RESUMO
Mammalian sperm-zona pellucida (ZP) interaction is mediated by sperm lectin-like proteins and ZP glycoproteins. We have previously reported the participation of binding sites for N-acetylglucosamine (GlcNAc) residues in human sperm function, including sperm interaction with the ZP. Additionally, previous results from our laboratory suggested that some of these events may be mediated by the glycosidase N-acetylglucosaminidase (beta-hexosaminidase, Hex, in mammals). In this study, we report the possible participation of Hex in human sperm-ZP interaction. Human recombinant Hex (hrHex) was obtained by expression in a stable transfected CHO cell line. When the recombinant enzyme was present during hemizona (HZ) assays, the number of sperm bound per HZ was significantly reduced. The same result was obtained when HZ were preincubated with hrHex. Additionally, the presence of a Hex-specific substrate during the HZ assay produced the same inhibitory effect. These results suggest the participation of a sperm Hex in the interaction with human ZP in vitro.
Assuntos
Interações Espermatozoide-Óvulo/fisiologia , Zona Pelúcida/fisiologia , beta-N-Acetil-Hexosaminidases/metabolismo , Acetilglucosamina/análogos & derivados , Acetilglucosamina/metabolismo , Acetilglucosamina/farmacologia , Animais , Células CHO , Cricetinae , Feminino , Humanos , Concentração de Íons de Hidrogênio , Himecromona/análogos & derivados , Himecromona/metabolismo , Himecromona/farmacologia , Masculino , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Proteínas Recombinantes/farmacologia , Interações Espermatozoide-Óvulo/efeitos dos fármacos , beta-N-Acetil-Hexosaminidases/genética , beta-N-Acetil-Hexosaminidases/farmacologiaRESUMO
The ability of strontium (Sr(2+)) to replace calcium (Ca(2+)) in maintaining human sperm function has still not been completely characterized. In the present study, acrosome reaction (AR) inducibility in response to human follicular fluid (hFF) was compared in spermatozoa incubated in either Ca(2+)- or Sr(2+)-containing media. Other events related to sperm capacitation, such as protein tyrosine phosphorylation and hyperactivation as well as zona pellucida (ZP) recognition under both conditions, were also analyzed. Spermatozoa incubated overnight in the presence of Sr(2+) were unable to undergo the AR when exposed to hFF. Nevertheless, when spermatozoa were incubated under this condition and then transferred to medium with Ca(2+), sperm response to hFF was similar to that of cells incubated throughout in the presence of Ca(2+). The sperm protein tyrosine phosphorylation patterns and the percentages of sperm motility and hyperactivation were similar after incubation in Ca(2+)- or Sr(2+)-containing media. Under both conditions, the same binding capacity to homologous ZP was observed. Similar results were obtained when EGTA was added in order to chelate traces of Ca(2+) present in Sr(2+) medium. From these results, it can be concluded that Sr(2+) can replace Ca(2+) in supporting capacitation-related events and ZP binding, but not hFF-induced AR of human spermatozoa.
Assuntos
Reação Acrossômica/efeitos dos fármacos , Líquido Folicular/fisiologia , Capacitação Espermática/efeitos dos fármacos , Espermatozoides/efeitos dos fármacos , Estrôncio/farmacologia , Cálcio/administração & dosagem , Cálcio/farmacologia , Ácido Egtázico/farmacologia , Feminino , Humanos , Cinética , Masculino , Fosforilação , Fosfotirosina/metabolismo , Motilidade dos Espermatozoides , Interações Espermatozoide-Óvulo , Espermatozoides/fisiologia , Zona Pelúcida/fisiologiaRESUMO
OBJECTIVE: To determine whether the use of a central assisted reproduction laboratory, with gamete transport to the facility (transport assisted reproduction), would decrease oocyte quality or performance in IVF-ET and intracytoplasmic sperm injection (ICSI). DESIGN: Retrospective clinical study. SETTING: Public and private fertility clinics. PATIENT(S): A total of 467 couples underwent transport IVF, whereas 108 underwent transport ICSI. A group of 60 couples underwent conventional IVF during the same period. All methods and protocols used were similar among centers. Oocyte pick-up was performed by ultrasound-guided vaginal puncture. INTERVENTION(S): Oocytes were transported under controlled conditions, from the site of follicular aspiration to a central laboratory. MAIN OUTCOME MEASURE(S): The fertilization and cleavage rates and clinical pregnancies were compared among the study populations. RESULT(S): The differences between the fertilization and cleavage rates of ova and the rates of clinical pregnancies produced by transport and conventional methods were not statistically significant. CONCLUSION(S): Gamete transport to a central laboratory was not harmful for oocytes or for the outcome of assisted reproduction. Transport makes the use of IVF and ICSI available to physicians who are not affiliated with an assisted reproduction program, reduces costs, and increases acceptability of the procedures to patients.
Assuntos
Fertilização in vitro/métodos , Microinjeções , Manejo de Espécimes/métodos , Adulto , Custos e Análise de Custo , Feminino , Fertilização in vitro/economia , Humanos , Masculino , Oócitos , Folículo Ovariano/citologia , Gravidez , Estudos Retrospectivos , Manejo de Espécimes/economia , SucçãoRESUMO
Glycosidic residues of the mammalian zona pellucida (ZP) are known to be involved in sperm binding, suggesting the presence of complementary carbohydrate binding sites on spermatozoa. However, in previous studies, in which sperm suspensions were incubated with monosaccharides, no inhibitory effect was observed. Results of studies in which sperm were treated shortly after swim-up suggest that the use of non-capacitated cells may explain the apparently conflicting results. In the present report, we studied the effect of preincubation of capacitated spermatozoa with different monosaccharides on their ability to bind to ZP. After 5 h under capacitating conditions, spermatozoa were incubated in medium with or without a monosaccharide, resuspended in fresh medium and used for hemizona (HZ) binding assay. When ZH were incubated with spermatozoa treated with N-acetyl-D-glucosamine, D-mannose, D-fucose, L-fucose or D-galactose, a significant decrease in the number of spermatozoa bound was observed (level of inhibition: 62, 58, 82, 68 and 48% respectively) while treatment of spermatozoa with D-glucose produced no inhibition. Sugar treatment neither altered sperm motility nor the rate of acrosome reaction. These results suggest that N-acetylglucosamine, mannose, fucose and galactose residues are involved in human sperm-zona pellucida binding in vitro.
Assuntos
Glicoconjugados/fisiologia , Interações Espermatozoide-Óvulo/fisiologia , Zona Pelúcida/fisiologia , Sítios de Ligação , Feminino , Glicoconjugados/química , Humanos , Técnicas In Vitro , Masculino , Monossacarídeos/metabolismo , Monossacarídeos/farmacologia , Capacitação Espermática , Interações Espermatozoide-Óvulo/efeitos dos fármacos , Espermatozoides/efeitos dos fármacos , Espermatozoides/fisiologia , Zona Pelúcida/químicaRESUMO
Neoglycoproteins with N-acetylglucosamine residues (BSA-GlcNAc) induced specifically the acrosome reaction (AR) in human spermatozoa. Our objective was to investigate the relationship between this phenomenon and the invitro fertilization (IVF) rate. Sperm suspensions from IVF protocols were incubated with BSA-GlcNAc (t), using calcium ionophore (i) or medium alone (c) as positive or negative controls. When the normalized AR percentage ratio (STIM) (% ARt-%ARc):(%ARi-%ARc) was compared with fertilization rate in 31 couples from our IVF programme, a positive correlation was found (r = 0.46, P < 0.01). The fertilization rate in patients with STIM > or = 0.2 was higher than in non-responders (STIM < 0.2); 72 +/- 7% compared with 5 +/- 3%. The overall predictive value of this test for adequate fertilization rate (> 30%) was 87%, sensitivity 91% and specificity 78%. False positives were 9% and false negatives 22%. For successful fertilization rates (> 60%), the results were: overall predictive value, 84%; sensitivity 100%; specificity 64%. False positives were 23% and no false negatives were found. The results indicated that the induction of AR in human spermatozoa by GlcNAc-neoglycoproteins could be used to predict their fertilizing ability in vitro.
Assuntos
Acetilglucosamina/farmacologia , Acrossomo/fisiologia , Fertilização in vitro , Soroalbumina Bovina/farmacologia , Espermatozoides/efeitos dos fármacos , Acrossomo/efeitos dos fármacos , Calcimicina/farmacologia , Combinação de Medicamentos , Feminino , Humanos , Masculino , Valor Preditivo dos Testes , Interações Espermatozoide-ÓvuloRESUMO
A polyclonal antiserum directed against human sperm coating proteins of epididymal origin (anti-KCl) was tested for its ability to alter sperm function. Spermatozoa from normal ejaculates were selected by swim-up separation and capacitated by overnight incubation at room temperature. Exposure of these cells to anti-KCl (0.39 mg protein/ml), prior to their use in the hamster ova penetration test, reduced the penetration of denuded oocytes by 65% (P less than 0.005). Significant inhibitions of lesser magnitude were observed at lower serum concentrations (to 0.098 mg/ml). In an effort to understand the mechanism of this inhibition, other sperm function parameters thought to be related to oocyte penetration were studied. The inhibitory effect was exerted without noticeable changes in sperm motility (determined by the percentage of motile cells and their linear velocity), and in the absence of major sperm agglutination. Anti-KCl did not inhibit the occurrence of spontaneous or induced (by human follicular fluid) acrosome reaction in capacitated spermatozoa. In contrast, exposure to anti-KCl reduced the ability of capacitated spermatozoa to bind tightly to the hamster oolemma. None of these effects were elicited by a control preparation obtained from pre-immune rabbit sera. Exposure of zona-free oocytes to the antiserum did not alter their penetrability by normal sperm. These results suggest that the antigens recognized by anti-KCl participate in some specific step of the sperm-ovum interaction.