A.P.A. Van Montfoort

Factors affecting the gene expression of in vitro cultured human preimplantation embryos

STUDY QUESTION What is the relative effect of common environmental and biological factors on transcriptome changes during human preimplantation development? SUMMARY ANSWER Developmental stage and maternal age had a larger effect on the global gene expression profile of human preimplantation embryos than the culture medium or oxygen concentration used in in vitro culture. WHAT IS KNOWN ALREADY Studies on mouse and bovine embryos have shown that different conditions in the in vitro culture of embryos can lead to changes in transcriptome profiles. For humans, an effect of developmental stage on the transcriptome profile of embryos has been demonstrated, but studies on the effect of maternal age or culture conditions are lacking. STUDY DESIGN, SIZE, DURATION Donated, good quality, day 4 cryopreserved human preimplantation embryos (N = 89) were randomized to be cultured in one of two culture media (G5 medium or HTF medium) and one of two oxygen concentrations (5% or 20%), with stratification for maternal age. Next to these variables, developmental stage after culture was taken into account in the analysis. PARTICIPANTS/MATERIALS, SETTING, METHODS Embryos that developed to morula or blastocyst stage during these 2 days whose amplified mRNA passed our quality control criteria for microarray hybridization were individually examined for genome-wide gene expression (N = 37). MAIN RESULTS AND THE ROLE OF CHANCE Based on the number of differentially expressed genes (DEGs), developmental stage (3519 DEGs) and maternal age (1258 DEGs) had a larger effect on the global gene expression profile of human preimplantation embryos than either tested culture medium (596 DEGs) or oxygen concentration (492 DEGs) used during in vitro culture. Interactions between the factors were found, indicating that culture conditions might have a different effect depending on the developmental stage or the maternal age of the embryos. Affected pathways included metabolism, cell cycle processes and oxidative phosphorylation. LIMITATIONS, REASONS FOR CAUTION Culture of embryos for only 2 days might have limited the effect on global gene expression by the investigated culture conditions. Earlier stages of development (Day 0 until Day 4) were not analyzed and these embryos might respond differently to the experimental conditions. The freezing and thawing procedures might have had an effect on gene expression. RT-PCR validation was not performed due to scarcity of the material. WIDER IMPLICATIONS OF THE FINDINGS Our results show that when studying gene expression in single human preimplantation embryos under various experimental conditions, one should take into account the confounding effect of biological variables, such as developmental stage and maternal age. This makes these experiments different from gene expression experiments where these variables can be tightly controlled, for example when using cell lines. STUDY FUNDING/COMPETING INTERESTS This study received no external funding and there were no competing interests.

Recent developments in genetics and medically assisted reproduction: from research to clinical applications

Two leading European professional societies, the European Society of Human Genetics and the European Society for Human Reproduction and Embryology, have worked together since 2004 to evaluate the impact of fast research advances at the interface of assisted reproduction and genetics, including their application into clinical practice. In September 2016, the expert panel met for the third time. The topics discussed highlighted important issues covering the impacts of expanded carrier screening, direct-to-consumer genetic testing, voiding of the presumed anonymity of gamete donors by advanced genetic testing, advances in the research of genetic causes underlying male and female infertility, utilisation of massively parallel sequencing in preimplantation genetic testing and non-invasive prenatal screening, mitochondrial replacement in human oocytes, and additionally, issues related to cross-generational epigenetic inheritance following IVF and germline genome editing. The resulting paper represents a consensus of both professional societies involved.

EP02.03: IVF culture media in relation to placental vascularisation: Electronic Poster Abstracts

Objectives: Optimum uterine blood flow is of pivotal importance needed for adequate implantation of the embryo and further development of the fetus. We hypothesise distribution of blood supply to the pregnant uterus is not uniform and dependant on placental location. Methods: A total of 62 singleton pregnancies reporting at the Prenatal Diagnostic Centre of the MUMC+ between September 2012-February 2018 and having their placenta located on either the right or left side of the uterus, were evaluated retrospectively. Experienced sonographers determined the location of the placenta in relation to the midline. Left and right uterine artery (UtA) pulsatility index (PI) were measured using a 4-8 mHz abdominal transducer. Percentiles of the PI corrected for gestational age (GA) were calculated offline. A minimum of two measurements per patient was performed during all pregnancy trimesters. Statistical analysis was performed using Student T-test. Results: In total, 184 uterine artery measurements were recorded between 10-38 weeks of gestation. The placenta was located right-sided in 77 cases (41.6%) and left-sided in 107 cases (58.4%). Mean left UtA-PI with a left-sided placenta was 1.1±0.6, compared to 1.4±0.7 with a right-sided placenta (p=0.002). Mean right UtA-PI with a left-sided placenta was 1.3±0.6, compared to 1.1±0.5 with a right-sided placenta (p=0.006). Furthermore, if we used the UtA-PI percentiles to correct for GA, differences remained statistically significant. Mean left UtA-PI showed in both leftand right-sided placenta a decrease of 0.04/week, compared to a mean right UtA-PI decrease of 0.03/week in both leftand right-sided placenta. No significant differences were found in birthweight or gestational age at delivery between rightor left-sided placentas. Conclusions: These findings indicate a raised resistance in contralateral uterine arteries when compared with their ipsilateral counterparts in case of placenta laterality. Further research is needed to answer the principal question if this finding is already a preconceptional occurrence leading to placental laterality.

OP21.02: Placental structure and vascularisation in IVF/ICSI- pregnancies compared to naturally conceived pregnancies: Short oral presentation abstracts

Methods: Interrectus distance (IRD) was measured at four locations (at the umbilical level, 3 cm above the umbilicus, 6 cm above the umbilicus, and 3 cm below the umbilicus) in 100 healthy nulliparous females and 99 postpartum females, while they remained relaxed and performed a head lift to activate the rectus abdominis muscles. Normal IRD values of the two groups were generated and subsequently compared. 99 postpartum females were subject to ultrasonographic examination of pelvic floor functions, thereby generating the hiatal area of the levator ani muscle and scores of pelvic floor dysfunction. The correlation of these values with diastasis recti was then examined. Results: We established ultrasonographic diagnostic criteria for diastasis recti as follows: IRD > 2 mm at 3 cm below the umbilicus, > 19 mm at the umbilicus, > 13 mm at 3 cm above the umbilicus, and > 7 mm at 6 cm above the umbilicus. The results revealed that IRD was positively correlated with body mass index (BMI) in the control group (r = 0.286) and the subject’s age in the patient group (r = 0.23). The incidence of pelvic floor dysfunction was approximately 82.8% among the patients with diastasis recti. Our data indicated that as the abdominal-pelvic pressure increases, IRD and pelvic floor dysfunction also increases. However, once the pressure achieves a certain threshold, pelvic organ prolapse relieves abdominal pressure, reducing the IRD. Conclusions: The IRD can be considered normal up to a width of 2 mm at 3 cm below the umbilicus, up to 19 mm at the umbilicus, up to 13 mm at 3 cm above the umbilicus, and up to 7 mm at 6 cm above the umbilicus in nulliparous women. The nonlinear correlation between diastasis recti and pelvic floor dysfunction may represent a dynamic balance of abdominal-pelvic biomechanics.