Ubiquitin-based sperm assay for the diagnosis of male factor infertility

Sperm morphology is correlated with fertility in men, yet the existing, subjective sperm morphology assays provide only a limited insight into patients' infertility. Here, we provide the experimental background for a new, objective and automated semen assay, based on the cross-reactivity of defective human spermatozoa with antibodies against a proteolytic marker peptide, ubiquitin. Using immunofluorescence and flow cytometry, we screened the spermatozoa from 17 infertility patients and two fertile donors for their cross-reactivity with anti-ubiquitin antibodies. Thirteen out of 17 patients, but neither of two fertile donors, displayed an increased binding of anti-ubiquitin antibodies to sperm surface, that reflected the occurrence of abnormalities in these samples and was corroborated by available clinical data. Highest correlation coefficient (r = -0.432) was obtained with the cleavage rate after IVF. The contribution of male factor was revealed in several couples previously diagnosed with idiopathic infertility. Ubiquitin-cross-reactive sperm-surface proteins thus seem to be a universal marker of semen abnormalities, including sperm head and tail defects and semen contaminants such as spermatids, leukocytes and cellular debris. We propose that the sperm-ubiquitin tag immunoassay (SUTI) may be a valuable diagnostic tool in treatment of male factor and idiopathic infertility.     

Studies on Sperm Survival and Motility in the Presence of Cytotoxic Sperm Antibodies

ABSTRACT: The effect of cytotoxic sperm antibodies and native complement in the serum and secretions from 40 fertile and 93 infertile couples on in vitro sperm survival and motion characteristics was studied. Sperm survival in vitro was unaffected by sera from fertile and infertile subjects without cytotoxic sperm antibodies and from infertile men with antibodies to control but not to autologous sperm. Sperm survival was reduced (P < .001) by sera from infertile men with antibodies to autologous sperm or to antologous and control sperm and from women with cytotoxic antibodies to sperm from both. Sera from fertile couples without sperm antibodies enhanced sperm swimming speed and motility index (P < .0001). Sera from infertile women with or without cytotoxic sperm antibodies did not affect sperm motility. Sperm survival and motility were reduced by seminal plasma from infertile men with cytotoxic antibodies to autologous and/or control sperm. Seminal plasma from fertile men enhanced sperm survival. Cervical mucus from infertile women with antibodies to autoimmune husbands' sperm or to husbands' and control sperm inhibited sperm motion, whereas cervical mucus from infertile women without sperm antibodies and women with antibodies to control sperm failed to have any effect. It is concluded that cytotoxic sperm antibodies developed through exposure to sperm antigens in autoimmune infertile men decrease in vitro sperm survival and/or motility.     

Prevalence of chromosomal abnormalities in phenotypically normal and fertile adult males: large-scale survey of over 10,000 sperm donor karyotypes

BACKGROUND: Sperm donors represent an appropriate population for evaluating the frequency of chromosomal abnormalities in phenotypically normal and fertile adult males. METHODS: A large multicentric retrospective study was made within the French CECOS (Centre d'Etude et de Conservation des ufs et du Sperme) for collecting cytogenetic, biological and familial data in sperm donors over a 25-year period. RESULTS: As a whole, 10,202 karyotypes have been recorded. Thirty-eight karyotype aberrations (0.37%) have been diagnosed including 21 balanced chromosomal rearrangements (0.2%). These results are in agreement with those obtained in most large-scale studies performed in unselected newborns. Semen parameters were known for all men carrying an abnormal karyotype and showed normal sperm counts, suggesting that these types of chromosomal aberrations have no or poor consequences on spermatogenesis. Available familial data did not reveal any particular history of malformations, mental retardation or fetal losses. CONCLUSION: This study is the first large-scale cytogenetic study made in normal and fertile males and shows that the frequency of chromosomal aberrations is not influenced by a previous normal fertility or by an uneventful familial history when compared to that found at birth.     

A trial to restore defective human sperm centrosomal function

BACKGROUND: In human fertilization, sperm centrosome function is essential for male and female pronuclear movement and fusion. In this study, we investigated the possibility of restoring human sperm centrosomal function in sperm exhibiting abnormalities in microtubule organization. METHODS: Semen was obtained from both a fertile donor and a patient with dysplasia of the fibrous sheath (DFS). Following heterologous ICSI using human sperm, we examined microtubules and chromatin configuration in bovine oocytes. Sperm were treated with dithiothreitol (DTT) prior to ICSI, while the oocytes were treated with the cytoskeletal stabilizer paclitaxel after ICSI. RESULTS: The combination of DTT and paclitaxel treatment induced microtubule organization in dead sperm from the fertile donor following heterologous ICSI. This treatment, however, was not effective for DFS sperm. In addition, expression of centrin, a protein functioning within the sperm centrosome, was reduced in DFS sperm from that of the normal levels observed in fertile donor sperm. CONCLUSION: These results indicate that sperm centrosomal function could be induced by the treatment of sperm with DTT before ICSI and of oocytes with paclitaxel after ICSI. DFS sperm are likely to exhibit such severe dysfunction of sperm centrosome that cannot be compensated for by this treatment; therefore, this method may be a practical way to discern the degree of sperm centrosomal dysfunction.     

Motion Characteristics of Spermatozoa From Men With Cytotoxic Sperm Antibodies

ABSTRACT: Semen samples from 55 fertile nonautoimmune and 44 infertile sperm autoimmune men were evaluated by computerized sperm cell motion analysis. Sperm counts (mean ± SEM, 59.6 ± 10.3 ± 10⁶ per ml), motility (39.0 + 4.6%), mean swimming speed (μm/sec, 26.5 ± 0.9), mean linearity (straight line distance of the cell track divided by the actual track length and multiplied by 10, 6.5 + 0.2), and motility index (% motility ± mean speed, 10.7 ± 1.4) in 23 men with significant titers of cytotoxic sperm antibodies in their serum and seminal plasma were less (p<0.0001) than those in the fertile controls. However, these parameters were comparable in 18 men with sperm antibodies in their seminal plasma but not in their serum, and the control group. Infertile men with serum cytotoxic sperm antibodies had more sperm cells swimming at 11–30 μm/sec, and fewer moving at 31 μm or higher; this was in contrast to results obtained from fertile men (p < 0.05). The percentages of sperm cells moving at 21–30 μm/sec were increased, while those moving at 51–60 μm/sec were decreased in men with seminal plasma sperm antibodies, versus controls. Spermatozoa with low linearities (≤6) were higher (p<0.05) in men with serum and seminal plasma cytotoxic sperm antibodies than in the fertile group. The numbers of sperm cells moving at a linearity of ten, on the other hand, were lower in the infertile men with sperm antibodies in the serum and seminal plasma (21.8 ± 2.7 versus 36.4 ± 1.3; p< 0.00001) or seminal plasma only (28.0 ± 1.5; p< 0.0001). We suggest that the spermatozoa of men with cytotoxic sperm antibodies in their serum and seminal plasma have poorer sperm motion characteristics than those of fertile men.     

Ultrastructural nuclear defects and increased chromosome aneuploidies in spermatozoa with elongated heads

BACKGROUND: Cellular and molecular mechanisms leading to elongated sperm heads are not known. We have analysed the nuclear status of spermatozoa with elongated heads. METHODS: Fourteen men with at least 30% of spermatozoa with an elongated nucleus were studied and compared with five fertile men as controls. Sperm morphology was analysed by a quantitative ultrastructural analysis. Sperm chromosomal content was assessed by three-colour fluorescence in-situ hybridization (chromosomes X, Y, 18). Y chromosome microdeletion and karyotype were analysed. RESULTS: Elongated sperm head rates of the patients were 46.9% (30-75 versus 0-2% in the control group) by light microscopy and 34.4% by electron microscopy. In all patients, the chromatin was poorly condensed in elongated sperm heads (50% of elongated nuclei). No anomalies of sperm biochemical markers were found. All the men showed normal karyotype (46,XY) and absence of Y chromosome microdeletion. Aneuploidy rates of gonosomes and chromosome 18 were significantly increased in patients (1.64- and 3.6-fold, P = 0.006 and 0.026, respectively). CONCLUSIONS: This study demonstrates that impaired chromatin compaction and slightly increased chromosome aneuploidies are found in spermatozoa with an elongated head, suggesting possible mechanisms such as meiotic non-disjunctions or spermiogenesis anomalies.     

Low proportions of sperm can bind to the zona pellucida of human oocytes

BACKGROUND: Sperm binding to the zona pellucida (ZP) is required for human fertilization. Under experimental conditions not limited by ZP binding sites, the cumulative numbers of sperm binding tightly to the ZP will asymptote with time to the total number of sperm in the insemination medium capable of binding. METHODS: Numbers of ZP-bound sperm were counted after groups of 10 oocytes were incubated with 2x10(4) motile sperm in 20 micro l droplets. The time-course of sperm binding was measured in three consecutive 2 h incubation periods using fresh oocytes for each period (n = 12). Using the kinetic theory of gases to model sperm-oocyte collision rates, the time-course results were extrapolated to give the total proportion of motile sperm capable of binding to the ZP. ZP binding of sperm after 4 h incubation was studied in 20 fertile and 20 normozoospermic subfertile men. RESULTS: The percentage of motile sperm capable of binding was for fertile men: mean 14% (range 8-25) and for the subfertile: 4.3% (range 0.1-13, P < 0.001). Sperm morphology correlated with the proportion of ZP-bound sperm. CONCLUSIONS: More than 75% of motile sperm from fertile men have no ability to bind to the ZP. This finding has important implications for improvement of semen analysis.     

Distinct Expression of Cytokines and Mitogenic Inhibitory Factors in Semen of Fertile and Infertile Men

PROBLEM: To assess the effect of seminal plasma (SP) of fertile and infertile men on leukocyte mitogenic response, and the capability of sperm cells to produce IL-1. METHODS: This study included four groups: fertile men (donors, normal), infertile men with azoospermia (azoo), oligo-terato-asthenozoospermia (OTA), and OTA with genital infection (OTA-inf). Mouse spleen cell proliferation in response to lipopolysaccharide (LPS) or Concanavalin-A (Con-A) was examined in the presence of SP from the above four groups. Supernatants (sup) and lysates (lys) of sperm cells from fertile and oligoteratoasthenospermic (OTA) men were evaluated for IL-1 bioactivity by specific bioassay. RESULTS: Seminal plasma (SP) of the four groups were shown to inhibit the mitogenic response of mouse spleen cells to LPS and Con-A. SP of fertile men was significantly more inhibitory than SP from infertile men. Sperm cells from fertile and OTA infertile men constitutively produced IL-1. Sperm cells of both groups produced similar levels of IL-1 as examined in the supernatants and lysates. CONCLUSIONS: Seminal plasma of fertile men had more inhibitory mitogenic activity than that of OTA. Sperm cells constitutively produce IL-1. It is possible that the factors involved in this inhibition are not only anti-proliferative immune factors. Cytokines and inhibitory factors of mitogenesis in the seminal plasma may be involved in the physiology and pathophysiology of sperm functions and thus affect male fertility.     

Effects of vasectomy on spermatogenesis and fertility outcome after testicular sperm extraction combined with ICSI

BACKGROUND: Each year 40,000 men have a vasectomy in the UK whilst another 2400 request a reversal to begin a second family. Sperm can now be obtained by testicular biopsy and subsequently used in assisted conception with ICSI. The study aims were to compare sperm yields of men post-vasectomy or with obstructive azoospermia (OA) of unknown aetiology with yields of fertile men and to assess any alteration in the clinical pregnancy rates after ICSI. METHODS: Testicular tissue was obtained by Trucut needle from men who had undergone a vasectomy >5 years previously or had OA from other causes and from fertile men during vasectomy. Seminiferous tubules were milked to measure sperm yields. Numbers of Sertoli cells and spermatids and thickness of the seminiferous tubule walls were assessed using quantitative computerized analysis. RESULTS and CONCLUSIONS: Sperm yields/g testis were significantly decreased in men post-vasectomy and in men with OA, relative to fertile men. Significant reductions were also observed in early (40%) and mature (29%) spermatid numbers and an increase of 31% was seen in the seminiferous tubule wall (basal membrane and collagen thickness) of vasectomized men compared with fertile men. Clinical pregnancy rates in couples who had had a vasectomy were also significantly reduced.     

The relationship between fertility potential measurements on cryobanked semen and fecundity of sperm donors

Sperm penetration assay (SPA) scores obtained from cryobanked semen were correlated with therapeutic insemination (TI) fecundity in a group of established sperm donors, thereby evaluating the efficacy of the SPA in screening donors for sperm banking. While the SPA has been used to separate fertile from infertile males, we altered assay conditions to use frozen semen and to distinguish performance among fertile donors. Three frozen ejaculates from 11 pregnancy-proven donors were analysed. Of 905 TI cycles, 275 recipients achieved 95 pregnancies. There were no significant relationships between fecundity and donor semen, washed sperm parameters, sperm recoveries or recipient age. A significant relationship was revealed between mean SPA scores (range 8.7-66.6 penetrations/ovum) and donor fecundity (range 0.04-0.16, P < 0.03). Sperm concentration was varied in an effort to establish the most sensitive test condition. Using 0.25x10(6) motile spermatozoa/ml, a highly significant relationship was observed (P < 0.002). The four donors with the lowest SPA scores achieved the four lowest fecundities. It is concluded that a modified SPA can be used on frozen donor semen to estimate donor fertility potential. If applied routinely in donor semen banking, poor quality applicants could be excluded, thereby increasing pregnancy rates while decreasing donor screening costs.     

Somatic cell apoptosis markers and pathways in human ejaculated sperm: potential utility as indicators of sperm quality

In this study we extended earlier work to determine whether sperm respond to somatic cell apoptotic stimuli and whether apoptotic phenotypes are significant indicators of human sperm quality. We evaluated ejaculated sperm from fertile donors and subfertile patients following purification of fractions of high and low motility. In unstimulated conditions, caspase enzymatic activity was higher in motile fractions from subfertile patients than in donors, and was higher in low motility fractions from both groups. Staurosporine, but not a Fas ligand or H2O2, significantly increased caspase activity, but only in high motility fractions. Procaspase-3, -7 and -9 and low levels of active caspase-3, -7 and -9 were identified by immunoblot analysis. Apoptosis-inducing factor (AIF) was present in all samples but poly ADP-ribose polymerase-1 (PARP-1) was not detected. Phosphatidylserine translocation was significantly increased only with H2O2 treatment. In ejaculates of both subfertile and fertile men, we demonstrated the presence and activation of several proteins that are key constituents of apoptosis-related pathways in somatic cells, which may serve as markers for sperm quality.     

Detection of fractalkine in human seminal plasma and its role in infertile patients

BACKGROUND: Fractalkine is a relatively newly discovered CX(3)C chemokine, which is a chemoattractant for T cells, monocytes and natural killer cells. Several reports have demonstrated the association between chemokine levels in seminal plasma and semen quality. The fractalkine levels in ejaculates from normal donors and infertile male patients with or without asthenozoospermia, were examined and correlated with sperm motility and morphology. METHODS AND RESULTS: Western blot analysis showed fractalkine protein to be present in the seminal plasma. Fractalkine titres in the seminal plasma of infertile men with asthenozoospermia (0.64 +/- 0.04 microg/ml; n = 58) were lower than those in patients without asthenozoospermia (0.94 +/- 0.10 microg/ml; n = 22, P < 0.01) and fertile donors (1.04 +/- 0.07 microg/ml; n = 10, P < 0.001). There was no significant difference between fractalkine levels in patients with and without leukospermia. No significant correlation was found between fractalkine and interleukin-8 levels in seminal plasma. Sperm motility was positively correlated (R(2) = 0.14, P < 0.001) with fractalkine concentration. The existence of CX(3)CR-positive leukocytes in semen was confirmed using specific primers for CX(3)CR. CONCLUSIONS: These results suggest that fractalkine is a chemokine associated with sperm motility and the migration of CX(3)CR-positive leukocytes into semen.     

Clinical adaptation of the sperm ubuquitin tag immunoassay (SUTI): relationship of sperm ubiquitylation with sperm quality in gradient-purified semen samples from 93 men from a general infertility clinic population

BACKGROUND: The proteolytic chaperone peptide ubiquitin accumulates in defective human spermatozoa. Immunodetection of ubiquitin in human sperm samples correlates with semen quality and male fertility. METHODS: Semen samples from 93 men from couples seeking infertility treatment were separated on a PureSperm density gradient and screened by immunofluorescence microscopy with anti-ubiquitin antibodies. The percentage of spermatozoa with head ubiquitylation was recorded and compared with clinical semen evaluation and embryo development data after IVF or ICSI. Subjects were divided into the following four groups based on the initial clinical diagnosis of the couples; group 1, male factor; group 2, idiopathic infertility; group 3, female infertility with neither partner having children previously; and group 4, female infertility with male partners having children from previous relationships. RESULTS: The percentage of sperm with ubiquitylated heads remaining after PureSperm separation in the respective groups was 4.0% (male factor), 2.5% (idiopathic infertility), 0.7% (female infertility and presumed fertile male) and 0.9% (female infertility with established fertile male). Negative correlations between sperm ubiquitin and several parameters reflective of embryo development after assisted fertilization were found within the male factor group. CONCLUSIONS: Use of this simplified ubiquitin-based sperm quality assay is feasible in a clinical environment. Since the gradient separation does not completely deplete the defective spermatozoa, the modified light microscopic sperm ubiquitin tag immunoassay could add a new level of stringency to the selection of human spermatozoa for ICSI.     

Time to pregnancy in relation to semen quality assessed by CASA before and after sperm separation.

BACKGROUND: In order to provide a reference for infertile men, we defined normal values of semen quality in a population of fertile men, using computer-assisted semen analysis (CASA) before and after sperm separation. Additionally, we investigated the relationship between semen quality and time to pregnancy (TTP). METHODS AND RESULTS: Semen samples were obtained from 315 proven fertile men. The median sperm concentration in fresh samples was 107 x 10(6)/ml (5-95 percentiles: 16-322 x 10(6)/ml), the median percentage of motile sperm cells was 65% (14-87%) and the median percentage of progressively motile cells was 37% (5-64%). After density gradient sperm separation, the median total sperm count was 46 x 10(6) (4-350 x 10(6)), the median percentage of motile sperm cells was 77% (16-95%) and the median percentage of progressively motile cells was 63% (11-84%). No significant associations were found between TTP and sperm counts or sperm motility, either before or after sperm separation. This may be due in part to the fact that the study comprised couples with proven fertility. CONCLUSION: We have defined semen parameters, including the results of density gradient separation, in a population of normal fertile men which may be of interest in the evaluation of semen samples from infertile men.     

Abnormalities for chromosomes 13 and 21 detected in spermatozoa from infertile men

Sperm samples from infertile men with oligozoospermia or teratozoospermiawere studied by multicolour fluorescence in-situ hybridization(FISH) using DNA probes for chromosomes 13 and 21. A total of90 809 sperm nuclei from nine infertile men and 182 799 spermnuclei from 18 control donors were analysed. There was a highlysignificant increase in the frequency of spermatozoa disomicfor chromosome 13 in infertile patients (0.28%) compared tocontrol donors (0.13%) (two-tailed Z statistic P <0.0001and for chromosome 21 (0.48% in infertile men versus 0.37% incontrols, P <0.0001). Also there was a significantly increasedfrequency of diploid spermatozoa in infertile men (0.85%) comparedto control donors (0.66%) (P <0.0001). Our previous studieson these same infertile patients demonstrated increased frequenciesof sperm disomy for chromosomes 1 and XY. This suggests thatinfertile men, who are prime candidates for intracytoplasmicsperm injection, may be at a very small increased risk of aneuploidoffspring.     

The high incidence of meiotic errors increases with decreased sperm count in severe male factor infertilities

BACKGROUND:The high frequency of aneuploidy sperm raises concerns that there may be an increased incidence of aneuploid offspring in ICSI programmes. In order to assess the role that chromosome complement plays in normal and abnormal fertility, detailed molecular cytogenetic studies must be done on sperm samples from men with normal and abnormal fertility. METHODS: To understand more clearly the cytogenetic make-up of sperm from oligoasthenoteratozoospermic (OAT) patients, multi-colour fluorescence in situ hybridization was used to determine numerical chromosome abnormalities. RESULTS: Increased aneuploidy frequencies for chromosomes 13, 18, 21, X and Y were detected in sperm from OAT patients. The frequencies of diploidy also increased. There were no differences in non-disjunction at meiosis I compared to meiosis II. Sperm count inversely correlated with the frequencies of diploidy, aneuploidies for chromosomes 13 and 21 in OAT patients. Twenty-two cycles of ICSI and 18 embryo transfers were performed in 20 couples. Only three cases achieved successful pregnancies. CONCLUSIONS: A higher incidence of meiotic errors and lower sperm counts was found in sperm from OAT patients.     

Increased levels of sperm ubiquitin correlate with semen quality in men from an andrology laboratory clinic population

BACKGROUND: Ubiquitin, a house-keeping protein that marks other proteins for proteasomal degradation, tags defective sperm during epididymal passage. To establish ubiquitin as a biomarker of human infertility, the present study examines the relationships between sperm ubiquitin content and clinical semen parameters among men from an infertility clinic population with varied aetiologies. METHODS: Anti-ubiquitin immunoreactivity was measured by flow cytometric sperm-ubiquitin tag immunoassay (SUTI) in sperm samples of 28 infertility patients and 15 fertile donors. Semen analyses were performed by computer-assisted semen analysis and World Health Organization morphology. RESULTS: Median values of ubiquitin-induced fluorescence had a strong negative correlation with sperm count (r = -0.63, P = 0.0003) and a positive correlation with % abnormal morphology (r = 0.55, P = 0.01). Infertility patients (n = 28) had significantly higher levels of sperm ubiquitin. Out of 28 patients, six reported possible occupational exposures to solvents, three were current smokers and six were ex-smokers. Within the patient group, men with known male factor infertility, those with self-reported occupational exposure to solvents and current smokers had the highest sperm ubiquitin levels. When men with jobs involving potential occupational exposure to solvents were combined with current smokers, the highest correlations were found between sperm ubiquitin and motility (r = -0.74), count (r = -0.82) and % sperm abnormalities (r = 0.73). CONCLUSIONS: Increased sperm ubiquitin was inversely associated with sperm count, motility and % normal morphology, supporting the use of ubiquitin as a biomarker of human semen quality. SUTI assay confirmed poor semen quality in all men with poor clinical semen parameters, but also was high in some patients with seemingly good clinical semen parameters. Occupational exposure to solvents and smoking may have contributed to high levels of sperm ubiquitin in some of these patients.     

Assessment of DNA integrity and morphology of ejaculated spermatozoa from fertile and infertile men before and after cryopreservation

Cryopreservation of human spermatozoa is extensively used in artificial insemination and IVF programmes. Despite various advances in cryopreservation methodology, the recovery rate of functional post-thaw spermatozoa remains mediocre, with sperm motility being significantly decreased after freezing. This aim of this study was to investigate the effects of cryopreservation on both DNA integrity and morphology of spermatozoa from fertile and infertile men. Semen samples were obtained from 17 fertile and 40 infertile men. All samples were prepared by discontinuous Percoll density centrifugation (95.0:47.5). Samples were divided into aliquots to allow direct comparison of fresh and frozen spermatozoa from the same ejaculate. Aliquots for cryopreservation were mixed with a commercial cryoprotectant and frozen by static phase vapour cooling before plunging into liquid nitrogen. Thawing was carried out slowly at room temperature. Sperm DNA integrity was determined using a modified alkaline single cell gel electrophoresis (comet) assay and sperm morphology analysed using the Tygerberg criteria. DNA of semen and prepared spermatozoa from fertile men was found to be unaffected by cryopreservation. In marked contrast, spermatozoa from infertile men were significantly damaged by freeze-thawing. Cryopreservation had a detrimental effect on morphology of semen and prepared samples from fertile and infertile men.     

Beneficial effect of microsurgical varicocelectomy on human sperm DNA integrity

BACKGROUND: Human sperm DNA damage may adversely affect reproductive outcomes, and the spermatozoa of infertile men possess substantially more DNA damage than that of fertile men. To date, there is no available treatment for men with high levels of sperm DNA damage. The objective of this study was to examine the effect of varicocelectomy on sperm DNA denaturation (DD, an index of sperm DNA damage) in infertile men with a clinical varicocele. METHODS. We reviewed the reports of 37 men who underwent microsurgical varicocelectomy at our institution from September 2001 to July 2002. Standard semen parameters and the percentage of spermatozoa with DD (monitored by flow cytometry analysis of acridine orange-treated spermatozoa) were assessed before and 6 months after varicocelectomy. RESULTS. The percentage of spermatozoa with DD decreased following varicocelectomy compared with pre-operatively (27.7 versus 24.6%, respectively, P < 0.05). Sperm concentration and the percentages of motile sperm and normal forms (WHO criteria) increased following varicocelectomy, but the difference did not reach statistical significance. CONCLUSIONS. Our data suggest that varicocelectomy can improve human sperm DNA integrity in infertile men with varicocele. These data represent the first report of improved sperm DNA integrity after therapy and further support the beneficial effect of varicocelectomy on human spermatogenesis.     

Use of antisperm antibodies in differential display Western blotting to identify sperm proteins important in fertility

BACKGROUND: Antisperm antibodies (ASA) may be an important cause of infertility but current tests for the detection of ASA have poor prognostic value. The inadequacy of current tests may reflect the inability of these tests to define the antigenic specificity of the sperm proteins with which the ASA react. Identification of the sperm proteins that ASA bind to is a necessary preliminary step to the development of more useful diagnostic tests for ASA. METHODS: A sensitive Western blotting technique was used to compare the antigenic specificities of ASA from men who were infertile (n = 6) with those who were fertile following vasectomy reversal (n = 3). Normal fertile men (n = 3) and infertile men with known ASA (n = 4) were also included in the analysis. RESULTS: All men, including the normal fertile controls, had ASA detectable in our system. Several sperm proteins were identified that react with ASA from infertile but not fertile men. Quantitative differences in the binding of ASA to some proteins were also demonstrated. Additionally, we demonstrated that normal motile sperm are coated with an antibody that appears to be bound to sperm by a non-antigenic mechanism. CONCLUSION: Sera from all men contained ASA, but clearly some of these did not cause infertility. Characterization of the proteins that are antigens for ASA from infertile but not fertile men may allow the development of more accurate tests for infertility-inducing ASA. The significance of immunoglobulin G coated on normal sperm remains to be determined.