Fine structure distribution of non-specific acid phosphatase in the head region of mouse spermatozoa from various regions of the male reproductive tract.
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The fine structure distribution of non-specific acid phosphatase was determined in the head region of mouse spermatozoa from the testes, the caput, corpus and cauda epididymidis and the ductus deferens. Enzymatic localization was achieved by the Gomori technique. The postacrosomal dense lamina, the nuclear side of the inner acrosomal membrane and the space between the plasmalemma and the outer acrosomal membrane showed reaction product in spermatozoa from the testis and caput epididymidis. Spermatozoa from the cauda epididymidis exhibited reaction product only between the plasmalemma and the outer acrosomal membrane. Spermatozoa from the corpus epididymidis and from the ductus deferens showed no reaction product in the head region. The changes observed in the distribution of acid phosphatase in the sperm head during epididymal transport may reflect maturational events. (+info)
Molecular aspects of mammalian fertilization.
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Mammalian fertilization is a highly regulated process, much of which are not clearly understood. Here we present some information in order to elaborate a working hypothesis for this process, beginning with the sperm modifications in the epidydimis up to sperm and egg plasmalemma interaction and fusion. We also discuss the still poorly understood capacitation process, the phenomenon of sperm chemo-attraction that brings the capacitated sperm to interact with the oocyte vestments and certain aspects of the acrosome reaction. (+info)
Behaviour of a sperm surface transmembrane glycoprotein basigin during epididymal maturation and its role in fertilization in mice.
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Basigin (bsg) is a transmembrane glycoprotein belonging to an immunoglobulin superfamily and is localized on the surface of the sperm tail. The behaviour of bsg during epididymal maturation and its role in fertilization were examined using an anti-bsg antibody. Spermatozoa from caput, corpus and cauda epididymides were immunostained by indirect immunofluorescence (IIF). Immunostaining revealed that bsg is localized on the principal piece of caput spermatozoa and the molecule was found on the middle piece during transit in the corpus and cauda epididymides. Concomitantly, the molecular mass of bsg was reduced from 37 kDa (testis) to 26 kDa (cauda epididymidis). IVF experiments were designed to assess the effect of anti-bsg antibody on the fertilization events. Anti-bsg antibody significantly inhibited primary binding to the cumulus-invested oocytes with intact zonae pellucidae in a dose-dependent manner. Consequently, the fertilization rate of cumulus-invested oocytes with intact zonae pellucidae was also inhibited. The bsg molecule was also detected on the head of live capacitated spermatozoa by IIF under IVF conditions. These findings indicate that testicular bsg is a glycosylated protein that undergoes molecular processing and deglycosylation during its transit in the epididymis. The bsg molecule that was detected on the sperm head after capacitation may facilitate the primary binding or might be involved in distinct events required for primary binding of spermatozoa to the zona pellucida during capacitation and sperm-cumulus interaction. (+info)
Efficacy and safety of a new vaginal contraceptive antimicrobial formulation containing high molecular weight poly(sodium 4-styrenesulfonate).
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Host cell infection by sexually transmitted disease (STD)-causing microbes and fertilization by spermatozoa may have some mechanisms in common. If so, certain noncytotoxic agents could inhibit the functional activity of both organisms. High molecular mass poly(sodium 4-styrenesulfonate) (T-PSS) may be one of these compounds. T-PSS alone (1 mg/ml) or in a gel (2% or 5% T-PSS) completely prevented conception in the rabbit. Contraception was not due to sperm cytotoxicity or to an effect on sperm migration. However, T-PSS inhibited sperm hyaluronidase (IC(50) = 5.3 microg/ml) and acrosin (IC(50) = 0.3 microg/ml) and caused the loss of acrosomes from spermatozoa (85% maximal loss by 0.5 microg/ml). T-PSS (5% in gel) also reduced sperm penetration into bovine cervical mucus (73% inhibition by 1 mg gel/ml). T-PSS (5% in gel) inhibited human immunodeficiency virus (HIV; IC(50)= 16 microg gel/ml) and herpes simplex viruses (HSV-1 and HSV-2; IC(50) = 1.3 and 1.0 microg gel/ml, respectively). The drug showed high efficacy against a number of clinical isolates and laboratory strains. T-PSS (5% in gel) also inhibited Neisseria gonorrhea (IC(50) < 1.0 gel/ml) and Chlamydia trachomatis (IC(50) = 1.2 microg gel/ml) but had no effect on lactobacilli. These results imply that T-PSS is an effective functional inhibitor of both spermatozoa and certain STD-causing microbes. The noncytotoxic nature should make T-PSS safe for vaginal use. T-PSS was nonmutagenic in vitro and possessed an acute oral toxicity of >5 g/kg (rat). Gel with 10% T-PSS did not irritate the skin or penile mucosa (rabbit) and caused no dermal sensitization (guinea pig). Vaginal administration of the 5% T-PSS gel to the rabbit for 14 consecutive days caused no systemic toxicity and only mild (acceptable) vaginal irritation. T-PSS in gel form is worthy of clinical evaluation as a vaginal contraceptive HIV/STD preventative. (+info)
Role of myometrial activity in sperm transport through the genital tract and in fertilization in sows.
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The effects of stimulation and suppression of uterine contractility at about the time of insemination on sperm distribution and fertilization in multiparous sows are described. For assessment of fertilization, sows were inseminated about 28 h before (synchronized) ovulation and killed at day 5 after ovulation (n = 53). For assessment of sperm distribution, sows were inseminated about 20 h before expected ovulation and were killed 12 h later (n = 26). At 10 min before insemination, sows received an intrauterine infusion of one of three solutions: (i) saline (control); (ii) 0.60 mg clenbuterol hydrochloride to suppress contractility; or (iii) 1 mg cloprostenol to stimulate contractility. Both clenbuterol and cloprostenol reduced median fertilization rate (P < 0.05) and median number of accessory sperm cells (P < 0.05). Distribution of sperm cells was also affected by treatments. Clenbuterol increased, and cloprostenol decreased, the number of sperm cells (P < 0.05) in the proximal 20 cm of the uterine horn and in the uterotubal junction. In addition, clenbuterol tended to increase and cloprostenol tended to decrease the number of sperm cells in the isthmus, although these effects were not significant. However, relative to the number of sperm cells in the uterus, clenbuterol treatment reduced the number of sperm cells in the uterotubal junction and oviduct, in contrast to cloprostenol. Cloprostenol increased the reflux of semen during insemination. It is hypothesized that suppression of uterine contractility increases transuterine transport time, reducing the ability of sperm cells to enter the uterotubal junction and the oviduct. Stimulation of uterine contractility above a certain level probably increases reflux and impedes transuterine transport of sufficient numbers of sperm cells. (+info)
Changes in flagellar bending during the course of hyperactivation in hamster spermatozoa.
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The motility pattern of mammalian spermatozoa changes during migration in the female genital tract and during incubation in vitro. This change in motility is termed hyperactivation. Hyperactivated spermatozoa swim vigorously in 'whiplash', 'figure-8' or 'small circle' trajectories. In this study, a quantitative analysis was carried out of the changes in the motility pattern of hamster spermatozoa during incubation to investigate the mechanism regulating hyperactivation. In the culture system used in this study, hyperactivation occurred 4 h after incubation. Several parameters in the analysis of sperm movement pattern were examined. Curvilinear velocity, average path velocity and straightness abruptly increased between 2 and 4 h. However, linearity, amplitude of lateral head displacement, beat cross frequency and average wavelength gradually changed with time. In the analysis of flagellar bending, the bend angles were measured after dividing images of the flagellum into short lengths. Flagellar bending changed in different manners in each region during incubation. The asymmetry in the direction of the curve of the head gradually increased with time in the first half of the flagellum. The flexibility, which was determined using the amplitude of bending and the rate of change in bend angles, abruptly decreased between 10 min and 1 h, and then increased between 2 and 4 h in the first half of flagellum. These results indicate that complex physiological changes occur before hyperactivation. (+info)
Immunohistochemical localization and expression of the hyaluronan receptor CD44 in the epithelium of the pig oviduct during oestrus.
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Hyaluronan is related to essential reproductive processes in pigs. Hyaluronan produced by cumulus cells builds, via specific cell surface receptors, an extracellular matrix responsible for cumulus cell cloud expansion during final oocyte maturation, a preparatory event for ovulation and fertilization. In addition, hyaluronan that has been localized in the pig oviduct both in the intraluminal fluid and on the surface of the lining epithelium of the preovulatory sperm reservoir, has proven beneficial during in vitro fertilization and embryo culture, thus indicating that it has a role in vivo. This study monitored the immunolocalization, protein determination and gene expression of the major cell surface hyaluronan receptor CD44 in the epithelial lining of the pig oviduct during selected stages of standing oestrus, in relation to spontaneous ovulation. The CD44 immunostaining in the lining epithelium was localized to the surface membrane and the supranuclear domain of mainly the secretory cells, particularly in the sperm reservoir of both treatment (inseminated) and control (non-inseminated) specimens. Up to four hyaluronan-binding protein (HABP) bands (60, 90, 100 and 200 kDa) were detected in the tubal epithelium, and the 200 kDa band was determined as CD44 by immunoblotting. The expression of CD44 mRNA was higher before than after ovulation (P < 0.05), most conspicuously in the uterotubal junction (UTJ). In addition, CD44 expression in the preovulatory UTJ and the ampullary-isthmic junction (AIJ) of control animals was higher than in those that were inseminated (P < 0.05 and P < 0.01 for UTJ and AIJ, respectively). The results demonstrate for the first time that the specific hyaluronan receptor CD44 is expressed by the oviduct epithelial cells during spontaneous oestrus, and is particularly abundant in the sperm reservoir before ovulation. Presence of spermatozoa in this segment seemed to downregulate the receptor. The variation in the expression of CD44 in relation to spontaneous ovulation and the presence of spermatozoa indicate that the hyaluronan CD44-signalling pathway may play a role in oviduct function during sperm storage and fertilization in pigs. (+info)
Effect of intrauterine insemination with spermatozoa or foreign protein on the mechanism of action of oestradiol in the rat oviduct.
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Previously, we showed that oestradiol accelerates oviductal egg transport through a non-genomic action involving oviductal protein phosphorylation in non-mated rats, and through a genomic action in mated rats. Thus, sensory stimulation, seminal fluid or sperm cells may be the source of signals that switch the mechanism of action of oestradiol in the oviduct to a genomic pathway. The present study examined the ability of spermatozoa to switch the mode of action of oestradiol in the absence of the sensory stimulation and seminal fluid provided by mating. Pro-oestrous rats were inseminated in each uterine horn with epididymal spermatozoa and 12 h later were injected subcutaneously with oestradiol and intrabursally with the mRNA synthesis inhibitor alpha-amanitin. The number of eggs in the oviduct, assessed 24 h later, showed that alpha-amanitin blocked the oestradiol-induced egg transport acceleration, indicating that the interaction of spermatozoa with the genital tract shifts the action of oestradiol from non-genomic to genomic. Other rats were inseminated with live or dead spermatozoa and then treated with the protein kinase inhibitor H-89, and oestradiol. Treatment with H-89 did not block the oestradiol-induced acceleration of egg transport in these rats, although dead spermatozoa did not enter the oviduct, indicating that the mere presence of spermatozoa in the uterus abrogated the non-genomic action of oestradiol in the oviduct. Treatment with H-89 also failed to prevent the acceleration of oviductal egg transport induced by oestradiol in rats inseminated with hamster spermatozoa or with BSA, whereas in rats inseminated with their own serum (autologous proteins), H-89 was able to prevent the effect of oestradiol. This finding reveals that the effect of insemination on the mode of action of oestradiol is neither species-nor sperm-specific and it is produced by foreign organic material. It can be concluded that the presence of spermatozoa or foreign protein in the uterus is one of the components of mating that is capable of switching the action of oestradiol in the oviduct from a non-genomic to a genomic mode. (+info)