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M H Sellens

Publications and source records attributed to M H Sellens.

14 recordsLinked to original sources
PubMed1994

Optimization of superovulation in the reproductively mature mouse.

Optimum gonadotropin doses and chronology were established for the induction of superovulation in sexually mature hybrid mice (BALB/cBy x C57BL/6By). A regime of 12 IU pregnant mares' serum gonadotropin (PMSG), followed 48 hr later by 20 IU human chorionic gonadotropin (hCG) administered 1 hr before the midpoint of the light cycle (1200), gave the maximum ovulatory response. There was no evidence that endogenous luteinizing hormone influenced the superovulation response to exogenous gonadotropins. Fewer than 50% of zygotes reached the blastocyst stage (90-93 hr post hCG), with the greatest rate of loss at the two- to four-cell stage. Litter size following superovulation was 19.6 +/- 0.9. There was no significant difference between the number of blastocysts observed and litter size. Similarly, counts of mature follicles in ovaries prior to hCG stimulation were not significantly greater than the number of secondary oocytes that subsequently ovulated. These data indicate that standard superovulation protocols may require fine-tuning to maximize productivity and confirm that embryo loss is greatest between the first cleavage division and blastocyst formation.

Analysis of Variance↗
PubMed1993

Preimplantation mouse embryos express Mhc class I genes before the first cleavage division.

We have used oligonucleotide primers complementary for polymorphic regions of the mouse H-2D gene in a highly sensitive polymerase chain reaction (PCR) assay to detect the transcription of maternal and paternal class I mRNAs in gametes and preimplantation embryos. Using congenic strains of mice differing only at the major histocompatibility loci, class I (H-2D) mRNA of both the maternal and paternal haplotypes was demonstrated in embryos from the one-cell zygote to the late blastocyst stage of development but could not be detected in vas deferens or in vitro capacitated sperm or in ovulated secondary oocytes. These data clearly show that both paternally and maternally inherited Major histocompatibility complex (Mhc) class I genes are transcribed from the earliest stages of embryonic development, and suggest that developmental regulation of expression of their protein products is principally at the post-transcriptional level.

Animals↗
PubMed1991

Free radical scavengers ameliorate the 2-cell block in mouse embryo culture.

Mouse zygotes from a strain combination which fails to undergo cleavage in vitro (the 2-cell block) were cultured in a variety of media, using either low (5%) or normal (20%) oxygen tension in the gas phase. Development beyond the 4-cell stage was only achieved in medium containing serum and with normal oxygen tension. When serum-free medium was supplemented with free radical scavengers, reduced glutathione (1 mM), but not histidine, catalase or superoxide dismutase, development to the morula or blastocyst stage was promoted in 50% of zygotes cultured with normal oxygen tension. These data suggest that the 2-cell block is a consequence, at least in part, of free radical damage incurred by embryos during collection and culture, and that medium supplementation with the radical scavenger, reduced glutathione, can improve embryo development in vitro.

Animals↗
PubMed1990

Lectin-induced abnormalities of mouse blastocyst hatching and outgrowth in vitro.

We have examined the role of cell surface glycoconjugates during mouse blastocyst maturation, hatching, attachment, and outgrowth by monitoring the influence of six lectins on blastocyst development in vitro. Two lectins, concanavalin A and wheat germ agglutinin were toxic to blastocysts at the concentrations used. Bandierea simplicifolia lectin 1 (BSL-1) induced abnormal growth, developmental arrest at the hatching stage, and some disruption of cell contacts. Culture with Lotus tetragonolobus lectin-1 (LTA-1) also disrupted cell contacts and caused developmental arrest. The remaining lectins, Dolichos biflorus agglutinin (DBA) and Ulex europaeus agglutinin (UEA), retarded blastocyst hatching and outgrowth but did not induce any major defects, although differentiation of the inner cell mass was limited by both. This study demonstrates that very low concentrations of lectins can disrupt blastocyst development, suggesting that exposed surface saccharide moieties may be involved in interactions between blastomeres and their environment.

Animals↗
PubMed1990

Mouse zygotes express endogenous lectins.

Cell interactions during mouse development have been shown to involve carbohydrate-containing macromolecules (glycoconjugates). We have therefore used a series of fluorescein-labelled synthetic glycoproteins to determine if mouse oocytes and zygotes also express sugar binding molecules (endogenous lectins) which might participate in such interactions. Unfertilized secondary oocytes did not express endogenous lectins at 4 degrees C but a low level of expression of fucose, mannose, and galactose-binding activity could be detected at 37 degrees C. In contrast, the zygote clearly expressed three classes of endogenous lectins, with preferential binding for i) fucose or mannose, ii) glucose or galactose, and iii) lactose. The expression of these lectins was much reduced at 4 degrees C and maximal binding at 37 degrees C was achieved only after 2 h incubation. We therefore conclude that a low level of endogenous lectin expression in the mouse oocyte is greatly enhanced after fertilisation and that, at both stages, expression, or the detection of expression, is markedly temperature dependent.

Animals↗
PubMed1987

Fucosylated glycoconjugates appear on mouse embryos during blastocyst formation.

Mouse embryos were analysed for expression of surface glycoconjugates using a panel of fluorescein-labelled lectins. Morulae and blastocysts but not early cleavage stages stained brightly with fucose binding lectins. By contrast, cleavage stages stained brightly with all the other lectins tested. These findings provide evidence for substantial reorganisation of the cell surface during blastocyst formation and are consistent with a role for fucosylated glycoconjugates in compaction.

Animals↗
PubMed1981

Protein and free amino acid content in preimplantation mouse embryos and in blastocysts under various culture conditions.

By using a rapid and sensitive method for determination of total protein and free amino acid contents we have shown that mouse preimplantation embryos developing in vivo and in vitro contain decreasing amounts of protein until the late blastocyst stage. Free amino acid contents in embryos developing in vivo drop following fertilization and then rise gradually until the late blastocyst stage. Embryos developing in vitro from the 2-cell stage have much lower levels of free amino acids than those in vivo. Embryos placed in culture at the blastocyst stage show a decrease in protein content within 24 h, but protein levels rise rapidly thereafter in medium containing serum and amino acids. In the same medium, total amino acid pools increase from the time the blastocysts are placed in culture. Blastocysts cultured in the absence of amino acids show a very slight net increase in protein content and free amino acid pools are eventually depleted. In the presence of amino acids but the absence of serum, there is only a modest increase in the protein content of cultured blastocysts, suggesting that, in addition to an exogenous supply of amino acids, other factors are required for optimal growth.

Amino Acids↗
PubMed1980

Effects of culture conditions on the developmental programme of mouse blastocysts.

We have investigated the developmental potential of mouse blastocysts cultured under a variety of conditions. A number of parameters were used as criteria for development and differentiation, namely hatching of blastocysts from the zona pellucida and their adhesion to the substratum, outgrowth and polyploidization of trophoblast cells, increase in cell number, protein content, beta-glucuronidase activity, appearance of lactate dehydrogenase A subunits, plasminogen activator production, and delta 5,3 beta hydroxysteroid dehydrogenase activity. Under optimal culture conditions, embryos grew relatively rapidly and expressed all the differentiative markers for which they were tested. Under less supportive conditions, the production of the markers was usually reduced quantitatively; the expression of some markers could also be considerably delayed or even totally prevented. In fact, embryos cultured in the least nutritive medium (one designed to support development only through pre-implantation stages) appeared to be in a state of metabolic quiescence closely resembling that of blastocysts in ovariectomy-induced delay. Overall, the results of our investigations lead us to propose that the expression of each of the aforementioned markers is probably under independent control and subject to intrinsic programming. Finally, the observation that some markers are produced by embryos in suboptimal media whereas others are not, suggests that the minimum metabolic level necessary for expression varies from one marker to another.

3-Hydroxysteroid Dehydrogenases↗
PubMed1978

Changes in the surface of the mouse blastocyst at implantation.

Implantation is a critical event, and perhaps the earliest one, in the maternal recognition of pregnancy. Information transfer from conceptus to mother might occur during, and subsequent to, implantation at the level of cell surface interaction. Therefore, attempts have been made both to identify the phases of implantation during which changes in the blastocyst surface occur and to characterized such changes. In vitro, blastocysts have been found to go through a series of discrete steps which are analogous to implantation in utero, and these steps can be retarded or prevented by the use of either suboptimal culture media or an inappropriate substratum. Morphological surface changes are not apparent when the blastocyst becomes adherent to the substratum; however, marked differences in blastocyst surface structure are revealed by scanning electron microscopy at the onset of trophoblast outgrowth. Studies at the molecular level implicate collagen as having a role in blastocyst adhesiveness, but other cell surface components are also likely to be involved.

Animals↗
PubMed1978

Major histocompatibility complex and non-major histocompatibility complex antigens on mouse ectoplacental cone and placental trophoblastic cells.

The expression of major histocompatibility complex and non-major histocompatibility complex antigens on mouse trophoblast cultured from two defined stages of development was investigated by the sensitive in vitro mixed haemadsorption assay. Outgrowths obtained 3 to 5 days after explanation of 7 1/2-day ectoplacental cones contained a mixed population of cells. Those with a giant cell morphology showed no haemadsorption with congenic H-2 antisera and were reactive with non H-2 antiserum only in the CBA strain. Other, smaller cells were reactive for both H-2 and non-H-2 in all strains examined except for C57BL, where the cells were nonreactive for H-2. Monolayer cultures of 13 to 14-day placental suspensions tested 24 hr after preparation were strongly reactive for both H-2 and non-H-2. The identity and alloantigenic status of the cells are discussed in relation to their function in maternal-foetal immunological interactions.

Animals↗
PubMed1976

Detection of alloantigens during preimplantation development and early trophoblast differentiation in the mouse by immunoperoxidase labeling.

An immunoperoxidase-labeling technique allowing visualization of antibody binding to the cell surface at the electron microscopical level has been employed an an analysis of H-2 and non-H-2 alloantigen expression on the early mouse embryo. The presence of non-H-2 antigenic determinants has been confirmed on eight-cell, morula, and blastocyst stages of development. Contrary to previous reports, however, low levels of H-2 antigen have also been detected on the blastocyst. This is the earliest stage at which H-2 has been shown to be expressed on the fertilized mouse egg and may reflect the greater resolution of the immunoperoxidase technique. Using two different models to study the critical peri-implantation stages, those of experimentally induced blastocyst activation and blastocyst outgrowth in vitro, it has been demonstrated that antigen loss occurs on the trophectoderm at the time of implantation, and that this is not necessarily dependent upon maternal influence. It is suggested that the loss may be an important factor in the prevention of maternal immune rejection during the establishment of the fetal allograft. The two major components of the early postimplantation conceptus display a striking differential in antigenic status. The embryonic sac shows a high degree of peroxidase labeling, while the ectoplacental cone trophoblast is unlabeled. These findings add support to the concept of antigenic neutrality of the early trophoblast and its role in the maintenance of a normal fetomaternal immunological equilibrium.

Animals↗