Sperm autoimmunity in vasectomized men and its relation to atherosclerotic coronary heart disease--report of the detection of circulating antisperm antibodies in 707 cases]

A study of sperm autoimmunization in 707 men and its relationship with atherosclerotic coronary heart disease (ACD) is reported. The following groups were studied: healthy vasectomized men (278), healthy nonvasectomized men (231), vasectomized men with ACD (987), and nonvasectomized men with AC (111). The gelatin agglutination test (GAT), tubeslide agglutination test (TSAT), and sperm immobilization test (SIT) were used to detect circulating antisperm antibodies. Sperm antibodies developed within 6 months postvasectomy and persisted for more than 20 years. There were no marked differences in the incidence of positive reaction and mean titer of antibodies between the healthy and AC vasectomized men. The results provide no evidence that the sperm autoimmunity after vasectomy will increase the incidence of coronary heart disease. (author's)     

输精管绝育者的精子免疫反应及其与冠心病的关系——707例精子循环抗体的测定

本文报告707例精子自身抗体的研究结果。其中健康男性输精管绝育者278例,未绝育者231例;男性冠心病人输精管绝育者87例,未绝育者111例。明胶凝集试验(GAT)、试管玻片凝集试验(TSAT)和精子制动试验(SIT)检测结果表明:精子抗体在手术后6月内就已产生,并持续20年以上。曾作输精管绝育术的健康男性和冠心病人二组比较,抗体的阳性率和滴度皆无差异显著性。没有证据表明输精管绝育术后的精子免疫反应会增加冠心病的发病率。     

Vasectomy and vasovasostomy in rhesus monkeys: the effect of circulating antisperm antibodies on fertility.

Rhesus monkeys develop agglutinating and complement-dependent antisperm antibodies after vasectomy. In order to study whether these antibodies affect fertility after vasovasostomy, 15 animals were given vasectomies and 6 months later vasovasostomies. Subsequently, each was mated with females of proven fertility. Five controls were given sham operations and similarly treated. During this period, each aimal was bled for serum to monitor the humoral immune response, ejaculated for semen analyses, and palpated for granuloma or fistula development. All control animals had a transient decrease in sperm density after sham vasectomy and vasovasostomy operations. The surgical procedures of vasectomy and subsequent vasovasostomy resulted in more animals having sperm of poor motility and quality. All of vasovasostomies were surgically successful in that sperm were again present in the ejaculate of each animal. The amount of sperm in the ejaculate could not be correlated with the ease of surgical procedure, presence or absence of macrophages in the ejaculum, motility, or forward progression. Only animals that had been vasectomized developed circulating antisperm antibodies. Sustained, elevated levels of antisperm antibodies most commonly occurred in monkeys that had high initial total sperm counts. Six of the experimental animals retained high levels of sperm-immobilizing antibodies after vasovasostomy. Of these, two were found to be infertile and two were classed as subfertile. Of the nine experimental animals without sustained antisperm antibody production, only one was classed as subfertile. This suggests that antisperm antibodies may in some cases impair the restoration of fertility after vasovasostomy.     

Immunological consequences of vasectomy and vasovasostomy in rhesus monkeys

The effects of circulating antibodies on fertility after vasovasostomy was studied in rhesus monkeys. After vasectomy there was rapid rise in antibody levels against spermatozoa; these reached a peak after 2 weeks, rather more rapidly than has been reported in man. Antibody levels declined after this period. Monkeys with high antibodies were also more likely to have granulomas and fistulas. After vasovasostomy several monkeys became fertile again with sperm counts in ejaculate reaching a high level at 3 months, although levels were lower than before vasectomy. Some males retained a high and sustained level of sperm immobilizing antibodies in plasma, sometimes for 18 months or more. These males produced fewer spermatozoa and were more likely to be subfertile or infertile after vasectomy. Almost all the monkeys without antibodies were fertile. Presence of antibodies at vasovasostomy was not a definite indication of subsequent infertility but did provide an indication of likelihood of continuing sterility. These are similar to observations of men undergoing vasectomy and vasosostomy.     

输精管结扎术后抗精子抗体在非射出精子上的存在

４８例正常育龄男子和２４例要求作输精管吻合术者，用免疫珠结合试验（ＩＢＴ）和精子－宫颈粘液接触试验（ＳＣＭＣ）分别检测其近睾端输精管液中的精子（非射出的精子）表面抗精子抗体的情况。结果显示：正常育龄男子非射出的精子上没有抗体存在；而输精管结扎术后非射出的精子上有抗精子抗体存在（ＩｇＧ：７９．４％，ＩｇＡ：３８．２％，ＳＣＭＣ：３５．５％）。这些抗体在输精管结扎术后不到１年就可发现，并保持到结扎术后３年以上。吻合术后１～３个月内，其精液中射出的精子上仍发现抗体存在。结果还提示：输精管结扎术后，不能以非射出的精子上抗体的水平去预测吻合术后射出的精子上抗体的水平；吻合术前血清抗体水平与非射出的精子上抗体水平并不一致。     

Occurrence of serum antisperm antibodies in patients with cystic fibrosis

OBJECTIVE: To determine if acquired obstruction of the vas deferens in men with cystic fibrosis (CF) induced the development of antisperm antibodies with genital tract obstruction similar to other men. DESIGN: Serum antisperm antibodies were assayed by an indirect immunobead test and an indirect immunofluorescence assay. Both homologous (human sperm/human zona) and heterologous (human sperm/zona-free hamster ova) sperm/egg interactions were evaluated in the presence of serum antisperm antibodies from patients with CF. SETTING: Cystic Fibrosis Clinic at the University of Oklahoma Health Sciences Center, a tertiary care referral center. PATIENTS: Fifteen CF patients (10 male and 5 female), 3 non-CF antisperm antibody-positive infertile patients (2 male and 1 female), 20 fertile controls (7 males and 13 females), and 9 fertile sperm donors were used. INTERVENTIONS: None. MAIN OUTCOME MEASURES: Serum antisperm antibody levels in patients with CF. In those patients with antisperm antibodies, determine effect of these sperm antibodies on sperm/egg interactions and complement-mediated events. RESULTS: Sera from 3 (30%) of 10 men with CF demonstrated immunoglobulin (Ig)G, IgA, and/or IgM antisperm antibodies, whereas sera from all 5 CF women and the 20 control sera were negative for antisperm antibodies. The maximal titers for IgG, IgA, and IgM antisperm antibody were 1:8, 192, 1:256, and 1:64, respectively. The immunobead binding, which was restricted to the sperm head and tail-tip or the midpiece and tail-tip, correlated with the indirect immunofluorescence pattern. Antisperm antibody-positive sera from men with CF impaired both the binding and penetration of human zonae and the penetration of hamster ova by human sperm. CONCLUSIONS: Similar to other men with congenital or acquired obstruction of their genital tract, antisperm antibodies may occur in some men with CF. Antisperm antibodies may contribute to immune sperm dysfunction in some men with CF by activated complement-mediated events and interfering with sperm/egg interactions.     

Human sperm protein encyclopedia and alloantigen index: mining novel allo-antigens using sera from ASA-positive infertile patients and vasectomized men

Anti-sperm antibodies (ASA) are an important cause of immunological infertility. The objective of this study was to identify immunodominant sperm antigens recognized by anti-sperm antibodies (ASA) in serum samples of infertile men, women and vasectomized men. High-resolution two-dimensional gel electrophoresis was employed to separate human sperm proteins using isoelectric focusing (IEF) or nonequilibrium pH gradient electrophoresis (NEPHGE), followed by PAGE and Western blotting. Serum samples from five infertile male and five infertile female subjects that contained ASA as assayed by the immunobead binding test (IBT), were analyzed by Western blotting using NEPHGE gels followed by enhanced chemiluminescence (ECL) to identify the basic sperm antigens reactive to the sera. Serum samples from five fertile male and five fertile female subjects that were ASA-negative by IBT were used as controls. Serum samples from six vasectomized men collected before vasectomy and at different time intervals until 6 months after vasectomy were analyzed by Western blotting using IEF gels. The ECL blots were analyzed to compare immunoreactivity between serum samples from fertile and infertile subjects and identify antigens unique to sera of the infertile subjects. Similarly, immunoreactivity between serum samples from pre- and post-vasectomy was compared to identify antigens unique to sera collected following vasectomy. Five allo-antigenic basic protein spots were recognized by sera from infertile males but not from fertile subjects. Five sperm iso-antigenic basic spots were recognized by infertile female subjects. Two among six of the vasectomized men's sera showed a difference in the Western blot profile 6 months after vasectomy, recognizing at least one new protein spot in each case when compared to pre-vasectomy sera. The acrosomal protein SP-10 was identified as an alloantigen recognized by a post-vasectomy serum. Molecular identities of the known allo- and iso-antigens identified in this study and in previous studies from this laboratory are reviewed and discussed.     

The clinical relevance of classes of immunoglobulins on spermatozoa from infertile and vasovasostomized males

Following reversal of vasectomy, conceptions occur even when antisperm antibodies are present in the seminal plasma, but this is most unusual in men with similar titers of such antibodies who are spontaneously infertile. To clarify the differences between antisperm antibodies occurring in infertile men and those associated with vasectomy reversal, we have studied 23 spontaneously infertile men and 22 men who underwent vasectomy reversal, all of whom had antisperm antibodies detected in seminal plasma by the same tray agglutination test. The class of antibody on spermatozoa was defined by a double-antibody technique using diluted rabbit anti-human IgG, IgM, or IgA or secretory component, followed, after washing, by 125I-labeled donkey anti-rabbit Ig. The results have shown that similar amounts of IgG and IgM were present on the spermatozoa, but infertile men had significantly more IgA and especially more secretory component than men who underwent vasectomy reversal. This was associated with significantly greater impairment of penetration of cervical mucus in the former group. It appears that the type of antibody on the spermatozoa may vary according to the stimulus for its production.     

Low fertility rate in vasovasostomized males and its possible immunologic mechanism.

Contradictory views have been expressed about the role of the various antisperm antibodies which develop after vasoligation. The present study was conducted in 50 normal fertile males, 50 vasectomized subjects and 25 subjects after recanalization of their vas deferens in order to investigate the development of various anti-sperm antibodies after vasectomy, along with their incidence, their persistence after successful relief of vaso-obstruction by vasovasostomy and their role in the causation of infertility in vasoanatomized normospermia males. Sperm agglutinating, immobilizing and haemagglutinating antibodies showed rises in titres with increase during the post-vasectomy period, indicating continuous antigenic stimulus. Age, post-operative complications and blood group did not seem to alter the results. 86% of subjects developed antisperm agglutinins, mostly tail-to-tail type (54.5%), 1-12 years after vasoligation, while only 2% of fertile men had circulating spermagglutinins. A lower incidence of positive sperm in the immobilization test than in the agglutination test suggests either that different antibodies are detected by these two tests or these tests have differing sensitivities. Of the 25 vasovasostomized subject, 13 (52%) cases became normospermic and 4 (16%) oligospermic while 8 (32%) remained azoospermics. Except for 3 oligospermic subjects, all had circulating spermagglutinins. Among the 13 normospermic vasovasostomized persons, a significant correlation was found between the titres of circulating antisperm agglutinins and autoagglutination of spermatozoa in their ejaculates; and also between the sperm immobilization values of their sera and the degree of their sperm motility. Three normospermic recanalized men, having low levels of sperm agglutinins and haemagglutinins with normal seminogram and no sperm immobilizing antibody, successfully impregnated their wives. Another 10 vasovasotomized infertile subjects had sperm agglutinins in significant titre; 5 showed positive sperm immobilization values, a similar number showed autoagglutination of sperm, while a decreased degree of motility of sperms was noted in 6 cases. Thus there was a significant correlation between the titres of anti-sperm antibodies and autoagglutination of spermatozoa, which might be an important cause of male infertility after successful anatomic relief of vasoobstruction. Histological studies of testicular biopsy showed normal spermatogenesis in azoospermic recanalized subjects, although they had high levels of antisperm antibodies. This suggests that these antibodies do not affect normal spermatogenesis, and sperm counts.     

Reconstruction of humoral response to sperm antigens in scid mice

OBJECTIVE AND DESIGN: Production of specific human antisperm antibodies by using human-SCID mice model with deposited peripheral blood lymphocytes. MATERIALS AND METHODS: Human peripheral blood lymphocytes (PBL's; CD8(+)-negative cell fraction) were grafted to the peritoneal cavity of severely-combined immunodeficient (SCID) mice at concentration of 20-35 x 10(6) cells per mouse. Lymphocytes were obtained from non-sensitized individual (to sperm antigen) and from in vivo primed males (vasectomized). Two sets of experiments were carried out, with 'native' (glycosylated) and enzymatically deglycosylated sperm antigenic extracts. In all applied variants, sperm antigens were administered with Complete and then with Incomplete Freund adjuvant to improve an immune response. RESULTS AND CONCLUSION: This approach allowed us to obtain better pronounced humoral antisperm response, specific to sperm deglycosylated antigens when PBL's were obtained from individuals in vivo sensitized to sperm (after vasectomy).     

Reduced T cell reactivity in vasectomized rhesus monkeys: association with histocompatibility type.

The capacity of peripheral lymphocytes from rhesus monkeys which had been vasectomized for 7 or 11 years to stimulate and respond to normal rhesus lymphocytes in mixed lymphocyte cultures (MLC) was tested to determine whether vasectomy affects immunologic reactivity. The ability to respond in MLC, a T cell function, was significantly reduced in the 11-year vasectomized animals and in two 7-year vasectomized animals. The ability to stimulate in MLC, a B cell function, was significantly increased in the 11-year vasectomized group. MLC reactivity of normal lymphocytes cultured in plasma from vasectomized animals and lymphocytes from vasectomized animals cultured in normal plasma was not altered, ruling out serum effects in the reduction of MLC responsiveness in these vasectomized animals. Seventy-five per cent of the vasectomized animals with markedly reduced MLC reactivity had the RhL-A determinates 19 and 24, indicating an association between the tendency toward reduced MLC reactivity after vasectomy and histocompatibility type.     

Consequences of autoimmunity to sperm antigens in vasectomized men.

The development by a large percentage of vasectomized men of sperm autoantibodies is discussed in this monograph chapter. The production of anti-sperm antibodies is attributed to: 1) granuloma formation; 2) increased permeability of epithelial barriers in the rete testis and epididymis; and 3) transport of phagocytic cells to regional lymph nodes. Individual variation in type of antibodies and response to antibody production is documented and is thought to depend on such factors as rate of sperm production, the structure of the blood-testis barrier, surgical technique, and expression of immune response genes. Morphological changes in spermatozoa and testes occurring after vasectomy may be induced by immunological mechanisms. Tests of cell-mediated immunity to sperm antigens are described, and more accurate tests are needed. Animal studies provide evidence that chronic immune stimulation can result in formation of circulating immune complexes, resulting in deleterious systemic effects, including damage to kidneys, blood vessels, and cells of the immune system. Further studies of the autoimmune orchitis phenomenon may aid in avoidance of immunologically mediated side effects of vasectomy.     

Cell-mediated immunity in vasectomized rhesus monkeys.

Cell-mediated immunity in rhesus monkeys that had been vasectomized for 2, 4, 7, or 11 years was measured by lymphocyte blastogenesis following stimulation with concanavalin A, phytohemagglutinin (PHA), and pokeweed mitogens. Several of the 7- and 11-year vasectomized animals had significantly reduced PHA reactivity when compared with control animals, and the percentage of animals with reduced PHA reactivity increased with time after vasectomy.     

Macrophages and infertility: enhancement of human macrophage-mediated sperm killing by antisperm antibodies

The mechanism by which antisperm antibodies inhibit fertility is not completely understood. Macrophages may play a role in mediating infertility by interacting with sperm and destroying gametes. Experiments were conducted evaluating the effect of antisperm antibody on the phagocytosis and lysis of sperm by human peritoneal macrophages in vitro. Sperm from a fertile man treated with sera from normal men and women or medium alone had 5 to 280 molecules of IgG/sperm, as determined by a 125I-labeled anti-human IgG monoclonal antibody assay. By contrast, sperm treated with sera containing antisperm antibodies had 310 to 1240 molecules of IgG/sperm. Peritoneal macrophages harvested from infertile women with tubal/adhesive problems mediated phagocytosis and lysis of 111In-labeled sperm which was enhanced by treatment of the sperm with sera containing antisperm antibodies (39.0% +/- 1.5% versus 76.3% +/- 3.2% phagocytosis, and 3.3% +/- 0.3% versus 23.3% +/- 2.3% lysis of sperm [control versus antibody-treated]). The likelihood of fertilization in couples with antisperm antibody may be determined not only by the antibody but also by the presence of genital tract macrophages capable of destroying the antibody-coated sperm.     

Biphasic production of antisperm autoantibodies follow vasectomy of the Lewis rat

Temporal changes in the specificity of post-vasectomy autoantibodies to SDS-PAGE separated sperm antigens were investigated in Lewis rats. Sera were obtained from nine vasectomized animals prior to vasectomy, every two weeks for 14 weeks, and less frequently thereafter, up to 41 weeks. Changes in antisperm autoantibodies over time were assessed by ELISA and western blot assay and compared to antisperm isoantiserum and normal Lewis rat serum. A "biphasic" pattern of autoantibody production over time was observed in a majority of individuals. This pattern was characterized by early phase autoantibodies, produced between 0 and 6 weeks after vasectomy, which bound antigens at the stacking, separating and ionic fronts and by late phase autoantibodies, produced after 4 weeks following vasectomy which bound antigens at 86, 63, 52, 43, 31 and 26 kDa. Previous work suggested that some high molecular weight autoantigens were disulfide-bonded polymers of the polypeptides at 86, 63, and 43 kba (Handley, et al., 1988). Indirect immunofluorescence with monospecific isoantisera to the 86 kDa autoantigen suggested that its corresponding high molecular weight polymer was located in the tail of cauda epididymal spermatozoa. This polymer possessed several characteristics of T cell independent autoantigens. These data show a change in the specificity of autoantibodies produced over time after vasectomy which may reflect a shift from T cell independent to T cell dependent autoantibody production by the Lewis rat.     

Immunological approach to male contraception

Sperm surface antigens may induce an immune response in mammals. In humans, the presence of antisperm antibodies has been noted in the blood, seminal plasma, cervical mucus, and follicular liquid. Because they provoke immobilization and/or agglutination of sperm and a diminution of the rate of fertilization in vitro, these antibodies are believed to be a factor in some unexplained infertility, as for example after surgical reversal of vasectomy. These observations have led to research oriented toward development of a nonhormonal contraceptive method based on the immunological capacities of some sperm antigens. Possible secondary effects and the modes of action remain poorly understood. Antisperm immunoglobulins in the male should be induced by epitopes that are specific to sperm excluding the proteins of the sperm membrane, so that they will be without effect on functions other than fertilization. The epitopes should be located on the sperm surface and should be of post-testicular origin because of the risk of orchitis posed by testicular antigens. The antisperm immunoglobulins should also be present in the area surrounding he sperm in the male genital tract because of the limited permeability of the tract to immunoglobulins and other large molecules. The antisperm immunoglobulins should not be inhibited in the male by the immunosuppressive activities of the seminal plasma or in the female by capacitation. In vivo and in vitro observations indicate that various functions may be altered by antisperm antibodies. The antibodies may immobilize the sperm, block interaction with the oocyte at the level of the zona pellucida, block adherence to the vitelline membrane, or cause anomalies in embryonic development. In rats, isoimmunization against epididymal cofactor of the sperm zona pellucida receptor results in reduced sperm mobility and reduced fertility. The same study was conducted in 12 rams selected for homogeneity of sperm characteristics. Isoimmunization of rams induced transient asthenospermia. Most sperm antigens involved in fertilization are poorly understood. Observations in the ram suggest that embryonic mortality following isoimmunization should be studied.     

Reduced levels of T suppressor/cytotoxic lymphocytes in semen from vasovasostomized men: relationship to sperm autoantibodies

The regulation of lymphocyte populations in semen and their function remains unknown. Using monoclonal antibodies and immunohistochemical techniques we compared the levels of mononuclear cells in ejaculates obtained from 14 men with intact vas deferens and 13 men who had undergone both a vasectomy and a microsurgical vasovasostomy. The presence of antisperm antibodies was also assessed by the immunobead binding assay. Semen from both groups contained about 10(3) T and B lymphocytes and monocytes. In men with intact vasa, T suppressor/cytotoxic cells predominated. In contrast, in vasovasostomized men the levels of T suppressor/cytotoxic cells were significantly reduced (P less than 0.005) and T helper/inducer cells predominated in their semen. Concentrations of seminal T helper/inducer lymphocytes, B lymphocytes and monocytes were similar in both groups. Antisperm antibodies were detected on sperm, in seminal fluid and/or in serum of all the vasovasovasostomy patients but in none of the controls, T suppressor/cytotoxic cells may limit the immune response to sperm within the male reproductive tract. Chronic obstruction resulting in damage to the integrity of the excurrent ducts may induce alterations in T cell regulation leading to a decrease in T suppressor/cytotoxic cells and create conditions permissive for the formation of autoantibodies to sperm-specific antigens.     

Increased frequency of autoantibodies in men with sperm antibodies

Autoantibodies to thyroid microsomes were more frequent in 102 infertile men with complement-dependent sperm-immobilizing activity (sperm immobilization test [SIT] ) in serum (11.8%) than in a control group of 277 men of comparable ages and semen quality without sperm antibodies (4.3%, P less than 0.05). Frequencies of organ-specific antibodies (antigastric parietal cell, antithyroglobulin, and antithyroid microsome) in 57 men with genital tract obstructions and positive SIT were similar to those for control subjects, and there were no significant differences in the frequencies of non-organ-specific autoantibodies (antinuclear antibody, rheumatoid factor, antimitochondrial, and anti-smooth muscle) in the three groups. Because in men without genital tract obstruction antithyroid microsomal autoantibodies were more common with sperm antibodies than without, the possibility of a genetic factor in the causation of sperm autoimmunity should be considered.     

Infertility in mice after unilateral vasectomy

The effects of unilateral vasectomy upon fertility and antisperm antibody production were studied using DBA/1J mice. Thirty-six males underwent either unilateral vasectomy, unilateral orchiectomy, or sham surgery. In vivo effects upon fertility were monitored by matings. Antisperm antibody titers were measured monthly. In vitro fertilization was performed in the presence of serum obtained 4 months postoperatively, and serum testosterone levels were also determined. After 3 months, only 1 male in the vasectomy group induced a pregnancy (1 of 12), while all but 1 of the males in the two control groups induced a pregnancy (20 of 21). The geometric mean antisperm antibody titer was 1:169 in the vasectomy group, while the orchiectomy and sham surgery groups had titers of 1:4 and 1:14, respectively (P less than 0.0001). The percentage of eggs fertilized in vitro in the presence of serum from experimental mice fell from 80% in the two control groups to 40% in the unilateral vasectomy group. Unilateral vasectomy induced infertility in DBA/1J mice and an antisperm antibody response. Sera containing these antibodies inhibited in vitro fertilization. This suggests that infertility after unilateral vasectomy may be immunologically mediated by antisperm antibodies.