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Biomedical subjects

C Mendoza

Publications and source records attributed to C Mendoza.

At least 91 records · Page 5Linked to original sources
PubMed1993

Fast acrosome reaction measure: a highly sensitive method for evaluating stimulus-induced acrosome reaction.

OBJECTIVE: To test a new method for evaluation of stimulus-induced acrosome reaction (AR). To determine whether this method, based on the definition of a specific staining pattern of recently reacted spermatozoa, brings an advantage of increased sensitivity as compared with a standard procedure. DESIGN: The hypothesis that a specific staining pattern with fluorescein-labeled Pisum sativum agglutinin corresponds to recently acrosome-reacted spermatozoa was tested by analyzing the frequency of this pattern at different time points after addition of ionophore A23187, an artificial AR inducer. The AR results obtained with a new method based on this relationship were compared with those obtained with a standard method of AR evaluation. SETTING: Private hospital, public research center, and a university-based research laboratory. PARTICIPANTS: Healthy sperm donors. MAIN OUTCOME MEASURE: Changes in the frequency of specific sperm-staining patterns. RESULTS: A specific sperm-staining pattern with P. sativum agglutinin was shown to be associated with a recently occurred AR, whereas the absence of staining was typical of reactions having occurred a longer time ago. This phenomenon was used to define a formula for fast AR measure after stimulus addition. An increased sensitivity provided by this formula as compared with a standard evaluation was demonstrated. CONCLUSIONS: Fast AR measure is a simple, easy-to-perform method for AR evaluation. It is particularly suitable for testing the effects of rapidly acting stimuli.

Acrosome↗
PubMed1993

Sperm treatment with pentoxifylline improves the fertilizing ability in patients with acrosome reaction insufficiency.

OBJECTIVE: To test whether pentoxifylline, a drug previously shown to sensitize spermatozoa from normal samples to the action of acrosome reaction stimuli, can be used to improve the fertilizing ability of spermatozoa from patients with acrosome reaction insufficiency. DESIGN: Prospective analysis of pentoxifylline effects on the acrosome reaction and zona-free egg penetration; retrospective comparison of IVF results with and without the use of pentoxifylline. SETTING: Private hospital, public research center, and university-based laboratory. PATIENTS: In vitro fertilization patients selected on the basis of a previous acrosome reaction test. INTERVENTIONS: None in the experimental part; IVF-ET in the clinical part of this study. MAIN OUTCOME MEASURES: Frequency of the acrosome reaction, rate of penetration of zona-free eggs, normal fertilization in IVF attempts. RESULTS: Pentoxifylline improves the acrosome reaction scores and zona-free egg penetration rates in patients with acrosome reaction insufficiency. Preliminary clinical experience shows an improvement of IVF results in these patients. CONCLUSIONS: In vitro pentoxifylline treatment of spermatozoa to be used in IVF improves the sperm fertilizing ability in patients with acrosome reaction insufficiency. However, the effect of pentoxifylline on the acrosome reaction should be tested individually in each patient before the application of the drug in this new indication.

Acrosome↗
PubMed1993

Solubilized human zona pellucida competes with a fucosylated neoglycoprotein for binding sites on the human sperm surface.

OBJECTIVE: To evaluate the implication of fucose binding sites on the sperm surface in the sperm-zona pellucida (ZP) binding by examining the ability of a fucosylated neoglycoprotein to compete with solubilized zona proteins for sperm binding. DESIGN: The binding of a fucosylated neoglycoprotein to spermatozoa was analyzed in relation to sperm capacitation and the actual acrosomal status. The effect of solubilized ZP material on the binding was examined. SETTING: Private hospital, public research center, and university department. PATIENTS, PARTICIPANTS: Healthy volunteers with normal sperm parameters. INTERVENTIONS: None. MAIN OUTCOME MEASURES: Fluorescence signal detected by fluorescence-activated cell sorting or fluorescence microscopy on spermatozoa incubated with fluorescein isothiocyanate-labeled fucosylated neoglycoprotein. RESULTS: There was a small subpopulation of spermatozoa that showed a strong binding activity for the fucosylated neoglycoprotein, and the size of this subpopulation increased with the time of exposure to capacitating conditions. The binding sites were located on the head (mainly on the acrosomal region) of acrosome-intact spermatozoa. The neoglycoprotein binding was competitively inhibited by solubilized ZP. CONCLUSIONS: The data show that a commercially available fucosylated neoglycoprotein competes with solubilized ZP proteins for binding sites on the sperm surface. Further research into the relationship between the neoglycoprotein binding and zona binding is warranted to evaluate the potential diagnostic usefulness of this neoglycoprotein.

Binding Sites↗
PubMed1993

Stimulation of protein tyrosine phosphorylation by a progesterone receptor on the cell surface of human sperm.

Mature human sperm initiate a rapid Ca2+ influx and the acrosomal exocytosis in response to progesterone. Recent evidence indicates that both events can be induced by antibody-mediated cross-linking of a sperm surface progesterone receptor. In many other systems in which signal is generated by receptor cross-linking, protein phosphorylation on tyrosine residues is involved in the signal transduction across the plasma membrane. In this study we examined whether tyrosine phosphorylation is implicated in the function of the sperm surface progesterone receptor, too. The effect of progesterone on the phosphorylation of proteins from a sperm membrane lysate was evaluated by in vitro kinase assay and by phosphoamino acid analysis using [gamma-32P]ATP as precursor. These experiments revealed a selective increase in the tyrosine phosphorylation of a 94-kilodalton phosphoprotein in the presence of progesterone. To decide whether the progesterone-induced increase in protein tyrosine phosphorylation is actually due to the hormone action on the cell surface, living sperm were treated with a cell-impermeant progesterone receptor agonist, and the resulting changes in the cellular level of phosphotyrosine proteins were examined. These experiments showed a clear relationship between the agonist binding and an increase in the phosphotyrosine concentration in the respective cells. This relationship was lost in the presence of genistein, which also efficiently inhibited the phosphorylation of the 94-kilodalton protein and the progesterone-induced acrosomal exocytosis. These results lead to the hypothesis that protein tyrosine phosphorylation is involved in signal transduction through the sperm surface progesterone receptor and may be implicated in nongenomic steroid effects in other cell types.

Adenosine Triphosphate↗
PubMed1992

Progesterone action through aggregation of a receptor on the sperm plasma membrane.

Rapid steroid effects, reported in several cell types, have pointed out the possibility of non-genomic mechanisms of action, presumably on cell surface receptors. Here we analyzed the effects of antibody-mediated aggregation of a novel type of progesterone receptor on the plasma membrane of human sperm cells. We report that aggregation of hormone-receptor complexes induces Ca2+ influx and a Ca(2+)-dependent exocytotic event in this system. These data suggest a possible mechanism for rapid steroid-induced events. Further research is warranted to examined if a similar mechanism is involved in rapid steroid effects in other cell types.

Biological Transport↗
PubMed1992

Selective expression of a progesterone receptor on the human sperm surface.

OBJECTIVE: To visualize progesterone (P) binding sites on the sperm surface, examine the relationship between hormone binding and hormone action (acrosome reaction), and determine the size of the hormone-responsive sperm subpopulation. DESIGN: Kinetic analysis of P binding was combined with the assessment of the hormone effect using a fluorescent acrosomal marker. SETTING: Private hospital, medical research center, and a university-based andrological laboratory. PATIENTS, PARTICIPANTS: Sperm samples were from healthy volunteers with normal spermiogram values. INTERVENTIONS: None. MAIN OUTCOME MEASURES: Progesterone binding was analyzed by fluorescence microscopy and flow cytometry using P coupled to fluorescein isothiocyanate-labeled bovine serum albumin. Tetramethylrhodamine isothiocyanate-labeled Pisum sativum agglutinin was used as acrosomal marker in double-labeling experiments. RESULTS: After in vitro capacitation, only few spermatozoa (approximately 10%) were able to bind P to the cell surface, but most of these cells subsequently generated the acrosome reaction in response to hormone binding. CONCLUSIONS: The expression of P receptor sites on the human sperm surface is a major factor controlling the P-induced acrosome reaction. Further studies are warranted to explore if defective expression of the receptor can compromise fertility.

Binding Sites↗
PubMed1992

Defective function of a nongenomic progesterone receptor as a sole sperm anomaly in infertile patients.

OBJECTIVE: To compare the function of a novel nongenomic progesterone (P) receptor on the human sperm surface (mediating the P-induced acrosome reaction) in spermatozoa from fertile donors and from infertile patients. To examine the possible implication of defective P receptor function as an etiologic factor in unexplained male infertility. DESIGN: Progesterone binding and P effects were assessed in sperm from infertile patients and compared with corresponding parameters for sperm from healthy donors. SETTING: Private hospital, medical research center, and a university-based andrological laboratory. PATIENTS, PARTICIPANTS: Sperm samples were from infertile patients (no pathology detected in their wives) attending our infertility clinic and from healthy sperm donors. INTERVENTIONS: None. MAIN OUTCOME MEASURES: Progesterone binding sites were visualized with a fluorescein-labeled protein-P conjugate. Indo 1-AM (a fluorescent indicator of intracellular free Ca2+) was used to measure P-induced Ca2+ influx. Progesterone-induced acrosome reaction was monitored after acrosomal staining with Pisum sativum agglutinin. RESULTS: Among 8 patient sperm samples with normal spermiogram values (of 53 examined), 5 showed a reduced percentage of P-binding spermatozoa and an abnormal response to the hormone in terms of Ca2+ influx and the acrosome reaction. CONCLUSIONS: Defective function of a sperm surface P receptor is described in some cases of male infertility. The observed fluorescence microscopic patterns of hormone binding may be used to further investigate receptor activity in unexplained male infertility.

Acrosome↗
PubMed1992

Effects of phosphodiesterase inhibitors caffeine and pentoxifylline on spontaneous and stimulus-induced acrosome reactions in human sperm.

OBJECTIVE: To determine whether the phosphodiesterase inhibitors caffeine and pentoxifylline influence the acrosome reaction in the conditions in which they are currently used as sperm movement enhancers. DESIGN: The frequency of acrosome reaction occurring spontaneously in capacitating media or induced by physiological (follicular fluid [FF]) and artificial (ionophore A23187) stimuli was compared in the presence and absence of the phosphodiesterase inhibitors. SETTING: Private hospital and research laboratory. PATIENTS, PARTICIPANTS: Patients undergoing routine semen examination before in vitro fertilization (no pathology detected) and healthy sperm donors. INTERVENTIONS: None. MAIN OUTCOME MEASURE: Percentage of acrosome-reacted sperm determined with the use of fluorescein-labeled Pisum sativum agglutinin as acrosomal stain. RESULTS: Caffeine alone augmented the frequency of acrosome reaction, but this effect was not observed with pentoxifylline alone. However, pentoxifylline increased sperm responsiveness to the acrosome reaction-inducing stimuli, FF and ionophore A23187. CONCLUSIONS: The promotion of spontaneous acrosome reaction may counteract the benefits from application of caffeine as motility stimulant. On the other hand, the sensitization to physiological acrosome reaction stimuli is expected to contribute to the improvement of sperm fertilizing ability by pentoxifylline and make this drug a potential candidate for the treatment of acrosome reaction anomalies.

3',5'-Cyclic-AMP Phosphodiesterases↗
PubMed1992

Influence of stabilization occlusal splints on sternocleidomastoid and masseter electromyographic activity.

The present work was conducted in order to determine the effect of stabilization occlusal splints on electromyographic (EMG) activity of sternocleidomastoid and masseter muscles, in subjects with tenderness to palpation in these muscles. A full-arch maxillary stabilization occlusal splint was made for each of 14 subjects. Tonic EMG activity, as well as during saliva swallowing and maximal voluntary clenching, was recorded with and without a stabilization occlusal splint inserted. Similar tonic, as well as maximal voluntary clenching EMG activity, with and without the stabilization occlusal splint, was observed. During saliva swallowing, the activity in both muscles was significantly lower with the stabilization occlusal splint. This suggests that daytime use of the stabilization occlusal splint might improve tenderness to palpation in the studied muscles, since the frequency of swallowing function is higher during waking hours.

Adult↗
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

Inhibition of natural killer activity in porcine mononuclear cells by African swine fever virus.

The coincubation at 37 degrees C for 24 hours of swine peripheral blood mononuclear cells with African swine fever virus inhibited in part the natural killer activity shown by cells incubated without the virus. This inhibition depended on the dose of the virus and on the time that cells were incubated with it. When the virus preparation was fractionated by ultracentrifugation, most of the inhibitory activity was found in the sedimented fraction, where viral particles were present; however, the loss of inhibitory activity in respect to the whole virus preparation indicated that some inhibitory activity was present in the supernatant fraction, probably as factors released by infected cells. Most of the inhibitory activity shown by the sedimented fraction was lost when the virus was inactivated by ultraviolet radiation, indicating an active role of virus infectivity in the inhibition.

African Swine Fever Virus↗
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

Inhibitory effect of African swine fever virus on lectin-dependent swine lymphocyte proliferation.

The incubation of swine peripheral blood mononuclear cells (PBMC) with African swine fever (ASF) virus preparations strongly inhibited the proliferative response of lymphocytes to PHA and other lectins. The inhibition, which persisted after inactivation of the virus by UV radiation, was dependent upon the dose and the time that virus preparations were present in cultures. When virus preparations were fractionated by ultracentrifugation, the inhibitory activity resulted to be soluble, whereas no activity was found in the sedimented viral fraction. However, the preincubation during 4 days of this sedimented fraction with swine PBMC, before the addition of the mitogen, restored the inhibitory activity. The results obtained suggest that the inhibition is mediated by one or more soluble factors released by swine PBMC after coincubation with ASF virus in a time dependent process. These factors show a molecular weight between 40 and 80 kDa by gel filtration chromatography. The inhibitory activity described in the present paper is an indication of inhibition of lymphocyte function produced by ASF virus which can help to understand how this virus escapes from the host immune system.

African Swine Fever↗
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

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↗
PubMed1988

[Effects of different methods of ovarian stimulation in adult rats].

The in vitro fecundation programs normally use different ovarian stimulation agents. The purpose of the present study was to compare the effects of some of these agents on indices of ovulation and serum levels of estradiol, progesterone, and FSH. Treatments studied consisted of clomiphene citrate, PMSG alone and in combination with clomiphene citrate, pure human FSH and epimestrol. The data obtained show PMSG alone and PMSG plus clomiphene citrate to be the most effective treatments, in terms of number of oocytes harvested. No differences were noted between serum levels of oestradiol, progesterone and FSH.

Animals↗
PubMed1987

Testosterone implants into the lateral septum of male rats, a positive effect on LH and FSH secretion.

After two days, testosterone implanted into the lateral septum increased serum levels of LH and FSH in male Wistar rats. As measured by RIA, LH in animals with testosterone implanted in them in comparison to those with an empty cannula was 0.220 +/- 0.015 vs. 0.111 +/- 0.019 ng/ml; p less than 0.001 and FSH was 3.20 + 0.21 vs. 1.50 + 0.21 ng/ml; p less than 0.001. Serum testosterone was not increased to a statistically significant extent by the implants (4.12 +/- 0.54 vs. 2.87 +/- 0.42 ng/ml; ns). It was concluded that testosterone or possibly one of its metabolites acting in the lateral septum facilitates the release of LH and FSH.

Animals↗