Impact of COVID-19 vaccination on seminal and systemic inflammation in men.

Expedited development of SARS-CoV-2 vaccines led to public concerns regarding impacts of the novel vaccine on gametes in patients seeking assisted reproduction. In cases of an acute intermittent illness or fever in men, it is often advised to postpone ART treatments so that efforts can be made to enhance wellbeing and improve sperm parameters. However, it is unknown whether sperm parameters are altered in the acute (24-72 hour) phase following COVID-19 vaccination. We performed a longitudinal cohort study of 17 normospermic male patients attending a fertility clinic for semen analysis. Semen and matched peripheral blood samples were collected prior to vaccination, within 46 + 18.9 hours of vaccine course completion (acute) and at 88.4 + 12 days (3 months) post-vaccination. No overall change from baseline was seen in symptoms, mean volume, pH, sperm concentration, motility, morphology or DNA damage in the acute or long phase. Seminal plasma was found to be negative for anti-SARS-CoV2 Spike antibody detection, and MCP-1 levels showed an acute but transient elevation post-vaccine, while IL-8 was marginally increased 3 months after completion of vaccination. A modest, positive correlation was noted between serum levels of the anti-inflammatory cytokine IL-10 and self-reported symptoms post-vaccine. Our findings are reassuring in that no significant adverse effect of vaccination was noted and provide evidence to support the current recommendations of reproductive medicine organisations regarding timing of vaccination during fertility treatment.

Update on the research on the antigens of anti-sperm antibodies over the last decade.

This review summarizes the advancements over a decade of research on antigens of anti-sperm antibodies (ASAs), which are key to male immune infertility. Despite the progress in assisted reproductive technologies, understanding the roles and mechanisms of ASAs and their antigens remains vital for immune infertility management. We conducted a comprehensive literature search on PubMed from January 2013 to December 2023 using the following keywords: "anti-sperm antibody," "sperm antigen," and "immune infertility." In this review, we focus on the discoveries in sperm antigen identification and characterization through proteomics, gene disruption technology, and immunoinformatics, along with the development of fertility biomarkers. Here, we discuss the clinical applications of improved ASA detection methods and the progress in the development of immunocontraceptive vaccines. The intersection of advanced diagnostic techniques and vaccine development represents a promising frontier in reproductive health. The findings also highlight the need for standardized ASA detection methods and a comprehensive molecular-level approach to understanding ASA-related infertility. These insights underscore the significance of ongoing reproductive immunology research in enhancing clinical fertility outcomes and contraceptive vaccine development.

A comprehensive insight into the contribution of epigenetics in male infertility; focusing on immunological modifications.

Numerous recent studies have examined the impact epigenetics-including DNA methylation-has on spermatogenesis and male infertility. Differential methylation of several genes has been linked to compromised spermatogenesis and/or reproductive failure. Specifically, male infertility has been frequently associated with DNA methylation abnormalities of MEST and H19 inside imprinted genes and MTHFR within non-imprinted genes. Microbial infections mainly result in male infertility because of the immune response triggered by the bacteria' accumulation of immune cells, proinflammatory cytokines, and chemokines. Thus, bacterially produced epigenetic dysregulations may impact host cell function, supporting host defense or enabling pathogen persistence. So, it is possible to think of pathogenic bacteria as potential epimutagens that can alter the epigenome. It has been demonstrated that dysregulated levels of LncRNA correlate with motility and sperm count in ejaculated spermatozoa from infertile males. Therefore, a thorough understanding of the relationship between decreased reproductive capacity and sperm DNA methylation status should aid in creating new diagnostic instruments for this condition. To fully understand the mechanisms influencing sperm methylation and how they relate to male infertility, more research is required.

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.

Evaluation of the relationships between oxidative stress, inflammation, hormonal status and sperm parameters in rats: Canonical correlation analysis.

Many endocrine or non-endocrine factors are involved in sperm production. Although reproductive hormones are very important for the initiation and maintenance of spermatogenesis, other factors, such as inflammation and oxidative stress, affect spermatogenesis. The aim of this study is to evaluate the relationships between sperm parameters and hormones, oxidative stress, and inflammation status. We conducted this study on 40 rats. Sperm parameters (motility, abnormal sperm rate, and dead sperm rate), oxidative stress (malondialdehyde, glutathione, glutathione peroxidase, and catalase), inflammation (NF-κß, TNF-α, IL-1ß, IL-6, and IL-10), and hormone parameters (follicle-stimulating hormone, luteinizing hormone, testosterone, melatonin, and corticosterone) were determined. Relationships between mentioned parameters were investigated by canonical correlation analysis. Canonical correlation coefficients for these data sets (sperm-oxidative stress, sperm-inflammation, and sperm-hormone parameters) were found to be strongly significant (rc= 0.875, p<0.001; rc= 0.868, p<0.001; rc= 0.886, p<0.001, respectively). The rate of explanation of oxidative stress, inflammation parameters and hormones by sperm parameters was 61.80 %, 56.10 % and 63.90 %, respectively. Canonical correlation analysis results have revealed that dead sperm rate is mostly related to nuclear factor-kappa beta (NF-κß), catalase, and corticosterone. CCA, which has taken into account the multiple relationships, has revealed that multidimensional evaluation of data sets can provide important and innovative information to researchers for the assessment of relationships between sperm, oxidative stress, inflammation, and hormone parameters.

Effects of antisperm antibodies post vasectomy reversal on pregnancy rates.

OBJECTIVE: To investigate the correlation between antisperm antibodies (ASAs), pregnancy rates, and the method of conception following vasectomy reversal. This is particularly relevant as patients undergoing vasectomy reversal often express concerns about the potential inhibitory effects of ASAs on achieving pregnancy. Additionally, the American Urological Association guidelines for vasectomy emphasize the need for further research to address this question. PATIENT AND METHODS: We conducted a retrospective analysis involving chart reviews and phone interviews with individuals who underwent vasectomy reversal at our institution between May 2015 and April 2023. Patients who underwent vasectomy reversal for reasons other than fertility, as well as those lacking postoperative semen analysis with ASA data, were excluded. We classified patients based on low (below 50%) or high (50% or above) ASA levels determined by their initial postoperative semen analysis. The primary outcome measured was the pregnancy rate, including details on the method of conception. RESULTS: A total of 145 patients were subjected to chart review. The median age at the time of surgery was 43 years, with a median obstruction interval of 7.7 years. The median age of their partners was 29 years. The majority (80%) of patients underwent bilateral vasovasostomy. Among them, 60 patients (41.4%) exhibited low (< 50%) ASA levels, while 85 (58.6%) had high (≥ 50%) ASA levels. Follow-up phone interviews were completed by 48 patients. Among them, the 19 men with low ASA levels, 13 (68.4%) achieved pregnancy, with 6 (31.6%) experiencing spontaneous conception. For the 29 men with high ASA levels, 21 (72.4%) achieved pregnancy, including 11 (38%) through spontaneous conception. The p-value from Fisher's exact test was 0.2. CONCLUSIONS: Our findings suggest that ASA levels do not show a significant association with either the pregnancy rate or the method of conception following vasectomy reversal.

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.

The enigmatic interplay of immune cells and abnormal spermatozoa through Mendelian randomization.

PURPOSE: Abnormal spermatozoa significantly impact reproductive health, affecting fertility rates, potentially prolonging conception time, and increasing the risk of miscarriages. This study employs Mendelian randomization to explore their potential link with immune cells, aiming to reveal their potential causal association and wider implications for reproductive health. METHODS: We conducted forward and reverse Mendelian randomization analyses to explore the potential causal connection between 731 immune cell signatures and abnormal spermatozoa. Using publicly available genetic data, we investigated various immune signatures such as median fluorescence intensities (MFI), relative cell (RC), absolute cell (AC), and morphological parameters (MP). Robustness was ensured through comprehensive sensitivity analyses assessing consistency, heterogeneity, and potential horizontal pleiotropy. The MR study produced a statistically significant p-value of .0000684, Bonferroni-corrected for the 731 exposures. RESULTS: The Mendelian randomization analysis revealed strong indications of a reciprocal relationship between immune cell pathways and sperm integrity. When examining immune cell exposure, a potential causal link with abnormal sperm was observed in 35 different types of immune cells. Conversely, the reverse Mendelian randomization results indicated that abnormal sperm might causally affect 39 types of immune cells. These outcomes suggest a potential mutual influence between alterations in immune cell functionality and the quality of spermatozoa. CONCLUSION: This study highlights the close link between immune responses and sperm development, suggesting implications for reproductive health and immune therapies. Further research may offer crucial insights into male fertility and immune disorders.

How sperm protects itself: A journey in the female reproductive system.

Sperm must pass a complex route in the female reproductive tract (FRT) to reach the fertilization site and join the oocyte. Thus, it should employ several mechanisms to survive against the female immune system, fertilize the oocyte, and successfully transmit paternal genes to the next generation. In addition to self-protection, sperm may be involved in the immune tolerance to the developing embryo and regulating the FRT for embryo implantation and subsequent pregnancy. Hence, this review intends to summarize the mechanisms that protect sperm in the FRT: including immunomodulatory factors that are carried by seminal plasma, cell-to-cell and molecular interaction of sperm with epithelial and immune cells of the FRT, high regulated secretions of inflammatory factors such as cytokines, chemokines, and growth factors, inducing immune tolerance to paternal antigens, and specialized expression of cell receptors and binding proteins. In most of these events sperm induces the FRT to protect itself by modulating immune responses for its own benefit. However, not all sperm in the semen are able to trigger the survival mechanisms and only high-quality sperm will overcome this challenge. A clear understanding of the molecular mechanisms that maintain sperm viability and function in the FRT can lead to new knowledge about infertility etiology and a new approach in assisted reproductive technologies for the preparation and selection of the best sperm based on the criteria that physiologically happen in-vivo.

Acute murine-betacoronavirus infection impairs testicular steroidogenesis and the quality of sperm production.

Although several testicular alterations promoted by coronavirus infection have been demonstrated, the extent, causes, and players of testicular pathogenesis are not totally understood. The present study aimed to investigate the short-term effects on male fertility of intranasally administered murine hepatitis virus strain 3 (MHV-3), a member of the genus Betacoronavirus, which causes a severe systemic acute infection. This mouse model might be used as a in vivo prototype for investigating the impact of betacoronavirus on the endocrine and exocrine testicular functions with the advantage to be performed in a biosafety level 2 condition. Herein, we performed virological, histopathological, and molecular studies regarding the testicular spermatogenesis and the spermatic quality analyses in an MHV-3-infected C57BL/6 mice. The main outcomes showed that MHV-3 infects mouse testis and induces a testicular inflammatory state, impairing the steroidogenic pathway. The infection led to several alterations in the testicular parenchyma, such as: seminiferous epithelium sloughing, retention of residual bodies, germ cell apoptosis, alterations in intercellular junction proteins, and worse spermatogenic parameters. Moreover, the levels of plasmatic testosterone as well as the quality of sperm production reduced. Therefore, the present data suggest that the viral/inflammatory impairment of the steroidogenic pathway and the consequent imbalance of androgen levels is critical in testicular pathology, disturbing the SC barrier function and the germ cell differentiation. Our study is important for comprehending the effects of beta coronavirus infections on testis function in order to develop treatments that could prevent virus-mediated male infertility.

Vaginal extracellular vesicles impair fertility in endometriosis by favoring Th17/Treg imbalance and inhibiting sperm activity.

Extracellular vesicles (EVs) play a key role in various diseases. However, their effect on endometriosis (EMs)-associated infertility is poorly understood. We co-cultured EVs from the female vaginal secretions with human sperm and also generated a mouse model of EMs by allogenic transplant to explore the effect of EVs on fertility. EVs from individuals with EMs-associated infertility (E-EVs) significantly inhibited the total motility (26.46% vs. 47.1%), progressive motility (18.78% vs. 41.06%), linear velocity (21.98 vs. 41.91 µm/s) and the acrosome reaction (AR) rate (5% vs. 22.3%) of human sperm in contrast to the control group (PBS). Furthermore, E-EVs dose-dependently decreased the intracellular Ca2+ ([Ca2+]i), a pivotal regulator of sperm function. Conversely, healthy women (H-EVs) increased human sperm motion parameters, the AR rate, and sperm [Ca2+]i. Importantly, the mouse model of EMs confirmed that E-EVs further decreased the conception rate and the mean number of embryo implantations (7.6 ± 3.06 vs. 4.5 ± 3.21) compared with the control mice by inducing the production of inflammatory cytokines leading to a Th17/Treg imbalance. H-EVs could restore impaired fertility by restoring the Th17/Treg balance. We determined the impact of EVs derived from the female genital tract on human sperm function and studied the possible mechanisms by which it affects fertility. Our findings provide a novel rationale to ameliorate EMs-associated infertility.

The effect of probiotics on immunogenicity of spermatozoa in couples suffering from recurrent spontaneous abortion.

BACKGROUND: Impaired spermatozoa immunogenicity can result in pregnancy complications such as recurrent spontaneous abortion (RSA). Given that spermatozoa contact with microbiota, it is possible that inappropriate microbiota composition in the reproductive tract could result in the alteration of spermatozoa antigenicity. Probiotics, as a representative of microbiota, may therefore have a beneficial effect on this altered immunogenicity. The objective of this study was to determine the effect of probiotics on spermatozoa immunogenicity. METHODS: Twenty-five fertile couples and twenty-five RSA couples were included in this study. Spermatozoa were purified and treated with probiotics. Untreated and probiotic treated spermatozoa were evaluated for human leukocyte antigen (HLA) class I & II expression by flow cytometry. Untreated and probiotic treated spermatozoa were also cocultured with the wife's peripheral blood mononuclear cells (PBMC) for 12 days. Then, the supernatant was assessed for IgG and APCA by enzyme-linked immunosorbent assay (ELISA) and complement-dependent cytotoxicity (CDC) assay respectively. RESULTS: Probiotic treatment of spermatozoa leads to an increase of HLA class I & II expression in both the fertile and RSA groups. The probiotic treatment resulted in a decrease in both IgG and APCA in the fertile group, but an increase in both IgG and APCA in the RSA group. CONCLUSIONS: The results of this study suggest that a supplementary probiotic treatment may be useful in couples suffering from RSA with an immunologic cause, because it improves disturbed HLA expression on spermatozoa and improves disturbed APCA and IgG production in the presence of spermatozoa.

Examination of testicular lumicrine regulation of activins and immunoregulatory genes in the epididymal caput.

BACKGROUND: Immunoregulatory genes encoding activin A (Inhba) and B (Inhbb), and indolamine 2,3-dioxygenase-1 (Ido1) are highly expressed in the murine caput epididymidis, which also has a network of intraepithelial mononuclear phagocytes. This environment is postulated to promote immunological tolerance to epididymal sperm. The factors regulating the immunoregulatory agents in the epididymal caput are poorly understood. OBJECTIVES: This study aimed to investigate the potential role of testicular lumicrine factors in regulating activin and other immune-related genes in the caput epididymidis. MATERIALS AND METHODS: The efferent ducts in adult C57/Bl6 mice were exposed and ligated bilaterally. Serum and tissues were collected seven days later. Animals with bilateral sham ligation and animals with no ligations (collectively referred to as the "intact" group) were used as controls. RESULTS: Pressure-induced seminiferous epithelial damage due to intratubular fluid accumulation was observed in all ligated testes. Testicular inhibin was significantly increased and testosterone was elevated in some animals following bilateral ligation, but serum testosterone, serum LH, and serum inhibin were normal. Ligation caused epithelial regression in the initial segment, with similar but less severe effects in other caput segments. Activin A staining by immunohistochemistry in the epithelium was reduced in bilateral ligation, particularly in the initial segment, with moderately reduced staining intensity in the rest of the caput. Inhba expression within the caput was not significantly affected by bilateral ligation, but Inhbb was reduced by more than 60%. Transcripts encoding the macrophage-specific receptor Cx3cr1 were significantly reduced following bilateral ligation, but other immune cell markers, Ido1, and inflammatory genes were unaffected. CONCLUSION: These data indicate that testicular lumicrine secretion regulates several genes that are preferentially expressed in the initial segment, but has marginal effects on genes such as those encoding activin A and IDO1, which are expressed more widely in the caput.

Influence of inguinal hernia repair on sperm autoimmunity: The largest single center experience.

BACKGROUND: Studies on immunological infertility after inguinal hernia correction are few and not very representative. Anti-sperm antibodies have been shown to reduce male fertility. Although the extent of infertility due to anti-sperm antibodies alone is not very clear, data indicates that about 8%-10% of infertile patients have immunological infertility DESIGN: This retrospective study includes all infertile male patients (n = 2258) who underwent mixed antiglobulin reaction tests and urologic examination from 2000 to 2020. Sperm quality (assessed by the number of spermatozoa, their motility, vitality, and normal form) was also evaluated. Among these patients, 191 had previously undergone unilateral or bilateral inguinal hernia surgery repair. The aim of the study is to evaluate if there is a higher incidence of positive mixed antiglobulin reaction test among patients undergoing inguinal hernioplasty compared to the unselected infertile population. RESULTS: Anti-sperm antibodies would seem to increase in both patients who performed general andrological surgery and groin hernia correction, respectively 3.48 (95% Confidence Interval: 1.70-7.10; p < 0.001) and 2.45 (95% Confidence Interval: 1.01-5.99; p < 0.05) times more than the unselected infertile population. CONCLUSIONS: Mixed antiglobulin reaction test could be useful in patients undergone previous scrotal surgery or hernia correction men, to avoid false unexplained infertility diagnoses and to direct the couple to assisted reproductive technology procedures. Basal evaluation of spermatozoa does not actually consider andrological surgery as an indication to autoimmunity investigation.

Hexavalent sperm-binding IgG antibody released from vaginal film for development of potent on-demand nonhormonal female contraception.

Nonhormonal products for on-demand contraception are a global health technology gap; this unmet need motivated us to pursue the use of sperm-binding monoclonal antibodies to enable effective on-demand contraception. Here, using the cGMP-compliant Nicotiana-expression system, we produced an ultrapotent sperm-binding IgG antibody possessing 6 Fab arms per molecule that bind a well-established contraceptive antigen target, CD52g. We term this hexavalent antibody "Fab-IgG-Fab" (FIF). The Nicotiana-produced FIF had at least 10-fold greater sperm-agglutination potency and kinetics than the parent IgG, while preserving Fc-mediated trapping of individual spermatozoa in mucus. We formulated the Nicotiana-produced FIF into a polyvinyl alcohol-based water-soluble contraceptive film and evaluated its potency in reducing progressively motile sperm in the sheep vagina. Two minutes after vaginal instillation of human semen, no progressively motile sperm were recovered from the vaginas of sheep receiving FIF Film. Our work supports the potential of multivalent contraceptive antibodies to provide safe, effective, on-demand nonhormonal contraception.

The Effects of Sperm and Seminal Fluid of Immunized Male Mice on In Vitro Fertilization and Surrogate Mother-Embryo Interaction.

The latest vaccination campaign has actualized the potential impact of antigenic stimuli on reproductive functions. To address this, we mimicked vaccination's effects by administering keyhole limpet hemocyanin (KLH ) to CD1 male mice and used their sperm for in vitro fertilization (IVF). Two-cell embryos after IVF with spermatozoa from control (C) or KLH-treated (Im) male mice were transferred to surrogate mothers mated with vasectomized control (C) or KLH-treated (Im) male mice, resulting in four experimental groups: C-C, Im-C, C-Im, and Im-Im. The pre-implantation losses were significantly lower in the Im-C group than in the C-Im group. At the same time, the resorption rates reduced markedly in the C-Im compared to the Im-C group. Embryo and placenta weights were significantly higher in the Im-Im group. Although the GM-CSF levels were lower in the amniotic fluid of the gestating surrogate mothers in the Im-Im group, they were strongly correlated with embryo mass. The number-size trade-off was only significant in the Im-Im group. This suggests a positive, cooperative effect of spermatozoa and seminal fluid from immune-primed males on embryo growth and the optimal distribution of surrogate mother maternal resources despite the negative impact of males' antigenic challenge on the IVF success rate.

Transmission of trained immunity and heterologous resistance to infections across generations.

Intergenerational inheritance of immune traits linked to epigenetic modifications has been demonstrated in plants and invertebrates. Here we provide evidence for transmission of trained immunity across generations to murine progeny that survived a sublethal systemic infection with Candida albicans or a zymosan challenge. The progeny of trained mice exhibited cellular, developmental, transcriptional and epigenetic changes associated with the bone marrow-resident myeloid effector and progenitor cell compartment. Moreover, the progeny of trained mice showed enhanced responsiveness to endotoxin challenge, alongside improved protection against systemic heterologous Escherichia coli and Listeria monocytogenes infections. Sperm DNA of parental male mice intravenously infected with the fungus C. albicans showed DNA methylation differences linked to immune gene loci. These results provide evidence for inheritance of trained immunity in mammals, enhancing protection against infections.

Identification of Arvicola terrestris scherman Sperm Antigens for Immune Contraceptive Purposes.

The cyclical proliferation of the wild fossorial rodent Arvicola terrestris scherman (ATS) is critical in mid-mountain ecosystems of several European countries. Our goal is to develop an immunocontraceptive vaccine to control their fertility, as a sustainable alternative to chemical poisons currently used. Indeed, these chemicals cause the death of ATS predators and animals sharing their ecosystem, and current laws progressively limit their use, making the development of a targeted vaccination strategy an interesting and efficient alternative. In order to identify species-specific sperm antigens, male and female ATS received subcutaneous injections of whole ATS spermatozoa to elicit an immune response. The analysis of the immune sera led to the identification of 120 immunogenic proteins of sperm cells. Of these, 15 were strictly sperm-specific and located in different regions of the male gamete. Some of these antigens are proteins involved in molecular events essential to the reproductive process, such as sperm-egg interaction, acrosomal reaction, or sperm motility. This approach not only identified a panel of immunogenic proteins from ATS sperm cells, but also demonstrated that some of these proteins trigger an immune response in both male and female ATS. These spermatic antigens are good candidates for the development of a contraceptive vaccine.

The Binding of Monoclonal and Polyclonal Anti-Z-DNA Antibodies to DNA of Various Species Origin.

DNA is a polymeric macromolecule that can display a variety of backbone conformations. While the classical B-DNA is a right-handed double helix, Z-DNA is a left-handed helix with a zig-zag orientation. The Z conformation depends upon the base sequence, base modification and supercoiling and is considered to be transient. To determine whether the presence of Z-DNA can be detected immunochemically, the binding of monoclonal and polyclonal anti-Z-DNA antibodies to a panel of natural DNA antigens was assessed by an ELISA using brominated poly(dG-dC) as a control for Z-DNA. As these studies showed, among natural DNA tested (Micrococcus luteus, calf thymus, Escherichiacoli, salmon sperm, lambda phage), micrococcal (MC) DNA showed the highest binding with both anti-Z-DNA preparations, and E. coli DNA showed binding with the monoclonal anti-DNA preparation. The specificity for Z-DNA conformation in MC DNA was demonstrated by an inhibition binding assay. An algorithm to identify propensity to form Z-DNA indicated that DNA from Mycobacterium tuberculosis could form Z-DNA, a prediction confirmed by immunoassay. Together, these findings indicate that anti-Z-DNA antibodies can serve as probes for the presence of Z-DNA in DNA of various species origin and that the content of Z-DNA varies significantly among DNA sources.

Production and characterization of a human antisperm monoclonal antibody against CD52g for topical contraception in women.

BACKGROUND: Approximately 40% of human pregnancies are unintended, indicating a need for more acceptable effective contraception methods. New antibody production systems make it possible to manufacture reagent-grade human monoclonal antibodies (mAbs) for clinical use. We used the Nicotiana platform to produce a human antisperm mAb and tested its efficacy for on-demand topical contraception. METHODS: Heavy and light chain variable region DNA sequences of a human IgM antisperm antibody derived from an infertile woman were inserted with human IgG1 constant region sequences into an agrobacterium and transfected into Nicotiana benthamiana. The product, an IgG1 mAb ["Human Contraception Antibody" (HCA)], was purified on Protein A columns, and QC was performed using the LabChip GXII Touch protein characterization system and SEC-HPLC. HCA was tested for antigen specificity by immunofluorescence and western blot assays, antisperm activity by sperm agglutination and complement dependent sperm immobilization assays, and safety in a human vaginal tissue (EpiVaginal™) model. FINDINGS: HCA was obtained at concentrations ranging from 0.4 to 4 mg/ml and consisted of > 90% IgG monomers. The mAb specifically reacted with a glycan epitope on CD52g, a glycoprotein produced in the male reproductive tract and found in abundance on sperm. HCA potently agglutinated sperm under a variety of relevant physiological conditions at concentrations ≥ 6.25 µg/ml, and mediated complement-dependent sperm immobilization at concentrations ≥ 1 µg/ml. HCA and its immune complexes did not induce inflammation in EpiVaginal™ tissue. INTERPRETATION: HCA, an IgG1 mAb with potent sperm agglutination and immobilization activity and a good safety profile, is a promising candidate for female contraception. FUNDING: This research was supported by grants R01 HD095630 and P50HD096957 from the National Institutes of Health.