfDNA level was an independent and significant predictive factor for pregnancy outcome (adjusted odds ratio = 0.69 [0.5; 0.96], p = 0.03). In multivariate analysis, the Receiving Operator Curve (ROC) analysis showed that the performance of FF cfDNA in predicting clinical pregnancy reached 0.73 [0.66-0.87] with 88% specificity and 60% sensitivity. CfDNA might constitute a promising biomarker of follicular micro-environment quality which could be used to predict IVF prognosis and to enhance female infertility management.
Subject(s)
DNA/analysis , Fertilization in Vitro , Follicular Fluid/chemistry , Adult , Biomarkers/analysis , Cell-Free System/chemistry , Female , Fertilization in Vitro/methods , Humans , Infertility, Female/metabolism , Infertility, Female/therapy , Ovarian Reserve , Ovulation Induction/methods , Polycystic Ovary Syndrome/metabolism , Pregnancy , Pregnancy Outcome , Prognosis , Prospective Studies , Sperm Injections, Intracytoplasmic , Young AdultABSTRACT
The majority of fluorescence in situ hybridization (FISH) studies on the meiotic segregation of Robertsonian translocations focus on the most common types, rob(13; 14) and rob(14; 21). Here we report the first study for carriers of rare Robertsonian translocations rob(13; 21) and rob(15; 22) combining analysis of meiotic segregation in sperm and blastomeres following pre-implantation genetic diagnosis (PGD). Dual-colour FISH was applied to nuclei from spermatozoa and blastomeres from PGD embryos using two subterminal contig probes for each translocation, and a second round with probes for chromosomes 16 and 18. Patient 1 had a rob(13; 21) and patient 2 had a rob(15; 22), and 86.3% and 87.5% of gametes respectively were consistent with meiotic segregation resulting in a normal or balanced chromosome complement. Analysis of embryos showed that for patient 1 and 2 respectively, 25% and 46% were balanced, and of the unbalanced embryos, 50% and 31% were mosaic or chaotic. Our patients with a rob(13; 21) and rob(15; 22) were found to have a similar meiotic segregation pattern to that for male carriers of the common Robertsonian translocations. The observed rate in unbalanced embryos being mosaic or chaotic may result in an increased risk of chromosomal abnormalities. Our results may help to improve the genetic counseling for carriers of rare Robertsonian translocations.
Subject(s)
Blastocyst/metabolism , Blastomeres/metabolism , Infertility, Male/genetics , Spermatozoa/metabolism , Adult , Blastocyst/pathology , Blastomeres/pathology , Chromosomes, Human, Pair 13/genetics , Chromosomes, Human, Pair 15/genetics , Chromosomes, Human, Pair 21/genetics , Chromosomes, Human, Pair 22/genetics , Genetic Counseling , Humans , In Situ Hybridization, Fluorescence , Male , Meiosis/genetics , Spermatozoa/pathology , Translocation, GeneticABSTRACT
OBJECTIVE: To directly study the meiotic segregation of a complex reciprocal translocation (CCR) as well as the occurrence of an interchromosomal effect. DESIGN: In situ sperm fluorescence in situ hybridization (FISH) analysis. SETTING: Department of Cytogenetics and INSERM research center. PATIENT(S): A male carrier of a balanced complex reciprocal translocation t(5;13;14)(q23;q21;q31). INTERVENTION(S): Sperm samples from the carrier and direct FISH analysis on sperm slide preparations. MAIN OUTCOME MEASURE(S): Meiotic segregation pattern determined on sperm nuclei and estimation of the incidence of unbalanced spermatozoa and an interchromosomal effect (ICE). RESULT(S): Only 27% of spermatozoa displayed a normal or balanced chromosome complement. The rate of unbalanced sperm was 69.4%, including different types of 3:3, 4:2, and 5:1 segregations. There was no evidence for the occurrence of an interchromosomal effect in autosomal chromosomes, but the gonosomes displayed a statistically significant increase in disomy rates. CONCLUSION(S): These results are consistent with the formation of a hexavalent configuration at the pachytene stage of meiosis and a high prevalence of imbalance production. The mechanisms of formation of CCRs must be examined with regard to these direct results and new molecular data on the formation of genomic rearrangements.