Impairment of spermatogenesis and sperm motility by the high-fat diet-induced dysbiosis of gut microbes.

OBJECTIVE: High-fat diet (HFD)-induced metabolic disorders can lead to impaired sperm production. We aim to investigate if HFD-induced gut microbiota dysbiosis can functionally influence spermatogenesis and sperm motility. DESIGN: Faecal microbes derived from the HFD-fed or normal diet (ND)-fed male mice were transplanted to the mice maintained on ND. The gut microbes, sperm count and motility were analysed. Human faecal/semen/blood samples were collected to assess microbiota, sperm quality and endotoxin. RESULTS: Transplantation of the HFD gut microbes into the ND-maintained (HFD-FMT) mice resulted in a significant decrease in spermatogenesis and sperm motility, whereas similar transplantation with the microbes from the ND-fed mice failed to do so. Analysis of the microbiota showed a profound increase in genus Bacteroides and Prevotella, both of which likely contributed to the metabolic endotoxaemia in the HFD-FMT mice. Interestingly, the gut microbes from clinical subjects revealed a strong negative correlation between the abundance of Bacteroides-Prevotella and sperm motility, and a positive correlation between blood endotoxin and Bacteroides abundance. Transplantation with HFD microbes also led to intestinal infiltration of T cells and macrophages as well as a significant increase of pro-inflammatory cytokines in the epididymis, suggesting that epididymal inflammation have likely contributed to the impairment of sperm motility. RNA-sequencing revealed significant reduction in the expression of those genes involved in gamete meiosis and testicular mitochondrial functions in the HFD-FMT mice. CONCLUSION: We revealed an intimate linkage between HFD-induced microbiota dysbiosis and defect in spermatogenesis with elevated endotoxin, dysregulation of testicular gene expression and localised epididymal inflammation as the potential causes. TRIAL REGISTRATION NUMBER: NCT03634644.

Reduction of fertility in male mice immunised with pSG.SS.C3d3.YL.Bin1b recombinant vaccine.

OBJECTIVES: To study the anti-fertility effect of a DNA vaccine using Bin1b as the target antigen in male mice. METHODS: A novel recombinant eukaryotic vector containing a fusion gene sequence of mouse Bin1b in tandem with three copies of C3d fragment (C3d3) was used to construct pSG.SS.C3d3.YL.Bin1b. The correct expression of the Bin1b-C3d3 protein was confirmed in transfected HEK293 cells by indirect immunofluorescence and western blot analysis. The fertility of immunised mice was determined by a mating experiment and sperm motility test. Anti-Bin1b antibody titres in sera were examined by ELISA assays. Binding activity of C3d3 fragment of the fusion protein was verified in C3d receptor-expressing Raji cells and flow cytometric analysis. RESULTS: Immunisation of pSG.SS.C3d3.YL.Bin1b recombinant DNA vaccine significantly decreased sperm motility and compromised fertility in male mice. ELISA results showed that the titres of anti-Bin1b IgG in sera of immunised mice increased markedly with the immunisation process. Further, the anti-fertility effect of pSG.SS.C3d3.YL.Bin1b was significantly better than that of pSG.SS.YL.Bin1b DNA vaccine and generated higher titres of anti-Bin1b antibody. CONCLUSIONS: Our results show that recombinant DNA vaccine targeting Bin1b can markedly reduce fertility in male mice, providing an alternative approach for birth control.

Advancements in the detection and implications of sperm-immobilizing antibodies in female infertility.

This review highlights over five decades of research on sperm-immobilizing antibodies (SI-Abs), which are crucial for understanding female infertility due to their effects on sperm motility and fertilization. Since the 1960s, Isojima et al. have made significant strides, notably with the Sperm Immobilization Test (SIT), which revolutionized the quantification of SI-Abs and their roles in infertility. Drawing from a comprehensive PubMed search on "the sperm immobilization test" and "sperm immobilizing antibody," our review underscores the critical insights gained into SI-Abs' impact on reproductive functions. SI-Abs result from the body's response to sperm antigens, potentially leading to infertility by affecting post-intercourse sperm function. However, the presence of anti-sperm antibodies does not guarantee infertility, indicating a complex relationship between these antibodies and reproductive outcomes. Isojima et al.'s pioneering studies paved the way for SIT and sperm immobilization titer (SI50), tools that have clarified the link between SI-Abs and infertility, focusing on disrupted sperm mobility and fertilization as key infertility mechanisms. Clinically, interventions such as in-vitro fertilization (IVF), which bypasses or eliminates SI-Abs, have improved pregnancy rates, whereas Freund's complete adjuvant therapy has deepened our understanding of infertility mechanisms. The SI50 value is crucial for predicting fertility treatment success and guiding therapeutic decisions based on antibody levels. In summary, the evolution of SI-Abs research has provided new hope for addressing infertility, significantly enriching the field of reproductive immunology, and highlighting the need for ongoing investigation.

Anti-tubulin beta 4A (TBB4A) antibody immobilized sperm in a complement-dependent manner in humans.

Sperm-immobilizing antibodies (SI-Abs) are detected in the sera of 3 % of infertile women. SI-Abs are occasionally produced as allogeneic antibodies against sperm, causing immune infertility. SI-Abs inhibit the passage of sperm through the female reproductive tract. Research on anti-sperm antibodies (ASA) remains of great importance for population control. We aimed to identify the antigens recognized by SI-Abs and elucidate the pathogenesis of immune infertility. Twelve sperm-immobilization test (SIT)-positive and fourteen SIT-negative sera were analyzed by two-dimensional electrophoresis and western blotting. Antigenic materials were extracted from well-motile sperm prepared using 0.1 % sodium dodecyl sulfate. In total, 22 different spots were detected in the 12 positive sera. Among these, three positive serum samples showed two positive signals with similar migration patterns. The significant positive spots were Mr: 49 K, pI: 5.1 and Mr: 51 K, pI: 5.6. All these positive spots were analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS); tubulin beta-4A (TBB4A) was identified from the spot Mr: 49 K, pI: 5.1. TBB4A is a major component of tubulin and constitutes the axoneme in the sperm tail and the centrosome in the sperm neck; it is generally located inside the cell. An authentic antibody against TBB4A showed a positive reaction in the sperm neck and tail regions in an immunofluorescence study. This antibody also inhibited sperm motility in a complement-dependent manner. Sperm membrane permeability reportedly changes during swimming and capacitation. We identified TBB4A as an antigenic molecule recognized by SI-Abs, which may be relevant to immunological contraception in the future.

The effect of 17α-ethynylestradiol on steroidogenesis and gonadal cytokine gene expression is related to the reproductive stage in marine hermaphrodite fish.

Pollutants have been reported to disrupt the endocrine system of marine animals, which may be exposed through contaminated seawater or through the food chain. Although 17α-ethynylestradiol (EE2), a drug used in hormone therapies, is widely present in the aquatic environment, current knowledge on the sensitivity of marine fish to estrogenic pollutants is limited. We report the effect of the dietary intake of 5 µg EE2/g food on different processes of testicular physiology, ranging from steroidogenesis to pathogen recognition, at both pre-spermatogenesis (pre-SG) and spermatogenesis (SG) reproductive stages, of gilthead seabream (Sparus aurata L.), a marine hermaphrodite teleost. A differential effect between pre-SG and SG specimens was detected in the sex steroid serum levels and in the expression profile of some steroidogenic-relevant molecules, vitellogenin, double sex- and mab3-related transcription factor 1 and some hormone receptors. Interestingly, EE2 modified the expression pattern of some immune molecules involved in testicular physiology. These differences probably reflect a developmental adjustment of the sensitivity to EE2 in the gilthead seabream gonad.

Effect of lycopene on semen quality, fertility and native immunity of broiler breeder.

1. The effect of drinking water supplementation with lycopene on the semen quality, fertility and immunity of broiler breeders was evaluated. 2. Broiler breeder males were individually caged from 25 to 42 weeks old and divided into two group: L group, treated birds (lycopene 0.5 g/l) and C group, control birds. Laying hens were divided into two groups and artificially inseminated. 3. Semen variables were evaluated and daily fertility recorded. Serum bactericidal activity was tested. 4. Semen production and viability were affected by lycopene supplementation. Serum bactericidal activity was better in L than in C group. The fertility rate curve of the L group displayed a positive trend.

Characteristics and Possible Role of Bovine Sperm Head-to-Head Agglutination.

Although sperm head-to-head agglutination has been reported in many mammalian species, the biological significance of this unique sperm-sperm interaction remains largely unknown. Here, we aimed to examine the functional characteristics of agglutinated bovine sperm to determine the possible role of sperm agglutination in the fertilization process. We initially examined temporal changes to the degree of head-to-head agglutination in culture, and found that bovine sperm agglutinated despite the lack of sperm agglutination inducers in medium. Sperm viability and motility were evaluated by SYBR14/PI and JC-1 staining, respectively, to identify the relationship between sperm agglutination and fertilizing ability. Agglutinated sperm had increased motility, viability, and intact mitochondrial function compared with unagglutinated sperm. Furthermore, we found that heparin significantly increased the percentage of unagglutinated sperm, but did not affect viability of both agglutinated and unagglutinated sperm, suggesting that sperm agglutination dictated the viability. In conclusion, agglutinated bovine sperm maintained viability and motility for a longer time than unagglutinated sperm. Thus, we propose that the head-to-head agglutination is a crucial sperm-sperm interaction to ensure the fertilizing ability of sperm.

Immunocontraceptive potential of the Ig-like domain of Izumo.

To investigate whether the Ig-like domain of sperm protein Izumo or the other part of the protein could be used as an immunocontraceptive antigen, three partially overlapping cDNA fragments (PA, PB, and PC), together covering entire mouse Izumo, were cloned, expressed, and purified. PB contains the whole Ig-like domain of mouse Izumo. The anti-PB antibody significantly inhibited the fusion of sperm with zona-free mouse eggs with no effect on sperm motility, while anti-PA and anti-PC antibodies virtually had no effect on sperm-egg fusion at the same concentration. Furthermore, in the presence of anti-PB antibody, the anti-sperm reactivity could be competitively inhibited by recombinant PB protein. The PB-specific antibody staining was restricted to the acrosome region in acrosome-reacted mouse spermatozoa by indirect immunofluorescence. Active immunization with the PB antigen sharply raised the antibody titers in mouse that were enough to cause a significant reduction in fertility compared to the PA and PC immunized groups. In conclusion, our data indicate that the Ig-like domain of Izumo plays an important role in the fertilization process, as verified by the dose-dependent reduction in fertilization rates in mouse IVF trials and mouse mating assay. These results indicate that the Ig-like domain of Izumo might be a new candidate for the development of a contraceptive vaccine.

Semen characteristics and inflammatory mediators in infertile men with different clinical diagnoses.

This study was aimed at investigating whether semen characteristics in different clinical diagnoses of infertility are associated with PMN elastase, IL-6, IL-8, IL-1beta and TNFalpha levels detected in seminal plasma. Sixty-eight patients were divided into groups according to their clinical diagnosis: idiopathic infertility (group I), varicocele with infections (group II), varicocele (group III), infections (group IV), controls (group V). Physical examination and scrotal Eco-color Doppler was used to detect the varicocele. Patients with positive bacteriological semen analysis were considered as having an infection of the male reproductive tract. Samples were examined by light microscopy and transmission electron microscopy (TEM). TEM data were quantified with a mathematical formula furnishing a fertility index and the percentage of sperm apoptosis, immaturity and necrosis. PMN elastase/alpha1-PI complex levels were determined by ELISA and IL-6, IL-8, IL-1beta, TNFalpha by Bio-Plex Cytokine assay. Sperm concentration (I-II: p < 0.005; III-IV: p < 0.0001), motility (I-IV: p < 0.0001) and the fertility index (I: p < 0.005; II-IV: p < 0.0001) were significantly lower in the groups vs. controls, whereas sperm pathologies, except for apoptosis, were significantly higher in group I and apoptosis and necrosis were higher in group III. An increase in immaturity (p < 0.005) with a decrease in necrosis (p < 0.005) were observed in group III vs. group IV. Significantly higher levels of inflammatory mediators were detected in groups III and IV vs. controls. Despite a broad relationship among different inflammatory mediators, no correlation was found among them and the semen parameters, including indices from TEM analysis. In conclusion, patients with idiopathic infertility showed altered semen quality and normal levels of inflammatory mediators. Genitourinary infection and varicocele induced an inflammatory effect which could play a detrimental role in spermatogenesis, revealed by a decrease in sperm motility and the fertility index, concomitant with an increase in immaturity mainly in varicocele and necrosis in infection.

The antibody against a nuclear autoantigenic sperm protein can result in reproductive failure.

To study whether the antibody against the testis form of the nuclear autoantigenic sperm protein (tNASP) could result in reproductive failure, we successfully cloned and expressed a 339-bp cDNA fragment of mouse tNASP (mtNASP). Using mouse as a model, recombinant mtNASP (rmtNASP) and a synthetic peptide, human tNASP(393-408) (htNASP(393-408)), were investigated for their antifertility effect. Active immunization with rmtNASP or the synthesized peptide raised high antibody titers in the immunized mice. Sperm-egg binding and fusion assay were carried out in 8-10-week-old BALB/c mice. Sperm-egg binding and in vitro fertilization of mouse oocytes were inhibited by co-incubation of zona-free mouse oocytes with capacitated mouse spermatozoa in the presence of varying concentrations of the antisera against rmtNASP. There was a significant antifertility effect in animals immunized with rmtNASP or the synthesized peptide. The effect on fertility in the mice immunized with the synthesized peptide was reversible. Our data indicate that active immunization with rmtNASP antigen may induce a strong antibody response that causes an inhibition of fertility.

Life-time exposure to waterborne copper IV: Sperm quality parameters are negatively affected in the killifish Poecilia vivipara.

In previous studies, we have shown that copper (Cu) is significantly accumulated in various tissues of killifish Poecilia vivipara following chronic exposure. Also, we showed that chronic metal exposure disrupted energy production and growth in this species. In the present study, we aimed to evaluate if chronic exposure to this metal could also affect reproductive parameters of P. vivipara males (sperm quality). In order to test that, newborn (<24 h-old) fish were exposed to two concentrations of waterborne Cu (5 and 9 µg/L) for 345 days. After exposure, fish were euthanized and the testes were collected for sperm analysis. We could observe that exposed animals had reduced sperm motility and period of motility. Also, the sperm of exposed fish had reduced plasma membrane integrity, mitochondrial functionality and DNA integrity when compared to sperm of control animals. It is suggested that the well-known association of Cu with elevated oxidative damage, endocrine disruption and energetic disturbance are involved with the observed outcomes. The results obtained in the present study show that chronic exposure to environmentally relevant concentrations of waterborne Cu caused reductions in all parameters used to evaluate sperm quality. Therefore, it is concluded that life-time exposure to this metal may disrupt fish reproduction and negatively affect the maintenance of its populations.

Dietary boron supplementation enhances sperm quality and immunity through influencing the associated biochemical parameters and modulating the genes expression at testicular tissue.

INTRODUCTION: Dietary boron improves immune and antioxidant status and calcium metabolism in mammals. However, till date the effects of dietary boron supplementation on male reproduction, especially on sperm production and sperm quality in farm animals are not documented. OBJECTIVE: The present study was aimed to investigate the influence of dietary boron on semen production, semen quality, immunity and molecular changes in the testis, blood and seminal plasma and to assess the interrelationship with other minerals in male goats. METHODOLOGY: The study was conducted in 21 adult male goats divided into 3 groups (control, boron and selenium supplemented groups, n = 7 each). In boron group, boron was supplemented at 40 ppm and in selenium group, selenium was supplemented at 1 ppm over and above the basal level. In control group, only the basal diet was fed without supplementary boron or selenium. The feeding trial was carried out for 60 days. Selenium was taken as a positive control for the dietary boron supplementation experiment. Following feeding trials, the sperm concentration, kinematics and functional attributes, immunity and molecular level changes in the testis, biomolecular changes in the blood and seminal plasma and also interrelationship with other minerals were studied. RESULTS: The average sperm concentration (million/ml) and the total sperm production (million/ejaculate) were significantly (p < 0.05) increased in boron supplemented group when compared to selenium and control groups. The boron levels in blood plasma (r = 0.65) and seminal plasma (r = 0.54) showed a positive correlation with sperm progressive motility. Blood and seminal plasma metabolic biomarker namely, aspartate aminotransferase (AST) (p < 0.01) was significantly lower in the boron and selenium supplemented group than control, while alanine aminotransferase (ALT) (p < 0.05) was significantly lower in the boron supplemented group than selenium and control group. There was a significant increase in the mRNA expression of serine proteinase inhibitor (SERPIN) and interferon γ (IFNγ) in the testis of boron supplemented than the control group. Boron supplementation up-regulated the immune-regulatory gene, interleukin 2 (IL2) and antioxidant gene, catalase (CAT) in the peripheral blood mononuclear cells (PBMC). On contrary, toll-like receptor 2 (TLR2) mRNA expression was significantly (p < 0.05) down-regulated in boron and selenium supplemented groups. CONCLUSION: The study revealed that dietary boron supplementation increased the sperm output, sperm motility and enhanced the immune and antioxidant defense capacity in male goats. The improved semen quality can be attributed to enhanced expression of testicular SERPIN, a crucial protein for the regulation of spermatogenesis process.

Application of computer-aided sperm analysis (CASA) for detecting sperm-immobilizing antibody.

PROBLEM: Since the 1970s, anti-sperm antibodies have been studied as a pathogenic factor contributing to infertility. The complement-dependent sperm-immobilization test (SIT) and quantitative SIT have been used as effective tools for detecting anti-sperm antibodies in clinical settings. These tests have been carried out traditionally by manually counting the number of motile sperm through eye estimation. METHOD OF STUDY: In this study, we developed a novel method using computer-aided sperm analysis. The results were compared with those obtained by the traditional method. RESULTS: The results were identical and 25 of 78 samples tested were positive and 53 samples were negative for sperm-immobilizing (SI) antibodies based on both methods. For SI-positive samples, the values of SI50 obtained using the two methods correlated closely with high co-efficiency. CONCLUSION: Using the novel method, manually counting the number of motile spermatozoa becomes unnecessary. The novel method presented here will increase the objectivity and convenience of using the SIT as a clinical indicator.

Human Papillomavirus Prophylactic Vaccination improves reproductive outcome in infertile patients with HPV semen infection: a retrospective study.

In this study we aimed to evaluate the effect on reproductive outcome of HPV vaccination in male subjects of infertile couples with HPV semen infection. In this single-center study, we retrospectively enrolled 151 infertile couples with detection of HPV in semen, attending our Hospital Unit of Andrology between January 2013 and June 2015, counseled to receive adjuvant HPV vaccination. Seventy-nine accepted vaccination (vaccine group) whilst 72 did not (control group). Our protocol of follow-up, aimed to evaluate HPV viral clearance, consisted in semen analysis, INNO-LiPA and FISH for HPV in semen cells after 6 and 12 months from basal evaluation. Spontaneous pregnancies, miscarriages and live births were recorded. Progressive sperm motility and anti-sperm antibodies were improved in the vaccine group at both time points (p < 0,05 vs control arm). Forty-one pregnancies, 11 in the control group and 30 in the vaccine group, were recorded (respectively 15% and 38,9%, p < 0,05) and resulted into 4 deliveries and 7 miscarriages (control group) and 29 deliveries and one miscarriage (vaccine group, p < 0,05 vs control group). HPV detection on sperms was predictive of negative pregnancy outcome. Adjuvant vaccination associated with enhanced HPV healing in semen cells and increased rate of natural pregnancies and live births.

Immunization with autologous CD46 generates a strong autoantibody response in rats that targets spermatozoa.

CD46, a membrane complement regulator, has been implicated as pathogen receptor, T cell activator and contributor to spermatozoa-egg interactions. In man, a role in the fertilization process was suggested by its localization on the acrosome. In rodents, CD46 is expressed only on the spermatozoal acrosome, suggesting an essential role at this site. This restricted expression led us to ask whether immunization with CD46 would generate anti-CD46 antibody responses that might target spermatozoa and influence fertility. We immunized male and female rats with rat CD46. Strong immune responses were generated in all rats and immune sera stained CD46 in testis extracts and in situ in testis and sperm. Incubation of spermatozoa with immune sera caused deposition of immunoglobulin and C3b in an acrosome pattern and reduced motility. We mated immune male rats with naïve females and female immune rats with naïve males. The incidence of pregnancy and number of fetuses were not different in matings involving immune male or female rats compared to controls. Testis sections from immune rats revealed no immunoglobulin deposition on CD46-positive sperm precursors, suggesting that acrosomal CD46 was inaccessible in this location. A minority of spermatozoa harvested from epididymis of immune rats had immunoglobulin and C3b bound to the acrosome, suggesting that anti-CD46, present in genital tract fluids, bound after acrosome reaction. These data demonstrate that the restricted expression of CD46 allows strong anti-CD46 responses in rats that target spermatozoa in vitro and in vivo. The anti-CD46 response did not influence fertility, perhaps reflecting the considerable redundancy for fertilization in rodents.

The expression and significance of CATSPER1 in human testis and ejaculated spermatozoa.

AIM: To investigate the distribution of cation channel of sperm 1 (CATSPER1) protein and the presence of CATSPER1 mRNA in human testis and ejaculated spermatozoa. The influence of anti-human CATSPER1 antibody upon human sperm motility was used to evaluate the function of human CATSPER1 and to estimate its possible use as a target for immunocontraception. METHODS: Human ejaculated sperm from normozoospermic donors (n = 12) and liquid nitrogen frozen human testis were used for the study of mRNA and protein expression of CATSPER1 by reverse transcription polymerase chain reaction (RT-PCR) and immunohistochemistry, respectively. Spermatozoa from normozoospermic donors (n = 12) were individually processed using a swim-up procedure and were then incubated with CATSPER1 antibody at final concentrations of 20, 4 and 0.8 microg/mL. After 1, 2 and 6 h incubation, progressive motility and fast progressive motility were measured by means of computer-assisted semen analysis. RESULTS: CATSPER1 transcript was detected in both human testis and each human ejaculated semen sample. CATSPER1 protein expressed in the membrane of spermatid and was localized in the principal piece of the sperm tail. The application of CATSPER1 antibody at all concentrations significantly inhibited both progressive motility and fast progressive motility after 1, 2 and 6 h incubation, and significant dose-dependent changes were observed. CONCLUSION: CATSPER1 is meiotically and post-meiotically expressed in human testis tissue. CATSPER1 mRNA in human ejaculated spermatozoa could be a more feasible target for study and infertility screening than testis biopsy. In addition, our results suggest that human CATSPER1 could be a possible target for immunocontraception.

Incidence of Sperm Surface Autoantibodies and Relationship with Routine Semen Parameters and Sperm Kinematics.

PROBLEM: Antisperm antibodies (ASA) are associated with male subfertility. However, results on sperm surface autoantibodies are controversial, the relationship between ASA and semen parameters (WHO, 2010) is unknown, and data on ASA and sperm kinematics are scarce. METHOD OF STUDY: A retrospective study carried out in men undergoing routine semen analysis (WHO 2010), ASA evaluation (direct SpermMAR(™) (IgG) test), and computer-assisted sperm analysis (CASA). RESULTS: A 2.6% and a 5.9% incidence of ASA-positive cases were found (cut-off 50% and 10%, respectively; n = 7492). ASA-positive samples had lower (P < 0.0001) sperm concentration, count, motility, and hypo-osmotic swelling (HOS) test score. HOS results did not correlate with sperm vitality in normozoospermic samples with high ASA levels. In unselected samples, ASA-positive samples (cut-off 50%) showed decreased sperm kinematics (VSL, VAP, LIN, ALH, STR, BCF, WOB), but in normozoospermic samples, ASA-positive and ASA-negative subgroups had similar CASA results. CONCLUSIONS: ASA evaluation is highly relevant in full semen assessment.

Effect of hepatitis B virus infection on sperm quality and oxidative stress state of the semen of infertile males.

PROBLEM: The effects of hepatitis B virus (HBV) infection on sperm quality and oxidative stress state of the semen of infertile males remain undetermined. METHOD OF STUDY: Normal males and 60 semen samples from infertile males (with or without HBV infection) were subjected to semen analysis. RESULTS: Semen volume, semen pH, sperm density, percentage of forward, movement of sperm, sperm activation rate, sperm survival rate, rate of normal sperm morphology of infertile males with HBV infection were significantly lower than those of infertile males without genital infection and of normal males (P<.05), while interleukin (IL)-17, IL-18, and malondialdehyde (MDA) levels in subjects with HBV infection were significantly higher than those of infertile males without genital infection and of normal males (P<.05). In patients with HBV infection, MDA level was found to be negatively correlated with semen quality, but positively correlated with semen IL-17 and IL-18 concentrations. CONCLUSIONS: HBV infection increased MDA level, induced abnormal expression of IL-17 and IL-18, and negatively affected male reproductive capacity, resulting in male infertility.

Target antigens for sperm-immobilizing antibodies found in infertile women.

Complement-dependent sperm-immobilizing antibodies (SI-Abs) have frequently been detected in the sera of infertile women. Analyses using monoclonal antibodies as well as patients' antisperm antibodies have shown that the carbohydrate moieties of sperm and seminal plasma are major epitopes of these antibodies. In the present study, we characterized the antigen molecules recognized by human monoclonal SI-Ab (MAb H6-3C4) derived from a patient with strong SI-Ab activity. Immunofluorescent staining showed that MAb H6-3C4 reacted with ejaculated human sperm and the epididymis but not with the testes and other somatic organs including the spleen. In Western blot analysis, MAb H6-3C4 reacted to antigen molecules of Mr 15-27 kDa present in the methanol-chloroform extracts of either ejaculated sperm or seminal plasma. Amino acid sequence analysis of the reactive molecule revealed that the amino acid sequence was identical to the core peptide of CD52, a GPI-anchored protein on lymphocytes. Furthermore, the epitope for MAb H6-3C4 was found to be the specific N-linked carbohydrate for the male reproductive tract. These results suggest that SI-Abs in some infertile women target a sperm-specific carbohydrate antigen on CD52 molecules which are secreted from the epididymis and coat the sperm surface.