Assessment of antisperm antibodies in a sample of Egyptian patients with hepatitis C virus infection

Association of hepatitis C virus (HCV) with autoimmune phenomena and impaired semen parameters has been previously reported. The aim of this study was to investigate the influence of HCV infection on the development of antisperm antibodies (ASAs) in HCV‐positive males. The study was conducted on 30 HCV‐infected individuals and 30 healthy control subjects. In both patients and control groups, liver enzymes and reproductive hormones were measured; computer‐assisted semen analysis (CASA) was performed; HCV‐RNA in serum was measured and IgG and IgA ASAs in semen were determined. Free testosterone, sperm concentration, progressive and total motility were significantly lower in HCV patients than in the control group, whereas ASAs of the IgG and IgA classes were significantly higher in HCV patients. However, correlations between viral load and the examined semen parameters and ASAs were nonsignificant. In conclusion, HCV may be responsible for the increased ASAs detected in HCV patients in the present study, possibly providing another plausible explanation for the decreased sperm motility reported in HCV patients. These findings could be of value in fertility management of HCV patients.     

Sperm antigens recognized by antisperm antibodies present in sera of infertile adults and prepubertal boys with testicular failure

Immunoblotting of a repertoire of sperm antigens reacting with antisperm antibodies present in sera of infertile adults and prepubertal boys with testicular failure was performed. In the subgroups selected for this study, 55% of examined infertile women, 65% of infertile men and 64% of prepubertal boys with gonadal failure gave positive results by Western blotting with extracted sperm antigens. Sperm antigens with molecular weights of 57, 58, 62, 63 and 66 kDa were the most immunodominant entities recognized by antisperm antibodies from prepubertal boys. No positive reactions were detected by Western blotting in a control population of fertile adults, whereas in a group of prepubertal healthy boys only one sample revealed reactivity against sperm antigens of 58 and 70 kDa.     

In vitro effects of nicotine on human spermatozoa

Washed human spermatozoa from 12 normozoospermic donors were treated with different concentrations of nicotine 0.1, 1.0, 5.0 and 10.0 mm and were compared to spermatozoa suspended in nutrient medium only (control). Computer‐aided sperm analysis was used to assess sperm kinematic properties after 30, 60, 120 and 180 min of incubation. Viability was assessed by means of a dye exclusion staining technique (eosin/nigrosin), while acrosome‐reacted cells were identified under a fluorescent microscope using fluorescein isothiocyanate–Pisum sativum agglutinin as a probe. Nicotine significantly reduced total motility, progressive motility, curvilinear velocity, amplitude of lateral head displacement, beat cross‐frequency, viability and caused spontaneous acrosome reaction at concentrations of ≥5.0 mm after 2 and 3 h of exposure. Nicotine concentrations of 0.1 and 1.0 mm had no significant effect (P < 0.05) on spermatozoa except that 1.0 mm significantly decreased (P < 0.05) sperm progressive motility at 2 and 3 h of incubation as well as viability after 3 h of incubation. This study concludes that the occurrence of high levels of nicotine in the body and seminal fluid might adversely affect fertilisation capacity of human spermatozoa through a mechanism that involves decreased motility, viability and premature induction of the acrosome reaction.     

A new method for isolation of human antisperm antibodies

Summary. A study was undertaken to isolate pure human antisperm antibodies from the sera of infertile couples. One hundred infertile couples attending the Infertility and IVF Unit (Beilinson Medical Center) because of unexplained infertility were tested (both partners) for antisperm antibodies. Sixty-eight experiments were performed with positive sera containing antisperm antibodies and normal donor sperm. These experiments were followed by experiments in order to elute pure human antisperm antibodies from the sperm surface. Three experiments were performed with human sperm which were found to be coated by antisperm antibodies, in order to directly elute these antibodies from the sperm surface. In all experiments we eluted antisperm antibodies of the IgG and IgA isotypes from the sperm surface. These antibodies were demonstrated in the eluate, in each case by either the indirect immunobead test, the radial immune diffusion assay, or the electrophoresis method. Control experiments were performed as follows: (i) normal donor sperm incubated with normal serum; (ii) normal donor sperm without serum incubation; (iii) normal donor lymphocytes incubated with serum containing antisperm antibodies; (iv) normal donor lymphocytes without serum incubation. No antisperm antibodies were obtained in any of these control experiments. Absorption and elution experiments can be used for the isolation of pure human antisperm antibodies, which may then be used for the production of anti-idiotypic antibodies to antisperm antibodies. The anti-idiotypic antibodies could be further utilized as antigen substitutes for the production of a contraceptive vaccine and/or for application in the treatment of spontaneous abortion and infertility.     

Antisperm antibodies and sperm motility: a study using timed exposure photomicrography

This study examined the effect of antisperm antibodies on sperm motility. Antisperm antibodies present in seminal plasmas with different sperm agglutinating titres were transferred passively to normal donor sperm, and the effects on movement characteristics (velocity of forward progression, amplitude of lateral head displacement, percentage progressive motility and percentage non-progressive motility) were analysed using timed exposure photomicrography. There was no significant association between sperm movement characteristics and the presence of titre of antisperm antibodies in seminal plasma. Furthermore, no differences were detected between those samples that possessed sperm agglutinating versus sperm immobilizing activity. These findings do not support the common belief that antisperm antibodies are a cause of poor sperm motility in semen.     

Flow cytometric determination of binding of monoclonal antibodies to human spermatozoa

Summary. Although several monoclonal antibodies to sperm antigens are available, only few systematical reports on the appearance of distinct antibody binding structures in spermatozoa of infertile patients are available. We studied the binding frequency of six monoclonal antibodies (TÜS-1, -2, -10, -17, -19, -20) by means of a flow cytometer in different semen samples and their possible relationship to other sperm parameters, in particular to the motility parameters as determined by computer-assisted semen analysis. The percentage of vital spermatozoa binding the different antibodies was 21.5% (TÜS-2) to 52.1% (TÜS-20). The percentage binding was higher in samples with normozoospermia than in those with oligo-asthenoteratozoospermia. A significant correlation occurred between sperm concentration and the percentage of spermatozoa stained by TÜS-2 and TÜS-17; between motile spermatozoa and those binding TÜS-17 and TÜS-20; the velocity average path and those binding TÜS-17; an inverse correlation occurred between the morphologically-abnormal spermatozoa and those binding TÜS-20; an inverse correlation between TÜS-17 and acrosin. We conclude from our results that although the antigens of the antibodies concerned are not yet characterized, the determination of antibody binding will give additional information on the functional status during capacitation and acrosome reaction of spermatozoa.     

不育相关AsAb的免疫印迹分析

用精子膜抗原Ｗｅｓｔｅｒｎｂｌｏｔ分析法，对７８例生育、１９８例不孕、２２例流产者及４４例中晚期妊娠孕妇血清抗精子抗体（ａｎｔｉｓｐｅｒｍａｎｔｉｂｏｄｉｅｓ，ＡｓＡｂ）进行研究。结果发现，抗分子量８５０００、５１０００、３００００精子抗原的ＡｓＡｂ与不孕相关；抗分子量８５０００、７９０００、６９０００，５１０００精子抗原的ＡｓＡｂ与女性不孕相关；不孕女性ＡｓＡｂ种类（在Ｗｅｓｔｅｒｎｂｌｏｔ上的反应条带数）明显多于生育女性。抗分子量８５０００、７９０００、５８０００精子抗原的ＡｓＡｂ与女性ＲＳＡ相关。本研究未发现不孕组男性、流产组男性及中晚期妊娠孕妇各种分子量精子抗原的ＡｓＡｂ检出率与生育组同性别相比有显著不同。所发现的这些ＡｓＡｂ可能与不育有密切关系，需要进一步研究和证实。     

精浆抗精子抗体阳性患者精子膜脂流动性变化的研究

为探讨抗精子抗体（ Ａｓ Ａｂ） 对男性不育患者的可能作用机制,用荧光偏振法测定了１８ 例精浆 Ａｓ Ａｂ 阳性患者与２０ 例 Ａｓ Ａｂ 阴性者的精子膜脂流动性（ Ｌ Ｆ Ｕ） 。结果表明, Ａｓ Ａｂ 阳性组 Ｌ Ｆ Ｕ 与对照组比较明显降低,（ Ｐ＜ ０ ．０１） ,提示对精子 Ｌ Ｆ Ｕ 的影响可能是 Ａｓ Ａｂ 引起免疫性不育的机制之一。     

Effect of isotypes of antisperm antibodies on semen quality

A direct immunobead test (IBT) was performed on 233 men who attended an immunological centre. Thirty-four (14.6%) of these men were found to be positive (greater than 20% binding) for antisperm antibodies (ASA). IgA, IgG and IgM were the most common sperm-associated immunoglobulins. In 50% of men with ASA asthenozoospermia, teratozoospermia, leukocytospermia or hypofunction of the seminal vesicles was observed. Semen parameters were altered most frequently when IgM was present in association with IgA and/or IgG. This suggests that there is an active inflammatory process in the reproductive tract, as evidenced by leukocytospermia, and this could be responsible for the abnormal semen parameters. ASA generation could be a consequence of this process rather than being the cause of the abnormal semen quality. If ASA do affect fertility, this could take place in the female reproductive tract.     

Pentoxifylline acts synergistically with A23187 to increase the penetration of zona-free hamster oocytes by cryopreserved human spermatozoa

The number of cryopreserved human spermatozoa which penetrated zona-free hamster oocytes afier stimulation with 2μmol A23187 per litre was increased by the hrther addition of 0.6 or 3.6 mmol pentoxifjrlline per litre. With spermatozoa prepared by washing by repeated centrifugation, the median numbers of sperm headd/egg were 1.9, 7.9 and 10.8 in the presence of 0, 0.6 or 3.6 mmol pentoxifylline per litre, respectively. A similar effect was observed with spermatozoa prepared on a Percoll gradient. As A23187 inhibited sperm motility, and this was exacerbated by pentoxifylline, the increased penetration rate of hamster oocytes cannot be explained by improved sperm motility. The number of spermatozoa stimulated to acrosome react by 2 μmol A23187 per litre was increased 3-fold by 3.6 mmol pentoxifylline per litre and 4-fold by 5 mmol caffeine per litre. These data suggest that CAMP may act synergistically with Ca²⁺ to stimulate the acrosome reaction. Pentoxifjrlline may improve the fertility of poor-quality human spermatozoa by enhancing their ability to respond to the Ca²⁺ signal produced by binding to the zona pellucida.     

Proteomic identification of sperm antigens using serum samples from individuals with and without antisperm antibodies

The aim of the study was to identify human sperm antigens reacting with polyclonal antisperm antibodies. Protein sperm extracts were subjected to electrofocusing, and next immune reactions (immunoblotting) were carried out with positive for antisperm antibodies and control (not containing antisperm antibodies) serum samples. Proteomic analysis of human sperm proteins resulted in identification of 80 sperm antigens that could be divided into three groups: antigens specific for patients with antisperm antibodies (32), antigens recognised by both infertile patients and control sera (35) and antigens detected by control serum samples only (13). Among antigens specific for infertile patients, there were 12 sperm entities known to be involved in fertilisation process. We have also characterised three protein entities identified only by sera of infertile women. Altogether, the proteomic analysis resulted in identification of 27 sperm entities not reported previously in human sperm proteome. Identified proteins are sperm antigens that could be potentially responsible for immunological infertility. The study also sheds new light on the sperm antigens in aspect of gender specificity. The investigation of human sperm proteome by the use of antisperm antibodies‐containing sera of infertile individuals not only may indicate new proteins but also can draft their immunological nature.     

Prevalence of antisperm antibodies by SpermMARtest in subjects undergoing a routine sperm analysis for infertility

To evaluate the prevalence of antisperm antibodies (ASA) attached to the sperm plasma membrane in male partners of infertile couples, the binding of latex particles to spermatozoa was investigated using SpermMARtest, included routinely in semen analysis. A total of 860 men were examined, who were referred consecutively for semen analysis. Of these, 750 men were referred because of infertility (0.6–10 years in duration) whereas 110 were volunteers with a history of previous fertility. Samples were assessed by the SpermMARtest kit using latex particles sensitized with human IgG. Sperm-latex binding was read after 3 min and samples scored as negative, positive or highly positive when < 10, > 10–40, or >40% binding occurred, respectively. Of the samples 132 (17.3%) were excluded because of azoo- or severe oligo-asthenozoospermia. IgG attached to spermatozoa were detected in nearly 13% of semen samples from the infertile population and in one of 110 fertile men (0.90/,). From the infertile group, 6.2% of samples showed > 40% binding, and 6.7% intermediate binding, with an overall ASA prevalence of 12.9% in subjects undergoing semen analysis for infertility.     

The incidence and influence upon fertility of antisperm antibodies in seminal fluid following vasectomy reversal

Seminal plasma samples from men undergoing vasovasostomy were analysed for antisperm antibodies using the indirect immunobead test. A pre-operative assessment showed antisperm antibodies of either IgA or IgG class to be present in 9/27 (33.3%) men. A significant increase (P less than 0.05) in the post-operative incidence of the antibodies was seen in the men who achieved patency (27/45, 60%) but not in those men for whom no sperm were seen in the ejaculate (4/10, 40%). After follow-up for a minimum of 1 year, conception rates for couples in which the male partner had achieved patency were similar in the groups with no antibodies detected post-operatively (12/18, 66.7%) or with IgA alone (2/3, 66.7%), but was reduced significantly in the presence of IgG (1/9, 11.1%; P less than 0.05) or IgA + IgG (3/15, 20.0%; P less than 0.01).     

Sperm antigens and reactivity of antisperm monoclonal antibodies in elisa

Summary. Several types of sperm antigenic suspensions as well as the whole sperm, either methanol-fixed or air-dried, were checked for intensity of binding to monoclonal antisperm antibodies with known characteristics of reactivity to sperm. The activity of sperm antigen—antibody binding was measured by elisa (enzyme-linked immunosorbent assay) and compared in several variations (parallely run) of the assay where different types of sperm antigen preparations were applied. The obtained results were then evaluated for statistical significance in Wilcox test. It was shown that antibody reactivity was markedly higher in experiments where the whole sperm was coated in a solid-phase in comparison to results obtained with adhered different sperm antigenic suspensions. However, one exception was noted, where the results from elisa, run with sperm organic extract, were (statistically) insignificantly lower than those obtained with the whole sperm. Therefore, organic sperm extracts (containing mostly glycolipids) can be a valuable alternative to screening for antisperm antibody activity and/or infertility background.     

In vitro effect of gold and silver nanoparticles on human spermatozoa

The cytotoxicity of Au/Ag nanoparticles (NPs) on human spermatozoa was investigated in vitro. Semen from donors were incubated (37 °C, 60′–120′) with 30, 60, 125, 250 and 500 μM Au/Ag‐NPs. Sperm motility was evaluated following WHO guidelines; sperm viability was assessed with eosin Y test. Au‐NPs were characterised and localised with field emission gun‐based scanning transmission electron microscope/energy dispersive spectroscopy and transmission electron microscopy. Both tested NPs exerted a significant dose‐dependent effect on motility and viability of human spermatozoa (P < 0.001). Ag‐NPs seem to show a slightly elevated toxicity although not significant (P > 0.05). Au‐NPs were localised in spermatozoa, whereas Ag‐NPs were undetectable. In conclusion, Au‐NPs and Ag‐NPs do not appear to be harmful for human spermatozoa up to high concentrations (250–500 μM) that are probably difficult to reach in vivo. It is mandatory to explore the genotoxic effect of NPs in germ cells.     

两种蛋白质对精子冷冻保护的影响

本文对蛋黄及白蛋白两种蛋白质在精子冷冻保护中的作用进行了比较，结果显示：尽管两种蛋白质对精子冷冻复苏后的活动率的影响没有明显的差异性，但在Ａ２３１８７诱导，ＦＩＴＣ－ＰＳＡ染色的精子顶体反应测试中，蛋黄显示出相对较好的保护作用；在临床人工授精效果的观察中，使用蛋黄保护剂的精子呈现了较高的怀孕率。