Cel Improve the yield of in vitro methodology to produce cattle embryos.Oocyte competence and maturation (D)A defined maturation medium (M199) has been used to investigate the effect of various components: single factors such as gonadotropins (FSH, LH) and growth factors (insulin family, EGF) as well as complex mixtures such as serum and follicular fluids from follicles of different sizes and health status. A fully defined maturation medium has been finally proposed (M199 + 10ng/ml EGF) that provides matured oocytes with high developmental potential in a reproducible manner. Calf oocytes have been chosen as a negative model for the study of oocyte developmental competence.Early embryo developmentOur study led to a better understanding of the ways through which oviduct might act in vivo and in vitro. Activin, EGF and TGFalpha were identified and localized in bovine oviduct mucosa. Their relevance to embryonic development was suggested by the presence of EGF/TGFalpha receptors on the bovine embryo from the zygote to the blastocyst (C). Two glycoproteins considered to have a positive role on embryo development (EGP, estrus associated glycoprotein and TIMP, tissue inhibitor of metalloproteinase) were also investigated but do not seem to play an important role on in vitro bovine embryo development (A).Serum-free conditioned media provided us with an interesting tool for the study of the effect of oviduct and BRL cell produced factors on embryo development.These media were biologically active: they supported development of the zygote to the blastocyst (15 to 30% blastocysts) stage (A,B). These blastocysts produced calves after transfer (31% born calves) (A, B, D) and could be frozen (20% born calves) (A).Our study clearly shows that somatic cells play an important active role in sustaining in vitro bovine embryo development both by detoxifying the basal medium and by secreting embryotrophic factors (A,D). These favourable effects do not depend on the stage of the oestrus cycle and do not seem to be oviduct specific (A, B, C, D, E).These embryotrophic factors were shown to increase blastocyst developmental rate when added to a defined medium (A). They were partially purified and some of their properties were characterised (A, B, E).The kinetics of development of the embryos in BOECM was related to their further development (A) and active transcription of the embryonic genome started at the 4 cell stage (A).The bovine homologue of the oct 4 transcription factor gene was identified, cloned, completely sequenced and mapped on chromosome 23 (C). However further analysis indicated that, in bovine, this gene is expressed ubiquitously contrary to the mouse where its expression is restricted to totipotent and multipotent cells.Finally, it was demonstrated that embryos generated in vitro are also able to produce biologically active trophoblastic vesicles without the requirement of transient exposure to a uterine environment (B, E).MAJOR SCIENTIFIC BREAKTHROUGHS:Identification of some factors (EGF) involved in oocyte maturation leading to the formulation of a chemically defined medium; detection of the expression of activin, TGFa and EGF in the oviduct; isolation and sequence of a clone of oct 4 gene; use of time-lapse cinematographic technique as a predictive tool of the embryonic developmental potential; marked improvement of embryo development in a defined medium by the addition of high molecular weight factors (>10 KDa) produced by oviduct cells; in vitro-derived trophoblastic tissue were shown to secrete large amounts of interferon tau and represents an important model for studies on peri-implantation physiology. Dziedzina nauki natural sciencesbiological sciencesdevelopmental biologymedical and health sciencesbasic medicinephysiologyagricultural sciencesanimal and dairy sciencedomestic animalsanimal husbandrynatural sciencesbiological sciencesgeneticschromosomesmedical and health sciencesclinical medicineembryology Program(-y) FP3-BIOTECH 1 - Specific research and technological development programme (EEC) in the field of biotechnology, 1990-1994 Temat(-y) 2.1 - Cellular regeneration, reproduction and development of living organisms Zaproszenie do składania wniosków Data not available System finansowania CSC - Cost-sharing contracts Koordynator UNIVERSITE CATHOLIQUE DE LOUVAIN Wkład UE Brak danych Adres Place De L'Universite 1 LOUVAIN-LA-NEUVE Belgia Zobacz na mapie Linki Strona internetowa Opens in new window Koszt całkowity Brak danych Uczestnicy (3) Sortuj alfabetycznie Sortuj według wkładu UE Rozwiń wszystko Zwiń wszystko Institut National de la Recherche Agronomique (INRA) Francja Wkład UE Brak danych Adres Domaine de l'Orfrasière Monnaie 37380 Nouzilly Zobacz na mapie Koszt całkowity Brak danych LUDWIG-MAXIMILIANS UNIVERSITY OF MUNICH Niemcy Wkład UE Brak danych Adres Geschwister-Scholl-Platz 1 80539 MUENCHEN Zobacz na mapie Koszt całkowity Brak danych THE UNIVERSITY OF MILANO Włochy Wkład UE Brak danych Adres Via Festa del Perdono 7 20122 MILANO Zobacz na mapie Koszt całkowity Brak danych
