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Approximately one-fifth of first trimester losses can be characterized by the onset of hypocomplementemia prior to ultrasonographic loss of embryonic viability. Monoclonal antibodies to the membrane complement regulatory protein, decay-accelerating factor (DAF), were bound to the surface of trophoblastic tissues, with the estimated number of DAF molecules in tissues obtained from hypocomplementemic women less than those from normocomplementemic women (approximately 10%). While trophoblastic tissue obtained from normocomplementemic women did not decomplement normal human sera, pretreatment of this tissue with antiDAF antibodies resulted in activation of the complement (C′) system via the alternate C′ pathway (ACP). If trophoblastic tissue from normocomplementemic women was pretreated with phospholipase C, decomplementation of human serum occurred. These data strongly suggest that trophoblasts evade the ACP with functional DAF, possibly through absorption of the molecule's lipophilic diacylglycerol membrane-anchored moiety into the outer lipid bilayer of the trophobast and may be a rational means for the utilization of immunotherapy in recurrent spontaneous aborters.

Mononuclear cell subpopulations from the peripheral blood (PB) and peritoneal fluid (PF) of fertile and infertile women were quantified by flow cytometry using a double-staining monoclonal antibody technique. No differences in the percentage distribution of mononuclear cells between fertile and infertile women were demonstrated when either the PB constituents or the PF components were compared to one another. When the mononuclear cell composition in the PB was compared with that in the PF however, the percentage of PF-activated T cells and monocytes was increased in both fertile and infertile women as opposed to that in the PB, while there was a decrease in T helper cells (fertile and infertile women) and natural killer cells (fertile women and infertile women with endometriosis). Even though shifts do exist in the immunocytes of PF from fertile and infertile women, evidence is lacking that the PF is an immunologic mediator of infertility.

Objective To study autoimmune aspects of endometriosis. Design Lymphoblast transformation and hemolytic plaque formation were used to assess specific T- and B-cell activity against endometrial antigens. Setting Military teaching hospital. Patients Ninety-four healthy women of reproductive age undergoing diagnostic laparoscopy as part of an evaluation for infertility or chronic pelvalgia were accordingly grouped into those with normal pelvic peritoneum (20), mild endometriosis (50), and severe endometriosis (24). Main Outcome Measure The study assessed the proliferative and humoral responses of lymphocytes from women with and without endometriosis to endometrial antigens and quantified the number of B-cell precursors, T-helper cells, and T-suppressor cells to these antigens. Results Unfractionated endometrial antigens were similarly blastogenic for lymphocytes from women with and without endometriosis. Despite equivalent numbers of B-cell precursors to these antigens, antiendometrial antibody responsiveness appears to have increased in women with mild endometriosis because of a decrease in T-suppressor cell activity and declined in women with severe endometriosis because of a further drop in T-suppressor cell activity and an increase in T-helper cell activity, as compared with women without endometriosis. Conclusions Taken together, these experiments support the possibility that pelvic endometriosis may result from a break in specific T-cell tolerance rather than nonspecific polyclonal activation of responder lymphocytes.

Natural killer (NK) cell activity was determined in 41 women during the first trimester of pregnancy. NK cytotoxicity was similar between the study subjects and nonpregnant controls and was not a reflection of a change in the circulating number of cells or density in culture. Although NK cells may play a role in maternal immune status in advanced pregnancy, NK cells do not appear to be crucially involved in the first trimester, when reproductive wastage usually occurs.

Immune responses in women with chlamydial genital tract infection who achieved a bacteriologic cure were prospectively compared to those women who had an incomplete clinical response. Local anti-chlamydia IgA antibody responses were significantly diminished in the outpatient treatment group who had an incomplete response, while circulating IgG responses did not differ significantly between groups. This is in contrast to both specific and non-specific lymphoproliferative responses which were similar between uninfected controls and women with a chlamydial genital tract infection, regardless of treatment outcome. Thus, the kinetics of successful outpatient treatment of chlamydial genital tract infections may have an immunologic component which can be measured at the time of initial evaluation, and may be predictive of the clinical response to adjunctive antibiotics.

Serum complement (C') activity in recurrent spontaneous aborters and primiparous controls with successful and unsuccessful pregnancies was quantified so as to define the dynamics of C' activation in early pregnancy loss. C' hemolytic activity was shown to be stable throughout the first trimester of pregnancy and did not differ from preconception levels in all of the successful pregnancies of recurrent aborters and controls and in the majority of pregnancy losses. However, 30% of recurrent aborters and 20% of controls with a pregnancy loss demonstrated activation of C' by the alternate pathway as early as the 7th week with a progressive decline in C' activity until abortion was clinically completed. Circulating levels of C3 dropped from 1.34 to 0.53 mg/dl, and factor B levels declined from 0.34 to 0.14 mg/dl in these hypocomplementemic women. Pregnancy loss is therefore associated with C' activation in a subset of both recurrent and nonrecurrent aborters and this occurs largely before loss of fetal viability.

This randomized prospective study was conducted to determine the length of time required for re-establishment of the reproductive axis following a 1st-trimester spontaneous abortion. The spontaneous gonadotropin secretion was significantly depressed during the first menstrual cycle after pregnancy loss, while the estradiol levels had normalized. Provocative testing revealed blunted gonadotropin release in the first menstrual cycle with return to normal during the first menstrual cycle after a spontaneous abortion. Endometrial biopsy specimens were also abnormal during the first menstrual cycle with normal histological characteristics by the second menstrual cycle. Therefore, restoration of the hypothalamic-pituitary- ovarian axis after a 1st-trimester loss is achieved within two menstrual cycles, as determined by return of normal pituitary function.

Using the Mishell and Dutton culture system an assay was developed for eliciting and quantifying sperm surface antigen (SSA)-specific immune responses in vitro. Peripheral blood mononuclear cells (PBMC) from sexually inexperienced and experienced women that were antisperm antibody (ASA) negative gave rise to a plaque-forming cell (PFC) response of similar magnitude (1,362 and 1,254 PFC/culture, respectively). PBMC from women that were ASA positive had lower PFC responses if cervical mucous antibodies were detected along with circulating antibodies (876 PFC/culture) than if they were not detected (1,512 PFC/culture). Limit dilution analysis was used to demonstrate that the level of the anticipated antibody response is regulated by T helper cell activity. The results of this investigation suggest that the antibody response to SSA is governed at least in part by both the secretory immune system and circulating T helper cells.

Objective To evaluate the effect of peritoneal fluid (PF) from patients with endometriosis on gamete interaction under hemizona assay (HZA) conditions. Design The HZA was used to study the effect of PF from patients with endometriosis on sperm binding to the zona pellucida using media and normal PF as controls. Setting The patients were collected from a university hospital infertility clinic. Patients Peritoneal fluid from 16 women being evaluated for infertility or sterilization who were found to have endometriosis at surgery was used. Three normal patients, who were being sterilized, had PF that was used as a control. Results Results suggest that there is a significant reduction in the number of tightly bound sperm to the zona surface in endometriosis specimens as reflected in the hemizona index and that this effect is directly related to the stage of the disease. Conclusions Although the sample is small, this methodology may help to elucidate one of the mechanisms responsible for endometriosis-associated infertility.

Antenatal administration of glucocorticoids is often used to facilitate fetal lung maturation in cases of prematurity; however, the effects of betamethasone on maternal immune function have not been established. Therefore maternal immune function was assessed with the use of in vitro techniques. Transient and incomplete suppression of the proliferative response to the T-cell mitogen phytohemagglutinin was demonstrated as early as 24 hours after administration of betamethasone. The magnitude and duration of suppression showed a corresponding increase with advancing gestational age, but these effects were not cumulative and were always short-lived (

Neuroendocrine disorders are rare. A case of hypothalamopituitary sarcoidosis is presented in which hyperprolactinemia and galactorrhea occurred in isolation.

Antisperm antibodies to sperm surface antigens in nulligravid women with primary upper genital tract infections were measured by the sperm mixed agglutination reaction assay. As many as 56% of women with a primary episode of pelvic inflammatory disease had antisperm antibodies. In addition, 69% of those women with no history of genital tract infection but with laparoscopic evidence of past pelvic infection had significant levels of circulating antisperm antibodies. Electroimmunoblots of sperm preparations probed with the sera of women who had either known or presumed upper genital tract infection revealed a uniformly recognized 69 kd antigen. In contrast, women with circulating antisperm antibodies before primary upper genital tract infection recognized up to five distinct sperm antigen determinants of 27, 54, 131, 146, and 174 kd. It is a distinct possibility that genital tract infections may lead to immunopotentiation of antisperm antibodies that could affect fertility.

It is reported that mice infected withTrypanosoma cruzi accumulate a suppressor substance (SS) in their sera which, when passively transferred to normal syngeneic recipients, can inhibit primary and secondary antibody responses of spleen and lymph-node cells to T-cell-dependent and -independent antigens; spleen cells were found to be suppressed earlier and to a greater degree than were lymph-node cells. Studies on the effects of the SS on normal cells in vitro also revealed that spleen cells were affected earlier than lymph-node cells, although there were no demonstrable differences in the maximum suppression effected by the SS between the two sources of lymphoid cells. Whereas normal mice could be passively immunosuppressed in vivo with the SS, it was found that serum from passively suppressed recipients did not retain measurable quantities of the SS in their sera, indicating that the substance was removed from circulation at an early stage or was diluted beyond effective concentrations. In in vivo transfers of the SS toH-2-similar and -dissimilar recipients it was found that the effectiveness of the SS related to theH-2 haplotype of the recipients.

C57BL/6 mice exhibit low parasitemias and often survive Trypanosoma cruzi infections, whereas C3H(He) mice die during the acute phase with relatively high parasitemias. The present study showed that both strains of mice develop nonspecific immunosuppression to challenge with sheep erythrocytes during the course of infection. Several major differences in immunosuppression-related phenomena between the two strains of mice were determined, yet there is no apparent relationship between immunosuppression and resistance to T. cruzi . Both the number of direct plaque-forming cells and the titer of 2-mercaptoethanol-sensitive agglutinating antibody were significantly lower on day 11 for C57BL/6 mice and day 9 for C3H(He) mice. The number of indirect plaque-forming cells and the titer of mercaptoethanol-resistant agglutinating antibody were reduced by day 36 of infection in C57BL/6 mice and 13 days postinfection in C3H(He) mice. In both strains the degree of humoral response suppression of mice increased concomitant with the period of infection, but was not correlated with the changes in spleen cell numbers. Preliminary experiments designed to explore the mechanism underlying the induction and maintenance of immunosuppression in this hostparasite model disclosed the presence of suppressor substance in the serum of T. cruzi -infected mice. The passive transfer of serum from infected mice to syngeneic recipients elicited a state of immunosuppression to sheep erythrocytes, but did not diminish anti-erythrocyte activity in allogeneic recipients. The induction of immunosuppression in normal mice was further found to be dependent on the interval between serum transfer and challenge with antigen. No quantitative differences existed between the magnitude of suppressed humoral responses in mice infected for varying lengths of time and recipients of serum collected from similarly infected mice.

In unidirectional mixed lymphocyte cultures containing (as responders, stimulators, or regulators) spleen cells from mice infected with Trypanosoma cruzi , alloantigen responses were less than in cultures containing normal spleen cells only. Depletion of plastic adherent cells from infected spleen cells (stimulators or regulators) reversed their inhibitory effect on normal spleen cells (responders); removal of adherent responder cells and/or B lymphocytes did not alter the low alloantigen responses of normal spleen cells (stimulated by infected spleen cells) or infected spleen cells (stimulated by normal spleen cells). Infected spleen cells were effective in regulating mixed lymphocyte cultures only when added at the initiation of the culture. Serum from infected mice suppressed mixed lymphocyte cultures containing responder spleen cells syngeneic to the serum donor if added up to 24 hr after initiation of cultures, whereas the “suppressor serum” had to be present at the initiation of cultures when responder cells were allogeneic to the serum donor. Cultures of infected spleen cells (whole or macrophage enriched) produced a factor which was suppressive when added to mixed lymphocyte cultures containing syngeneic responder cells at initiation. It is proposed that the serum suppressor substance regulates cell-mediated immune responses directly by suppressing the response-potential of cells and indirectly by triggering the release of a factor from adherent splenic cells which induces a hyporesponsive state in T lymphocytes.

During the course of Trypanosoma cruzi infection in C57BL/6 mice, which are relatively resistant to the parasite, the hosts developed antibody activity against previously unencountered antigens. The anti-sheep erythrocyte and antitrinitrophenyl antibody levels increased rapidly from Day 7 of infection, reached a peak by the 21st day, and were maintained at this level through 120 days postinfection in these mice. In contrast, highly susceptible C3H(He) mice did not have demonstrable antibody responses to SRBC or TNP during the 24-day infection period. Autoantibody activity against the selfantigens presented on isologous erythrocytes or thymocytes, however, were reduced in infected C57BL/6 mice. No significant reduction in autoreactivity to the self-antigens on erythrocytes or thymocytes was observed in C3H(He) mice infected with T. cruzi although a trend of reduced autoresponsiveness toward erythrocytes appeared to be developing by the time of death. C57BL/6 mice immunized with sheep erythrocytes as neonates and infected with T. cruzi as adults, or adult mice primed with low doses of sheep erythrocytes prior to infection, had elevated antibody responses to sheep erythrocytes unless the mice were immunized with sheep erythrocytes during the course of infection, in which case suppression of the response against sheep erythrocytes resulted. The nonspecific synthesis of immunoglobulins in infected C57BL/6 mice was, in part, a result of the lymphocyteactivating properties of T. cruzi-associated antigens. The T. cruzi-associated antigens induced proliferative and differentiative responses in spleen cells in vitro. It is proposed that the T. cruzi-associated antigens differentially affect lymphocytes capable of responding to antigen and those lymphocytes previously stimulated by antigen.

Attempts to immunize susceptible hosts against infection with Trypanosoma cruzi have generally not met with a high degree of success. In the present study, we used the antimetabolite mitomycin C to produce nonreplicating, attenuated culture forms of T. cruzi. Attempts to immunize highly susceptible C3H(He) mice with single or multiple inoculi of mitomycin C-treated trypanosomes 2 weeks before injection of infective blood-form trypomastigotes did not, however, lead to greater longevity in immunized mice over nonimmunized, control mice. It was determined that a single injection of 10(7) attenuated parasites induced a transient suppression of the immune response to sheep erythrocytes which was maximum on day 7, less on day 14, and undetectable by the 21st day after immunization. This immunosuppression to a heterologous antigen did not, however, appear to be the cause of the failure of the vaccination procedure to elicit protective immunity since mice immunized once with 10(7) mitomycin C-treated trypanosomes and challenged 30, 60, or 90 days later exhibited no greater longevity than control mice.

Mice that differ in susceptibility to Trypanosoma cruzi were shown to differ in susceptibility to Aeromonas hydrophila as well when infection with A. hydrophila succeeded an established infection with T. cruzi. Challenge with A. hydrophila had no effect on longevity in T. cruzi-infected mice. Mice which are highly susceptible to T. cruzi (C3H(He)) exhibited increased resistance to an intravenous challenge with A. hydrophila on the 7th day of infection; then resistance waned, and by the 21st day of infection, C3H(He) mice were more susceptible than uninfected mice to challenge with A. hydrophila. Mice that are relatively resistant to T. cruzi (C57BL/6) expressed increased resistance to an intravenous challenge with A. hydrophila from the 7th through the 28th day of infection, although no increase in resistance occurred from the 7th day forward. Neither C3H(He) nor C57BL/6 mice infected with T. cruzi developed altered susceptibility to an intraperitoneal challenge with A. hydrophila. While the growth of A. hydrophila in the blood, liver, and spleen increased during the first 36 hr in normal mice, marked retardation of growth occurred at the times of increased resistance in both strains of mice. In C3H(He) mice, bacterial growth returned to normal as resistance was lost. The degree of resistance to A. hydrophila in mice increased as the T. cruzi infection inoculum increased from 102 to 104; however, at the highest dose tested (5 x 104), both strains of mice showed lowered resistance, yet were still more resistant than uninfected mice. Even though resistance to A. hydrophila increased and decreased in C3H(He) mice, and increased and plateaued in C57BL/6 mice, titers of agglutinating antibody to A. hydrophila continued to increase throughout the course of infection with T. cruzi in both strains of mice. Passive suppression of mice with the T. cruzi-induced suppressor substance prior to challenge with A. hydrophila rendered C57BL/6 mice more resistant than normal mice, had no effect on the susceptibility of C3H(He) mice, but lowered the anti-A. hydrophila agglutinating antibody titers in both strains of mice.

Serum from mice infected with Trypanosoma cruzi-suppressed (=SSS) lipopolysaccharide (LPS)-, phytohemagglutinin (PHA)-, and concanavalin A (Con A)-induced lymphoblast transformation when added to cultures of spleen cells or lymph node cells. This serum maximally suppressed blastogenic responses in spleen cell and lymph node cell cultures that contained supportive fetal bovine serum concentrations of 2% and 4%, respectively. Preincubation of lymphoid cells with SSS for 18 to 48 hr prior to initiation of the blastogenesis assay led to suppression of LPS-, PHA-, and Con A-induced proliferation at the optimal concentration of supportive fetal bovine serum (5%), whereas adsorption of lymphoid cells with SSS at 4 C for 30 min before stimulation with mitogen led to suppression of LPS-induced proliferation in spleen cells only. There was a close temporal correspondence between the induction and manifestation of suppression to the T-cell mitogens (PHA and Con A), but the manifestation of suppression preceded the induction of suppression to the B-cell mitogen (LPS) by approximately 12 hr. The SSS-induced suppression of proliferative responses, except in spleen cell cultures stimulated with LPS, was shown to be dependent on the presence of macrophages during the preincubation and stimulation phases of the assay system. The combined results of experiments in which macrophages were preincubated with SSS, or in which macrophages from the spleen were cultured with lymphocytes from the lymph nodes and vice versa (before and after preincubation with SSS), clearly demonstrated the presence of SSS-activated suppressor cells in the spleen, but not in the lymph nodes. Furthermore, the activation of these suppressor macrophages was reliant upon interactions with splenic lymphocytes.

Trypanosoma cruzi releases a factor (SCAF) when grown in vitro which decomplements normal mouse, human, and guinea pig sera. The production and potency of SCAF was dependent on the density of cultured parasites, parasite viability and proliferative capacity, and duration of culture. The in vitro interaction between SCAF and serum complement (C') occurred rapidly and was complete within 30 min of mixing. The administration of SCAF to normal mice resulted in up to 50% reduction in hemolytic C' activity, whereas SCAF had no effect on the C' levels in mice infected with T. cruzi for more than 10 days. The active moiety of SCAF was shown to be a nonproteinaceous substance(s) with a molecular weight of approximately 23,000 daltons. Protozoan and metazoan parasites have been shown to activate serum complement

Spleen cells from normal mice and mice infected with Trypanosoma cruzi undergo mitogen-induced lymphoblast transformation in different ways. Some of these differences may be interpreted as suppression of the blastogenic response, whereas others cannot. Close examination of the culture conditions has enabled us to discriminate between in vivo-induced suppression and in vitro-determined (artifactual) suppression. Regardless of the type of suppression observed, it was shown to be dependent on duration of culture, length of labeling period, dose of mitogen, density of spleen cells, and method of expressing lymphoblast transformation data. The titration of mitomycin C-treated spleen cells from infected mice into cultures of normal spleen cells revealed two, separate, regulatory activities of spleen cells from infected mice as follow: (1) an enhancing effect which appeared late in infection and was obtained with low input of cells, and (2) a suppressive effect which appeared early in infection and was obtained with high input of cells. The enhancing effect was mediated by cells with properties characteristic of T-lymphocytes, whereas macrophages were found to be responsible for the suppressive effects.

Serum from mice infected with Trypanosoma cruzi (SSS) is known to interact with normal spleen cells to induce an immunosuppressed condition and activate splenic suppressor cells. The induction of immunosuppression by SSS was shown to be independent of, and precede the activation of, suppressor cells. Suppressor-cell activation, however, was demonstrable only after the induction of immunosuppression. Furthermore, mice that were given two aliquots of SSS at different intervals of time, exhibited suppression of humoral responses of similar duration and magnitude, regardless of the SSS transfer regimen, whereas both the length and degree of suppressor-cell activity was critically dependent on the interval of time between SSS transfers. SSS interacted with spleen cells via a trypsin-sensitive membrane site which was regenerable within a 4- to 5-hr period, yet the suppressive effects of SSS on spleen cells following interaction was resistant to treatment with trypsin. The interaction between SSS and spleen cells during brief adsorption protocols leads to immunosuppression only because extensive washing of SSS-treated spleen cells did not reverse the immunosuppression process, but did prevent the development of detectable suppressor cells. The phenomenon of suppressor-cell activation was further distinguished from immunosuppression in that supernates from culture of spleen cells derived from SSS-treated mice or T. cruzi-infected contained a factor that activated suppressor cells, but did not directly induce a state of suppression in the responding cell population.

Complement (C') levels in 2 strains of mice that differ in susceptibility to Trypanosoma cruzi were monitored during the course of infection. C3H(He) mice, which are highly susceptible to the parasite, exhibited a progressive decrease in C' and died on about day 24 with approximately 43% of the normal level of C'. C57BL/6 mice, which are more resistant and often survive experimental infections, underwent a reduction of C' to approximately 15% of normal by day 30, a level that persisted until the experiment was terminated on day 45. Additional experiments performed to investigate the parasite's role in the induction of this hypocomplementemia revealed that intact culture-form trypanosomes (live or formalin-treated), crude sonicates of the parasite, or supematants from cultured trypanosomes could decomplement normal serum by 30-70%. Selective chelation of Mg" and/or Ca`+ suggested that viable trypanosomes and trypanosome culture supernatants activate C', primarily via the classical pathway. Collateral studies explored the relationship between the hypocomplementemic state and T. cruzi-induced immunosuppression. Transfer of serum (known to be suppressive for humoral responses) from infected, syngeneic mice into normal recipients did not result in a significant reduction in C' levels. While the significance of lowered C' levels during experimental Chagas' disease is unclear, it does not appear that hypocomplementemia plays a direct role in host susceptibility or the expression of parasite-induced immunosuppression. Krettli and Brener (1976), Kierszenbaum and Howard (1976), and Culbertson and Kolodny (1938) clearly demonstrated that passive transfer of anti-Trypanosoma cruzi antiserum provides some degree of protection to infected recipients. Presumably this resistance is because of antibody, although the participation of antibody in immune rejection of T. cruzi has not been determined. Antibody could be effective in this protozoan infection as an opsonin, as a cytophilic antibody to enhance macrophage destruction of the parasite, and/or as a lytic agent in cooperation with the complement (C') system. From in vitro studies it is known that all stages of T. cruzi are susceptible to antibodymediated complement dependent lysis suggesting that this immune manifestation of antibody could contribute to in vivo destruction of the parasite (Anziano et al., 1972; Budzko, Pizzimenti, and Kierszenbaum, 1975; Kierszenbaum, 1976; Nogueira, Bianco, and Cohn, 1975). If antibody-mediated complement dependent lysis of T. cruzi is an important, if not primary, mechanism of protective immunity in the mammalian host, then, clearly, an understanding of the contribution of C' during the course of infection is as necessary as information on levels Received for publication 31 March 1978. of specific antibody developed against the para

In vitro antisheep erythrocyte (SRBC) and antitrinitrophenyl (TNP) antibody responses of spleen cells obtained from C57BL/6 mice infected with Trypanosoma cruzi were reduced as early as 6 days postinfection and not detectable after 18 days of infection. Lymph node cells had normal antibody responses to SRBC and TNP in vitro until the 11th day of infection, after which responses were diminished. By day 31 of infection, lymph node cells were unresponsive to both SRBC and TNP in vitro. Not only were the antibody responses of spleen and lymph node cells to T-cell-dependent and -independent an- tigens progressively reduced as the period of infection increased, but in addition, the effect of lymphoid cell density and antigen dose on antibody production underwent several sequential changes. As the infection advanced, low densities of cultured lymphoid cells and low doses of antigen were ineffective in eliciting a detectable immune response, whereas high densities of lymphoid cells and high doses of antigen resulted in responses approximately equivalent to that observed with normal cells under the same conditions. Results of cell mixing studies have shown that a plastic-adherent, macrophagelike cell plays a major role in the suppressed humoral responses observed in this host-parasite system. A number of viral, bacterial, protozoal, and metazoal infections of mammals effect states of nonspecific immunosuppression in their hosts. In some of these host-parasite relation