Journal of andrology最新文献

ABSTRACT: The accurate measurement of semen fertilizing potential is of great importance in determining the acceptability of processed semen for breeding purposes. A good sperm preparation technique results in a sample with high viability and motility and also takes into account other parameters such as the capacitation and apoptotic state which could compromise the ability to fertilize an oocyte. In this study, we investigate the effects of 4 sperm preparation techniques (a dextran/swim-up procedure, discontinuous Percoll density gradient centrifugation, sucrose washing, and filtration) on ram sperm quality parameters. Besides the evaluation of viability and the capacitation state, we also analyzed the apoptotic status of the sperm samples by assessing the phosphatidylserine translocation and caspase-3 and −7 activities. This is the first report, to our knowledge, that evidences the presence of active caspases in ram sperm. The results confirm the better ability of the dextran/swim-up procedure to select nonapoptotic spermatozoa, in addition to viable and noncapacitated sperm, compared with other sperm preparation methods. This should be considered to enhance results of artificial insemination techniques in ovine reproduction.

The total sperm count (number of spermatozoa per ejaculate) rather than sperm concentration (number of spermatozoa per unit volume of semen) is the more important semen variable related to fertility. It reflects testicular volume (Handelsman et al, 1984; Andersen et al, 2000; Behre et al, 2000), and thus is a measure of total testicular sperm output (MacLeod and Wang, 1979), which is directly related to the chances of pregnancy after coitus. The concentration of spermatozoa in the ejaculate, however, depends on the extent of dilution of epididymal spermatozoa by secretions of the prostate and seminal vesicles occurring at ejaculation and is therefore influenced by the secretory capacity of the accessory sex glands. This is an important distinction, for when comparing semen quality from older and younger men, sperm concentrations do not differ, yet semen volume is reduced in the older men, and so the total number of spermatozoa per ejaculate is lower in the older men (Ng et al, 2004; Nieschlag et al, 1982). The total number of spermatozoa per ejaculate is obtained by multiplying the concentration of spermatozoa by the semen volume. The latter is best measured by weighing (Eliasson, 2003), assuming a density of 1.0 g/mL (Auger et al, 1995; Jorgensen et al, 1997, 2001; Brazil et al, 2004), but alternative methods, such as collection into graduated cylinders (Behre et al, 2000), pipetting from the collection vessel (Mortimer 1994; Jorgensen et al, 1997), and pouring from the collection vessel into a graduated tube (Jorgensen et al, 1997), are in current practice.

Two recent studies have found that pipetting semen from the collection vessel leads to an underestimation of about 0.5 mL (range 0.3–0.8 mL; Brazil et al, 2004; Iwamoto et al, 2006) compared with weighing, but no data are available about losses incurred when pouring semen into graduated cylinders. Because the area of contact with the sides of the collection vessel while decanting semen into a graduated cylinder is likely to be far larger than that during pipetting, retention within the vessel could be much larger, leading to a larger underestimation of volume with this method. In this study, new data are obtained on the loss of semen volume during decanting to a cylinder and previously published results on losses because of pipetting, and the density of semen is reanalyzed together with additional data.

This study has shown that a consistent and significant reduction in the volume of semen is obtained when a pipette or a graduated cylinder is used to measure liquefied semen transferred from its collection vessel. These losses cannot be accounted for by evaporation because samples were capped during liquefaction at room temperature and pipetted or decanted immediately after weighing. It could be that with particularly viscous samples, transfer would result in even lower volumes because more would be retained on the side of the decanting vessel and some might adhere to the sides

ABSTRACT: To explore the biological characteristics of the recombinant zona pellucida 3 (ZP3) peptides of rhuZP3a^22∼176 and rhuZP3b^177∼348, we examined whether rhuZP3a^22∼176 or rhuZP3b^177∼348 trigger the acrosome reaction (AR) of human spermatozoa and we investigated the underlying mechanism. The assessment of AR was performed using chlortetracycline staining. The intracellular free calcium concentration ([Ca²⁺]i) in Fura-2/AM-loaded human sperm was monitored with a spectrofluorophotometer. We found that rhuZP3a^22∼176 and rhuZP3b^177∼348 were capable of eliciting AR at different concentrations. With the addition of either peptide, the [Ca²⁺]i level was raised to a peak with or without a plateau. The AR could be inhibited by pertussis toxin (PTX), EGTA, and pimozide (a T-type calcium channel blocker), whereas verapamil was less effective in this regard. The results of the present study suggest that peptides rhuZP3a^22∼176 and rhuZP3b^177∼348 have a role similar to human ZP3, and that the mechanism of the response to the peptides involves influx of calcium, the G protein pathway, and a T-type calcium channel.