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Association between recurrent spontaneous abortions and circulating IgG antibodies to sperm tails in women
Journal of Reproductive Immunology, 15 (1989) 151-- 158
151
Elsevier Scientific Publishers Ireland Ltd.
JRI 00594
Association between recurrent spontaneous abortions and circulating IgG antibodies to sperm tails in women S t e v e n S. W i t k i n a n d A n u C h a u d h r y Immunology Division, Department of Obstetrics and Gynecology, Cornell University Medical College, New York (U.S.A.) (Accepted for publication 15 March 1989)
Summary The isotype and regional specificity of antisperm antibodies in the circulation of women with recurrent spontaneous abortions was examined. There was a statistically significant association (P < 0.005) between the presence of IgG tail-directed antisperm antibodies and a history of unexplained recurrent spontaneous abortion. These antibodies were detected in 36.4% of 44 women with recurrent abortions and 14.6°70 o f 616 female partners of infertile marriages. In contrast, no differences in IgG sperm head-directed antibodies or in IgA and IgM antisperm antibodies were observed between the two groups. Husbands of women in the miscarriage or infertile groups had similar semen evaluations. Antisperm antibodies may be a marker for defective immunosuppression in women with recurrent miscarriages. Alternatively, exposure of sperm-sensitized pregnant women to sperm may activate the maternal immune system to respond to paternal antigens present on the embryo. Key
words:
antisperm
antibodies;
recurrent spontaneous
abortion;
infertility.
Correspondence to: Dr. Steven S. Witkin, Cornell University Medical College, Department of Obstetrics & Gynecology, 515 East 71st Street New York, New York 10021, U.S.A. 0165-0378/89/$03.50 © 1989 Elsevier Scientific Publishers Ireland Ltd. Published and Printed in Ireland
152
Introduction
Antisperm antibodies in women's sera reduce the probability that deposited sperm will reach the oviduct a n d / o r fertilize the ovum (Bronson et al., 1984; Witkin and David, 1988). However, the role, if any, of antisperm antibodies in post-fertilization pregnancy loss is still unclear. Female rabbits and cattle, after being immunized with sperm, have an increased incidence of spontaneous abortion in subsequent pregnancies (Menge, 1970). Both polyclonal (O'Rand, 1977) and monoclonal (O'Rand et al., 1984) antibodies to rabbit sperm react with antigens present on pre-implanatation embryos. In two studies, approximately 50°70 of women with a history of early recurrent spontaneous abortions were positive for circulating sperm agglutinating or immobilizing antibodies (Jones, 1976) or cytotoxic or hemagglutinating antibodies (Mathur et ai., 1987). In another investigation, however, no relation between a history of one to four spontaneous miscarriages and the presence of sperm agglutinating antibodies could be demonstrated (Ingerslev and Ingerslev, 1980). Haas et al. (1986) also noted an association between antisperm antibodies in women and a poor pregnancy prognosis. In their patients, the incidence of sperm immobilizing antibodies but not sperm agglutinating antibodies or antibodies to sperm extracts detected by radioimmunoassay, was greater in recurrent aborters than in infertile women. In the most recent study, antisperm antibodies detected by enzyme-linked i m m u n o s o r b a n t assay were identified in 7 of 16 women (43070) with early stage miscarriages (Witkin and David, 1988). The present study was initiated to evaluate and compare the incidence and regional specificity of antisperm antibodies in infertile women and women with a history of recurrent spontaneous abortions. Sperm antibody detection was by the i m m u n o b e a d test (IBT), which allows the determination of both the isotype and the location on the sperm of any antisperm antibodies detected. Moreover, the IBT is specific for antibodies reactive with the surface of living motile sperm. This assures that antibodies reactive to internal sperm components, present in many fertile individuals or in children, (Tung et ai., 1976) and, therefore, irrelevant to infertility, will not be evaluated. Materials and methods
Subjects Six hundred and sixty women, all of the patients referred to our laboratory for antisperm antibody testing from January to November 1988, were studied. Forty four of the females were referred due to a recent history of two or more consecutive and unexplained miscarriages in the first trimester of pregnancy; the remaining 616 women were unable to achieve a concep-
153
tion. Extensive clinical evaluations had failed to reveal an endocrine or microbiological cause for the recurrent miscarriages.
Sperm antibody testing A standardized semen analysis (Witkin and David, 1988) was performed on each male partner and antibodies bound to the surface of their isolated motile sperm were measured by the IBT (Bronson et al., 1984) using isotypespecific reagents. A positive test was defined as immunobead binding to at least 40070 of the sperm. This level of binding was the minimum never achieved in fertile controls. Data were also analyzed using 30070 or 50°70 binding to sperm as the cutoff point for a positive test. Sera of the female partners were diluted 1 : 10 in PBS and incubated for 60 min at 37°C with 5 x 106 motile sperm isolated from their husband's ejaculate by the swim-up technique, in a final volume of 1.0 ml. In the miscarriage group, sperm from a fertile donor was used in three cases where husband's sperm were poorly motile and in five cases where sperm contained bound antibodies. The incidence of sperm antibody detection was unchanged in both the infertile and miscarriage groups when donor sperm was substituted for husband's sperm. The incubated sperm were washed three times with PBS containing 5% bovine serum albumin and subjected to the IBT. The isotype and regional binding specificities of the immunoglobufins were determined.
Statistics Differences in the various parameters were evaluated by X2 analysis, using the Yates correction factor. Results
Among the women with a history of recurrent abortions, 63.6% (28 of 44) were positive for antisperm antibodies in their sera. This was significantly higher (P < 0.005) than the 37.8 % incidence (233 of 616) observed in the 616 infertile women. Similar differences were obtained using at least 30% (P < 0.025) or 50°70 (P < 0.05) binding of antibody to sperm as the minimum required to define antisperm antibody-positive sera (Table 1). The relation between antisperm antibodies and numbers of miscarriages -was evaluated (Table 2). Although the small numbers precluded statistical analysis, the incidence of antisperm antibodies was higher in women who had a greater number of miscarriages. Both women who had five miscarriages (100070) and seven of the eight women (87.5%) with at least four consecutive miscarriages had antisperm antibodies in their sera. Seven of the 11 women (63.6%) with three previous miscarriages and 14 of the 25 women (56.0070) with two miscarriages were positive for antisperm antibodies.
154 TABLE 1 Antisperm antibodies in sera of infertile women and women with recurrent miscarriages. Antisperm antibodies in sera from 616 female partners of infertile marriages and from 44 women with at least two consecutive, spontaneous early stage miscarriages were detected by the immunobead test. % Antibodypositive sperm
No. positive patients (%) Infertile
Miscarriage
P value
• 30 • 40 • 50
303 (49.2) 233 (37.8) 180 (29.2)
30 (68.2) 28 (63.6) 20 (45.5)
< 0.025 < 0.005 < 0.05
The isotypes of antisperm antibodies present in sera of infertile women and in women with recurrent miscarriages were compared (Table 3). The incidence of IgG antisperm antibodies was significantly higher (P < 0.005) in women with a history of recurrent miscarriages (40.9070) than in the infertile women (19.2o70). In marked contrast, IgM and IgA antisperm antibodies occurred with similar frequencies in the two groups. The regional binding specificities of the antisperm antibodies present in the two groups of women were also compared (Table 4). Women with recurrent miscarriages had a higher incidence (P < 0.005) of IgG antibodies directed to the principal piece of sperm tails (36.4070) than did the infertile women (14.6070). No significant differences in the occurrence of IgG antibodies to sperm heads and in IgA and IgM head and tail-directed antibodies were observed between the two groups. For each antibody isotype, taildirected antibodies were more prevalent than head-directed antibodies in both groups of women. An evaluation of ejaculated sperm from partners of women with and without a history of recurrent spontaneous abortions (Table 5) revealed no differences in the incidence of low sperm count (~<20 × 106), low motility
TABLE 2 Incidence of circulating antisperm antibodies in women with two--five recurrent miscarriages. No. of miscarriages
No. of patients
No. of antibodies (%)
2 3 4 5 2--5
25 11 6 2 44
14 (56.0) 7 (63.6) 5 (83.3) 2 (100) 28 (63.6)
155 TABLE 3 Isotypes of antisperm antibodies in infertile women and in women with recurrent miscarriages. Antibodies
Total IgG IgA IgM
No. positive (%) Infertile
Miscarriage
P value
233 (37.8) 118 (19.2) 88 (14.3) 126 (20.5)
28 (63.6) 18 (40.9) 11 (25.0) 10 (22.7)
< 0.005 < 0.005 n.s. n.s.
(<50¢/0 motile sperm), abnormal morphology (>~50~/0 sperm with head or tail defects), sperm agglutination, or antibodies on the surface of ejaculated sperm. Discussion The availability of an easy and accurate radioimmunoassay for measuring serum levels of human chorionic gonadotropin (hCG) has made possible the diagnosis of pregnancy even before the first missed menstrual period. Many pregnancies are never diagnosed clinically but can be proven to have occurred due to increased serum hCG levels ("chemical pregnancies"). Thus, the line between infertility and pregnancy loss is not as distinct now as it was earlier. Women who present with infertility and regular menstrual cycles, infertility with irregular menstrual cycles, and recurrent early miscarriages may be part of a continuous spectrum of women with inefficient reproduction. A considerable overlap may, therefore, exist between factors that cause infertility and those that cause early stage pregnancy losses. TABLE 4 Specificity of antisperm antibodies in infertile women and in women with recurrent miscarriages. Isotype and location IgG-Heads IgG-Tails IgA-Heads IgA-Tails IgM-Heads IgM-Talls
P value
No. positive (%) Infertile
Miscarriages
44 (7. I) 90 (14.6) 24 (3.9) 69 (I 1.2) 17 (2.8) 103 (16.7)
4 (9. I) 16 (36.4) 3 (6.8) 8 (18.2) 1 (2.3) 7 (15.9)
n.s. < 0.005 n.s. n.s. n.s. n.s.
156 TABLE 5 Sperm analysis of partners of infertile women and women with miscarriages. No. positive (%)
Sperm count g 2 x 107 Motility g50¢/0 Abnormal morphology Sperm agglutination Antibodies on sperm
Infertile
Miscarriages
P value
78 (12.7) 165 (26.8) 40 (6.5) 17 (2.8) 54 (8.8)
4 (9.1) 11 (25.0) 2 (4.5) 3 (6.8) 5 (11.4)
n.s. n.s. n.s. n.s. n.s.
In addition to their role in infertility, antisperm antibodies can also be associated with recurrent spontaneous abortions. In the present study the incidence of IgG antibodies to sperm tail antigens was more than twice as high in sera of women with at least two early stage pregnancy losses than in sera from female partners with unexplained infertility. We realize that this study only analyzed a selected subpopulation of women with early pregnancy loss. W o m e n with known causes for their spontaneous abortions would not have been referred to our laboratory for testing. Nevertheless, the data demonstrates that a large percentage of women with unexplained early stage miscarriage will be positive for a particular isotype and specificity of circulating antisperm antibody. Whether antisperm antibodies are directly involved in pregnancy failure or, instead, merely serve as a marker for other factors that negatively influence embryo survival is still open to speculation, The reports of sperm antigen conservation on the blastocyst or placenta of n o n - h u m a n mammals has led to the suggestion that reactivity with these antigens may be the mechanism by which antisperm antibodies affect the developing fetus (O'Rand, 1977; O ' R a n d et al., 1984; Menge et al., 1978). Alternatively, Clark et al. (1987) has suggested that spontaneous abortions may occur by immune mechanisms that are not antigen-specific. In this regard, exposure to sperm in women sensitized to sperm, but not in women negative for antisperm antibodies, has been shown to induce lymphocyte activation and interferon g a m m a production (Witkin, 1988). Since interferon g a m m a can induce inappropriate major histocompatibility complex antigen expression on human trophoblasts (Feinman et al., 1987) as well as activate cytotoxic monocytes (Le et al., 1983) and lymphocytes (Farrar et al., 1981), exposure of spermsensitized pregnant women to sperm would render the embryo at increased risk to attack by the maternal i m m u n e system. Sperm antibodies could also serve as a marker for a deficiency in maternal genital tract immunosuppressor mechanisms. Antisperm antibody form-
157
ation in some women is associated with a failure of their i m m u n e system to respond to immunosuppressive factors in sperm or seminal fluid (Witkin, 1988a; Witkin, 1989). A similar failure of the female immune system to suppress the response to paternal antigens on the fetus could lead to early stage pregnancy loss (Rocklin et al., 1976). Data has also been presented demonstrating that women with recurrent spontaneous abortions due to an immune abnormality have a higher incidence of antisperm antibodies than do women with recurrent abortions not associated with immune problems (Haas et al., 1986; Beer et al., 1987). This reinforces the likelihood that sperm antibodies serve as a marker for defective immunosuppression rather than being directly involved in binding to embryo antigens that are crossreactive with sperm. On a practical level, it may be worthwhile to screen female partners of infertile couples, women with unexplained spontaneous abortions, and women who failed in vitro fertilization for circulating antisperm antibodies. Avoidance of exposure to sperm following conception in these women may be beneficial in lowering the spontaneous abortion rate due to genital tract immune system activation. Studies to evaluate this possibility are now in progress.
Acknowledgement The technical assistance of Elena Kissin and Ermelinda Montalvo and manuscript preparation by Sherry Holzman are gratefully acknowledged. The studies were supported by NIH grant HD 21909.
References Beer, A.E., Quebbeman, J.F. and Semprini, A.E. (1987) Immunopathological factors contributing to recurrent and spontaneous abortion in humans. In: Spontaneous and Recurrent Abortion (Bennett, M.J. and Edmonds, D.K., eds.) pp. 90--108. Blackwell Scientific Publications, Oxford. Bronson, R., Cooper (3. and Rosenfeld, D. (1984) Sperm antibodies: their role in infertility. Fertil. Steril. 42, 171--183. Clark, A., Croy, B., Wegmann, T.G. and Chaouat, G. (1987) Immunological and paralmmunological mechanisms in spontaneous abortion: recent insights and future directions. J. Reprod. Immunol. 12, 1--12. Farrar, L., Johnson, M. and Farrar, J. (1981) Regulation of the production of immune interferon and cytotoxic T lymphocytes by interleukin 2. J. Immunol. 126, 1120--1125. Feinman, A., Kliman, J. and Main, K. (1987) HLA antigen expression and induction by interferon in cultured human trophoblasts. Am. J. Obstet. Gynecol. 157, 1429--1434. Haas, G.G., Kubota, K., Quebbeman, J.F., Jijon, A., Menge, A.C. and Beer, A.E. (1986) Circulating antisperm antibodies in recurrently aborting women. Fertil. Steril. 45,209--215. Ingerslev, H.J. and Ingerslev, M.L. (1980) Clinical findings in infertile women with circulating antibodies against spermatozoa. Fertil. Steril. 33, 514--520. Jones, W.R. (1976) Immunological aspects of infertility. In: Immunology of Human Reproduction (Scott, J.S. and Jones, W.R., eds). pp. 375---414. Academic Press, London.
158
Le, J., Prensky, W., Yip, Y.K., Chang, Z., Hoffman, T., Stevenson, H.C., Balazs, I., Sadlik, J.R. and Vflcek, J. (1983) Activation of human monocyte cytotoxicity by natural and recombinant immune interferon. J. Immunol. 131, 2821--2826. Mathur, S., Neff, M.R., Williamson H.O., Genco, P.V., Rust, P.F. and Glassman, A.B. (1987) Sperm antibodies and human leukocyte antigens in couples with early spontaneous abortions. Int. J. Fenil. 32, 59--65. Menge, A.C. (1970) Immune reactions and infertility. J. Reprod. Fertil. (Suppl.) 10, 171--186. O'Rand, M.G. (1977) The presence of sperm-specific surface isoantigens on the egg following fertilization. J. Exp. Zool. 202, 267--273. O'Rand, M.G., Irons, G.P. and Porter, J.P. (1984) Monoclonal antibodies to rabbit sperm autoantigens. 1. Inhibition of in vitro fertilization and localization on the egg. Biol. Reprod. 30, 721--729. Rocklin, R.R., Kitzmfller, J.L., Carpenter, C.B., Garroy, M.R. and David, J.R. (1976) Absence of an immunologic blocking factor from the serum of women with chronic abortions. New Eng. J. Med. 295, 1209--1212. Tung, K.S.K., Cooke, W.D. Jr., McCarthy, J.A. and Robiataille, P. (1976) Human sperm antigens and antisperm antibodies II. Age related incidence of antisperm antibodies. Clin. Exp. lmmuno125, 73-79. Witkin, S.S. and David, S.S. (1988) The effect of sperm antibodies on pregnancy outcome in a subfertile population. Am. J. Obstet. Gynecol. 158, 59--62. Witkin, S.S. (1988) Production of interferon gamma by lymphocytes exposed to antibody coated spermatozoa: a mechanism for sperm antibody production in females. Fertil. Steril. 50, 498--502. Witkin, S.S. (1988a) Mechanisms of active suppression of the immune response to spermatozoa. Am. J. Reprod. Immunol. Microbiol. 17, 61--64. Witkin, S.S. (1989) Failure of sperm-induced Immunosuppression: association with antisperm antibodies in females. Am. J. Obstet. Gynecol. in press.
151
Elsevier Scientific Publishers Ireland Ltd.
JRI 00594
Association between recurrent spontaneous abortions and circulating IgG antibodies to sperm tails in women S t e v e n S. W i t k i n a n d A n u C h a u d h r y Immunology Division, Department of Obstetrics and Gynecology, Cornell University Medical College, New York (U.S.A.) (Accepted for publication 15 March 1989)
Summary The isotype and regional specificity of antisperm antibodies in the circulation of women with recurrent spontaneous abortions was examined. There was a statistically significant association (P < 0.005) between the presence of IgG tail-directed antisperm antibodies and a history of unexplained recurrent spontaneous abortion. These antibodies were detected in 36.4% of 44 women with recurrent abortions and 14.6°70 o f 616 female partners of infertile marriages. In contrast, no differences in IgG sperm head-directed antibodies or in IgA and IgM antisperm antibodies were observed between the two groups. Husbands of women in the miscarriage or infertile groups had similar semen evaluations. Antisperm antibodies may be a marker for defective immunosuppression in women with recurrent miscarriages. Alternatively, exposure of sperm-sensitized pregnant women to sperm may activate the maternal immune system to respond to paternal antigens present on the embryo. Key
words:
antisperm
antibodies;
recurrent spontaneous
abortion;
infertility.
Correspondence to: Dr. Steven S. Witkin, Cornell University Medical College, Department of Obstetrics & Gynecology, 515 East 71st Street New York, New York 10021, U.S.A. 0165-0378/89/$03.50 © 1989 Elsevier Scientific Publishers Ireland Ltd. Published and Printed in Ireland
152
Introduction
Antisperm antibodies in women's sera reduce the probability that deposited sperm will reach the oviduct a n d / o r fertilize the ovum (Bronson et al., 1984; Witkin and David, 1988). However, the role, if any, of antisperm antibodies in post-fertilization pregnancy loss is still unclear. Female rabbits and cattle, after being immunized with sperm, have an increased incidence of spontaneous abortion in subsequent pregnancies (Menge, 1970). Both polyclonal (O'Rand, 1977) and monoclonal (O'Rand et al., 1984) antibodies to rabbit sperm react with antigens present on pre-implanatation embryos. In two studies, approximately 50°70 of women with a history of early recurrent spontaneous abortions were positive for circulating sperm agglutinating or immobilizing antibodies (Jones, 1976) or cytotoxic or hemagglutinating antibodies (Mathur et ai., 1987). In another investigation, however, no relation between a history of one to four spontaneous miscarriages and the presence of sperm agglutinating antibodies could be demonstrated (Ingerslev and Ingerslev, 1980). Haas et al. (1986) also noted an association between antisperm antibodies in women and a poor pregnancy prognosis. In their patients, the incidence of sperm immobilizing antibodies but not sperm agglutinating antibodies or antibodies to sperm extracts detected by radioimmunoassay, was greater in recurrent aborters than in infertile women. In the most recent study, antisperm antibodies detected by enzyme-linked i m m u n o s o r b a n t assay were identified in 7 of 16 women (43070) with early stage miscarriages (Witkin and David, 1988). The present study was initiated to evaluate and compare the incidence and regional specificity of antisperm antibodies in infertile women and women with a history of recurrent spontaneous abortions. Sperm antibody detection was by the i m m u n o b e a d test (IBT), which allows the determination of both the isotype and the location on the sperm of any antisperm antibodies detected. Moreover, the IBT is specific for antibodies reactive with the surface of living motile sperm. This assures that antibodies reactive to internal sperm components, present in many fertile individuals or in children, (Tung et ai., 1976) and, therefore, irrelevant to infertility, will not be evaluated. Materials and methods
Subjects Six hundred and sixty women, all of the patients referred to our laboratory for antisperm antibody testing from January to November 1988, were studied. Forty four of the females were referred due to a recent history of two or more consecutive and unexplained miscarriages in the first trimester of pregnancy; the remaining 616 women were unable to achieve a concep-
153
tion. Extensive clinical evaluations had failed to reveal an endocrine or microbiological cause for the recurrent miscarriages.
Sperm antibody testing A standardized semen analysis (Witkin and David, 1988) was performed on each male partner and antibodies bound to the surface of their isolated motile sperm were measured by the IBT (Bronson et al., 1984) using isotypespecific reagents. A positive test was defined as immunobead binding to at least 40070 of the sperm. This level of binding was the minimum never achieved in fertile controls. Data were also analyzed using 30070 or 50°70 binding to sperm as the cutoff point for a positive test. Sera of the female partners were diluted 1 : 10 in PBS and incubated for 60 min at 37°C with 5 x 106 motile sperm isolated from their husband's ejaculate by the swim-up technique, in a final volume of 1.0 ml. In the miscarriage group, sperm from a fertile donor was used in three cases where husband's sperm were poorly motile and in five cases where sperm contained bound antibodies. The incidence of sperm antibody detection was unchanged in both the infertile and miscarriage groups when donor sperm was substituted for husband's sperm. The incubated sperm were washed three times with PBS containing 5% bovine serum albumin and subjected to the IBT. The isotype and regional binding specificities of the immunoglobufins were determined.
Statistics Differences in the various parameters were evaluated by X2 analysis, using the Yates correction factor. Results
Among the women with a history of recurrent abortions, 63.6% (28 of 44) were positive for antisperm antibodies in their sera. This was significantly higher (P < 0.005) than the 37.8 % incidence (233 of 616) observed in the 616 infertile women. Similar differences were obtained using at least 30% (P < 0.025) or 50°70 (P < 0.05) binding of antibody to sperm as the minimum required to define antisperm antibody-positive sera (Table 1). The relation between antisperm antibodies and numbers of miscarriages -was evaluated (Table 2). Although the small numbers precluded statistical analysis, the incidence of antisperm antibodies was higher in women who had a greater number of miscarriages. Both women who had five miscarriages (100070) and seven of the eight women (87.5%) with at least four consecutive miscarriages had antisperm antibodies in their sera. Seven of the 11 women (63.6%) with three previous miscarriages and 14 of the 25 women (56.0070) with two miscarriages were positive for antisperm antibodies.
154 TABLE 1 Antisperm antibodies in sera of infertile women and women with recurrent miscarriages. Antisperm antibodies in sera from 616 female partners of infertile marriages and from 44 women with at least two consecutive, spontaneous early stage miscarriages were detected by the immunobead test. % Antibodypositive sperm
No. positive patients (%) Infertile
Miscarriage
P value
• 30 • 40 • 50
303 (49.2) 233 (37.8) 180 (29.2)
30 (68.2) 28 (63.6) 20 (45.5)
< 0.025 < 0.005 < 0.05
The isotypes of antisperm antibodies present in sera of infertile women and in women with recurrent miscarriages were compared (Table 3). The incidence of IgG antisperm antibodies was significantly higher (P < 0.005) in women with a history of recurrent miscarriages (40.9070) than in the infertile women (19.2o70). In marked contrast, IgM and IgA antisperm antibodies occurred with similar frequencies in the two groups. The regional binding specificities of the antisperm antibodies present in the two groups of women were also compared (Table 4). Women with recurrent miscarriages had a higher incidence (P < 0.005) of IgG antibodies directed to the principal piece of sperm tails (36.4070) than did the infertile women (14.6070). No significant differences in the occurrence of IgG antibodies to sperm heads and in IgA and IgM head and tail-directed antibodies were observed between the two groups. For each antibody isotype, taildirected antibodies were more prevalent than head-directed antibodies in both groups of women. An evaluation of ejaculated sperm from partners of women with and without a history of recurrent spontaneous abortions (Table 5) revealed no differences in the incidence of low sperm count (~<20 × 106), low motility
TABLE 2 Incidence of circulating antisperm antibodies in women with two--five recurrent miscarriages. No. of miscarriages
No. of patients
No. of antibodies (%)
2 3 4 5 2--5
25 11 6 2 44
14 (56.0) 7 (63.6) 5 (83.3) 2 (100) 28 (63.6)
155 TABLE 3 Isotypes of antisperm antibodies in infertile women and in women with recurrent miscarriages. Antibodies
Total IgG IgA IgM
No. positive (%) Infertile
Miscarriage
P value
233 (37.8) 118 (19.2) 88 (14.3) 126 (20.5)
28 (63.6) 18 (40.9) 11 (25.0) 10 (22.7)
< 0.005 < 0.005 n.s. n.s.
(<50¢/0 motile sperm), abnormal morphology (>~50~/0 sperm with head or tail defects), sperm agglutination, or antibodies on the surface of ejaculated sperm. Discussion The availability of an easy and accurate radioimmunoassay for measuring serum levels of human chorionic gonadotropin (hCG) has made possible the diagnosis of pregnancy even before the first missed menstrual period. Many pregnancies are never diagnosed clinically but can be proven to have occurred due to increased serum hCG levels ("chemical pregnancies"). Thus, the line between infertility and pregnancy loss is not as distinct now as it was earlier. Women who present with infertility and regular menstrual cycles, infertility with irregular menstrual cycles, and recurrent early miscarriages may be part of a continuous spectrum of women with inefficient reproduction. A considerable overlap may, therefore, exist between factors that cause infertility and those that cause early stage pregnancy losses. TABLE 4 Specificity of antisperm antibodies in infertile women and in women with recurrent miscarriages. Isotype and location IgG-Heads IgG-Tails IgA-Heads IgA-Tails IgM-Heads IgM-Talls
P value
No. positive (%) Infertile
Miscarriages
44 (7. I) 90 (14.6) 24 (3.9) 69 (I 1.2) 17 (2.8) 103 (16.7)
4 (9. I) 16 (36.4) 3 (6.8) 8 (18.2) 1 (2.3) 7 (15.9)
n.s. < 0.005 n.s. n.s. n.s. n.s.
156 TABLE 5 Sperm analysis of partners of infertile women and women with miscarriages. No. positive (%)
Sperm count g 2 x 107 Motility g50¢/0 Abnormal morphology Sperm agglutination Antibodies on sperm
Infertile
Miscarriages
P value
78 (12.7) 165 (26.8) 40 (6.5) 17 (2.8) 54 (8.8)
4 (9.1) 11 (25.0) 2 (4.5) 3 (6.8) 5 (11.4)
n.s. n.s. n.s. n.s. n.s.
In addition to their role in infertility, antisperm antibodies can also be associated with recurrent spontaneous abortions. In the present study the incidence of IgG antibodies to sperm tail antigens was more than twice as high in sera of women with at least two early stage pregnancy losses than in sera from female partners with unexplained infertility. We realize that this study only analyzed a selected subpopulation of women with early pregnancy loss. W o m e n with known causes for their spontaneous abortions would not have been referred to our laboratory for testing. Nevertheless, the data demonstrates that a large percentage of women with unexplained early stage miscarriage will be positive for a particular isotype and specificity of circulating antisperm antibody. Whether antisperm antibodies are directly involved in pregnancy failure or, instead, merely serve as a marker for other factors that negatively influence embryo survival is still open to speculation, The reports of sperm antigen conservation on the blastocyst or placenta of n o n - h u m a n mammals has led to the suggestion that reactivity with these antigens may be the mechanism by which antisperm antibodies affect the developing fetus (O'Rand, 1977; O ' R a n d et al., 1984; Menge et al., 1978). Alternatively, Clark et al. (1987) has suggested that spontaneous abortions may occur by immune mechanisms that are not antigen-specific. In this regard, exposure to sperm in women sensitized to sperm, but not in women negative for antisperm antibodies, has been shown to induce lymphocyte activation and interferon g a m m a production (Witkin, 1988). Since interferon g a m m a can induce inappropriate major histocompatibility complex antigen expression on human trophoblasts (Feinman et al., 1987) as well as activate cytotoxic monocytes (Le et al., 1983) and lymphocytes (Farrar et al., 1981), exposure of spermsensitized pregnant women to sperm would render the embryo at increased risk to attack by the maternal i m m u n e system. Sperm antibodies could also serve as a marker for a deficiency in maternal genital tract immunosuppressor mechanisms. Antisperm antibody form-
157
ation in some women is associated with a failure of their i m m u n e system to respond to immunosuppressive factors in sperm or seminal fluid (Witkin, 1988a; Witkin, 1989). A similar failure of the female immune system to suppress the response to paternal antigens on the fetus could lead to early stage pregnancy loss (Rocklin et al., 1976). Data has also been presented demonstrating that women with recurrent spontaneous abortions due to an immune abnormality have a higher incidence of antisperm antibodies than do women with recurrent abortions not associated with immune problems (Haas et al., 1986; Beer et al., 1987). This reinforces the likelihood that sperm antibodies serve as a marker for defective immunosuppression rather than being directly involved in binding to embryo antigens that are crossreactive with sperm. On a practical level, it may be worthwhile to screen female partners of infertile couples, women with unexplained spontaneous abortions, and women who failed in vitro fertilization for circulating antisperm antibodies. Avoidance of exposure to sperm following conception in these women may be beneficial in lowering the spontaneous abortion rate due to genital tract immune system activation. Studies to evaluate this possibility are now in progress.
Acknowledgement The technical assistance of Elena Kissin and Ermelinda Montalvo and manuscript preparation by Sherry Holzman are gratefully acknowledged. The studies were supported by NIH grant HD 21909.
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