Preimplantation Genetic Diagnosis Cycles Research Articles

Effect of low-molecular-weight heparin in women undergoing frozen-thawed embryo transfer cycles: a retrospective cohort study

BackgroundRecurrent pregnancy loss (RPL) and recurrent implantation failure (RIF) during in vitro fertilization (IVF) treatment are still tough problems without effective treatments; thus, they are important research topics. There is controversy on whether low molecular weight heparin (LMWH) improves pregnancy outcomes in women with unexplained RPL and RIF. Moreover, currently, there is a paucity of reports on the role of LMWH in the entire population undergoing frozen-thawed embryo transfer (FET) cycles. This study aimed to estimate the effects of LMWH on pregnancy outcomes in women undergoing FET cycles.MethodsThere were 1881 female patients included in the study. Of the 1881 patients, 107 underwent preimplantation genetic diagnosis cycles, which were analyzed individually. The patients were divided into two groups: the LMWH group received injections of 4100 IU/d LMWH from the day of transfer until 14 ± 2 days posttransplant, the control group was the comparison group (without LMWH use). The baseline characteristics and reproductive outcomes of the patients were reviewed.ResultsOf the 1774 women with normal FET cycles, no significant differences were found in the number of embryos implanted (1.31 ± 0.02 vs. 1.28 ± 0.02), embryo implantation rate, biochemical pregnancy rate, clinical pregnancy rate, live birth rate, late abortion rate, and ectopic pregnancy rate between the two groups. The LMWH group had a higher early abortion (17.8% [76/427] vs. 12.5% [55/439], p = 0.030). In the sub-group analysis, among the patients who underwent more than four transfers, the LMWH group had a lower late abortion rate (1.7% [1/60] vs. 13.2% [7/53], p = 0.043). Similarly, of the 107 women who underwent preimplantation genetic diagnosis cycles, the reproductive outcomes were comparable between the two groups.ConclusionIn the general population and PGD patients, LMWH did not improve pregnancy outcomes. Therefore, the routine use of LMWH is not recommended for early treatment.

Effect of Parity on Placental Histopathology Features in Singleton Live Births Following IVF.

The aim of this study was to evaluate the effect of parity (primipara vs multipara) on the histopathology of the placenta in singleton live births following in vitro fertilization. We conducted a retrospective cohort study evaluating data of all IVF resulted live births from one university affiliated hospital during 2009-2017. All patients had the placenta sent for pathological evaluation. Exclusion criteria were history of miscarriage or elective termination of pregnancy, abnormal uterine cavity findings, previous uterine surgery, in vitro maturation cycles, gestational carrier cycles, oocyte recipient cycles, preimplantation genetic diagnosis cycles, and multiple pregnancies. The outcomes measured included anatomical, inflammation, vascular malperfusion, and villous maturation placental features. A multivariate analysis was conducted to adjust the results for factors potentially associated with placental pathology features. A total of 395 live births were included in the final analysis and were allocated to the study groups according to parity: primipara (n = 273) and multipara (n = 122). After adjustment for potential confounding factors, multiparity was found to be significantly associated with delayed villous maturation (OR 4.9; 95% CI 1.2-19.8) and primiparity was significantly associated with maternal vascular malperfusion (OR 0.6; 95% CI 0.3-0.8). We showed that parity has an impact on placental histopathological changes which in turn may affect perinatal outcome.

GENETIC SEGREGATION OF MITOCHONDRIAL 3697>GA MUTATION IN PGD CYCLES

To explore the dynamics and inheritance of m.3697G>A mutation and the performance of preimplantation genetic diagnosis (PGD) for mutant carriers. To analyze whether the cumulus cells surrounding oocytes can be used as a noninvasive PGD indicator for maternal inherited mtDNA mutation. The study encompassed a pedigree of four m.3697G>A carriers, including one asymptomatic PGD patient and one LS patient. Next generation sequencing (NGS) was used to detect m.3697G>A heteroplasmy in blood samples from the pedigree. Twelve cumulus oocyte complexes (COCs) were collected in the first PGD cycle and 11 COCs in the second cycle from the patient. Cumulus cells were isolated from COCs. Trophectoderm (TE) cells were biopsied from blastocysts on day 5 or day 6. The mutation load of TE biopsies, arrested embryos, blastomeres, polar bodies, the degenerate oocyte and corresponding cumulus cells were analysed via NGS. From 23 COCs, 20 oocytes were fertilized successfully via intracytoplasmic sperm injection. On day 5 or day 6 post-fertilization, 15 blastocysts were obtained and biopsied. The mutation load of the 15 TE biopsies (ranged from 15.2% to 100%), allowing selection of embryos for implantation. In the first cycle, a blastocyst with mosaic chromosomal copy number and mutation load of 31.7% was opted for transfer. However, the patient failed to yield a clinical pregnancy. In the second PGD cycle, a euploid blastocyst with mutation load of 53.9% was selected to transfer, which enable her to carry a successful pregnancy. Now she is in the stage of second trimester. Mutation load of cumulus cells may not correlate well with TE mutation load. This study first reported that m.3697G>A mutation showed a random pattern of inheritance in PGD embryos, it is almost impossible to produce zero or very low heterogenous embryos. But PGD could be applied to reduce risk of inheritance of a high mutation load and to yield successful pregnancy. Cumulus cells are not a suitable predictor of late-stage embryo mutation load.

Effect of newborn gender on placental histopathology and perinatal outcome in singleton live births following IVF

Research question: Does newborn gender affect placental histopathology pattern and perinatal outcome in singleton live births following IVF treatment?Design: Retrospective cohort study evaluating data of all live births from one academic tertiary hospital following IVF treatment during 2009–2017. All patients had placentas sent for pathological evaluation irrelevant of maternal and fetal complications status. Exclusion criteria were abnormal uterine cavity findings, previous uterine surgery, in-vitro maturation cycles, gestational carrier cycles, oocyte recipient cycles, preimplantation genetic diagnosis cycles and multiple pregnancies. The primary outcomes included anatomical, inflammation, vascular malperfusion and villous maturation placental features. The secondary outcomes included fetal, maternal, perinatal and delivery complications. A multivariate analysis was conducted to adjust the results for factors potentially associated with placental pathology features.Results: A total of 1057 live births were included in the final analysis and were allocated to the study groups according to fetal gender: males (n = 527) and females (n = 530). After adjustment for potential confounding factors, male gender was significantly associated with villous agglutination (odds ratio [OR] 9.8; 95% confidence interval [CI] 1.4–78.2), avascular villi (OR 4.1; 95% CI 1.3–12.6) and maternal vascular malperfusion (OR 1.8; 95% CI 1.2–2.7). Female gender was significantly associated with bilobed placenta (OR 0.2; 95% CI 0.06–0.8) and subchorionic thrombi (OR 0.5; 95% CI 0.3–0.9). The prevalence of adverse fetal, maternal and delivery outcomes was similar between the groups.Conclusions: Newborn gender has a significant impact on the placental histopathology pattern, which can contribute to the development of adverse perinatal outcomes.

Cumulative outcome of pre-implantation genetic diagnosis for sickle cell disease: a 5-year review.

To review the cumulative outcome of pre-implantation genetic diagnosis (PGD) cycles performed for prevention of sickle cell disease (SCD). Couples referred for PGD for SCD between April 2012 and October 2017 were included. Ovarian stimulation was performed using a short gonadotrophin-releasing hormone (GnRH) antagonist protocol and follicle-stimulating hormone injections. The GnRH agonist was used to trigger oocyte maturation. Oocytes were fertilised using intracytoplasmic sperm injection. Trophectoderm biopsy was performed on day 5 or 6 followed by vitrification. Genetic testing was done using pre-implantation genetic haplotyping. A total of 60 couples started 70 fresh PGD cycles (mean 1·2cycles/couple) and underwent a total of 74 frozen-embryo-transfer (FET) cycles (mean 1·3FET/couple). The mean (SD) female age was 33(4·4)years and the mean (SD) anti-müllerian hormone level was 22·9(2·8)pmol/l. The cumulative live-birth rate was 54%/PGD cycle started and 63%/couple embarking on PGD. The rate of multiple births was 8%. The cumulative outcome of PGD treatment for prevention of SCD transmission is high and PGD treatment should be offered to all at-risk couples.

Preimplantation genetic diagnosis for retinoblastoma survivors: a cost-effectiveness study.

This study aimed to investigate the cost-effectiveness of preimplantation genetic diagnosis (PGD) for the reproductive choices of patients with heritable retinoblastoma. The study modelled the costs of three cycles of in-vitro fertilization (IVF) and PGD across all uptake rates of PGD, number of children affected with retinoblastoma at each uptake rate and the estimated quality-adjusted life years (QALYs) gained. Cost-effectiveness analysis was conducted from the Australian public healthcare perspective. The intervention was the use of three cycles (one fresh and two frozen) of IVF and PGD with the aim of live births unaffected by the retinoblastoma phenotype. Compared with the standard care pathway (i.e. natural pregnancy), IVF and PGD resulted in a cost-saving to 18 years of age of AUD$2,747,294 for a base case of 100 couples with an uptake rate of 50%. IVF and PGD resulted in fewer affected (n = 56) and unaffected (n = 78) live births compared with standard care (71 affected and 83 unaffected live births), and an additional 0.03 QALYs per live birth. This modelling suggests that the use of IVF and PGD to achieve an unaffected child for patients with heritable retinoblastoma resulted in an overall cost-saving. There was an increase in QALYs per baby across all uptake rates. However, in total, fewer babies were born following the IVF and PGD pathway.

64. CAN FISH PROBE STRATEGIES INFLUENCE MOSAIC EMBRYO RATES IN ROBERTSONIAN TRANSLOCATION PGD?

Introduction Robertsonian translocation (RT) results from centric fusion of two acrocentric chromosomes. RT carriers are phenotypically normal, but they are known to be at increased risk of repeated miscarriages or of pregnancies resulting in the birth of a child with congenital anomalies, mental retardation. Preimplantation Genetic Diagnosis (PGD) is therefore a solution for RT carriers. A suitable probe strategy on cleavage stage allows us to differentiate balanced embryos, unbalanced embryos, mosaic and chaotic embryos. According to ESHRE recommendations, only balanced embryos are considered for transfer. However, transfer of some mosaic embryos has resulted in live births. We performed the first comparative analysis between a two or a three probes FISH strategies, in order to enhance the safety and security of embryos transferred and to establish the best reliable and low-cost strategy for RT PGD management. Material and methods Retrospective analysis of 253 preimplantation genetic diagnosis (PGD) cycles were performed on 91 men and 42 women carriers of different Robertsonian translocations t(13;14), t(14;21), t(14;15), t(15;21), t(13;15), t(13;21), t(14;22), t(15;22), t(21;22), t(13;13). The first strategy involved two relevant probes use. Each probe is located within the terminal long arm. The second strategy required a third probe addition on one of the two translocated chromosomes. The additional probe is located close to centromeric regions. Result(s) Regardless of probe strategies, 253 PGD cycles were performed in the aim to exclude imbalances in both carriers. On a total of 1,549 biopsied cells, 1,458 were successfully analyzed. Among these, 232 were transferred. Clinical pregnancy was achieved in 108 patients, and 85 healthy babies were delivered. No significant difference was observed in the balanced embryo rates (40.5% and 40.4% in a two probes and a three probe strategies respectively). Among these balanced embryos, diagnostic established on single or two mononucleate cells, wasn't affected by the probe strategies. However, a significant difference (p Conclusion(s) According to the present findings, concerning balanced embryo rates, the two probes strategy seems more suitable for Robertsonian translocation PGD analysis, firstly because it led to similar results as the three probes strategy. Secondly, this study also highlighted an increase mosaic embryo rate observed with the two probes strategy. Recent data indicated that mosaic embryos may represent a second category to transfer after balanced embryos. Obviously, balanced embryos must be transferred in priority. However in absence of balanced embryos, transfer of a mosaic embryo should be considered. In this case, genetic counseling about risks and potential benefits is of utmost importance to ensure informed decision-making by patients.

SAT-279 Can Preimplantation Genetic Diagnosis Be Used for Monogenic Endocrine Diseases?

Background: Preimplantation genetic diagnosis (PGD) is currently used for over 400 monogenic diseases. Some endocrine conditions that occur due to monogenic defects are either life threatening or can cause severe morbidities; thus, PGD may be an option to avoid the occurrence of such diseases. But a high cost and limited availability of licensed IVF centers can be a barrier for many patients who might otherwise benefit from PGD. Objective: To review the literature for endocrine monogenic disorders that have been prevented with PGD. Methods: An initial search in PubMed/Medline search was done to identify monogenic endocrine conditions using MeSH term “preimplantation diagnosis” and other searches using “preimplantation genetic diagnosis (PGD)”, plus “endocrine” or “monogenic” anywhere in the title or abstract. Studies were eligible for consideration if they included use of PGD for any known monogenic endocrine disease. Studies were omitted if the disease was non-endocrine or PGD was not used for embryo selection. Eleven articles (1999-2018) reported 15 cases using PGD for monogenic endocrine diseases. These were Congenital adrenal hyperplasia (n=2), X-linked adrenoleukodystrophy (n=2), Congenital lipoid adrenal hyperplasia (n=1), Persistent hyperinsulinemic hypoglycemia of infancy (n=4), Pseudohypoparathyroidism type 1a (n=1), Hypoparathyroidism-retardation-dysmorphism syndrome (n=2) and Multiple endocrine neoplasia types 1 (n=1) and 2a (n=2). Techniques used for PGD included PCR, whole genome amplification, FISH and more recently next generation sequencing. Clinical and outcome data of these cases were reviewed with respect to number of PGD cycles, successful pregnancy rates, live births and their genetic status. Results: Fifteen couples underwent 32 PGD cycles (ranging from 1-9 per couple), of which 17 cycles resulted in a pregnancy. Seven couples underwent a single PGD cycle. Four of these 7 couples had successful pregnancies each resulting in live births (all singletons), 1 couple was unable to achieve pregnancy, 1 chose to terminate the pregnancy as the karyotype detected a Turner fetus (45XO) and outcome data was not reported in one. The remaining 8 couples underwent multiple PGD cycles (2-9 per couple) and all had successful pregnancies in at least one cycle, resulting in 16 livebirths (6 singletons and 5 twins). Amongst the total live births (n=20), 60% were genetically unaffected for the tested monogenic disorder and 40% were carriers of the autosomal recessive gene mutation for which PGD was performed. Conclusion: Preimplantation genetic diagnosis may be a potential tool for preventing the inheritance of severe monogenic endocrine diseases in future generations. The current use of PGD in endocrine disorders is rare, but provides a promising option on a case by case basis provided the optimal resources are available.

Preimplantation embryos sex ratios in couples with four or more children of same sex, what should be expected from a preimplantation genetic diagnosis cycle?

Our aim was to assess the preimplantation embryos’ sex ratios in couples with four or more children of same sex, undergoing sex selection for nonmedical reasons. We conducted a cohort-historical study of all consecutive patients admitted to the IVF-PGD program in a large tertiary center. We reviewed the computerized files of all consecutive women admitted to our IVF for sex selection for nonmedical reasons. Patients and their PGD cycle characteristics were compared according to the desired sex of their embryo and the mode of fertilization. Nine patients underwent a total of 19 PGD cycle attempts during the study period. Of the 77 embryos with complete molecular diagnosis, 41 revealed a male embryo and 36 a female embryo. Thirty-five percent of all the diagnosed embryos were of the desired sex. For couples desiring a boy, IVF cycles achieved a higher ratio of the desired embryonal sex compared to ICSI (52% vs 18.7%, p = .03). For couples desiring a girl, ICSI cycles had a higher percentage of the desired embryonal sex compared to IVF cycles (38% vs 23%). Moreover, 29.7% of ICSI and 40% of IVF embryos achieved the desired sex. In conclusion, PGD for sex selection results in a relatively low percentage (35%) of embryos demonstrating the desired sex. Nonetheless, selecting the mode of fertilization (ICSI/IVF) might improve the success rate. Further studies are required to explore the appropriate and cost-effective method for sex selection for nonmedical reasons.

Analysis of chromosomes of embryos derived from translocation carriers during preimplantation genetic diagnosis cycles

To explore the effect of chromosomal translocations on the composition of embryonic chromosomes and its mechanism. For 52 couples with one partner carrying a chromosomal translocation, results of next generation sequencing of all embryos derived from 61 cycles were divided into different groups based on the type of translocations, gender of the carrier, and maternal age. Effect of parental chromosomal translocations on the composition of embryonic chromosomes of each group was analyzed. A significant difference was found between carriers of reciprocal and Robertsonian translocations in terms of proportion of abnormal embryos and structurally normal chromosomes (63.3% vs. 27.5%, and 1.1% vs. 0.3%, respectively). Compared with male carriers, there was an increase in the rate of abnormalities for female carriers (67.2% vs. 58.3% for reciprocal translocations, and 45.5% vs. 13.8% for Robertsonian translocations). The risk for chromosomal abnormality also increased with the maternal age. No significant difference was found in the proportion of abnormal embryos between carriers divided by involvement of acrocentric chromosomes or terminal chromosomal breakpoints. The types of parental translocation, gender of carrier, maternal age, and interchromosomal effect have certain effect on the composition of embryonic chromosomes.

Post-warming degree of blastocoel re-expansion is the best prognosticator for live birth rates in vitrified-warmed single blastocyst transfer cycles

Recent studies have presented contradictory results regarding significance of Inner-cell-mass (ICM)/Trophectoderm (TE) gradation and degree of blastocoel expansion/re-expansion for increased live-birth rates. Also, few studies have reported combined results from fresh/frozen and single/double transfer cycles. This study aimed at individually assessing the relevance of grades of ICM, TE and degree of post-warm blastocoel re-expansion on live-birth rates exclusively in vitrified-warmed single-blastocyst transfer cycles. Retrospective study (n=216) vitrified-warmed elective single blastocyst transfer cycles. Oocyte donation, Embryo-donation, assisted hatching and preimplantation genetic diagnosis cycles were excluded. All blastocysts were graded as per Gardner and Schoolcraft method of classification as 1-6 for degree of expansion and grades A/B/C for ICM and TE. Natural-cycle endometrial preparation with hormone supplementation followed by luteal-phase support with micronized progesterone. Endometrial response was noted by ultrasound. Single blastocyst-transfer was done 3 hours after warming of vitrified blastocyst. Warmed blastocyst was compared with pre-vitrified blastocyst for degree of expansion, gradation of ICM/TE. β-hCG level measured on day8 of transfer indicated pregnancy. Live-birth was the primary outcome measure. Women were classified into Live-birth (LB; n=74) and non-pregnant (NP; n=142) groups. Age, BMI, infertility period, number of oocytes retrieved, rate of formation of good quality cleavage-stage and blastocyst-stage embryos and survival rates post-warming did not differ significantly between the two groups. Same brand of embryo-transfer catheter and same brand and volume of media was used for transfer. However, degree of re-expansion was significantly higher in LB group than in NP group (Mean ± SD: 3.6 ± 1.0 v 3.2 ± 0.94, P=0.0068). The Fisher-Exact test odds-ratio for achieving a live-birth with 3-4 degree of re-expansion was 2.78 (p=0.0016) whereas the odds ratio was much lower (0.36) with any other degree of re-expansion. Although ICM grade was higher/better in Live-birth group than in non-pregnant group, the difference remained statistically non-significant (Odds ratio 1.25, p=0.82). No significant difference was observed in TE grades between the two study groups (p=0.17). A notable difference was also observed in endometrial echopattern (p=0.03) although the endometrial thickness remained comparable between the groups. Thus, post-warming degree of re-expansion is the single most promising predictive factor for live birth rates in such cycles. Re-expansion of blastocoel to an expanded grade 3/4 is superior to ICM and TE cell gradation for better live-birth rates in vitrified-warmed single blastocyst transfer cycles. However, larger multicentric trials may be required to establish it as robust predictor of live-birth.

Comparison of two different next generation sequencing (NGS) systems, VeriSeq with whole genome amplification and FAST-Seqs with PCR for preimplantation genetic testing for aneuploidy (PGT-A)

VeriSeq with whole genome amplification (VeriSeq) and FAST-SeqS with PCR (FAST-SeqS) are two different methods of PGT-A and the former is reported as a more sensitive method for determination of aneuploidy including mosaicism. Here we compared the two PGT-A methods to determine if these would affect clinical outcome. Retrospective study by a private IVF center. 2,121 blastocysts from 414 cases were biopsied and NGS performed by VeriSeq or FAST-SeqS method. Preimplantation genetic diagnosis (PGD) cycles were excluded. All blastocysts were vitrified after trophectoderm biopsy. Endometrial thickness was measured at day 12 of frozen embryo transfer (FET) cycle. For pregnancy outcome, 367 FET cycles were analyzed. The statistical analysis was performed using t-test, chi-square test or z-test. A p value of <0.05 was considered statistically significant. 1,200 blastocysts biopsy from 233 cases and 921 blastocysts biopsy from 181 cases were tested by VeriSeq and FAST-SeqS, respectively. Average age of patients was not significantly different. No call rate of biopsy was not different (1.4%). Euploid rate of VeriSeq was significantly lower than that of FAST-SeqS (50.7% vs 63.0%, p=0.0028) and mosaicism rate of VeriSeq was significantly higher than that of FAST-SeqS (8.8% vs 4.0%, p<0.0001). Partial (segmental) aneuploidy rate was not significantly different (8% vs 7%). Pregnancy outcomes were summarized in Table 1. Perhaps due to increased sensitivity in methodology, the euploid rate was lower and the mosaicism rate was higher with VeriSeq. However, this had no effect on numerous markers of clinical success in IVF including ongoing pregnancy/live birth rates.Table 1Pregnancy outcomesVeriSeqFAST-SeqSAve. age of patients34.0±4.134.0±3.4Endometrial thickness (mm)9.3±2.39.6±2.4Ave. number of embryo transferred1.2±0.41.1±0.3Pregnancy rate67.3% (142/211)69.2% (108/156)Clinical pregnancy rate61.1% (129/211)66.7% (104/156)Ongoing/live birth rate52.6% (111/211)59.0% (92/156)Implantation rate53.2% (132/248)60.3% (105/174)Miscarriage rate21.8% (31/142)14.8% (16/108)the result is not significantly different Open table in a new tab

Multiple displacement amplification as the first step could increase diagnosis efficiency of preimplantation genetic diagnosis for α-thalassemia

To evaluate whether the use of multiple displacement amplification (MDA) as the first step could increase the diagnosis efficiency of preimplantation genetic diagnosis (PGD) for α-thalassemia. Retrospective study. We retrospectively analyzed 632 PGD cycles of α-thalassemia that were included in our center between January 2014 to December 2017. In this study, we divided patients into two groups based on two different detection methods. Embryos were biopsied on day 3 post-insemination or at the blastocyst stage. For the polymerase chain reaction (PCR) group(n=313), fluorescent gap PCR were performed directly after cells lysis. For the MDA+ PCR group(n=319), whole genome of single cell(s) were directly amplified using MDA and its products were diluted 1: 100 and then detected by fluorescent gap PCR for α-thalassemia. a total of 5441 embryos were diagnosed, of which 3064 embryos were diagnosed in the PCR group and 2377 embryos were diagnosed in the MDA+PCR group. No statistical differences were found in the baseline characteristics between two groups. However, a higher overall diagnostic efficiency was observed in the MDA+PCR group(96.51% versus 86.16%, P <0.01). Moreover,the rate of embryos available for transfer was significantly higher in the MDA+PCR group than in the PCR group (71.62% versus 62.23%, P<0.01). In addition, there was a statistically significant difference in the carrier rates between the two groups (MDA+PCR group:46.38% versus PCR group:31.74%, P <0.01). Furthermore, misdiagnosis rate was lower in the MDA+PCR group than in the PCR group, but it was not statistically significant (0% versus 1.84%, P=0.317). Besides, the implantation rate was significantly higher in the MDA+PCR group than in the PCR group (50.36% versus 40.23%, P <0.01). Clinical pregnancy rate, miscarriage rate, ectopic pregnancy rate, and live birth rate were similar in both groups. When MDA is used as the first step, diagnosis efficiency may be increased. It is highly effective for PGD for α-thalassemia and deserve wide usage.

Outcomes of Preimplantation Genetic Diagnosis Cycles by Fluorescent In situ Hybridization of Infertile Males with Nonmosaic 47,XYY Syndrome.

Background:The 47,XYY syndrome could result in fertility problems. However, seldom studies reported comprehensive researches on the embryonic development and pregnancy outcomes of these patients. This study aimed to evaluate the clinical outcomes of nonmosaic 47,XYY patients performed with fluorescent in situ hybridization (FISH) and preimplantation genetic diagnosis (PGD) treatment.Methods:This was a retrospective study. Between January 2012 and May 2017, 51 infertile males with nonmosaic 47,XYY syndrome underwent FISH-PGD were included in the study. According to sex chromosomal FISH results, embryos were classified as normal signal, no nuclei fixed, no signal in fixed nuclei, suspensive signal, and abnormal signal groups, respectively. The incidence of each group, the fixation rate, and hybridization rate were calculated. Embryonic development and pregnancy outcomes were also analyzed. The measurement data were analyzed with Student's t-test. The comparison of categorical data was analyzed with the Chi-square test and Fisher's exact test when expected cell count was <5.Results:The 53 PGD cycles with 433 embryos were analyzed. The fixation rate was 89.6%, while the hybridization rate was 96.4%. There were 283 embryos with two sex chromosomal signals with clear diagnosis (65.4%). The numbers of no nuclei fixed, no signal in fixed nuclei, suspensive signal, and abnormal signal groups were 45 (10.4%), 14 (3.2%), 24 (5.5%), and 67 (15.5%), respectively. Embryos with abnormal signals were abandoned. The number of good-quality embryos was 210 (57.4%), including implanted embryos on day 4/day 5 and cryopreserved. The rates of good-quality embryos in the no nuclei fixed (22.2%), no signal in fixed nuclei (28.6%), and suspensive signal groups (33.3%) were comparable (P > 0.05), and were significantly lower than the normal signal group (66.4%, P < 0.001). The clinical pregnancy rates of fresh and frozen embryos transferred cycles were 70.6% and 85.7%, respectively.Conclusions:Among embryos with a clear diagnosis of sex chromosome, about one-fifth showed abnormal signals. Embryos with two sex chromosomal signals are more likely to develop into good-quality ones. The application of the PGD by FISH may help to improve the clinical outcomes.

Importance of embryo aneuploidy screening in preimplantation genetic diagnosis for monogenic diseases using the karyomap gene chip

We investigated the incidence of aneuploidy in embryos from couples carrying monogenic diseases and the effect of embryo aneuploidy screening on the monogenic disease preimplantation genetic diagnosis (PGD). From November 2014 to April 2017, 36 couples carrying monogenic diseases were enrolled. The karyomap gene chip technique was used to analyze the blastocysts from the subjects and select normal embryos for transfer. A total of 43 single-gene PGD cycles were performed. A total of 687 eggs were obtained and 186 blastocysts were biopsed. After analysis via karyomap chip, 175 blastocysts received diagnostic results. In our monogenic disease PGD, 66.8% (117/175) of the embryos were diagnosed as normal or non-pathogenic (silent carriers), and 33.2% (58/175) of the embryos were diagnosed as abnormal or pathogenic. For preimplantation genetic screening (PGS), the aneuploidy rate of embryos was 22.9% (40/175). Among embryos diagnosed as normal for monogenic diseases, 26.5% (31/117) of the embryos were aneuploid and could not be transferred. Thus, approximately 1/4 of normal or non-pathogenic blastocysts diagnosed based on monogenic disease PGD were aneuploid, indicating the necessity and importance of embryo aneuploidy screening during PGD for monogenic diseases.

Clinical outcomes in carriers of complex chromosomal rearrangements: a retrospective analysis of comprehensive chromosome screening results in seven cases

To evaluate the clinical outcomes in carriers of complex chromosomal rearrangements (CCRs). Case series. An institute for reproductive and stem cell engineering. Seven couples with CCRs. Assisted reproduction with preimplantation genetic diagnosis (PGD). PGD results, embryo rating, pregnancy outcomes. In cases 1, 2, 3, 4, 5, and 6, each woman underwent one cycle of PGD. Case 7 underwent two PGD cycles. We obtained 51 blastocysts from seven couples with CCR, of which 47 were eligible for biopsy; only 3 (5.9%) were normal/balanced, and 2 (3.9%) conceptions resulted. One healthy baby girl was born (the other was not yet born at the time of publication). Karyotyping revealed that the healthy baby girl was 46,XX. Although the patient with both a balanced translocation and a CCR (case 7) had 12 embryos available for biopsy, all were chromosomally unbalanced. It is interesting that 22 (57.9%) of the total 38 blastocysts were of high quality for type A CCRs, and 2 (15.4%) of the total 13 blastocysts were of high quality for type B CCR at day 6 after fertilization. The chances of identifying normal/balanced blastocysts in patients with CCR are <6%; the chances of a pregnancy are <4%. Greater complexity CCRs result in fewer transplantable embryos. Moreover, CCRs of greater complexity have a lower rate of high quality blastocysts than CCRs of less complexity.

Preimplantation genetic diagnosis of X-linked Charcot-Marie-Tooth disease by indirect linkage analysis

ObjectiveTo present methodical approach of preimplantation genetic diagnosis (PGD) as an option for an unaffected pregnancy in reproductive-age couples who have a genetic risk of the X-linked dominant peripheral neuropathy Charcot-Marie-Tooth type 1 disease. Patients and methodsWe performed PGD of X-linked Charcot-Marie-Tooth type 1 disease using haplotyping/indirect linkage analysis, when during analysis we reach to exclude embryos that carry a high-risk haplotype linked to the causal mutation p.Leu9Phe in the GJB1 gene. ResultsWithin the PGD cycle, we examined 4 blastomeres biopsied from cleavage-stage embryos and recommended 3 embryos for transfer. Two embryos were implanted into the uterus; however, it resulted in a singleton pregnancy with a male descendant. Three years later, the couple returned again with spontaneous gravidity. A chorionic biopsy examination of this gravidity ascertained the female sex and a pericentric inversion of chromosome 5 in 70% of the cultivated foetal cells. ConclusionUsing indirect linkage analysis, PGD may help to identify genetic X-linked defects within embryos during screening, thereby circumventing the potential problems with abortion.

Thirteen years' experience of 893 PGD cycles for monogenic disorders in a publicly funded, nationally regulated regional hospital service.

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.

Clinical outcomes after preimplantation genetic diagnosis of patients with Corino de Andrade disease (familial amyloid polyneuropathy)

The aim of this study was to determine whether patients with transthyretin-related hereditary amyloidosis (V30M), after transplantation or under tafamidis treatment, have normal gamete reproductive capacity. A retrospective analysis was carried out of all preimplantation genetic diagnosis (PGD) cycles performed in patients with the V30M mutation. The groups analysed were: total cases with V30M, female cases with V30M and male cases with V30M. Detailed demographic, stimulation, embryological, clinical and newborn outcomes were evaluated. Comparisons revealed that patients have a high likelihood of achieving a live birth per PGD treatment cycle (48%). This is the first large report on patients with the V30M mutation treated with PGD. The high rate of live birth obtained should represent a strong stimulus for patients to use PGD as it proved to be effective and safe. As a neurodegenerative disease that leads to death, it is of maximum importance that it could be eradicated using PGD in order to definitively avoid the transmission of the disease.

Preliminary analysis of numerical chromosome abnormalities in reciprocal and Robertsonian translocation preimplantation genetic diagnosis cases with 24-chromosomal analysis with an aCGH/SNP microarray.

The aim of this study was to determine whether an interchromosomal effect (ICE) occurred in embryos obtained from reciprocal translocation (rcp) and Robertsonian translocation (RT) carriers who were following a preimplantation genetic diagnosis (PGD) with whole chromosome screening with an aCGH and SNP microarray. We also analyzed the chromosomal numerical abnormalities in embryos with aneuploidy in parental chromosomes that were not involved with a translocation and balanced in involved parental translocation chromosomes. This retrospective study included 832 embryos obtained from rcp carriers and 382 embryos from RT carriers that were biopsied in 139 PGD cycles. The control group involved embryos obtained from age-matched patient karyotypes who were undergoing preimplantation genetic screening (PGS) with non-translocation, and 579 embryos were analyzed in the control group. A single blastomere at the cleavage stage or trophectoderm from a blastocyst was biopsied, and 24-chromosomal analysis with an aCGH/SNP microarray was conducted using the PGD/PGS protocols. Statistical analyses were implemented on the incidences of cumulative aneuploidy rates between the translocation carriers and the control group. Reliable results were obtained from 138 couples, among whom only one patient was a balanced rcp or RT translocation carrier, undergoing PGD testing in our center from January 2012 to June 2014. For day 3 embryos, the aneuploidy rates were 50.7% for rcp carriers and 49.1% for RT carriers, compared with the control group, with 44.8% at a maternal age <36years. When the maternal age was ≥36years, the aneuploidy rates were increased to 61.1% for rcp carriers, 56.7% for RT carriers, and 60.3% for the control group. There were no significant differences. In day 5 embryos, the aneuploidy rates were 24.5% for rcp carriers and 34.9% for RT carriers, compared with the control group with 53.6% at a maternal age <36years. When the maternal age was ≥36years, the aneuploidy rates were 10.7% for rcp carriers, 26.3% for RT carriers, and 57.1% for the control group. The cumulative aneuploidy rates of chromosome translocation carriers were significantly lower than the control group. No ICE was observed in cleavage and blastocyst stage embryos obtained from these carriers. Additionally, the risk of chromosomal numerical abnormalities was observed in each of the 23 pairs of autosomes or sex chromosomes from day 3 and day 5 embryos. There was not enough evidence to prove that ICE was present in embryos derived from both rcp and RT translocation carriers, regardless of the maternal age. However, chromosomal numerical abnormalities were noticed in 23 pairs of autosomes and sex chromosomes in parental structurally normal chromosomes. Thus, 24-chromosomal analysis with an aCGH/SNP microarray PGD protocol is required to decrease the risks of failure to diagnose aneuploidy in structurally normal chromosomes.