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Endogenně produkovaný sulfan v reprodukčním traktu samice ve vztahu k fertilizaci
BRICHCÍN, Jiří
The discovery of the mechanisms responsible for the proper process of fertilization and early embryonic development helps in the development of assisted human reproduction and technologies in livestock breeding. One of the regulatory mechanisms involved in reproduction is the gasotransmitter hydrogen sulfide, whose cellular signaling in germ cells is not fully described yet. According to the hypothesis, hydrogen sulfide is released in the female reproductive tract, where it physiologically affects sperm and is necessary for fertilization and the development of pronuclei. The aim was to verify the expression of hydrogen sulfide-releasing enzymes and the existence of a time-space gradient of hydrogen sulfide within the estrous cycle of the mouse (time axis) and to compare the expression profile of the fallopian tube ampulla and uterotubal junction (space axis). The experiments were performed on a laboratory mouse model (Mus musculus). The expression of hydrogen sulfide-releasing enzymes was studied by western blot. Hydrogen sulfide production was determined by colorimetry. The role of hydrogen sulfide in the fertilization process was studied by in vitro fertilization with hydrogen sulfide donor-treated sperm and by immunocytochemical staining of the zygotes. The expression of the known hydrogen sulfide-releasing enzymes, i.e. cystathionine--lyase (CTH), cystathionine-ß-synthase (CBS), and 3-mercaptopyruvate sulfurtransferase (3-MPST), was detected within the ovary and fallopian tube. Any enzymes did not show statistically significant differences in estrus and diestrus expression. Moreover, analogous results were found in the spatial axis; both the fallopian tube and the uterotubal junction expressed CTH, CBS, and 3-MPST enzymes, but no statistical differences were noted. Also, this applies to the production of hydrogen sulfide by these tissues. The effect of hydrogen sulfide on the fertilization process was analyzed according to lamin B1, a paternal pronucleus development marker. The performed analysis did not confirm the influence of the hydrogen sulfide donor on the development of the zygote pronuclei. The work showed that hydrogen sulfide is enzymatically released by the tissues of the female reproductive tract, regardless of the phase of the estrous cycle or the location in the fallopian tube. There is assumed a physiological influence of maternal-born hydrogen sulfide to sperm, which is capable of fertilization and leads to pronuclei development.
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Gasotransmise v epigenetických regulacích gametogeneze a embryogeneze
BRICHCÍN, Jiří
As the problems with the reproduction of livestock and humans starting to increase, the need for knowledge of mechanisms involved in regulating the correct process of gametogenesis and embryogenesis also rises. For the experimental part of this work, two components from two different, mostly separately explored fields, i.e. gasotransmitters and epigenetic mechanisms, which are necessary for the correct process of gametes production and early embryonic evolution were chosen. Hydrogen sulfide was chosen from the series of gasotransmitters, and histone deacetylase Sirtuin 1 (SIRT1) was chosen as its possible substrate. Confirmation of the presence of these components was carried out on oocytes and embryos of laboratory mice (Mus musculus).
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