Incubator type affects human blastocyst formation and embryo metabolism: a randomized controlled trial.

STUDY QUESTION: Does the type of incubator used to culture human preimplantation embryos affect development to the blastocyst stage and alter amino acid utilization of embryos in assisted reproduction? SUMMARY ANSWER: Culturing embryos in a time lapse system (TLS) was associated with a higher Day 5 blastocyst formation rate and altered amino acid utilization when measured from Day 3 to Day 5 compared to the standard benchtop incubator. WHAT IS KNOWN ALREADY: Culture environment is known to be important for the developing preimplantation embryo. TLSs provide a stable milieu allowing embryos to be monitored in situ, whereas embryos cultured in standard benchtop incubators experience environmental fluctuations when removed for morphological assessment. STUDY DESIGN, SIZE, DURATION: A prospective clinical trial randomizing 585 sibling embryos to either the TLS (289 embryos) or the standard benchtop incubator (296 embryos) over a 23-month period in a UK University Hospital Fertility Clinic. PARTICIPANTS/MATERIALS, SETTING, METHODS: Participants were aged 42 years or under, had an antral follicle count of ≥12 and ≥6 2 pronucleate zygotes. Zygotes were cultured individually in 25 µl of medium. Randomized embryos were graded and selected for transfer or cryopreservation on Day 5. For those embryos produced by women who underwent stimulation with recombinant FSH injections and were triggered with hCG, spent medium was collected on Day 5 for amino acid analysis by high pressure liquid chromatography. Clinical pregnancy was defined as the presence of a foetal heart beat on ultrasound scan at 7 weeks. MAIN RESULTS AND THE ROLE OF CHANCE: Overall, blastocyst formation rate on Day 5 was significantly higher in embryos cultured in the TLS (55%) compared to the standard incubator (45%; P = 0.013). Similarly, there was an increase in the number of blastocysts suitable for cryopreservation in the TLS (31%) compared to the standard incubator (23%; P = 0.032). There was a significant difference in the utilization of 12 amino acids by blastocysts cultured from Day 3 to Day 5 in the TLS compared to the standard incubator. Embryos cultured in the TLS displayed an increased total amino acid utilization (P < 0.001) and reduced amino acid production (P < 0.001) compared to those in the standard incubator. Irrespective of incubator used, embryos fertilized by ICSI depleted significantly more amino acids from the medium compared to those fertilized by conventional IVF. There was no difference in the mean score of blastocysts transferred, or the clinical pregnancy rate after transfer of embryos from either of the incubators. LIMITATIONS, REASONS FOR CAUTION: The study was not powered to discern significant effects on clinical outcomes. WIDER IMPLICATIONS OF THE FINDINGS: The metabolism and development of preimplantation embryos is impacted by the type of incubator used for culture. Further research is required to investigate the long-term implications of these findings. STUDY FUNDING/COMPETING INTEREST(S): NIHR Southampton Biomedical Research Centre Commercial and Enterprise Incubator Fund funded this study. The TLS was provided on loan for the study by Vitrolife. The authors declare no conflict of interests. TRIAL REGISTRATION NUMBER: ISRCTN73037149. TRIAL REGISTRATION DATE: 12 January 2012. DATE OF FIRST PATIENT'S ENROLMENT: 21 January 2012.

Stem cell marker TRA-1-60 is expressed in foetal and adult kidney and upregulated in tubulo-interstitial disease.

The kidney has an intrinsic ability to repair itself when injured. Epithelial cells of distal tubules may participate in regeneration. Stem cell marker, TRA-1-60 is linked to pluripotency in human embryonic stem cells and is lost upon differentiation. TRA-1-60 expression was mapped and quantified in serial sections of human foetal, adult and diseased kidneys. In 8- to 10-week human foetal kidney, the epitope was abundantly expressed on ureteric bud and structures derived therefrom including collecting duct epithelium. In adult kidney inner medulla/papilla, comparisons with reactivity to epithelial membrane antigen, aquaporin-2 and Tamm-Horsfall protein, confirmed extensive expression of TRA-1-60 in cells lining collecting ducts and thin limb of the loop of Henle, which may be significant since the papillae were proposed to harbour slow cycling cells involved in kidney homeostasis and repair. In the outer medulla and cortex there was rare, sporadic expression in tubular cells of the collecting ducts and nephron, with positive cells confined to the thin limb and thick ascending limb and distal convoluted tubules. Remarkably, in cortex displaying tubulo-interstitial injury, there was a dramatic increase in number of TRA-1-60 expressing individual cells and in small groups of cells in distal tubules. Dual staining showed that TRA-1-60 positive cells co-expressed Pax-2 and Ki-67, markers of tubular regeneration. Given the localization in foetal kidney and the distribution patterns in adults, it is tempting to speculate that TRA-1-60 may identify a population of cells contributing to repair of distal tubules in adult kidney.

Society for Reproductive Biology Founders' Lecture 2003. The making of an embryo: short-term goals and long-term implications.

During early development, the eutherian mammalian embryo forms a blastocyst comprising an outer trophectoderm epithelium and enclosed inner cell mass (ICM). The short-term goal of blastocyst morphogenesis, including epithelial differentiation and segregation of the ICM, is mainly regulated autonomously and comprises a combination of temporally controlled gene expression, cell polarisation, differentiative cell divisions and cell-cell interactions. This aspect of blastocyst biogenesis is reviewed, focusing, in particular, on the maturation and role of cell adhesion systems. Early embryos are also sensitive to their environment, which can affect their developmental potential in diverse ways and may lead to long-term consequences relating to fetal or postnatal growth and physiology. Some current concepts of embryo-environment interactions, which may impact on future health, are also reviewed.

The embryo and its future.

The preimplantation mammalian embryo from different species appears sensitive to the environment in which it develops, either in vitro or in vivo, for example, in response to culture conditions or maternal diet. This sensitivity may lead to long-term alterations in the characteristics of fetal and/or postnatal growth and phenotype, which have implications for clinical health and biotechnological applications. We review the breadth of environmental influences that may affect early embryos and their responses to such conditions along epigenetic, metabolic, cellular, and physiological directions. In addition, we evaluate how embryo environmental responses may influence developmental potential and phenotype during later gestation. We conclude that a complex of different mechanisms may operate to associate early embryo environment with future health.