Biomedical subjects

Juraj Koppel

Publications and source records attributed to Juraj Koppel.

5 recordsLinked to original sources

PubMed2005

Apoptotic processes during mammalian preimplantation development.

The paper provides a review of the current state of knowledge on apoptosis during normal preimplantation development based on the literature and on the authors' own findings. Information is focused on the occurrence and the characteristics of spontaneous apoptotic processes. Reports concerning the chronology and the incidence of programmed cell death in mouse, cow, pig and human embryos in early preimplantation stages up to the blastocyst stage are summarized. In addition, specific attributes of the apoptotic process in mammalian preimplantation development are provided, including the description of both morphological and biochemical features of cell death.

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PubMed2005

Expression of beta adrenergic receptors in mouse oocytes and preimplantation embryos.

Accumulating evidence indicates the role of endogenous catecholamines in mammalian embryogenesis. We searched public databases containing nucleotide sequences derived from mouse preimplantation cDNA libraries and found a partial sequence homology between a cDNA clone from mouse blastocysts and the mouse beta 2-adrenergic receptor sequence. No significant sequence homology was found for other mouse adrenergic and dopamine receptors. Using RT-PCR, we showed that beta 2-adrenoceptor is transcribed not only at blastocyst stage but also at earlier stages of preimplantation development as well as in oocytes. Moreover, we demonstrated that transcripts encoding both isoforms of the beta 3-adrenoceptor (beta 3a- and beta 3b-) are expressed in mouse oocytes and preimplantation embryos as well. We did not detect the beta 1-adrenoceptor transcript either in oocytes or in preimplantation embryos. Using an antibody against the mouse beta 2-adrenergic receptor, we showed that the receptor protein is expressed in oocytes and preimplantation embryos; in blastocysts, the immufluorescence labeling was stronger in the inner cell mass than in throphectodermal cells. The cell number of the in vitro cultured mouse preimplantation embryos exposed to isoproterenol (a potent beta adrenoceptor agonist) was lower than in control embryos, suggesting that activation of beta adrenergic receptors by appropriate agonist concentration can influence cell proliferation in mouse pre-implantation embryos. Thus, our results indicate that beta adrenergic receptors are expressed in mouse oocytes and preimplantation embryos and that ligands for the receptors can affect the mouse embryo even in the very early stages of development.

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PubMed2004

Inhibitory effect of IGF-I on induced apoptosis in mouse preimplantation embryos cultured in vitro.

Insulin-like growth factor I (IGF-I) has been shown to promote mammalian early embryo development. Increased cell division or decreased cell death have been proposed as two main possible mechanisms in its effect. Here we examine the nature of this promoting effect in a model situation. Camptothecin (0.01 microg/ml) and actimomycin D (0.005 microg/ml) were used to induce apoptosis. Four-cell mouse embryos were cultured in vitro to blastocyst stage in the temporary (15 h) presence or absence of apoptotic inductors and in the permanent presence or absence of IGF-I (100 ng/ml). Embryos were assessed by morphological triple staining (Hoechst 33342, propidium iodide, Calcein AM) and comet assay on Day 5, 120 h after administration of hCG. The number of nuclei, the blastocyst formation, the proportion of embryos containing fragmented DNA and the percentage of apoptotic and secondary necrotic nuclei were assessed. Both inductors of apoptosis significantly increased the percentage of apoptotic and secondary necrotic cells and reduced total cell counts (camptothecin, P>0.001; actinomycin D, P>0.001). When IGF-I was added to the culture medium in the presence of an apoptosis inductor, apoptosis incidence was significantly decreased (P<0.001). The addition of IGF-I into control samples also decreased the percentage of apoptotic and secondary necrotic cells. In contrast, IGF-I addition had no significant influence on embryo development (P>0.05). Our data suggest a primary role for IGF-I as an apoptotic survival factor in mouse preimplantation embryos in specific conditions.

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PubMed2004

Serotonin localization and its functional significance during mouse preimplantation embryo development.

Serotonin is a neurotransmitter functioning also as a hormone and growth factor. To further investigate the biological role of serotonin during embryo development, we analysed serotonin localization as well as the expression of specific serotonin 5-HT1D receptor mRNA in mouse oocytes and preimplantation embryos. The functional significance of serotonin during the preimplantation period was examined by studying the effects of serotonin on mouse embryo development. Embryo exposure to serotonin (1 microM) highly significantly reduced the mean cell number, whereas lower concentrations of serotonin (0.1 microM and 0.01 microM) had no significant effects on embryo cell numbers. In all serotonin-treated groups a significant increase in the number of embryos with apoptotic and secondary necrotic nuclei was observed. Expression of serotonin 5-HT1D receptor mRNA in mouse oocytes and preimplantation embryos was confirmed by in situ hybridization showing a clearly distinct punctate signal. Immunocytochemistry results revealed the localization of serotonin in oocytes and embryos to the blastocyst stage as diffuse punctate cytoplasmic labelling. It appears that endogenous and/or exogenous serotonin in preimplantation embryos could be involved in complex autocrine/paracrine regulations of embryo development and embryo-maternal interactions.

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PubMed2003

Induced cell death of preimplantation mouse embryos cultured in vitro evaluated by comet assay.

The occurrence of apoptosis in mouse preimplantation embryos was analyzed using DNA staining (Hoechst 33342, PI) for the visualization of nuclear changes and by the comet assay, a single-cell gel electrophoresis assay, modified for the analysis of blastocysts. Mouse preimplantation embryos isolated 56 h after superovulation were cultured in vitro for 64 h. Apoptosis was induced by treatment with camptothecin and actinomycin D during the first 15 h of culture. After culture in vitro, a number of embryos were stained and analyzed using morphological criteria. The remaining embryos were examined using the comet assay for the detection of DNA fragmentation. The proportion of damaged embryos in experimental groups, in comparison to controls, was dependent on the dose of apoptosis inductor. At high doses (camptothecin, microg/ml and actinomycin D, 0.05 microg/ml) over 90% (chi-square test, P<0.001) of embryos had apoptotic comets, at medium doses (camptothecin, 0.01 microg/ml and actinomycin D, 0.005 microg/ml) comets appeared only in 30-70% of embryos (camptothecin, P<0.01 and actinomycin D, P<0.001). At low doses (camptothecin, 0.001 microg/ml and actinomycin D, 0.0005 microg/ml) the increase in damaged embryos was not statistically significant. Hoechst/PI staining showed a higher percentage of damaged blastomeres at high doses. Morphological changes correlated with the outcome of the comet assay. Our results show that comet assay is an appropriate method for studying apoptosis in preimplantation embryos, and it appears to be more sensitive than the classically used morphological analyses.

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