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J Tesarik

Publications and source records attributed to J Tesarik.

At least 127 records · Page 7Linked to original sources
PubMed1992

Effect of pentoxifylline on sperm movement characteristics in normozoospermic and asthenozoospermic specimens.

Preliminary evidence has suggested that the phosphodiesterase inhibitor pentoxifylline augments the fertilizing potential of asthenozoospermic sperm samples, presumably by improving sperm movement. Here, we used computer-assisted sperm movement analysis to compare the effects of pentoxifylline in normozoospermic and asthenozoospermic specimens. The study focused on the following issues: the changes in individual movement characteristics in response to pentoxifylline, the rapidity of the response, the effect of sperm capacitation on the response, the persistence of the response after drug removal and the variability of responses among asthenozoospermic individuals. Data obtained show that (i) pentoxifylline increases the curvilinear velocity, path velocity, straight-line velocity, lateral head displacement, beat cross frequency and sperm hyperactivation in both normozoospermic and asthenozoospermic specimens, (ii) pentoxifylline does not modify the percentage of motile spermatozoa, (iii) the pentoxifylline effect reaches a maximum within 10 min of treatment in fresh semen as well as in capacitated sperm suspensions and persists for at least 2 h after drug removal and (iv) pentoxifylline improves the movement characteristics in most asthenozoospermic individuals. Results are discussed with regard to methods of therapeutic application of pentoxifylline as an enhancer of sperm movement in assisted reproductive technology.

Humans↗
PubMed1992

Distinction between true acrosome reaction and degenerative acrosome loss by a one-step staining method using Pisum sativum agglutinin.

When western blots of human sperm proteins solubilized by acid extraction (presumably mainly acrosomal proteins) or by sodium dodecyl sulfate (SDS) were probed with biotin-conjugated Pisum sativum agglutinin (PSA), distinct sets of proteins were labelled in both preparations. When smears of human spermatozoa were treated with methanol either for 30 s or for 15 min and then exposed to FITC-conjugated PSA, the resulting fluorescence pattern essentially depended on the time of methanol treatment. With the longer treatment, fewer spermatozoa showed selective acrosomal labelling and more were labelled uniformly throughout, without a clear predilection for a single sperm region. With the shorter time of methanol treatment, the poorly topographically differentiated, whole-cell labelling was typical of dead spermatozoa as confirmed by a close correlation between the percentages of spermatozoa showing this type of labelling and of those stained supravitally with Hoechst 33258. The preferential whole-cell labelling of dead spermatozoa with PSA is considered to be due to increased availability of the nonacrosomal set of PSA-reactive sites in dead spermatozoa after a short treatment with methanol, whereas this treatment is probably not sufficient to expose most of these sites when applied to living spermatozoa. The simplicity of the staining protocol makes this method feasible in routine work in a number of clinical and research applications.

Acrosome↗
PubMed1991

Mechanism of maturation and nature of carbohydrate chains of boar sperm acrosin.

The acrosin zymogen proacrosin exists in two molecular forms which are believed to be single-chain polypeptides. During autoactivation in a cell-free system, the 55 and 53 kDa zymogens are sequentially converted into the 49, 36, 31 and 25 kDa forms. A similar mechanism of maturation was revealed, when the calcium ionophore A23187 was added to suspensions of boar spermatozoa. The 49 kDa form has been identified as the first active acrosin form in the maturation cascade. However, this form is indistinguishable from the 53 kDa zymogen in SDS-PAGE at nonreducing conditions. Two carbohydrate chains were evidenced on the acrosin molecule. The chain attached to the Asn3 of the acrosin light chain was enzymatically cleaved without loss of acrosin activity. By contrast, the carbohydrate chain linked to the acrosin heavy chain could be cleaved only after acrosin denaturation. Based on the susceptibility of acrosin to endoglycosidases F and H, a biantennary structure of both carbohydrate chains is proposed.

Acrosin↗
PubMed1991

Progression of oocyte maturation from metaphase I to metaphase II is disturbed by previous immunological interference with cumulus cell function.

The effects of an antibody preparation reacting with preovulatory mouse cumuli oophori (anticumulus Ig) on oocyte maturation in vivo and in vitro were studied. Continuous presence of anticumulus Ig in culture medium did not impair oocyte maturation in vitro. Similarly, no effect on oocyte maturation in vivo was observed when anticumulus Ig was given to females superovulated with pregnant mare serum gonadotropin (PMSG) and human chorionic gonadotropin (hCG) at the time of hCG treatment. However, when administered earlier, anticumulus Ig brought about serious disturbances of oocyte meiotic competence, since only immature oocytes were ovulated after anticumulus Ig injection at the time of PMSG treatment and as much as 70% of the ovulated oocytes were immature when the antibody was applied 24 hr later. Previous absorption of anticumulus Ig with isolated cumulus cells removed the inhibitory effect of this preparation on oocyte meiotic competence to the same extent as absorption with whole cumuli oophori, despite the persistence of a strong reactivity of the cumulus cell-absorbed antibody preparations with the cumulus intercellular matrix. The ability of oocytes obtained from antibody-injected animals to mature in vitro was also considerably impaired when the injection was made at the time of PMSG treatment. In all cases the maturation defect concerned the progression of meiosis from metaphase I to metaphase II, while the ability of oocytes to undergo germinal vesicle breakdown (GVBD) was unchanged.(ABSTRACT TRUNCATED AT 250 WORDS)

Absorption↗
PubMed1991

Expression of D-mannose binding sites on human spermatozoa: comparison of fertile donors and infertile patients.

OBJECTIVE: The hypothesis that defective expression of D-mannose binding sites (presumptive elements of the sperm-zona pellucida binding mechanism) is related to male infertility was tested. DESIGN: Experiments were performed on sperm samples from two groups of men classified, respectively, as fertile and infertile, based on their reproductive history. SETTING: The study was carried out in an andrologic laboratory of a University Hospital. PATIENTS, PARTICIPANTS: Fertile men were healthy sperm donors; infertile men were patients with fertility problems. INTERVENTIONS: D-mannose binding sites were visualized by fluorescence microscopy using a mannosylated neoglycoprotein probe. MAIN OUTCOME MEASURES: The hypothesis as reported in the objective section was formulated before data collection and was not modified thereafter. RESULTS: Sperm from fertile men displayed a characteristic pattern of changes in the expression of D-mannose binding sites during in vitro capacitation. This pattern was altered in sperm from infertile men. CONCLUSIONS: If the relationship between defective expression of D-mannose binding sites and decreased sperm fertilizing ability is validated by parallel tests of sperm-zona binding, it may be used for development of chemical tests replacing the current ones using human zonae pellucidae.

Binding Sites↗
PubMed1990

Assembly of the nucleolar precursor bodies in human male pronuclei is correlated with an early RNA synthetic activity.

The nucleolar precursor bodies (NPBs) are electrondense, homogeneous, and finely filamentous intranuclear structures occurring in mammalian zygotes during the early postfertilization development and subsequently transforming into nucleoli. In this study we addressed the question if the assembly of the NPBs is correlated with adenosine incorporation into the pro-nuclear chromatin, an event previously reported to precede the beginning of the pronuclear DNA synthetic phase and suggested to reflect an early pronuclear RNA synthesis. The degree of polyspermy was manipulated artificially, and the effects of the resulting changes in the nucleocytoplasmic ratio on adenosine incorporation and the NPB assembly were analyzed using a quantitative autoradiographic examination. The NPB assembly occurred only in pronuclei which incorporated [3H]-adenosine. This [3H]adenosine labeling of pronuclei was not due to DNA replication, but it could be removed by treatment of egg sections with hot trichloracetic acid before autoradiography, a method which quantitatively extracts nucleic acids. With increasing nucleocytoplasmic ratio, the capacity of developing pronuclei to incorporate adenosine was lost progressively and this phenomenon was closely correlated with impairment of the NPB assembly. These findings suggest that an early pronuclear RNA synthesis is required for the development of the NPBs.

Adenosine↗
PubMed1990

Zona pellucida penetrability of metaphase I and II human oocytes after aging and salt treatment.

Human oocytes that failed to fertilize in vitro were examined for the degree of meiotic maturity and reinseminated either in the living state or after treatment with a concentrated salt solution as used for storage of human zonae pellucidae for diagnostic tests of sperm function. No dependence of zona pellucida penetrability on the oocyte maturity status was observed in either treatment group. Zonae pellucidae of metaphase I and metaphase II oocytes were equally penetrable after the salt treatment, whereas penetration occurred only exceptionally when the oocytes were living. These results show that, after in vitro aging and subsequent salt treatment, eventual differences in oocyte maturity status will not influence results of zona pellucida penetration tests. On the other hand, successful penetration of salt-stored zonae pellucidae by spermatozoa does not necessarily imply that these spermatozoa are able to penetrate the zona pellucida of a fresh living oocyte.

Cell Survival↗
PubMed1990

Effect of follicular fluid on sperm movement characteristics.

Human spermatozoa previously washed from seminal plasma and incubated in capacitating medium were exposed to human follicular fluid (FF) for different time periods and their movement characteristics were examined by computer-assisted digital image analysis. No differences from control samples were observed after short exposures (15 minutes and 2 hours). When the exposure was prolonged to 6 hours, spermatozoa incubated in the presence of FF displayed significant differences in some movement characteristics as compared with samples incubated without FF supplement. When only rapidly moving sperm subpopulations were considered, these effects of FF were similar to those produced by a short incubation with the solubilized cumulus oophorus matrix as described previously. After 24 hours of incubation, there was a remarkable beneficial effect of FF on motility preservation. Possible physiological and clinical implications are discussed.

Body Fluids↗
PubMed1990

Reversible inhibition of fertility in mice by passive immunization with anticumulus oophorus antibodies.

The mucified cumulus oophorus represents an outer enveloping layer around ovulated mammalian oocytes. This coat in its definitive expanded form appears late in the preovulatory development as a result of intensive secretion of intercellular matrix by cumulus cells. We have shown recently that antibodies to the cumulus matrix inhibit human fertilization in vitro. This study was undertaken to assess, in an animal model, the effects of anticumulus oophorus antibodies on fertility by use of different passive immunization protocols. A purified anticumulus immunoglobulin fraction was prepared from hyperimmune rabbit serum and administered at different times before and after mating to mice superovulated with equine chorionic gonadotropin (eCG) and human chorionic gonadotropin (hCG). A dose-dependent negative effect of this anticumulus antibody preparation on the number of fertilized eggs recovered from the oviducts of treated animals was observed when the antibodies were given before mating. High antibody doses also interfered with oocyte maturation and ovulation if applied on the day of eCG treatment, but no effects on these processes were found when the antibodies were given on the day of hCG treatment. The antifertility effect of anticumulus antibodies was reversible and the antibodies did not affect postfertilization development. These findings make cumulus oophorus antigens serious candidates for the development of a contraceptive vaccine.

Animals↗
PubMed1990

Acrosin activation follows its surface exposure and precedes membrane fusion in human sperm acrosome reaction.

Evidence has accumulated suggesting multiple roles of acrosin in fertilization, including its participation in early steps of gamete recognition and binding. However, the implication of acrosin in many of these processes is not compatible with its presumptive sequestration within the sperm acrosome until a late phase of the acrosome reaction. In an earlier study (J. Tesarik, J. Drahorad, J. Peknicova, 1988, Fertil. Steril. 50, 133-141), we reported the binding of an anti-acrosin monoclonal antibody (MO-AKR.1) to the plasma membrane overlying the acrosome of human spermatozoa starting the acrosome reaction. In this study, we characterized further this antibody with regard to its reactivity with different forms of acrosin and found that it recognizes specifically an active form of this enzyme and does not react with its proenzyme form. MO-AKR.1 was thus used as a probe for in situ analysis of acrosin activation during the acrosome reaction. When suspensions of living spermatozoa were incubated with MO-AKR.1 and with another monoclonal antibody (T6) directed to an intra-acrosomal cytoskeletal protein, significantly more spermatozoa reacted with the former antibody than with the latter; this indicated that some of the spermatozoa showing acrosin immunoreactivity carried activated acrosin on the cell surface, while their acrosome was still impermeable to intra-acrosomal-directed probes. The size of this particular sperm subpopulation was increased by the action of follicular fluid (a natural acrosome reaction inducer), but not ionophore A23187 (an artificial acrosome reaction inducer); it corresponded to the proportion of spermatozoa showing acrosin immunoreactivity on the plasma membrane but neither intra-acrosomal staining nor perceptible membrane perturbations when examined by immunoelectron microscopy. When spermatozoa were pre-incubated with protease inhibitors before the addition of acrosome reaction-inducing agents, the percentage of cells binding MO-AKR.1 was markedly reduced. These data suggest that limited acrosin activation on the sperm plasma membrane is an early event in the physiological acrosome reaction.

Acrosin↗
PubMed1990

Effects of prolonged administration of anti-cumulus oophorus antibody on reproduction in mice.

Mice were passively immunized over 6 weeks with contraceptive doses of an anti-cumulus oophorus antibody preparation. The persistence of oestrus, accompanied by complete inhibition of conception, was observed throughout treatment. After cessation of treatment, fertility was restored within 2 weeks. Histological examination did not reveal any depletion of the ovarian oocyte stock. These results warrant further research into the nature of cumulus antigens and their use in active immunization studies.

Animals↗
PubMed1989

Development of human male pronucleus: ultrastructure and timing.

The process of human male pronuclear formation was studied using an experimental model based on in vitro inseminated human zona-free eggs prepared from oocytes that failed to fertilize in a clinical in vitro fertilization program. The main ultrastructural changes in penetrated sperm nuclei transforming pronuclei were used to define four stages of pronuclear development. The first two stages, representing partial (Stage 1) and total (Stage 2) sperm chromatin decondensation, appeared as early as 1 hr after mixing of gametes. This rapid initial phase was followed by a more lengthy array of events leading to transformation of decondensed sperm nuclei into fully developed male pronuclei (Stages 3 and 4). Stage 3 was characterized by reformation of the nuclear envelope, reorganization of chromatin, and the assembly of nucleolar precursors. It was not completed until 12 hr after in vitro insemination when fully developed male pronuclei (Stage 4) were first observed. In some eggs pronuclei did not reach Stage 4 at all. The results of this study provide a morphological background for further research into molecular aspects of human male pronuclear development and its regulation.

Cell Nucleus↗
PubMed1989

The potential diagnostic use of human zona-free eggs prepared from oocytes that failed to fertilize in vitro.

Human oocytes that failed to fertilize in a clinical in vitro fertilization (IVF) program were freed from the zona pellucida and reinseminated with spermatozoa from subfertile patients or from fertile donors. Penetration results were, in general, significantly worse when patients spermatozoa were used as compared with donor spermatozoa. When inseminated with donor spermatozoa, oocytes from cases of male infertility gave better penetration results as compared with those from cases of idiopathic infertility with no apparent sperm defect. No differences in penetration results were found between oocytes aged 1 day and 2 days in culture before zona removal and reinsemination, even though less pronuclei reached full functional maturity in the more aged group of eggs. These results indicate that human zona-free eggs prepared from oocytes after unsuccessful IVF attempts may be used to assess the fertilizing ability of human spermatozoa. Moreover, when used with donor spermatozoa, this system may help to determine the contribution of the oocyte to fertilization failure.

Female↗
PubMed1989

Human sperm--egg interactions and their disorders: implications in the management of infertility.

The present era of infertility management is characterized by an increasing interest in defects of sperm--egg interactions. When the spermatozoon meets the egg, it must first react with the enveloping coat which controls sperm access to the cell surface of the oocyte. Adequate sperm motility is a major condition for penetration through the egg investments. The ability of spermatozoa to respond to physiological stimuli by a timely acrosome reaction is another necessary prerequisite for this event, as well as for sperm fusion with the oocyte. The clinical significance of current laboratory tests evaluating different aspects of gamete function is discussed and a practical guide is given for their indication and interpretation and for the use of their results in choosing rational therapy.

Acrosome↗
PubMed1989

Appropriate timing of the acrosome reaction is a major requirement for the fertilizing spermatozoon.

This study was undertaken to determine the site of the acrosome reaction of spermatozoa penetrating into freshly inseminated human oocytes. The inseminated oocytes were treated with an anti-acrosin monoclonal antibody and the bound antibody was visualized at the ultrastructural level with the use of a second peroxidase-conjugated antibody. Quantitative analysis of serial thin sections cut throughout the specimens showed that the number of spermatozoa within the zona pellucida (potentially fertilizing ones) corresponded to the number of acrosin deposits associated with acrosomal ghosts on the zona pellucida surface. As it is known that a large acrosin bundle is liberated from a spermatozoon at a well-defined point of the acrosome reaction, these findings indicate that the acrosome reaction of the fertilizing spermatozoon must be exactly synchronized with its penetration through the egg vestments, apparently by the action of specific acrosome reaction-promoting substances in the oocyte/cumulus complex. These results represent a theoretical basis for evaluation of direct and indirect laboratory tests for human sperm acrosome reaction. 'Good' sperm samples should display elevated levels of acrosome-reacted spermatozoa after the administration of an appropriate stimulus and low levels of spontaneous acrosome reactions.

Acrosin↗
PubMed1989

Developmental control of the human male pronucleus by ooplasmic factors.

The nature of oocyte cytoplasmic factors controlling the development of the male pronucleus was investigated by inseminating human, zona-free oocytes at metaphase of the 1st and 2nd meiotic division. Oocytes at metaphase of the 2nd meiotic division could support the full structural and functional development of male pronuclei, whereas the vast majority of those at metaphase of the 1st meiotic division failed to do so. This suggests that oocyte cytoplasmic factors required for male pronuclear formation do not develop fully until the oocyte reaches the 2nd meiotic metaphase. When increasing numbers of spermatozoa entered one oocyte, the transformation of sperm nuclei into pronuclei was impaired progressively. The later stages of pronuclear development were particularly sensitive to polyspermy. These factors should be taken into consideration in the development of techniques of micromanipulation-assisted insemination.

Autoradiography↗