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J Tesarík

Publications and source records attributed to J Tesarík.

At least 37 records · Page 2Linked to original sources
PubMed1988

Subcellular immunochemical localization of acrosin in human spermatozoa during the acrosome reaction and zona pellucida penetration.

The changes in acrosin immunoreactivity in human spermatozoa undergoing spontaneous or chemically induced acrosome reactions were studied by electron microscopic immunocytochemistry with an acrosin-specific monoclonal antibody. Migration of limited amounts of acrosin to the sperm surface was the earliest event characterizing the beginning of the acrosome reaction. The acrosome of such spermatozoa remained morphologically intact, swelled, or showed intraacrosomal vesiculation without any disruption of the plasma and acrosomal membrane integrity. Massive release of acrosin coincided with the fusion of the plasma and outer acrosomal membranes. However, even fully acrosome-reacted spermatozoa always retained some acrosin on the exposed inner acrosomal membrane and in the equatorial segment of the acrosome. This residual acrosin was also detected on spermatozoa within the zona pellucida of human oocytes inseminated in vitro, while the previously released bulk of acrosin remained attached to the surface of the zona pellucida at the site of sperm entry. These findings are compatible with multiple functions of acrosin in human sperm-egg interaction, including sperm-zona pellucida binding, dispersal of acrosomal contents, and facilitation of zona pellucida penetration.

Acrosin↗
PubMed1988

The role of cumulus cell-secreted proteins in the development of human sperm fertilizing ability: implication in IVF.

Cumulus cells surrounding pre-ovulatory human oocytes were found to secrete a variety of proteins which became firmly associated with the cumulus intercellular material. Antibodies raised against human cumuli oophori completely blocked fertilization in vitro by impairing the sperm-zona pellucida interaction. A group of glycoproteins of high mol. wt were identified as the main cumulus cell secretory products. These proteins showed a marked affinity for human spermatozoa and were potent stimulators of the conversion of human and boar proacrosin into acrosin and of human sperm acrosome reaction. Another fraction of proteins of human cumulus intercellular matrix with an apparent mol. wt of approximately 25,000 daltons was also found to stimulate significantly the acrosome reaction of human spermatozoa, although this fraction had no proacrosin-converting activity. These results indicate that proteins secreted by pre-ovulatory human cumulus cells have an indispensable role in the development of human sperm fertilizing ability. This effect seems to be realized by a concerted action of different types of cumulus-derived proteins just prior to and during the sperm-zona pellucida interaction. Disorders of cumulus cell secretory activity may account for some cases of idiopathic infertility and repeated IVF failures.

Antibodies↗
PubMed1988

Zona pellucida resistance to sperm penetration before the completion of human oocyte maturation.

Human oocytes exposed to capacitated spermatozoa in vitro when at metaphase of the 1st meiotic division (metaphase I) were not penetrated, even though some subsequently progressed to metaphase of the 2nd meiotic division (metaphase II). When the non-penetrated oocytes that had reached metaphase II during the incubation with spermatozoa were freed from the zona pellucida and reinseminated, two or more pronuclei developed in most of them. By contrast, no penetration was observed when the oocytes were reinseminated in the zona-intact state. When compared with metaphase II oocytes, metaphase I oocytes had a similar zona-binding capacity for spermatozoa, but fewer spermatozoa were found within the zona. These results indicate that the zona pellucida of human oocytes undergoes important maturational changes during the transition from metaphase I to metaphase II. Ultrastructural and previous histochemical findings suggest that these changes involve secretions from both the oocyte and cumulus cells and that the increased zona resistance at metaphase I may be due to relative insufficiency of cumulus cell-secreted 'softening' factors. If the integrity of the cumulus oophorus is disturbed at this stage, this condition becomes irreversible.

Female↗
PubMed1988

Activation of proacrosin by a locally produced component of human follicular fluid.

Components of human follicular fluid were separated on Sepharose 6B columns and the effects of different fractions on the conversion of pig proacrosin to acrosin were examined. A high-molecular-weight fraction (Mr greater than 3,000,000) of follicular fluid was a potent stimulator of this reaction. The proacrosin converting activity was absent in the corresponding fraction of blood serum. The acceleration of proacrosin activation was dependent on the concentration of material with proacrosin converting activity. The results indicate that human follicular fluid contains a high-molecular-weight component of local origin which is capable of accelerating proacrosin in a dose-dependent manner.

Acrosin↗
PubMed1987

Ultrastructural and autoradiographic observations on multinucleated blastomeres of human cleaving embryos obtained by in-vitro fertilization.

Human embryos from the 2-cell to the morula stage developing in vitro after monospermic fertilization were incubated with [3H]thymidine or [3H]uridine and those possessing multinucleated blastomeres were examined by conventional transmission electron microscopy and by light- and electron-microscope autoradiography. Labelled DNA was present in all nuclei showing normal ultrastructural appearance, in pseudonuclei lacking nucleoli and their precursors and often demonstrating an unusual chromatin organization, free in the cytoplasm in structures resembling aggregates of dense chromatin and in small cytoplasmic vesicles in the close vicinity of these aggregates. The labelling with [3H]thymidine was not detected in about 50% of the cytoplasmic chromatin aggregates, suggesting that this extranuclear DNA was no longer replicated. Signs of extrusion of pseudonuclei to the extracellular space were occasionally observed. RNA synthesis could not be detected on free cytoplasmic chromatin, while [3H]uridine was incorporated into all nuclei and most pseudonuclei of multinucleated blastomeres of embryos at the 8-cell and morula stages. However, the major outburst of transcriptional activity and the ultrastructural cytoplasmic changes typical of the assumption of embryonic genome expression, occurring normally at the 8-cell stage, were observed only exceptionally.

Autoradiography↗
PubMed1987

High-resolution autoradiographic localization of DNA-containing sites and RNA synthesis in developing nucleoli of human preimplantation embryos: a new concept of embryonic nucleologenesis.

Human embryos from the 2-cell to the morula stage, obtained by in vitro fertilization, were incubated with [3H]thymidine or [3H]uridine so as to achieve labelling of all replicating nuclear DNA and the newly synthesized RNA, respectively. The label was localized in different structural components of developing nucleoli using electron microscopic autoradiography. Careful study of the relationship between the structural pattern and nucleic acid distribution made it possible to define four stages of embryonic nucleologenesis. Homogeneous nuclear precursors (i) consist of nucleolar matrix elements appearing as filaments of 3 nm thickness, (ii) do not contain recently replicated DNA and (iii) lack RNA synthetic activity. Penetration of DNA into these bodies is a key event leading to their transformation into heterogeneous nucleolar precursors. In addition to the 3 nm matrix filaments, two types of 5 nm fibrillar components can be recognized in them. The denser type contains DNA and is the site of nucleolar RNA synthesis, while the more loosely arranged 5 nm fibrils are not labelled with [3H]thymidine and apparently represent the newly produced pre-rRNA detached from the transcribing rDNA filament. Compact fibrillogranular nucleoli are characterized by the first appearance of the granular component and reduction of the nontranscribing part of the fibrillar component, both indicating the activation of the machinery for rRNA processing. Finally, the granular component is most evident in reticulated nucleoli, occupying mostly the inner parts of their nucleolonema, while the transcription sites tend to be located at the nucleolar periphery. Our findings advocate a unique concept of embryonic nucleologenesis, different from any other nucleolar event during the cell cycle of differentiated cells. This developmental pattern is characterized by a gradual activation of rRNA synthesis and processing, mediated by progressive association of rDNA and, later on, the newly formed pre-rRNA with pre-existing nucleolar matrix elements that are originally topically separated from nucleolar organizer regions. This model may have a general validity in early animal embryos despite some interspecies variability in the timing of individual steps and resulting structural peculiarities.

Autoradiography↗
PubMed1986

Selective association of some hamster-egg-synthesized proteins with decondensing human sperm chromatin.

Zona-free hamster eggs were fertilized in vitro with human spermatozoa in a culture medium enriched with either 3H-arginine or 3H-tryptophan. Autoradiography was used to investigate decondensing sperm heads and all pronuclei for the presence of newly synthesized, 3H-labelled proteins. In the case of 3H-arginine-labelled proteins, an intense accumulation of radioactivity was detected in all autoradiograms of chromatin structures. On the other hand, no comparable accumulation was seen for 3H-tryptophan-labelled proteins up to the progressed-pronucleus stage. It is concluded that, as a part of changes of the nucleoproteins in decondensing sperm chromatin, there is an accumulation in the male (as well as in the female) pronucleus of basic nuclear proteins synthesized by the egg during fertilization. Since non-histone, 3H-tryptophan-labelled proteins were not incorporated in the same way, these 3H-arginine-labelled proteins accumulating in pronuclear chromatin during the earliest phase of pronucleus formation are probably histones.

Animals↗
PubMed1986

Late preovulatory synthesis of proteoglycans by the human oocyte and cumulus cells and their secretion into the oocyte-cumulus-complex extracellular matrices.

Light- and electron-microscope autoradiography using 3H-glucosamine and 3H-fucose as precursors was employed to investigate proteoglycan synthesis and secretion by late preovulatory human oocytes and cumulus cells. Both the oocyte and cumulus cells were found to be important cellular sources supplying proteoglycans to the oocyte-cumulus-complex extracellular matrices, i.e., the zona pellucida and the cumulus intercellular matrix. Both the oocyte and cumulus cells were shown to secrete labelled proteoglycans into the zona pellucida. Labelled proteoglycans were also detected in the cumulus intercellular matrix. Chase experiments revealed the labelled molecules to be relatively closely associated with both the zona pellucida and the cumulus intercellular matrix. Staining with chromic acid and phosphotungstic acid showed proteoglycan material to penetrate from the cumulus intercellular matrix into pores of the zona pellucida. This material is thought to be a structural equivalent of the newly synthesized proteoglycans secreted by cumulus cells and migrating into the zona pellucida (as detected by autoradiography). It is concluded that newly synthesized proteoglycans secreted by the oocyte and cumulus cells in the late preovulatory period are a component of the microenvironment in which fertilization takes place.

Autoradiography↗
PubMed1986

Nucleologenesis in the human embryo developing in vitro: ultrastructural and autoradiographic analysis.

Nucleolar development during cleavage of human embryos in vitro was analyzed using combined autoradiographic and ultrastructural approaches. Human 2- to 4-cell embryos lack functionally active nucleoli. They possess nucleolus-like bodies formed by a homogeneous mass composed of densely packed fine fibrils. The nucleolus-like bodies contain no recently replicated embryonic DNA and do not show any detectable RNA synthesis. Their transformation into nucleoli is a relatively rapid process characterized by progressive infiltration of these bodies by adjacent chromatin, followed by the beginning of RNA synthesis and processing reflected by the appearance of the first nucleolar granules. These changes take place in 6- to 8-cell embryos, probably starting after the third cleavage division. Finally, most frequently in 10- to 12-cell embryos, typical nucleolar structure is established as a result of intranucleolar differentiation giving rise to distinct fibrillar and granular components as well as to nucleolar interstices. It is suggested that this pattern of nucleologenesis, different from that pertinent to mouse embryos (M. Geuskens and H. Alexandre (1984). Cell Differ. 14, 125-134), might be related to the relatively late activation of embryonic rRNA genes transcription and to the expressed rapidity of this process.

Blastomeres↗
PubMed1986

Activation of nucleolar and extranucleolar RNA synthesis and changes in the ribosomal content of human embryos developing in vitro.

RNA synthetic activity of human 2-16-cell embryos developing in vitro was studied by [3H]uridine light-microscope autoradiography. Parallelly cut thin sections were examined in the electron microscope. The first extranucleolar RNA synthesis was detected in 4-cell embryos, but nucleoli were never labelled until the 3rd cleavage (6-8-cell embryos). In 6-cell embryos the nucleolar labelling was mostly confined to a narrow peripheral zone. In later cleavage stages most of the blastomeres showed intensive labelling of nucleoli and extranucleolar chromatin. However, rather low levels of extranucleolar RNA synthesis and the absence of nucleolar activity were often seen even in blastomeres of fully compacted morulae. The activation of nucleolar RNA synthesis entailed a noticeable increase in the number of ribosomes (estimated by electron microscope morphometry) that followed a marked drop during the period between the 2-cell and 8-cell stages. The results indicate that the concentration of ribosomes in the preovulatory oocyte is a major factor of its developmental potential.

Autoradiography↗