Semen characteristics and fertility tests required for storage of spermatozoa.

This review addresses critical issues in the selection of semen donors who are very fertile. Traditional semen parameters have been employed and are still used to assess pre- and post-freeze samples in order to discriminate between donors of high and low fecundity. The most predictive factor is the number of motile spermatozoa per straw and the number of motile spermatozoa inseminated. Nevertheless, no absolute standards for fertile samples can be derived from the basic semen examination. The employment of sperm function testing such as the hamster penetration test or computerized motility analysis has been shown to enhance moderately the prediction of fertility of semen samples however, further studies are necessary to determine if these improvements are clinically useful. The need to determine with a high degree of confidence the fecundity of donor semen is enhanced by limitations in the number of pregnancies allowed per donor. The recent publication of league tables in the UK has put extra pressure in clinics to use highly fertile donors. Spermatozoa are also cryostored for patients prior to cancer treatment. With the development of intracytoplasmic sperm injection every sample produced by cancer patients can be stored irrespective of the quality. However, several factors need to be elucidated to maximize the fertility of those patients. The establishment of regional centres in Europe will be a good starting point to deal with many of the issues raised in this review.

The polypeptide backbone of recombinant human zona pellucida glycoprotein-3 initiates acrosomal exocytosis in human spermatozoa in vitro.

Human gamete interaction is of fundamental biological importance, yet the molecular interactions between spermatozoa and the zona pellucida are poorly understood. Surprisingly, the role of the polypeptide backbone of zona pellucida glycoprotein 3 (ZP3), the putative ligand for spermatozoa activation, has been largely overlooked. Purified recombinant human ZP3 was expressed in Escherichia coli as a C-terminal fusion to the dimeric glutathione S-transferase (GST) from Schistosoma japonicum and was shown to induce acrosomal exocytosis in live, capacitated human spermatozoa. The level of exocytosis is comparable with that obtained using purified, glycosylated, recombinant human ZP3 [van Duin, M., Polman, J.E.M., DeBreet, I.T.M., Van Ginneken, K., Bunschoten, H., Grootenhuis, A., Brindle, J. and Aitken, R.J. (1994). Biol Reprod. 51, 607-617]. These data imply that the polypeptide chain of human ZP3 contributes to recognition of spermatozoa during acrosomal exocytosis in vitro.

A double-blind randomized placebo cross-over controlled trial using the antioxidant vitamin E to treat reactive oxygen species associated male infertility.

OBJECTIVE: To determine the effectiveness of the in vivo administration of vitamin E as treatment for reactive oxygen species-associated male infertility. SETTING: University-based center for reproductive medicine. DESIGN: Double-blind randomized placebo cross-over controlled trial. PATIENTS, PARTICIPANTS: Thirty healthy men with high levels of reactive oxygen species generation in semen and a normal female partner. INTERVENTIONS: Patients were allocated to two groups according to the blinded randomization. Each patient received either 600 mg/d of vitamin E (Ephynal, 300 mg tablets; F. Hoffman-La Roche Ltd., Basle, Switzerland) (order A) or identical placebo tablets (order B) for 3 months. Then after a 1-month wash-out period the patients were crossed-over to the other treatment. MAIN OUTCOME MEASURES: Improvement in the in vitro function of the spermatozoa measured by conventional semen analysis, computerized motility assessment, determination of reactive oxygen species generation, binding to the zona pellucida of the unfertilized human oocyte in a competitive zona binding assay, development of hyperactivated motility (both spontaneous and in the presence of 20% of the natural agonist, human follicular fluid) and pregnancy. RESULTS: Rise in the blood serum vitamin E levels after treatment accompanied by improvement in one of the sperm function tests: the zona binding assay. The zona binding ratio for order A improved from 0.2 (range 0 to 0.5) before treatment to 0.5 (range 0.1 to 1.0) after treatment, the corresponding values for order B were 0.2 (range 0 to 1.0) before treatment and 0.3 (range 0.1 to 0.7) after treatment. CONCLUSION: Oral administration of vitamin E significantly improves the in vitro function of human spermatozoa as assessed by the zona binding test.

The contribution of a hidden male factor to unexplained infertility.

OBJECTIVE: To assess sperm function in patients with unexplained infertility in comparison with normal fertile men. DESIGN: Prospective study. SETTING: Infertility outpatient clinic and donor insemination program, University Department of Obstetrics and Gynecology at the Jessop Hospital for Women, Sheffield, United Kingdom. PATIENTS: Nineteen patients with unexplained infertility and nineteen normal fertile men from a donor insemination program. MAIN OUTCOME MEASURES: Monitoring of reactive oxygen species generation; computerized assessment of the hyperactivated motility pattern of the spermatozoa under capacitating conditions, with and without the addition of follicular fluid (FF); evaluation of sperm-zona pellucida (ZP) binding. RESULTS: Reactive oxygen species generation was not significantly different between the study and control group. Follicular fluid-induced hyperactivation (HA) was significantly lower in patients with unexplained infertility than in the donors, but no difference was found comparing spontaneous HA (without FF) between the two groups. Follicular fluid-induced HA was completely absent in 39% of infertile men and only one fertile man. A low sperm-ZP binding ratio (< or = 0.3) was observed in 28% (5/18) of patients with unexplained infertility. In all, nine infertile men (47%) had 0% FF-induced HA and/or low sperm-ZP binding ratio. CONCLUSIONS: Assessing FF-induced HA and sperm-ZP binding may have clinical significance in distinguishing a subgroup of men with unexplained infertility with sperm function abnormalities in which the fertilizing capacity of the spermatozoa might be reduced. These patients cannot be identified by the conventional semen analysis. Both tests may also be useful in planning the appropriate treatment for couples with unexplained infertility.

Origin of reactive oxygen species in human semen: spermatozoa or leucocytes?

Peroxidative damage induced by reactive oxygen species (ROS) has been proposed as one of the major causes of defective sperm function. In previous studies of the production of ROS in semen, the contribution of contaminating leucocytes was not assessed. We determined the levels of ROS in 60 semen samples from men attending our infertility clinic and demonstrated by performing extraction experiments with antibody-coated magnetic beads that, within this unselected population of patients, leucocytes were the major source of ROS in the low-density Percoll fraction. Of the sperm motion parameters examined using computerized semen analysis, beat-cross frequency was the only one significantly affected by the ROS in semen.

Seminal white blood cells in men with urethral tract infection. A monoclonal antibody study.

Monoclonal antibodies were used to detect seminal leucocytes in 19 men with urethritis. A wide range in the number and type of leucocytes between individuals was documented (median 2.06 x 10(6)/ml, range 0.6-29.89 x 10(6)/ml); 22% of the men had less than 1 x 10(6) leucocytes/ml. The results suggest that the threshold of greater than or equal to 1 x 10(6)/ml proposed by the World Health Organisation to indicate genital tract infection is unsuitable for men with urethritis.

Detection of white blood cell populations in the ejaculates of fertile men.

Using a now established, reliable technique for detecting white blood cells in human semen, we have examined the ejaculates of a fertile group of men. We have also highlighted the need for strict recruitment criteria to be established to enable accurate comparisons to be drawn between different patient groups and results from different laboratories.

Seminal peroxidase positive cells are not an adequate indicator of asymptomatic urethral genital infection.

The relationship between asymptomatic urethral infection and seminal white blood cells, as detected using the peroxidase enzyme system, was examined. Eighty-four semen donors were tested. Twenty-four (29%) were diagnosed as having an active urethral genital infection. There was no statistical relationship between the total number of concentrations of peroxidase-positive cells and a urethral genital infection. Further studies should concentrate on the subtypes of seminal leucocytes and their surface receptors using monoclonal antibodies.