Non-invasive Embryo Assessment: Altered Individual Protein Profile in Spent Culture Media from Embryos Transferred at Day 5.

In order to improve ART outcome, non-invasive embryo assessment is gaining more and more attention. Therefore, the aim of this study is to determine the consecutive implantation potential via the secretome between blastocysts with or without implantation and to analyse possible interactions between these differentially expressed proteins. In this prospective study, 69 spent culture media from blastocysts transferred at day 5 were collected from patients undergoing IVF/ICSI treatment in a single IVF centre between April 2015 and November 2018 after informed consent and analysed individually. Exclusion criteria were the absence of informed consent, PCOS, endometriosis and maternal age > 42 years. Dependent on the treatment outcome, media were subsequently divided into two groups: from embryos who implanted successfully (n = 37) and from embryos without implantation (n = 32). Ninety-two proteins were measured simultaneously using the proximity extension assay (PEA) technology with the Olink® CVD III panel employing oligonucleotide-labelled antibodies. Statistical analysis was performed using the Kolmogorov-Smirnov test, Student's t test, the Mann-Whitney U test and Fisher's exact test. Media from implanted blastocysts showed significantly higher expression of EPHB4, ALCAM, CSTB, BMH, TIMP4, CCL24, SELE, FAS, JAM-A, PON3, PDGF-A, vWF and PECAM-1 compared with media from blastocysts without subsequent implantation. The highest relative expression change could be demonstrated for PECAM-1 and TIMP4. PECAM-1, SELE and vWF were co-expressed. Especially EPHB 4, SELE, ALCAM, MCP-1, CCL24, FAS, JAM-A and PDGF-A have already been described in early embryonic development and metabolism. Therefore, these proteins together with PECAM-1 indicate possible biomarkers for non-invasive embryo assessment in the future. However, due to the innovative methodology, defining a threshold for the use as biomarkers remains to be assessed.

Intraindividual Embryo Morphokinetics Are Not Affected by a Switch of the Ovarian Stimulation Protocol Between GnRH Agonist vs. Antagonist Regimens in Consecutive Cycles.

Background: The impact of controlled ovarian stimulation (COS) during medically assisted reproduction (MAR) on human embryogenesis is still unclear. Therefore, we investigated if early embryonic development is affected by the type of gonadotropin-releasing hormone (GnRH) analog used to prevent a premature LH surge. We compared embryo morphology and morphokinetics between GnRH agonist and antagonist cycles, both involving human chorionic gonadotropin (hCG)-trigger. To reduce possible confounding factors, we used intraindividual comparison of embryo morphokinetics in consecutive treatment cycles of the same patients that underwent a switch in the COS protocol. Methods: This retrospective cohort study analyzed morphokinetics of embryos from patients (n = 49) undergoing a switch in COS protocols between GnRH agonists followed by GnRH antagonists, or vice versa, after culture in a time-lapse incubator (EmbryoScope®, Vitrolife) in our clinic between 06/2011 and 11/2016 (n = 49 GnRH agonist cycles with n = 172 embryos; n = 49 GnRH antagonist cycles with n = 163 embryos). Among time-lapse cycles we included all embryos of the two consecutive cycles before and after a switch in the type of COS in the same patient. In-vitro fertilization (IVF) or intracytoplasmic sperm injection (ICSI) was performed and embryos were imaged up to day 5. Data were analyzed using Mann-Whitney U test or Fisher's exact test. The significance level was set to p = 0.05. Patients with preimplantation genetic screening cycles were excluded. Results: The mean age (years ± standard deviation) of patients at the time of treatment was 35.7 ± 4.3 (GnRH agonist) and 35.8 ± 4.0 (GnRH antagonist) (p = 0.94). There was no statistically significant difference in the number of oocytes collected or the fertilization rate. The numbers of top quality embryos (TQE), good-quality embryos (GQE), or poor-quality embryos (PQE) were also not different in GnRH agonist vs. antagonist cycles. We found no statistically significant difference between the analyzed morphokinetic parameters between the study groups. Conclusions: Our finding supports the flexible use of GnRH analogs to optimize patient treatment for COS without affecting embryo morphokinetics.

Serum levels of Pentraxin 3 differ significantly at the time of blastocyst transfer depending on implantation success: a pilot study.

OBJECTIVE: Many approaches try to identify the underlying molecular mechanisms to detect new potential biomarkers for successful artificial reproductive therapies. One factor has been described as a possible regulator of inflammation during implantation: Pentraxin 3 (PTX3), which seems to be essential for female fertility on one hand, but whose overexpression has been described in many obstetric complications based on abnormal placentation on the other hand. Therefore, we investigated if serum levels of PTX3 at the time of embryo transfer differ between women with an ongoing pregnancy compared to those without implantation. METHODS/DESIGN: During in vitro fertilization cycles of 51 patients, PTX3 levels at the time of embryo transfer were compared between patients without implantation (n = 26) and those with ongoing pregnancy (n = 25) using an enzyme-linked immunosorbent assay. Statistical analysis was performed using the Kolmogorov-Smirnov test, Fisher's exact test and Student's t test RESULTS: No significant differences were found concerning possible confounders (patients age, smoking pattern, embryo quality, number of embryos transferred and prior IVF attempts). Patients without implantation presented with significantly higher serum levels of PTX3 at the time of embryo transfer compared to women who became pregnant (0.781 ± 0.074 ng/ml vs. 0.578 ± 0.055 ng/ml, p < 0.05). CONCLUSIONS: PTX3 could present as a possible biomarker for ART success. The main limitation of this pilot study is its small sample size that needs validation with a larger study population.

Randomized, open trial comparing a modified double-lumen needle follicular flushing system with a single-lumen aspiration needle in IVF patients with poor ovarian response.

Study question: Is a modified double-lumen aspiration needle system with follicular flushing able to increase the mean oocyte yield by at least one in poor response IVF patients as compared to single-lumen needle aspiration without flushing? Summary answer: Follicular flushing with the modified flushing system did not increase the number of oocytes, but increased the procedure duration. What is known already: Most studies on follicular flushing were performed with conventional double-lumen needles in patients who were normal responders. Overall, these studies indicated no benefit of follicular flushing. Study design size, duration: Prospective, single-centre, randomized, controlled, open, superiority trial comparing the 17 G Steiner-Tan Needle® flushing system with a standard 17 G single-lumen aspiration needle (Gynetics®); time frame February 2015-March 2016. Participants/materials setting methods: Eighty IVF patients, 18-45 years, BMI >18 kg/m2 to <35 kg/m2, presenting with ≤ five follicles >10 mm in both ovaries at the end of the follicular phase were randomized to either aspirating and flushing each follicle 3× with the Steiner-Tan-Needle® automated flushing system (n = 40) or a conventional single-lumen needle aspiration (n = 40). Primary outcome was the number of cumulus-oocyte-complexes (COCs). Procedure duration, burden (Depression Anxiety and Stress Scale; DASS-21) and post-procedure pain were also assessed. Main results and the role of chance: Flushing was not superior with a mean (SD) number of COCs of 2.4 (2.0) and 3.1 (2.3) in the Steiner-Tan Needle® and in the Gynectics® group, respectively (mean difference -0.7, 95% CI: 0.3 to -1.6; P = 0.27). Likewise no differences were observed in metaphase II  oocytes, two pronuclear oocytes, number of patients having an embryo transfer and DASS 21 scores. The procedure duration was significantly 2-fold increased. Limitations reasons for caution: Testing for differences in the number of patients achieving an embryo transfer or differences in pregnancy rate would require a much larger sample size. Wider implications of the findings: The use of follicular flushing is unlikely to benefit the prognosis of patients with poor ovarian response. Study funding/competing interest(s): The Steiner-Tan Needles® and the flushing system were provided for free by the manufacturer. K.v.H. has received personal fees from Finox and non-financial support from Merck-Serono; M.D. has received personal fees from Finox and non-financial support from Merck-Serono. A.S.-M. has received personal fees and non-financial support from M.D., Ferring, Merck-Serono, Finox, TEVA. G.G. has received personal fees and non-financial support from M.D., Ferring, Merck-Serono, Finox, TEVA, IBSA, Glycotope, as well as personal fees from VitroLife, NMC Healthcare LLC, ReprodWissen LLC and ZIVA LLC. Trial registration number: NCT 02365350 (clinicaltrials.gov). Trial registration date: Sixth of February 2015. Date of first patient's enrolment: Ninth of February 2015.

Self-operated endovaginal telemonitoring: a prospective, clinical validation study.

OBJECTIVE: To study the comparability of self-operated endovaginal telemonitoring (SOET) with conventional two-dimensional transvaginal sonography (2D-TVS) monitoring during assisted reproductive technology (ART) cycles. DESIGN: Single center, observational, single-blinded cohort study. SETTING: University-affiliated in vitro fertilization center. PATIENT(S): A total of 60 women undergoing ART cycles. INTERVENTION(S): Explanation, training, and use of SOET system, and measurements of follicular and endometrial diameter with SOET and 2D-TVS. MAIN OUTCOME MEASURE(S): Correlation of the total number of follicles >10 mm measured by SOET versus conventional 2D-TVS. RESULT(S): In 16 cases (26.7%) the images were judged unsuitable for analysis. In these excluded cases the body mass index (BMI) was statistically significantly higher (29.3 vs. 24.4 kg/m(2)). The total number of follicles >10 mm was highly similar comparing SOET with conventional 2D-TVS (r = 0.91). For the concordance of whether more than 19 follicles or more than 25 follicles >10 mm were present, we found agreement between the methods in 43 of 44 cases (κ = 0.88) and 43 of 44 cases (κ = 0.85), respectively. For concordance on predefined human chorionic gonadotropin administration criteria, agreement was found in 39 of 44 cases (κ = 0.734). CONCLUSION(S): The incidence of SOET videos not suitable for analysis seems to be associated with higher BMI. Otherwise, SOET showed good agreement with conventional 2D-TVS both for follicles and endometrium measurements. More importantly we also found good concordance regarding the cutoffs relevant for clinical decisions.

Plants Release Precursors of Histone Deacetylase Inhibitors to Suppress Growth of Competitors.

To secure their access to water, light, and nutrients, many plant species have developed allelopathic strategies to suppress competitors. To this end, they release into the rhizosphere phytotoxic substances that inhibit the germination and growth of neighbors. Despite the importance of allelopathy in shaping natural plant communities and for agricultural production, the underlying molecular mechanisms are largely unknown. Here, we report that allelochemicals derived from the common class of cyclic hydroxamic acid root exudates directly affect the chromatin-modifying machinery in Arabidopsis thaliana. These allelochemicals inhibit histone deacetylases both in vitro and in vivo and exert their activity through locus-specific alterations of histone acetylation and associated gene expression. Our multilevel analysis collectively shows how plant-plant interactions interfere with a fundamental cellular process, histone acetylation, by targeting an evolutionarily highly conserved class of enzymes.