Hamster Oocyte Penetration Research Articles

Hepatitis B virus surface protein induces sperm dysfunction through the activation of a Bcl2/Bax signaling cascade triggering AIF/Endo G-mediated apoptosis.

BackgroundHepatitis B virus (HBV) was found to exist in semen and male germ cells of patients with chronic HBV infection. Our previous studies demonstrated that HBV surface protein (HBs) could induce sperm dysfunction by activating a calcium signaling cascade and triggering caspase‐dependent apoptosis. However, the relationship between sperm dysfunction caused by HBs and caspase‐independent apoptosis has not been investigated.ObjectivesTo evaluate the effects of HBs exposure on sperm dysfunction by activating caspase‐independent apoptosis.Materials and methodsSpermatozoa were exposed to HBs at concentrations of 0, 25, 50, and 100 μg/mL for 3 h. Flow cytometry, qRT‐PCR, immunofluorescence assay, ELISA, and zona‐free hamster oocyte penetration assays were performed.ResultsWith increasing concentrations of HBs, various parameters of the spermatozoa changed. The number of Bcl2‐positive cells declined and that of both Bax‐positive cells and Apaf‐1–positive cells increased. The transcription level of Bcl2 increased and that of both Bax and Apaf‐1 declined. The average levels of AIF and Endo G declined in mitochondria and increased in the cytoplasm and nucleus. The sperm DNA fragmentation index increased. The mean percentages of live spermatozoa declined and that of both injured and dead spermatozoa increased; and the sperm penetration rate declined. For the aforementioned parameters, the differences between the test and the control groups were statistically significant.ConclusionHBs exposure can activate the Bax/Bcl2 signaling cascade that triggers AIF/Endo G–mediated apoptosis, resulting in sperm DNA fragmentation, sperm injury, and death, and a decrease in the sperm fertilizing capacity. This new knowledge will help to evaluate the negative impact of HBV on male fertility in HBV‐infected patients.

Supplementation of cryopreservation medium with TAT-Peroxiredoxin 2 fusion protein improves human sperm quality and function

To investigate the potential effects of TAT-PRDX2 protein supplementation to the cryopreservation medium on post-thaw sperm quality and function. Invitro prospective study. Medical university hospital. Fifty normozoospermic, 50 asthenozoospermic, and 50 oligoasthenozoospermic men undergoing semen analysis for couple infertility. Each semen sample was divided into three aliquots: fresh, cryopreserved control (without additive), and cryopreserved with TAT-PRDX2 protein. Sperm motility, viability, mitochondrial potential, and DNA damage as well as reactive oxygen species (ROS) levels and lipid peroxidation were analyzed. Acrosome reaction and zona-free hamster oocyte penetration tests were performed to assess the fertilization ability of cryopreserved spermatozoa. In normozoospermic and asthenozoospermic groups, the addition of 150μg/mL TAT-PRDX2 significantly reduced intracellular ROS and malondialdehyde levels and enhanced post-thaw sperm motility and viability when compared with the cryopreserved control of the respective groups but did not produce any significant protective effect in the oligoasthenozoospermic group. Mitochondrial potential was significantly increased, whereas DNA fragmentation was significantly decreased, after TAT-PRDX2 supplementation only in the asthenozoospermic group when compared with the cryopreserved control. Although the penetration rate and the penetration index were not markedly improved, TAT-PRDX2 supplementation obviously reduced spontaneous acrosome reaction and increased calcium ionophore-induced acrosome reaction in the normozoospermic and asthenozoospermic groups. TAT-PRDX2 protein effectively exerted cryoprotective effects on spermatozoa by reducing intracellular ROS level and thereby improved post-thaw sperm quality and function, especially for asthenozoospermic samples. TAT-PRDX2 protein is a promising additive for developing a new and highly efficient semen cryoprotectant.

Aged men share the sperm protein PATE1 defect with young asthenozoospermia patients.

Does a defect in the human sperm-located protein prostate and testis expressed 1 (PATE1) exist in both aged men and young asthenozoospermia patients? A defect in sperm PATE1 exists in both aged men and young asthenozoospermia patients, and an antibody against PATE1 can decrease human sperm motility and zona-free hamster oocyte penetration. Both aged men and young asthenozoospermia patients have poor sperm quality. The PATE1 protein seems to mediate sperm-egg interactions; however, the mechanisms are still unknown. This was a case-control study including 60 young fathers (aged 28-32 years) and 60 aged fathers (68-72 years old) who donated semen by masturbation after 7 days of sexual abstinence. Comparative sperm proteome analysis from the young fathers and aged fathers was performed to discover key proteins. The target protein PATE1 was chosen and validated by western blotting and immunohistochemistry. Quantitative assessment of sperm PATE1 protein was performed on sperm from 60 young fathers, 60 aged fathers and 110 young asthenozoospermia patients. Furthermore, an antibody against PATE1 assay was used to test whether PATE1 participated in sperm motility and penetration of zona-free hamster egg. Samples were pooled and separated by two-dimensional gel electrophoresis followed by identification by matrix-assisted laser desorption/ionization time of flight mass spectrometry. Western blotting and immunohistochemistry were used to validate the confidence of proteomic data. Sperm immunofluorescence quantification experiments disclosed whether the aged men indeed shared the same PATE1 defect with 110 young asthenozoospermia patients. The sperm motility test and penetration of zona-free hamster egg assay were performed for PATE1. Twenty-two sperm proteins with significant differential expression between young adults and aged men were identified (P < 0.05, mean ratio >1.5), including 13 proteins with decreased expressions with aging. Based on bioinformatics, PATE1 was chosen for further study, and exhibited similar changes in expression level and localization on sperm from aged men and young asthenozoospermia patients. Antibody blocking revealed that PATE1 was involved in sperm-egg penetration and sperm motility. Before any clinical application of PATE1 as a biomarker for the diagnosis of male infertility, more cases should be used to evaluate confidence in this approach. This study revealed a common molecular basis underlying the decline in sperm quality in the natural aging process and in young men with asthenozoospermia. The data should greatly contribute to the development of molecular evaluation of sperm quality, and the diagnosis and treatment of asthenozoospermia. This work was supported by grants from the National Natural Science Foundation of China (NO. 81300533, 81370013 and 81000277) and Shandong Provincial Natural Science Foundation, China (ZR2013HQ002, ZR2014HQ068). The authors declare no competing financial interests.

Factors affecting sperm fertilizing capacity in men infected with HIV.

Studies on the sperm-fertilizing capacity of HIV-seropositive men show conflicting results for reasons that are not yet clear. The aim of this study was to investigate the effects and relationships of some factors such as patient age, CD4(+) cells count, fathering offspring, concomitant sexually transmitted diseases (STD), and receipt of highly active anti-retroviral therapy (HAART) on sperm fertilizing capacity. Semen samples were collected from 33 HIV-seropositive men. Data on the above factors were acquired from a self-designed questionnaire. Computer-assisted sperm analysis, a hypo-osmotic swelling, and zona-free hamster oocyte penetration tests were performed according to criteria of the World Health Organization. CD4(+) cells in peripheral blood were examined using a flow cytometric (FCM) analyzer. Sperm vitality, sperm motility (grades a + b), total sperm motility, and sperm penetration rates were significantly higher in patients whose CD4(+) counts were ≧350/µl than in those whose CD4(+) counts were <350/µl (P < 0.05), and the parameters mentioned above were also significantly correlated with CD4(+) cell number (all P < 0.05). Significant differences in total sperm count and sperm tail swelling rate between patients co-infected with STD and without STD were observed (P < 0.05). Sperm penetration rate in patients receiving HAART was significantly higher than in those not receiving HAART (P < 0.05). Blood CD4(+) cell counts are an important indicator for evaluating sperm fertilizing capacity of HIV-seropositive men. After receiving HAART, the sperm penetration rate of HIV-seropositive men can be improved.

Adhesion molecules and matrix proteins on human spermatozoa.

Ejaculated spermatozoa and spermatogenic cells express alpha- and beta-chains of beta 1, 3 and 4 integrins as well as their ligands fibronectin and laminin in an extended intra- and interindividual variation and in different patterns of location. The mRNA transcripts of these molecules were detectable by nested polymerase chain reaction in the spermatozoa. The conclusion of a functional competence of these adhesion molecules (AM) was supported by their relation to the results of the zona-free hamster oocyte penetration (HOP) test, the in vitro fertilization of human oocytes and cell attachment assays. AM labelling was influenced by the disintegration of the sperm plasma membrane, especially in seminal plasma, by progesterone, human follicular fluid and microorganisms, but was barely modified by sperm cryopreservation. Despite substantial advances in the knowledge about sperm adhesion molecules, many questions remain to be answered.

Assessment of sperm function in fertile and infertile men.

The sperm function of fertile men (control), infertility patients (experimental), and men with varicocele were compared. The bioassays used were the follicular fluid-induced acrosome reaction, the binding to the zona pellucida, and the penetration of zona-free hamster oocytes. The percentage (mean +/- SEM) of reacted spermatozoa was 35 +/- 3 in the control, 22 +/- 1 in the experimental, and 22 +/- 3 in the varicocele. The minimum value of acrosome reaction in control men was 20%. The mean number of zona-bound spermatozoa was 250 +/- 30 in the control, 160 +/- 28 in the experimental, and 196 +/- 44 in the varicocele. The minimum number of zona bound spermatozoa in control men was 50. The mean number of hamster oocytes penetrated was 50 +/- 8 in the control, 19 +/- 3% in the experimental, and 10 +/- 3 in the varicocele. The minimum number of oocytes penetrated in control men was 6%. In the experimental group, 22 men had a normal sperm function, 58 had 1 or 2 bioassays below the minimum (relative dysfunction), and 10 had all bioassay below the minimum (abnormal sperm function). The results of these bioassays could help to reclassify the infertile men in several subgroups.

Relationship between sperm apoptosis signalling and oocyte penetration capacity

Human sperm have been documented to display apoptosis-like features such as externalization of phosphatidylserine (EPS), disruption of the transmembrane mitochondrial potential (MMP) and activation of caspases. Our aim was to evaluate possible association between activation of the apoptosis cascade in human sperm and its oocyte penetration capacity using the zona free hamster oocyte penetration assay (SPA). Semen specimens from 76 unselected donors were subjected to double density gradient centrifugation followed by incubation under capacitating conditions for 3 h and SPA. Apoptosis signalling was monitored by assessment of EPS, disruption of MMP and activation of caspase-3 by flow cytometry. Semen samples with subnormal SPA values (<20% penetrated oocytes) contained significantly higher amounts of spermatozoa with EPS, disrupted MMP and activated caspase-3 compared with those samples with normal SPA values (>20% penetrated oocytes, p < 0.01). All three apoptosis markers showed a significantly negative correlation with the percentage of penetrated oocytes (p < 0.01). Apoptosis-related signalling appears to have a negative association with sperm-oocyte penetration. The exclusion of sperm presenting with those apoptosis-related features during assisted reproduction may improve success rates of procedures such as intrauterine insemination and in vitro fertilization.

Meiotic segregation analysis in male translocation carriers by using fluorescent in situ hybridization

Balanced reciprocal and Robertsonian translocations are the most common structural chromosome abnormalities in humans, with incidences of 0.7 and 1.23 per 1000. These translocations can affect fertility and/or pregnancy outcome because of possibly impaired production of gametes with an unbalanced zygote caused by the parental arrangement. Fertility problems in male translocation carriers are because of various degrees of sperm alterations that are directly related to the disturbance of the meiotic process. Investigation of human sperm chromosomes was performed by karyotyping spermatozoa after penetration of zona-free hamster oocytes, karyotype analysis now being possible to analyse the segregation patterns by using fluorescent in situ hybridization (FISH). Here, we document the results of meiotic segregation analysis for four Robertsonian and four reciprocal translocation carriers by FISH. In the sperm of Robertsonian translocation males, the majority of spermatozoa were normal/balanced. On the other hand, males with reciprocal translocations demonstrated a high rate of unbalanced spermatozoa of about 50% on meiotic segregation, with an unusually high rate (23.5%) of 3 : 1 segregation. This knowledge can be used for genetic counselling of families with these types of translocations.

The Role of Protein C Inhibitor in Human Reproduction

Protein C inhibitor (PCI) is a heparin-dependent serine protease inhibitor found in human plasma, urine, and other body fluids. In blood plasma, PCI is present at approximately 0.08 microM and inactivates activated protein C and other coagulation and fibrinolytic enzymes. In seminal plasma, PCI is present at 2.2 to 3.7 microM. The main sources of seminal PCI are the seminal vesicles, where it remains fully active. Following ejaculation, PCI completely looses its activity in approximately 2 hours, when it partially complexes with prostate-specific antigen, two plasminogen activators (urokinase-type and tissue-type plasminogen activators), and tissue kallikrein. PCI is also present in an active form in follicular fluid at approximately 0.1 microM. Purified functionally active human blood plasma-derived as well as inactive semen-derived PCI inhibited both binding and penetration of zona-free hamster oocytes by human sperm. The binding inhibition by PCI was dose dependent. A concentration of 0.04 microM PCI (approximately 100-fold lower than that present in seminal plasma) inhibited 50% of the binding and penetration ability. Given that capacitated sperm used for in vitro fertilization usually contains more than 0.05 microM of PCI, fertilization rates might be significantly reduced. All of these data suggest that PCI is involved in human reproduction at several steps, including the fertilization process.

Selection of Nonapoptotic Spermatozoa As a New Tool for Enhancing Assisted Reproduction Outcomes: An In Vitro Model1

Magnetic cell sorting (MACS) using annexin V-conjugated microbeads eliminates apoptotic spermatozoa based on the externalization of phosphatidylserine residues. The procedure delivers two sperm fractions: annexin V-negative (nonapoptotic) and annexin V-positive (apoptotic). Our aim was to determine whether the sperm fertilizing potential can be improved by selecting a nonapoptotic fraction using MACS. Semen samples (n = 35) were subjected to separation on a density gradient followed by MACS. Extent of apoptosis was assessed by measuring levels of activated caspase 3 using fluorescein-labeled inhibitors of caspase, alterations in mitochondrial membrane potential (MMP) using a lipophilic cationic dye, and DNA fragmentation using terminal deoxynucleotidyl transferase-mediated fluorescein-dUTP nick end labeling assay. The sperm fertilization potential was assessed using hamster oocyte penetration assay and hamster oocyte-intracytoplasmic sperm injection (ICSI). Annexin V-negative sperm displayed superior quality in terms of high motility, low caspase 3 activation, MMP integrity, and small extent of DNA fragmentation. Annexin V-negative sperm demonstrated higher oocyte penetration capacity but comparable sperm chromatin decondensation (SCD) following ICSI. Conversely, the annexin V-positive sperm presented with poor quality and fertilization potential. The oocyte penetration rate was negatively correlated with apoptotic marker expression, whereas SCD following ICSI was only associated with apoptosis on sperm-damaged membranes. We conclude that apoptosis appears to impact sperm-oocyte penetration rate; however, it does not seem to affect early stages of fertilization such as SCD in spermatozoa of healthy donors. The selection of nonapoptotic sperm by MACS may be used to enhance results of in vitro fertilization by increasing sperm-oocyte penetration.

The Relationship Between the Sperm Deformity Index (SDI), Apoptosis and Sperm Penetration Capacity

The Relationship Between the Sperm Deformity Index (SDI), Apoptosis and Sperm Penetration Capacity

Human testicular protein TPX1/CRISP-2: localization in spermatozoa, fate after capacitation and relevance for gamete interaction

Testicular protein Tpx-1, also known as CRISP-2, is a cysteine-rich secretory protein specifically expressed in the male reproductive tract. Since the information available on the human protein is limited to the identification and expression of its gene, in this work we have studied the presence and localization of human Tpx-1 (TPX1) in sperm, its fate after capacitation and acrosome reaction (AR), and its possible involvement in gamete interaction. Indirect immunofluorescence studies revealed the absence of significant staining in live or fixed non-permeabilized sperm, in contrast to a clear labelling in the acrosomal region of permeabilized sperm. These results, together with complementary evidence from protein extraction procedures strongly support that TPX1 would be mainly an intra-acrosomal protein in fresh sperm. After in vitro capacitation and ionophore-induced AR, TPX1 remained associated with the equatorial segment of the acrosome. The lack of differences in the electrophoretic mobility of TPX1 before and after capacitation and AR indicates that the protein would not undergo proteolytical modifications during these processes. The possible involvement of TPX1 in gamete interaction was evaluated by the hamster oocyte penetration test. The presence of anti-TPX1 during gamete co-incubation produced a significant and dose-dependent inhibition in the percentage of penetrated zona-free hamster oocytes without affecting sperm motility, the AR or sperm binding to the oolema. Together, these results indicate that human TPX1 would be a component of the sperm acrosome that remains associated with sperm after capacitation and AR, and is relevant for sperm-oocyte interaction.

Platelet activating factor improves the in vitro penetration of zona free hamster eggs by buffalo ( Bubalus bubalis) spermatozoa

Twelve buffalo bulls of Murrah breed, selected on the basis of their conception rates, were classified into low-, moderate- and high-fertility groups. Frozen semen was thawed and treated with 200 μM platelet activating factor (PAF) for 15 min at 37 °C and 5% CO 2. In both treated and control (no PAF) semen samples (five replicates per bull), the following were assessed: motility, acrosome reaction (AR) evaluation (for 10 replicates of each bull), and zona-free hamster oocyte penetration test—to determine aspects of fertilization in vitro, viz., sperm attached per ovum (SA/O), fertilization percent (FP), fertilization index (FI), and polyspermic ova (PO). There was an effect of group ( P < 0.01) on all parameters; all except motility were increased by PAF treatment. However, the group X treatment interaction was not significant for any parameter. The overall mean values of motility, AR, SA/O, FP, FI, and PO, for controls, treated spermatozoa and (net change) were: 42.89 ± 0.85, 36.65 ± 0.85, (−6.24); 28.94 ± 0.46, 61.44 ± 0.58, (32.50); 126 ± 2, 145 ± 2, (19); 74.21 ± 1.59, 89.11 ± 1.18, (14.90); 0.79 ± 0.02, 1.10 ± 0.03, (0.31) and 5.22 ± 1.22, 21.69 ± 1.88, (16.47)%, respectively. In conclusion, PAF significantly increased the AR and other aspects of fertilization, despite a small reduction in motility.

Meiotic segregation of translocations during male gametogenesis.

Balanced reciprocal and Robertsonian translocations are the most common structural chromosomal abnormalities in humans. Generally, they are without consequence for the carrier, but for various degrees of oligoasthenoteratozoospermia in men. As these carriers can produce a significant percentage of gametes with an unbalanced combination of the parental rearrangement, there is a more or less significant risk, according to cases, of chromosomal imbalances for their offspring. Therefore, techniques were developed to study the meiotic segregation of these translocations in males. Direct investigation of human sperm chromosomes became possible by karyotyping spermatozoa after penetration of zona-free hamster oocytes and, more recently, using fluorescent in situ hybridization (FISH). This paper reviews the results obtained using these techniques in Robertsonian and reciprocal translocations. The studies on spermatozoa from translocation carriers help the comprehension of the mechanisms of the meiotic segregation. They should be integrated in the genetic exploration of the infertile men, in order to give them a personalized risk assessment of unbalanced spermatozoa, specially as a correlation was found recently between the percentage of abnormal spermatozoa and that of abnormal embryos.

Human spermatozoa as a model for studying membrane receptors mediating rapid nongenomic effects of progesterone and estrogens

In the past few years, besides the classical genomic effects of steroid hormones, a plethora of so called rapid non genomic effects have been described in different cell types, which are too rapid to be due to activation of gene expression. Although some of these effects might involve the same nuclear steroid receptors acting on different cellular signalling, others have been ascribed to poorly characterized membrane receptors. Several rapid nongenomic effects of progesterone (P) and estrogens (E) have been recently demonstrated in human spermatozoa. They seem to be mediated by the steroid binding to specific receptors on plasma membrane different from the classical ones. In particular, P has been demonstrated to stimulate calcium influx, tyrosine phosphorylation of sperm proteins, including extracellular signaling regulated kinases, chloride efflux and cAMP increase, finally resulting in activation of spermatozoa through induction of capacitation, hyperactivated motility and acrosome reaction. Conversely, E, by acting rapidly on calcium influx and on protein tyrosine phosphorylation, seem to modulate sperm responsiveness to P. Several attempts have been used to characterize the putative membrane receptors for P (mPR) and E (mER) in spermatozoa, however their isolation still remains elusive. However, in the past few years our laboratory has obtained several evidences supporting the existence and functional activity of mPR and mER in human spermatozoa. To characterize these membrane receptors, we used two antibodies directed against the ligand binding domains of the classical receptors, namely c262 and H222 antibodies for PR and ER respectively, hypothesizing that these regions should be conserved between nongenomic and genomic receptors. In western blot analysis of sperm lysates the antibodies detected a band of about 57 kDa for PR and of 29 kDa for ER, excluding the presence of the classical receptors. On live human spermatozoa, both antibodies were able to block the calcium and AR response to P and E respectively, whereas, antibodies directed against different domains of the classical PR and ER were ineffective.. Moreover, c262 antibody also blocks in vitro human sperm penetration of hamster oocytes. Taken together all these data strongly support the existence of mPR and mER different from the classical ones, mediating rapid effects of these steroid hormones in human spermatozoa.

Evidence that P36, a human sperm acrosomal antigen involved in the fertilization process is triosephosphate isomerase

P36 is one of the immunodominant sperm antigens identified by antibodies eluted from the spermatozoa of infertile men. In a previous study, we isolated and characterized this auto-antigen as a glycoprotein with several isoforms. Specific rabbit antibodies were produced to investigate sperm topography and the role of P36 in the fertilization process and we showed that P36 is present on the equatorial segment of acrosome-reacted spermatozoa and is involved in sperm-binding and the penetration of zona-free hamster oocytes. In the present study, we demonstrated, by means of immunofluorescence and electron microscopy, that P36 is present all over the acrosomal membranes of non-reacted spermatozoa. We also investigated the role of P36 in the acrosome reaction and sperm binding to the zona pellucida (ZP). The exposure of capacitated spermatozoa to rabbit anti-P36 antibodies had no effect on primary fixation to the ZP, but inhibited secondary binding to the ZP and the Ca2+ ionophore-induced acrosome reaction. These results suggest that P36, an acrosomal antigen, is involved in several steps of the fertilization process. On two-dimensional Western blots, human anti-sperm antibodies (ASA) and rabbit anti-P36 antibodies recognized five to six isoforms of P36, all 36/37 kDa in size, with a pI between 5.1 and 5.7. Two major spots were identified as human triosephosphate isomerase (TPI) by MALDI-TOF mass spectrometry. Anti-TPI antibodies were shown to react with the isoforms recognized by human and rabbit anti-P36 antibodies. We also demonstrated the presence of TPI in human sperm heads. Further studies are underway to establish whether there is a sperm-specific isoform of TPI and its role in sperm function.

Chromosome complements in 695 sperm from three men heterozygous for reciprocal translocations, and a review of the literature.

Sperm chromosome complements were analysed in three men heterozygous for reciprocal translocations. A total of 695 sperm were karyotyped after in vitro penetration of hamster oocytes: 275 sperm from t(7;20)(q33.2;p13), 268 from t(3;11)(q25.3;q25) and 152 from t(15;22)(q26.1;q11.2). All possible 2:2 and 3:1 meiotic segregations were observed for all three translocations. The frequencies of alternate, adjacent 1, adjacent 2, and 3:1 segregations were 38%, 40%, 16%, and 5% for t(7;20); 48%, 46%, 6%, and 1% for t(3;11); and 34%, 40%, 22%, and 4% for t(15;22), respectively. Within the alternate segregation, the number of normal sperm was not significantly different from the number of sperm carrying a balanced form of the translocation for any of the translocations, as theoretically expected. The percentage of sperm with an unbalanced form of the translocation was 62% for t(7;20), 52% for t(3;11) and 66% for t(15;22). Sperm chromosome complements were observed in all three translocations that could be attributed to crossing-over in the interstitial segment, although nondisjunction at anaphase II could also account for the complements. There was no evidence for an interchromosomal effect in any of the translocations since the frequencies of numerical abnormalities unrelated to the translocation were within the normal range of control donors. Data from a total of 27 reciprocal translocations studied by sperm chromosomal analysis were reviewed.

Effects of metronidazole, ipronidazole, and dibromochloropropane on rabbit and human sperm motility and fertility

Treatment of protozoal pathogens in the reproductive system with chemical agents exposes flagellated sperm cells to potential toxicants. A widely used antiprotozoal agent is metronidazole. Its effect on rabbit and human sperm was compared with a more soluble 5-nitroazole compound, ipronidazole, and with a systemic environmental toxicant, dibromochloropropane (DBCP). The percentages of motile rabbit and human sperm incubated with the compounds, the velocity of sperm, migration of sperm in polyacrylamide gel, young born in rabbits, and penetration of hamster oocytes by treated human sperm were measured in seven experiments. Up to 10 mg/ml metronidazole and 1 mg/ml DBCP had little effect on most sperm characteristics. However, 10 mg/ml metronidazole and 5 mg/ml of ipronidazole increased attachment of human sperm to hamster oocytes, but oocyte penetration was unaffected. Rabbit sperm exposed to 5 mg/ml ipronidazole were infertile. No oocytes were penetrated by DBCP-treated human sperm.

Chlamydia trachomatis infection in male partners of infertile couples: incidence and sperm function.

Chlamydia trachomatis infection is one of the most common sexually transmitted diseases. Its effect on male fertility, however, is still controversial. In this study, 284 male partners of infertile couples consulting the Center of Studies in Reproductive Biology (CEBRE) were analyzed. The incidence of C. trachomatis infection among male partners of infertile couples was 38.6%. There were no significant differences between infected and noninfected infertile men in any of the sperm parameters assessed (sperm concentration, motility and morphology). The results of the three bioassays developed to evaluate sperm physiology, namely spermatozoa-zona pellucida binding, acrosome reaction stimulated with human follicular fluid and zona-free hamster oocyte penetration, showed no differences between infected and noninfected men. Electron microscopy studies suggest that spermatozoa are active agents in the dissemination of the chlamydial infection; they could be acting as 'vehicles' for the pathogens. These, and other results, suggest that the possible effect of C. trachomatis on male fertility is not due to alterations in sperm 'quality' or function, but rather to the transmission of the disease to female partners, causing inflammatory processes and promoting the generation of antisperm antibodies.

Induction of the acrosome reaction and zona-free hamster oocyte penetration by a bull with complete teratospermia versus a half brother with normal sperm.

A fertile bull producing normal sperm and a sterile half brother exhibiting 100% teratospermia were available to study an induced sperm acrosome reaction and oocyte penetration. Pedigree analysis indicated that this condition was inherited. Experiments were undertaken to study the induction of the acrosome reaction using dilaurylphosphatidylcholine (PC12) liposomes, because this procedure was previously established to be highly correlated with bull fertility. The sperm from each bull were incubated with several PC12 concentrations for varying time periods. The initial percentages of sperm from the sterile bull with intact, partially intact, and lost acrosomes were 67%, 18%, and 14%, respectively, vs 82%, 13%, and 5% for the fertile bull (P < .05). After incubation for 15 minutes with 50 microM PC12 liposomes the corresponding values were, respectively, 51%, 26%, and 19%; and 60%, 28%, and 12%. Thus, the differences after induction of the acrosome reaction, although significant (P < .05), were small. The number of sperm adhered to each oocyte averaged 22 and 10, respectively, for the fertile and sterile bulls, whereas 74% of the fertile bull sperm and only 11% of the sterile bull sperm penetrated oocytes. Mixing the sperm-oocyte complex during incubation and increasing the sperm concentration during incubation to compensate for differences in sperm motility did not markedly affect oocyte penetration by teratogenic sperm, which is consistent with this bull being sterile. In other studies, microinjection of this type of sperm was demonstrated to induce fertilization, so the consequences of using sperm with hereditary defects in assisted reproductive programs to overcome human male sterility may be a concern.