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J. Laurincik, D. Rath and H. Niemann

To elucidate the developmental differences occurring after in vitro fertilization (IVF) of pig oocytes matured either in vitro (n = 1934) or in vivo (n = 1128), the present experiment investigated the morphological changes from penetration to the two-cell stage. Oocytes were examined every 2–4 h from 2 to 32 h after in vitro insemination to study sperm penetration, male and female pronucleus formation, synkaryosis and first cleavage. The penetration rate was significantly higher (P < 0.05) for in vivo matured oocytes (69.8%) than for in vitro matured oocytes (35.0%). Penetration of spermatozoa into the ooplasm was first recorded 6 h (in vitro matured oocytes) and 4 h (in vivo matured oocytes) after addition of the spermatozoa to the oocytes. For both in vivo and in vitro matured oocytes, 2 h were required for sperm head decondensation. However, maximum sperm head decondensation occurred 2 h later in in vitro matured oocytes. Within 6 h, 41.7 ± 5.6% of the in vivo matured oocytes had completed second meiotic division, whereas only 20.8 ± 6.5% of the in vitro matured oocytes reached this developmental stage (P < 0.01). For in vitro matured oocytes, male pronucleus formation was retarded 2–4 h after onset of insemination and development of the female pronucleus was enhanced compared with in vivo matured oocytes. Synchronized opposing pronuclei were observed 14 h after insemination in in vitro matured oocytes and after 8 h in in vivo matured oocytes. Synkaryosis was first observed at 16 and 18 h in in vivo and in vitro matured oocytes, respectively. First cleavage was observed 32 h (in vitro matured oocytes) and 28 h (in vivo matured oocytes) after insemination. It is concluded that under our IVF conditions, oocytes matured in vitro display lower penetration and cleavage rates and asynchronous pronucleus development, as well as delayed cleavage, compared with oocytes matured in vivo.

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D Rath, S Barcikowski, S de Graaf, W Garrels, R Grossfeld, S Klein, W Knabe, C Knorr, W Kues, H Meyer, J Michl, G Moench-Tegeder, C Rehbock, U Taylor and S Washausen

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D Rath, S Barcikowski, S de Graaf, W Garrels, R Grossfeld, S Klein, W Knabe, C Knorr, W Kues, H Meyer, J Michl, G Moench-Tegeder, C Rehbock, U Taylor and S Washausen

Pre-selection of spermatozoa based on the relative DNA difference between X- and Y-chromosome bearing populations by flow cytometry is an established method that has been introduced into commercial cattle production. Although several important improvements have increased the sort efficiency, the fertilising ability of sexed spermatozoa based on offspring per insemination is still behind farmers' expectations. The main stress factors, especially on mitochondria, that reduce the lifespan of spermatozoa are described, and new technical as well as biological solutions to maintain the natural sperm integrity and to increase the sorting efficiency are discussed. Among these methods are the identification of Y-chromosome bearing spermatozoa by bi-functionalised gold nanoparticles and triplex hybridisation in vivo as well as new laser-controlled deflection system that replaces the deflection of spermatozoa in the electrostatic field. Additionally, as well as a new nonsurgical transfer system of spermatozoa into the oviduct of cows has been developed and allows a significant reduction of spermatozoa per transfer. Altogether, the improvements made in the recent years will allow a broader use of sex-sorted spermatozoa even in those species that require more cells than cows and sheep.

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C Passaro, D Tutt, S Bagés-Arnal, C Maicas, R Laguna-Barraza, A Gutierrez-Adán, J A Browne, D Rath, S K Behura, T E Spencer, T Fair and P Lonergan

The aims of this study were (i) to investigate changes in the global transcriptome of bovine endometrial explants induced by exposure to blastocysts, (ii) to investigate if male and female blastocysts elicit a differential response in the endometrial transcriptome in vitro and (iii) to determine whether bovine endometrium responds to the presence of murine embryos. In Experiment 1, endometrial explants from the same uterus were cultured for 6 h with or without 20 in vitro-produced bovine blastocysts. In Experiment 2, endometrial explants were cultured with male or female bovine blastocysts produced in vitro by IVF either using sex-sorted semen or conventional unsorted semen followed by embryo sexing based on a biopsy. In Experiment 3, endometrial explants were cultured alone or in the presence of bovine blastocysts (n = 25) or murine blastocysts (n = 25). Following culture, explants were snap frozen and stored at −80°C until RNA extraction, qPCR or RNA-Seq. Culture with bovine blastocysts increased endometrial expression of 40 transcripts, all of which were interferon-tau induced. Culture with male or female bovine blastocysts increased transcript abundance of five classic interferon-stimulated genes (MX1, MX2, ISG15, OASY1, RSAD2) in explants; however, there was no difference in abundance of transcripts previously reported to be related to embryonic sex (IFNAR1, IFNAR2, CTGF, ARTN, SLC2A1, SLC2A5). Exposure to murine blastocysts did not elicit any detectable change in transcript abundance. These findings, coupled with our previous data, indicate that very local, interferon-tau-induced changes in endometrial gene expression occur in response to blastocysts; whether such changes play any role in subsequent pregnancy recognition remains to be established.