Embryo drop-out rates in preimplantation genetic testing for aneuploidy (PGT-A): a retrospective data analysis from the DoLoRes study.

PURPOSE: To evaluate the decline in transferable embryos in preimplantation genetic testing for aneuploidy (PGT-A) cycles due to (a) non-biopsable blastocyst quality, (b) failure of genetic analysis, (c) diagnosis of uniform numerical or structural chromosomal aberrations, and/or (d) chromosomal aberrations in mosaic constitution. METHODS: This retrospective multicenter study comprised outcomes of 1562 blastocysts originating from 363 controlled ovarian stimulation cycles, respectively, 226 IVF couples in the period between January 2016 and December 2018. Inclusion criteria were PGT-A cycles with trophectoderm biopsy (TB) and next generation sequencing (NGS). RESULTS: Out of 1562 blastocysts, 25.8% were lost due to non-biopsable and/or non-freezable embryo quality. In 10.3% of all biopsied blastocysts, genetic analysis failed. After exclusion of embryos with uniform or chromosomal aberrations in mosaic, only 18.1% of those originally yielded remained as diagnosed euploid embryos suitable for transfer. This translates into 50.4% of patients and 57.6% of stimulated cycles with no euploid embryo left for transfer. The risk that no transfer can take place rose significantly with a lower number of oocytes and with increasing maternal age. The chance for at least one euploid blastocyst/cycle in advanced maternal age (AMA)-patients was 33.3% compared to 52.1% in recurrent miscarriage (RM), 59.8% in recurrent implantation failure (RIF), and 60.0% in severe male factor (SMF). CONCLUSIONS: The present study demonstrates that PGT-A is accompanied by high embryo drop-out rates. IVF-practitioners should be aware that their patients run a high risk of ending up without any embryo suitable for transfer after (several) stimulation cycles, especially in AMA patients. Patients should be informed in detail about the frequency of inconclusive or mosaic results, with the associated risk of not having an euploid embryo available for transfer after PGT-A, as well as the high cost involved in this type of testing.

Comment on the recent PGDIS Position Statement on the Transfer of Mosaic Embryos 2021.

The worldwide demand of preimplantation genetic testing for aneuploidy (PGT-A) is still growing. However, chromosomal mosaic results greatly challenge the clinical practice. The recently published PGDIS Position Statement on the Transfer of Mosaic Embryos is the third PGDIS position statement on how to deal with embryos diagnosed as chromosomal mosaics (CM) and, one of many attempts of different societies and working groups to provide a guideline for clinicians, laboratories, clinics, and genetic counselors. But still, as in previous statements, many issues remained unresolved. Moreover, from our point of view, the question how to deal with embryos diagnosed as CM, consisting of two or more karyological cell lines cannot be separated from all the other aspects of PGT-A including its accuracy. The paucity of clearcut indications for PGT-A and evidence of benefit as well as an overall cost-benefit assessment is given below.

Diagnosing chronic endometritis: when simplification fails to clarify.

Reproductive immunology has grown in importance in recent years and has even developed into a discipline of its own within the field of reproductive medicine. Many aspects of reproductive failure such as repeated implantation failure or recurrent miscarriages are, meanwhile, seen as a consequence of aberrant expression of immunological factors. This is reflected by the increasing number of tests for assessing and quantifying different immune cell types as well as by a wide range of immune therapies offered to a clientele consisting of desperate patients requesting additional 'IVF tools': first, what is still usually disregarded is the enormous plasticity and fluctuation of most immune cells in the genital tract; second, their still poorly characterized functions in the endometrial cycle: further, their partially unknown role in embryo implantation and in establishing a pregnancy; and third, the fact that one of the fundamental hypotheses of reproductive immunology-of note-the Medawar concept or 'Medawar's Paradox' of semi-allogeneic graft embryo, is partially based on an erroneous assumption, i.e. the immunologic rejection and tolerance of an embryo. In the present opinion article, we comment on the diagnostic procedures and therapy approaches for chronic endometritis within the scope of reproductive medicine.

Medical research and reproductive medicine in an ethical context: a critical commentary on the paper dealing with uterine lavage published by Munné et al.

A recent study published in Human Reproduction claimed that uterine lavage offers a non-surgical, minimally invasive strategy for the recovery of human embryos from fertile women who do not want or need IVF for medical reasons but who desire preimplantation genetic testing (PGT) for embryos. To prove this hypothesis, the researchers recruited dozens of young Mexican women. The prospective oocyte donors underwent ovarian stimulation to induce the production of multiple mature oocytes. Subsequently, these women were inseminated by donor semen. A few days later, the developing embryos were collected by uterine lavage (uterine flushing) and subjected to genetic testing for aneuploidies (PGT-A). Oocyte donors with persistently elevated hCG levels, indicating the implantation of one or more embryos after uterine lavage, had to undergo uterine curettage and/or treatment with methotrexate. A critical opinion paper discussing the aforementioned study was published by De Santis and colleagues and has raised critical issues that are largely technical in nature. However, this opinion paper neglects-from our point of view-critical issues of the Mexican study regarding ethical principles and moral standards in human research. These aspects are summarized below.

Suboptimal endometrial-embryonal synchronization is a risk factor for ectopic pregnancy in assisted reproduction techniques.

RESEARCH QUESTION: What are the main risk factors associated with ectopic pregnancy and what is the true incidence of ectopic pregnancies in an IVF programme? DESIGN: Retrospective single-centre study of 12,429 blastocyst transfers (8182 fresh and 4247 frozen embryo transfers) conducted between January 2010 and December 2017. IVF outcome was analysed, and ectopic pregnancy risk evaluated according to patient's characteristics and assisted reproductive technology treatment factors. RESULTS: Of 5061 patients reporting a positive pregnancy test, 43 were diagnosed with ectopic pregnancy (0.85%). Neither female age (36.7 versus 35.8 years), body mass index, quality of transfer nor stimulation protocol affected the ectopic pregnancy rate, but history of previous ectopic pregnancy (OR 3.26; P = 0.0080), tubal surgery, or both (OR 6.20; P < 0.0001) did. The incidence of ectopic pregnancy was increased in women with uterine malformations (OR 3.85; P = 0.0052), uterine pathologies (OR 5.35; P = 0.0001), uterine surgeries (OR 2.29; P = 0.0154) or sub-optimal endometrial build-up (OR 4.46 to 5.31; P < 0.0001). Transfer of slow-developing blastocysts (expressed by expansion) significantly increased the risk of ectopic pregnancy (OR 2.59; P = 0.0102). CONCLUSIONS: Unfavourable uterine environment, including uterine pathologies, uterine or tubal surgery and suboptimal endometrial build-up were related to ectopic pregnancy. Low expansion grade of blastocysts was identified as an additional putative risk factor for ectopic pregnancy, indicating the importance of proper embryonal-maternal synchronization. The overall ectopic pregnancy rate after blastocyst transfer was low, comparable with reported ectopic pregnancy rates in spontaneous conceptions. Proper evaluation of tubal and uterine pathologies, optimizing endometrial preparation and the transfer of expanded blastocysts in a frozen embryo transfer cycle, might be beneficial.

Impact of high magnification sperm selection on neonatal outcomes: a retrospective study.

PURPOSE: The aim of this study was to compare the effect of the deselection of spermatozoa presenting vacuole-like structures using IMSI (intracytoplasmic morphologically selected sperm injection) with ICSI (intracytoplasmic sperm injection) by means of neonatal outcomes. METHODS: In a retrospective two-center analysis, a total of 848 successful IMSI or ICSI cycles ending with a live birth, induced abortion, or intrauterine fetal death (IUFD) were included. RESULTS: The IMSI and ICSI groups included 332 and 655 babies or fetuses, respectively. The parents were older in the IMSI group than in the ICSI group (mothers were 35.1 vs 32.9 years, and fathers were 39.1 vs 36.2 years). The multiple pregnancy rate was higher in the IMSI group. The mean pregnancy duration and mean birth weight were almost identical in both groups. There was no significant difference in major congenital malformations between the two groups. However, this rate was decreased in the IMSI group compared to that in the ICSI group (1.8 vs 3.2%), the difference being mainly found in singletons (1.4 vs 3.3%). Boys were more often affected than girls in both groups. The percentages of chromosomal abnormalities did not differ between the IMSI and ICSI groups (0.6 and 0.8%). The reported congenital malformations mainly affected the heart, urogenital, and musculoskeletal systems. CONCLUSIONS: In the present study, the malformation rates observed in the IMSI and ICSI groups were not significantly different, even if slightly lower after IMSI. However, the observed difference followed the same trends observed in previous reports, indicating the possible impact of IMSI on decreasing congenital malformation occurrences. This highlights the necessity to prospectively evaluate the impact of IMSI on neonatal outcome after IVF treatment.

Intrauterine administration of human chorionic gonadotropin does not improve pregnancy and life birth rates independently of blastocyst quality: a randomised prospective study.

BACKGROUND: Successful embryo implantation depends on a well-timed maternal-embryonic crosstalk. Human chorionic gonadotropin (hCG) secreted by the embryo is known to play a key role in this process and to trigger a complex signal transduction cascade allowing the apposition, attachment, and invasion of the embryo into the decidualized uterus. Production of hCG was reported to be dependent on blastocyst quality and several articles suggested that intrauterine hCG injection increases pregnancy and implantation rates in IVF patients. However, no study has as yet analysed birth rates as final outcome. Our objective was to determine whether clinical outcome after blastocyst transfer can be improved by intrauterine injection of hCG and whether this is dependent on blastocyst quality. METHODS: A prospective randomised study was conducted in two settings. In cohort A, hCG application was performed two days before blastocyst transfer. In cohort B, the administration of hCG occurred just prior to embryo transfer on day 5. For both cohorts, patients were randomised to either intrauterine hCG application or to the control group that received culture medium. Clinical outcome was analysed according to blastocyst quality of transferred embryos. RESULTS: The outcome of 182 IVF-cycles (cohort A) and 1004 IVF-cycles (cohort B) was analysed. All patients received a fresh autologous blastocyst transfer on day five. Primary outcomes were pregnancy rates (PR), clinical pregnancy rates (cPR), miscarriage rates (MR), and live birth rates (LBR). No improvement of clinical outcome after intrauterine hCG administration on day 3 (cohort A) or day 5 (cohort B) was found, independently of blastocyst quality transferred. The final outcome in cohort A: LBR after transfer of top blastocysts was 50.0 % with hCG and 53.3 % in the control group. With non-top blastocysts, LBR of 17.1 % (hCG) and 18.2 % (control) were observed (n.s.). In cohort B, LBR with top blastocysts was 53.3 % (hCG) and 48.4 % (control), with non-top blastocysts it came to 28.7 % (hCG) and 35.0 % (control). The differences between the groups were statistically not significant. Furthermore, we investigated a possible benefit of hCG administration in correlation with female age. In both age groups (<38 years and ≥ 38 years) we found similar LBR after treatment with hCG vs. medium. A LBR of 47.1 % vs. 48.7 % was obtained in the younger group and 26.6 % vs. 30.8 % in the older group. CONCLUSIONS: In contrast to previous studies indicating a substantial benefit from intrauterine hCG application in cleavage stage embryo transfers, in our study we could not find any evidence for improvement of clinical outcome in blastocyst transfer cycles, neither with top nor with non-top quality morphology.

Massage therapy improves in vitro fertilization outcome in patients undergoing blastocyst transfer in a cryo-cycle.

CONTEXT: Massage therapy is increasingly used to relieve physical and mental discomfort and is suggested as a safe therapeutic modality, without any significant risks or any known side effects. Although a multitude of complementary therapies, such as acupuncture, are applied in reproductive medicine, no information is available with regard to the application of massage as an adjuvant therapy in assisted-reproduction techniques (ARTs). OBJECTIVES: This study was intended to assess the effectiveness of a deep relaxation (andullation) therapy based on oscillating vibrations when used prior to embryo transfer (ET) in in vitro fertilization (IVF) cryo-cycles. DESIGN: The research team designed a retrospective, observational study. Participants willing to undergo the massage treatment were allocated to the intervention (andullation) group. SETTING: The study was performed at the IVF Centers Prof. Zech-Bregenz in Bregenz, Austria. PARTICIPANTS: A total of 267 IVF patients, with a mean age of 36.3 y, participated in this single-center study. INTERVENTION: All patients receiving a transfer of vitrified and warmed blastocysts between January and December 2012 were included in the evaluation. Prior to ET, the andullation group received a standardized program of therapy-a 30-min, deep relaxation massage on an oscillating (vibrating) device, whereas the control group did not. OUTCOME MEASURES: To determine efficacy, the primary outcomes that the study measured were (1) pregnancy rates (PRs), by testing urine and obtaining a positive ß-human chorionic gonadotropin (ß-hCG); and (2) ongoing, pregnancies (oPR), by observation of fetal heartbeat and birth rates (BR) as well as miscarriage rates. The patients' medical histories and types of infertility as well as the quality of the embryo transfers (ETs) were evaluated. RESULTS: In patients using the massage therapy prior to ET, significantly higher PRs, oPRs, and BRs were observed compared with the control group-PR: 58.9% vs 41.7%, P<.05; oPR: 53.6% vs 33.2%, P<.01; and BR: 32.0% vs 20.3%, P<.05. No differences were detected among groups for patients' ages, hormonal substitution protocols, endometrium structures and buildups, quality of transferred embryos, or quality of transfers. No adverse effects were noted in the massage group. CONCLUSIONS: The research team's results suggested that andullation therapy prior to blastocyst transfer in a cryo-cycle improves embryo implantation, most likely due to a reduction in stress (ie, a relaxation effect on patients), a reduction in uterine contractions, and, probably, an enhancement of the blood flow in the abdominal region. These findings provide support for use of andullation as a complementary therapy for ART.

Outpatient follicle monitoring: a plea for standardization in ultrasound based follicle monitoring and data transfer.

BACKGROUND: The complexity of assisted reproductive technology (ART) increased during the last decades. New scientific and medical findings as well as the statutory requirements for improving the safety and the outcome of ART were the main impetus for its development. While therapy planning is done and ART is used by the IVF centers, the medical support and monitoring of patients is conducted by referring gynecologists. Reported follicle measurements by the gynecologist allow the adoption of the therapy plan. Most notably, the crucial aspect is processing and interpretation of ultrasound scan (US). The results of the received US, the transfer of data between IVF center(s) and referred physician(s) as well as the subjective interpretation often culminate in interpretation and logistical problems. This might increase the error probability with considerable detriments for the patients and ART outcome. METHODS: The follicle monitoring was performed using Voluson I ultrasound system combined with SonoAVC(®) software. Results were communicated via DICOM language to DynaMed(®) software, a medical program for managing an IVF center with seamless integration of all processes needed for an accurate and precise workflow. RESULTS: In this study, no loss of data was detected. All data were integrated by DynaMed(®) software and were recallable in a fast and easy manner. CONCLUSION: The broad usage of Voluson I ultrasound SonoAVC(®) software and communication of the results via Picture Archiving and Communication System (PACS) server between the IVF center and local gynecologist would provide more assistance for the patients and consequently the ART outcomes can be improved.

Transfer of blastocysts with deviant morphological and morphokinetic parameters at early stages of in-vitro development: a case series.

Time-lapse imaging is increasingly applied as an adjunct to reproductive medicine. The gained information of the morphological and morphokinetic variables before the onset of transcription are supposed to be good predictors for the selection of the best embryo for transfer and are often seen in line with clinical outcomes. This retrospective case series investigated the outcome of transferred blastocysts that did not fulfil the proposed embryo scores at early cleavage or at later stages of development. The observations were made by time-lapse imaging. This study reports the birth of 16 healthy children after day-5 blastocyst transfer, of which at least one of the transferred embryos originated from deviant morphology and/or kinetic cleavage patterns. This case series suggests that some blastocysts derived from embryos with poor conventional morphological score and/or suboptimal morphokinetics can be successfully transferred and might result in live births. Such results might raise awareness that discarding embryos based only on early events is not a suitable approach to give patients the chance to conceive. In conclusion, to date only the transfer of viable embryos after culturing them until day 5 guarantees optimal embryo selection and helps to prevent embryo wastage.

Sperm head vacuoles are not affected by in-vitro conditions, as analysed by a system of sperm-microcapture channels.

Since the introduction of the motile sperm organelle morphology examination, there has been increasing recognition of the fact that the presence of large nuclear vacuoles might have deleterious effects on embryo development. Nevertheless, one fundamental question still being debated is whether specific in-vitro conditions during the handling of semen have an impact on vacuole formation. This study's objective was to analyse whether incubation temperature (20, 37°C) or oxidative stress stimulates the formation of nuclear vacuoles. Furthermore, it examined whether vacuoles disappear in the presence of an acrosome reaction inducer. Therefore, a system of sperm-microcapture channels was developed to permit the observation of the same living spermatozoa over a period of 24h. Neither incubation at 37°C nor induction of oxidative stress led to de-novo formation of nuclear vacuoles. Induction of the acrosome reaction using calcium ionophore A23587 did not lead to any modifications in the proportion of spermatozoa with vacuoles or to the disappearance of pre-existing vacuoles. According to these observations, it is concluded that nuclear vacuoles on the sperm head are already produced at earlier stages of sperm maturation and are not induced or modulated by routine laboratory environments. The examination of spermatozoa at very high magnification has led to the increasingly widespread recognition that the presence of large vacuoles in the human sperm head has deleterious effects on embryo development. One fundamental question, however, still remains: do specific conditions in the laboratory during the preparation and the handling of semen have an impact on vacuole formation? Our initial objective was to analyse whether different incubation temperatures (20, 37°C) and the induction of oxidative stress lead to the formation of sperm head vacuoles. Furthermore, we examined whether vacuoles disappear in the presence of an acrosome reaction inducer. In order to do this we developed a system of sperm-microcapture channels, which permits the observation of the same living spermatozoa over a period of 24h. Incubation at 37°C or induction of oxidative stress did not lead to the formation of any new vacuoles. After inducing the acrosome reaction, we did not detect any modification in the proportion of vacuolated spermatozoa. According to our observations, different temperatures or environmental conditions in the laboratory have no impact on the formation or disappearance of vacuoles. We conclude that sperm head vacuoles are already produced at earlier stages of sperm maturation.

The combination matters--distinct impact of lifestyle factors on sperm quality: a study on semen analysis of 1683 patients according to MSOME criteria.

BACKGROUND: Poor sperm quality can negatively affect embryonic development and IVF outcome. This study is aimed at investigating the influence of various lifestyle factors on semen quality according to MSOME (motile sperm organelle morphology examination) criteria. METHODS: 1683 male patients undergoing assisted reproductive technologies (ART) in our clinic were surveyed about their age, BMI (body mass index), ejaculation frequency, nutrition, sports, sleeping habits and social behavior. Semen samples were collected and evaluation of semen parameters according to MSOME and WHO criteria was performed. Results were grouped and statistically analyzed. RESULTS: Although single parameters had minor effects on sperm parameter, the combination of age, BMI, coffee intake, ejaculatory frequency and duration of sexual abstinence were identified as factors having a negative effect on sperm motility. Additionally, we could demonstrate that MSOME quality was reduced. The negative impact of age, BMI and coffee intake on sperm quality could be compensated if patients had a high ejaculation frequency and shorter periods of sexual abstinence. CONCLUSIONS: Combinations of adverse lifestyle factors could have a detrimental impact on sperm, not only in terms of motility and sperm count but also in terms of sperm head vacuolization. This negative impact was shown to be compensated by higher ejaculation frequency and a shorter period of sexual abstinence. The compensation is most likely due to a shorter storage time in the male gonads, thus reducing the duration of sperms' exposure to reactive oxygen species (ROS).

Effects of embryo transfer quality on pregnancy and live birth delivery rates.

BACKGROUND: To analyze the effects of embryo transfer (ET) quality on clinical pregnancy (CPR) and live birth delivery rates (LBDR). METHODS: In a retrospective study at a single, private infertility center between November 2005 and December 2009 one thousand fifty-five day-3 and day-5 ETs following IVF/ICSI/IMSI were evaluated. We analyzed the impact of an atraumatic ET with a soft catheter (ET 1), after external guidance (ET 2), after probing of the cervix with a stylet (ET 3), or after grasping the portio vaginalis with a tenaculum (ET 4) on CPR and LBDR. RESULTS: The use of external guidance showed a significantly reduced LBDR as compared to an atraumatic ET (26.0% vs. 32.5%). The lowest CPR and LBDR were found in ET 4. The application of stylets in cases of difficult ETs was superior to the use of external guidance. No differences in miscarriages between ET 1-4 were noted. CONCLUSIONS: Besides embryo culture and patient history, the quality of an ET might also have an important impact on pregnancy outcome. Techniques to ensure an atraumatic ET, such as mechanic uterine cavity length measurements, before starting treatment might help identify patients at risk for a difficult ET and lead to modified treatments, such as the primary use of a stylet. Limitation of study: retrospective analysis.

Dietary supplementation of antioxidants improves semen quality of IVF patients in terms of motility, sperm count, and nuclear vacuolization.

BACKGROUND: This study aimed to investigate the influence of an oral antioxidative supplementation on sperm quality of in vitro fertilization (IVF) patients, as analyzed by sperm motility according to the WHO criteria and motile sperm organelle morphology examination (MSOME). METHODS: Semen samples were collected from 147 patients before undergoing an IVF/intracytoplasmic morphologically-selected sperm injection (IMSI) cycle and 2 - 12 months after an antioxidative supplementation. Semen analysis was evaluated according to WHO and MSOME criteria. Spermatozoa were grouped according to the size of nuclear vacuoles within the sperm's heads. Patients were divided into oligoasthenoteratozoospermic (OAT) and non-OAT men. Between first and second semen analysis, patients were supplemented orally with an antioxidative preparation. RESULTS: After the antioxidative therapy we observed a significant reduction in the percentage of immotile sperm cells in the patients. Additionally, the percentage of class I spermatozoa according to MSOME criteria was significantly higher after antioxidative supplementation. In OAT patients the percentage of class I sperm was found to be increased, although not significantly. However, we observed a drastic improvement in sperm motility as well as in total sperm count in this group. CONCLUSION: The results demonstrated a considerable improvement in semen quality, notably in OAT patients. Considering the putative relationship between semen quality on the one hand and reactive oxygen species on the other, the observed changes in the sperm parameters indicate that a decline in semen quality, and even subtle morphological changes, might be associated with oxidative stress. Our findings suggest that an antioxidative and micronutrient supplementation has a remarkable benefit for IVF patients having restricted sperm parameters, in particular.

Dopamine agonist bromocriptine for the prevention of ovarian hyperstimulation syndrome.

The aim of this retrospective study is to investigate the frequency and severity of ovarian hyperstimulation syndrome and the pregnancy rate in a patient collective at risk who received bromocriptine treatment.