Biomedical subjects

E G Arts

Publications and source records attributed to E G Arts.

5 recordsLinked to original sources

PubMed1997

Protein involvement in the fusion between the equatorial segment of acrosome-reacted human spermatozoa and liposomes.

Artificial membranes (liposomes) can interact with the equatorial segment (ES) of human spermatozoa, provided that the acrosome reaction (AR) has occurred [Arts, Kuiken, Jager and Hoekstra (1993) Eur. J. Biochem. 217, 1001-1009]. Using fluorescently labelled liposomes, this interaction can be seen as either punctate fluorescence in the ES (lip-ESp), reflecting only bound liposomes, or as diffuse fluorescence in this region (lip-ESd), indicating that the liposomes have fused with the ES membrane. Only equatorial segments that still contain constituents of the acrosomal matrix have the capacity to bind liposomes and eventually to fuse with them. Since the exposure of such intact equatorial segments is the exclusive result of induction of the AR under physiological conditions, these results imply that liposomes can be used for the rapid detection of acrosome-reacted spermatozoa. The lip-ESp and lip-ESd patterns were shown to be reflections of two distinct properties of the ES. Proteolytic treatment after AR completely inhibited the formation of a lip-ESd pattern, whereas formation of the lip-ESp pattern was only marginally inhibited by the proteolytic treatment. The same results were obtained using anti-sperm antibodies, which did not react with acrosome-intact spermatozoa. Proteolytic treatment of spermatozoa before AR induction had no effect on the fusion capacity of the ES after subsequent AR, which implies that the putative fusion protein is not accessible before AR. Thus fusion of liposomes with the ES of human spermatozoa is mediated by a sperm protein(s), whereas the lip-ESp pattern is not likely to represent the liposome-binding stage that precedes the fusion step.

Acrosome↗

PubMed1994

Evidence for the existence of lipid-diffusion barriers in the equatorial segment of human spermatozoa.

Liposomes consisting of negatively charged phospholipids interact almost exclusively with the equatorial segment (ES) of human spermatozoa provided the cells have undergone the acrosome reaction (AR) [Arts, Kuiken, Jager and Hoekstra (1993) Eur. J. Biochem. 217, 1001-1009]. Using fluorescently tagged liposomes, this interaction can be observed by fluorescence microscopy, showing either a diffuse fluorescence in the ES region (pattern ESd, presumably reflecting membrane-incorporated lipids as a result of fusion) or a punctate fluorescence (pattern ESp, representing adhering liposomes). These distribution patterns remain unchanged during prolonged incubation, up to 40 min. Not only do these observations suggest the existence of fairly specific liposomal binding sites, associated with the ES region, but also that a barrier to lipid lateral diffusion seems to exist in the ES membrane. Using liposomes that contain fluorescent lipid analogues in either both leaflets or in the inner leaflet only, we demonstrate that this putative barrier entails both membrane leaflets. Treatment with EDTA caused fluorescence to spread from the ES towards other membrane domains. Since only spermatozoa displaying pattern ESd were affected by the chelator, the randomization was not caused by EDTA-induced fusion activity. Therefore, this observation provides further evidence that in spermatozoa displaying pattern ESd the fluorescent lipid analogues were incorporated in the ES membrane as a result of fusion. Furthermore, these experiments support the view of the existence of a transmembranous block to lipid lateral diffusion in the ES, the stability of which may be governed by bivalent cations.

Acrosome↗

PubMed1994

A new method to detect acrosome-reacted spermatozoa using biotinylated soybean trypsin inhibitor.

OBJECTIVE: To develop a method to detect acrosome-reacted spermatozoa on human zonae pellucidae using only commercially available reagents and without need for sperm fixation. DESIGN: Sperm head labeling with biotinylated soybean trypsin inhibitor (SBTI-biotin) was compared with results of a known method using fluorescein isothiocyanate (FITC)-conjugated Pisum sativum agglutinin. The SBTI-biotin method was applied to sperm bound to human zonae pellucidae. SETTING AND SUBJECTS: Healthy sperm donors with normal semen characteristics were recruited by the Laboratory for Reproductive Medicine in a university medical center. MAIN OUTCOME MEASURES: Soybean trypsin inhibitor-biotin binding patterns on nonfixed spermatozoa were visualized with avidin-Texas Red. The development in time of various patterns upon induction of acrosome reaction (AR) in suspension with Ca(2+)-ionophore A23187 was noted and compared with P. sativum agglutinin FITC labeling patterns. Soybean trypsin inhibitor-biotin labeling patterns of spermatozoa bound to human zonae pellucidae were determined. RESULTS: Soybean trypsin inhibitor-biotin bound specifically, via the SBTI-moiety, to an acrosomal factor as soon as AR started. Sperm-head labeling patterns could be assigned to defined stages of the AR process. The results were highly correlated to those obtained with P. sativum agglutinin FITC. The end point of the AR in suspension and on zonae pellucidae was SBTI-biotin binding confined to the equatorial segment. CONCLUSION: The SBTI-biotin method can be used to detect nonfixed acrosome-reacted spermatozoa both in suspension and on zonae pellucidae.

Acrosome↗

PubMed1993

Fusion of artificial membranes with mammalian spermatozoa. Specific involvement of the equatorial segment after acrosome reaction.

The fusogenic properties of bovine and human spermatozoa membranes were investigated, using phospholipid bilayers (liposomes) as target membranes. Fusion was monitored by following lipid mixing, as revealed by an assay based on resonance-energy transfer. In addition, fusion was visualized by fluorescence microscopy, using fluorescent lipid vesicles. Cryopreserved bovine sperm fused with liposomes before induction of the acrosome reaction, fluorescence being located in essentially all spermatozoa membrane domains. Fresh bovine and human spermatozoa fused with liposomes only after the induction of the acrosome reaction, as triggered by calcium ionophore A23187 or zonae pellucidae (proteins), while the fluorescence distribution was mainly restricted to the equatorial segment (ES). However, with spermatozoa that had undergone a freeze/thawing cycle, domains other than ES also became labeled. Hence, the redistribution of the lipid probes over the entire membrane occurring during lipid mixing with cryopreserved bovine sperm is probably related to membrane perturbations caused by long-term cryopreservation. Fusion with liposomes was governed by spermatozoa factors and required the presence of acidic phospholipids like cardiolipin and phosphatidylserine in the liposomal bilayer. Incorporation of the zwitterionic lipid phosphatidylcholine in the vesicles inhibited the fusion reaction. Fusion was pH dependent. The results indicate that the ES is the primary domain of spermatozoa membranes that harbours the fusogenic capacity of sperm. Liposomes appear a valuable tool in further characterizing the properties of this domain, which has been claimed [Yanagimachi, R. (1988) in The physiology of reproduction (Knobil, E. & Neill, J., eds) pp. 135-185, Raven Press, New York] to represent the putative, initial fusion site for the oocyte.

Acrosome↗

PubMedDate not supplied

Variation in semen parameters derived from computer-aided semen analysis, within donors and between donors.

The development of computer-aided semen analysis (CASA) has made it possible to study sperm motility characteristics objectively and longitudinally. In this 2-year study of 8 sperm donors, we used CASA to measure 7 semen parameters (concentration, percentage of motile spermatozoa, curvilinear velocity, average path velocity, straight-line velocity, amplitude of lateral head displacement, and beat/cross frequency). The frequency distributions of the 7 parameters in the semen samples of each donor were investigated. All parameters but one were normally distributed; concentration was distributed log-normally. Variation within individual donors and between donors was studied. Analysis of variance demonstrated that variation between donors was not explained by the longitudinal variation within individual donors. Variations in motility characteristics between donors were substantial, which may make motility characteristics of limited value as a tool for establishing fertility. Strong correlations were found between the 7 parameters, partly because by definition, motility characteristics are interdependent. Fisher's discriminant analysis demonstrated that each donor appeared to have his own set of semen characteristics and, more specifically, his own motility signature. From this data set it can be predicted that in order to find population means among sperm, it may be more efficient to measure more subjects than to increase the number of samples per subject.

Humans↗