The JI
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     
 

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow Request Permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Blois, S. M.
Right arrow Articles by Arck, P. C.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Blois, S. M.
Right arrow Articles by Arck, P. C.
The Journal of Immunology, 2004, 172: 5893-5899.
Copyright © 2004 by The American Association of Immunologists

Depletion of CD8+ Cells Abolishes the Pregnancy Protective Effect of Progesterone Substitution with Dydrogesterone in Mice by Altering the Th1/Th2 Cytokine Profile1

Sandra M. Blois*,{dagger}, Ricarda Joachim*, Judith Kandil*, Ricardo Margni{dagger}, Mareike Tometten*, Burghard F. Klapp* and Petra C. Arck2,*

* Charité, Department of Internal Medicine, Biomedizinisches Forschungszentrum, Humboldt University of Berlin, Berlin, Germany; and {dagger} Humoral Immunity Studies Institute-National Council of Scientific and Technological Research, University of Buenos Aires, Buenos Aires, Argentina


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 References
 
One of the most remarkable immunological regulations is the maternal immune tolerance toward the fetal semiallograft during pregnancy, which has been referred to as immunity’s pregnant pause. Rejection of the semiallogeneic trophoblast cells must be selectively inhibited and pathways presumably include Th2 cytokines unopposed by Th1 cytokines. Steroid hormones, including progesterone, have similar effects. Low levels of progesterone and Th2 cytokines and high levels of Th1 cytokines are attributable for increased abortions in mammalians, which may be triggered by psychoemotional stress. Thus, the aim of the present study was to provide experimental evidence for the mechanism involved in the mediation of immune responses by endocrine signals during pregnancy and stress-triggered pregnancy failure. DBA/2J-mated CBA/J female mice were randomized in three groups: 1) control females, 2) mice exposed to stress on gestation day 5.5, and 3) mice exposed to stress and substituted with dydrogesterone, a progestogen with a binding profile highly selective for the progesterone receptor on gestation day 5.5. On gestation days 7.5, 9.5, and 10.5, mice of each group were sacrificed, and the frequency of CD8+ cells and cytokine expression (IL-4, IL-12, TNF-{alpha}, IFN-{gamma}) in blood and uterus cells was evaluated by flow cytometry. Additionally, some mice were depleted of CD8 cells by injection of mAb. We observed that progesterone substitution abrogated the abortogenic effects of stress exposure by decreasing the frequency of abortogenic cytokines. This pathway was exceedingly CD8-dependent, because depletion of CD8 led to a termination of the pregnancy protective effect of progesterone substitution.


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 References
 
Often, the embryo has been referred to as "the mating product of histoincompatible individuals," and–consequently–was compared with a semiallogeneic graft having to be tolerated by the maternal immune system over the full gestational period (1).

The spatial adjacencies between fetal and maternal tissues guarantee nourishment of the fetus and the placenta, which is composed of fetal and maternal structures and an area for intense exchanges for nutrients and oxygen (1). In contrast, due to the presence of paternal Ags expressed by the placenta, rejection via the maternal immune system has to be avoided (2, 3, 4, 5). Published data indicate that–during pregnancy–the maternal immune system appears to be "aware" of the fetal Ags, thus, tolerance mechanisms have evolved during evolution to ensure the maintenance of the fetoplacental graft (6).

However, rejection of the fetus and subsequent miscarriages are a frequent pregnancy complication affecting 15–40% of human pregnancies (7, 8). Inevitably, the fetoplacental graft may thus either be tolerated or rejected by the maternal immune system. The pathways of fetal tolerance appear to be plural and matter at diverse times in gestation. Intriguingly, these tolerance mechanisms act systemically on the maternal immune system and locally at the placental level. Indeed, the decision of survival or rejection of the fetus is particularly dependent on the events endangering the mother, prevalently environmental parameters such as exposure to toxic substances, infectious agents, or psychoemotional stress (9).

Systemic effects of fetal tolerance are first and foremost mediated by immunoactive hormones, e.g., progesterone, the hormone of pregnancy. In humans, progesterone-receptor antagonists promptly induce abortion if given before 7 wk of gestation (10). Likewise, surgical removal of the source of progesterone, the corpus luteum, results in pregnancy loss in rodents (11, 12, 13, 14), suggesting that adequate progesterone production is critical to the maintenance of pregnancy. Interestingly, psychoemotional stress is known to generally inhibit the female reproductive system primarily through inhibition of progesterone (15, 16).

Locally at the fetomaternal interface, numerous mechanisms are involved and prevalently intertwined, e.g., expression of MHC class I is reduced on the syncytiotrophoblast, and the specific expression of HLA-G and HLA-E may block the cytolytic activity of NK cells (17, 18). Further, indoleamine 2, 3 dioxygenase, which catabolizes tryptophan, can be detected at the fetomaternal interface and has recently been reported to famish the local maternal immune response by depriving the T cells of tryptophan and/or by inhibiting lymphocyte proliferation (19, 20).

Recent evidence, much of it emerging from research on animal models, indicates important roles for T cells and cytokines in causing pregnancy failure (4, 21, 22, 23, 24). It has been proposed that an inflammatory response mediated by macrophages, NK cells, and T cells with increased levels of Th1 cytokines may mediate abortions (25, 26). Based on the observations of deleterious effects of Th1-type cytokines on mammalian pregnancy and on observations that pregnancy appears to be associated with a down-regulation of cell-mediated immunity manifested as reduced delayed-type hypersensitivity, it has been suggested that the Th1/Th2 balance could be a critical and a complex factor in the tolerance of the conceptus (8).

Data have indicated that a shift of Th1 to Th2 cytokines and suppression of NK cell cytolytic activity may be under the control of an immunomodulatory protein known as progesterone-induced blocking factor (PIBF)3 (27). PIBF is secreted by T cells, predominantly {gamma}{delta} TCR+ and CD8+ (24), upon interaction of progesterone with the progesterone receptors on such cells (28).

In successful pregnancies after immunotherapy (using the male partner’s blood as the source of leukocytes), systemic CD8+ progesterone-receptor+ cells were found to be significantly increased compared with preimmunotherapy (29). Because much of our understanding of early human pregnancy is inferred from studies in animals, we recently performed a pilot study to investigate the effect of the progesterone derivative dydrogesterone (6-dehydro-retroprogesterone), a progesterone with a binding profile highly selective for the progesterone receptor (30), in stress-triggered murine abortion (31) and observed that stressed animals present lower systemic levels of progesterone and PIBF and a reduced expression of progesterone receptor at the fetomaternal interface (30). Injection of dydrogesterone increased levels of plasma PIBF in stressed mice, but did not affect progesterone levels.

Based on the notion that stress perception decreased levels of progesterone which in turn may result in a Th1>Th2 predominance with consecutive abortion, we now investigated–upon confirming that stress during early gestation (day 5.5) boosts the abortion rate, which can be abrogated by the progesterone derivative dydrogesterone–whether 1) the percentage of CD8+ uterine cells was affected in stressed mice with and without dydrogesterone treatment over a period of 5 days after stress exposure, 2) the relative numbers of Th1 (TNF-{alpha}, IFN-{gamma}, IL-12) and Th2 (IL-4) cytokine-producing cells in the uterus would be affected by stress or dydrogesterone treatment, respectively, and 3) depletion of CD8+ cells by injection of anti-Lyt 2.1 Ab would abolish the protective effect of dydrogesterone on the abortion rate in stressed mice by affecting the Th1/Th2 cytokine profile.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 References
 
Animals

Mice were purchased from Charles River Breeding Laboratories (Sulzfeld, Germany) and maintained in an animal facility with a 12 h light/dark cycle. Animal care and experimental procedures were followed according to institutional guidelines and conformed to the requirements of the state authority for animal research conduct (LaGetSi, Berlin, Germany). DBA/2J-mated CBA/J female mice were randomized in different groups of females: 1) control mice that received s.c. injection of sterile filtered sesame oil (200 µl) on gestation day 5.5. The mice were sacrificed either on gestation day 7.5 (n = 7), 9.5 (n = 7), 10.5 (n = 7), or 13.5 (n = 7) for evaluation of the abortion rate and number of implants; 2) mice that were exposed to sound stress on gestation day 5.5 and also received a s.c. injection of 200 µl of sterile filtered sesame oil. Again, the mice were sacrificed either on gestation day 7.5 (n = 7), 9.5 (n = 7), 10.5 (n = 7), or 13.5 (n = 7) for evaluation of the abortion rate and number of implants; 3) mice that were exposed to stress on gestation day 5.5 and received a s.c. injection of dydrogesterone (provided by Solvay Pharmaceutical, Hannover, Germany) at a concentration of 1.25 mg/200 µl of sterile filtered sesame oil on gestation day 5.5, before exposure to sound stress was commenced. As in 1) and 2), the mice were sacrificed either on gestation day 7.5 (n = 7), 9.5 (n = 7), 10.5 (n = 7), and 13.5 (n = 7) for evaluation of the abortion rate and number of implants; 4) in a second experiment, an additional group of mice (n = 20) was treated as in 3), and divided into two subgroups (n = 10 each) which received s.c. injection of anti-Lyt 2.1 (cat. no. 558733) (85 µg/200 ml PBS) CD8 depletion on day 6.5 or PBS alone. Because we knew from the previous experiments on the mice described in 1)-3) that gestation day 7.5 was a crucial day with respect to changes of the uterine cytokine equilibrium, we exclusively focused on the presence of cytokines on this particular day in the CD8 depletion experiments in five mice per subgroup in addition to the evaluation of the abortion rate and number of implants on day 13.5 (n = 5 for each subgroup).

Application of stress

The CBA/J mice were exposed to sound stress for the duration of 24 h starting on gestation day 5.5. The sound stress was emitted by a rodent repellant device (Conrad Electronics, Berlin, Germany) at a frequency of 300 Hertz in intervals of 15 s. The stress device was placed into the mouse cage so that the mice could not escape the sound perception.

Abortion rate

Mice were sacrificed by neck dislocation on day 13.5, the uteri were removed, and the total number of implantations and resorbing sites (signs of abortion) was recorded. The abortion rate was calculated as percentages of resorption sites calculated from total number of implantations based on the following formula: % abortion rate = A/(A + V) x 100, whereby A means the number of dead placental units and V means viable placental units.

Blood cells

On gestation days 7.5, 9.5, and 10.5, mice of the respective groups were narcotized and blood cells were obtained by retro-orbital punction and collected in tubes containing heparin. After treatment with ammonium chloride lysis buffer for 10 min to deplete erythrocytes, the cells were washed twice with sterile PBS.

Blood cells were used for determination of phenotype (surface expression) and cytokine expression on cells by flow cytometry.

Uterus cell isolation

Uteri were removed and uterus cells were isolated by the method described by Marquez et al. (32), with some modifications. In brief, the uterus was washed with sterile PBS, carefully cut into small pieces, and collected in tubes containing HBSS and digested for 20 min at 37°C under slight agitation in HBSS with 1 mM DTT (cat. no. D-0632; Sigma-Aldrich, Munich, Germany). After that, the isolated cells were collected in a fresh tube through a 100-µm net (BD Biosciences, San Francisco, CA) and washed with RPMI 1640 complemented with 10% FCS. The procedure was repeated twice, with HBSS medium containing no DTT. Mononuclear cells were purified by Lympholyte/M (cat. no. GTS5030XK; Cedarlane Laboratories, Hornby, Ontario, Canada) gradient centrifugation. These cells were used to characterize surface and intracellular expression by flow cytometry.

Flow cytometric analysis

For flow cytometry, the blood and uterus cells were incubated for 3 h with brefeldin A (106 cells/ml medium with 1 µl of Golgi Plug, 55-2301KZ; BD PharMingen, Heidelberg, Germany) in RPMI 1640 with FCS in a humidified incubator at 37°C with 5%CO2. Flow cytometry was performed using our standard protocol (31): briefly, uterus cells were washed twice with FACS buffer (PBS supplemented with 1% BSA (cat. no. A-9418; Sigma-Aldrich) and 0.1% sodium acide (Sigma-Aldrich)). Cells were then incubated for 30 min at room temperature (RT) with FITC-labeled Ab against CD8-{alpha} (cat. no. 553030). After the cells were washed and fixed using Fix solution (BD Biosciences, Erembodegem, Belgium), they were incubated for 30 min at RT in the dark. Subsequently, the cells were washed and permeabilized, using FACS Permeabilizing Solution (BD Biosciences), followed by incubation with intracellular Ab PE-labeled TNF-{alpha} (cat. no. 554419), IL-4 (cat. no. 554435), IFN-{gamma} (cat. no. 554412), IL-12 (cat. no. 554479), and IL-10 (cat. no. 554467) for 30 min at RT in the dark. As a control, cells were stained with the corresponding isotype-matched mAb. All mAbs were purchased from BD Biosciences. The cells were then washed and read. The acquisition was performed using a FACSCalibur (BD Biosciences). Instrument compensation was set in each experiment using single-color stained samples. Data were analyzed by using CellQuest software. Flow cytometry results were expressed as the percentage of cells positive for the surface marker evaluated.

Statistical analysis

Statistical significance was determined using the nonparametric Mann-Whitney U test. Significance was set at p < 0.05.


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 References
 
Stress-triggered increase of the abortion rate could be abrogated by the progesterone derivative dydrogesterone

As depicted in Fig. 1A, we reproduced our previous findings that exposure to stress on gestation day 5.5 significantly boosts the abortion rate in DBA/2J-mated CBA/J females, as analyzed on gestation day 13.5, when resorption = abortion sites are visible (31). Further, substitution of progesterone with 1.25 mg of dydrogesterone on gestation day 5.5 significantly abrogated the stress-triggered increase of abortions. Neither stress nor dydrogesterone treatment or CD8 depletion had a significant effect on the number of implantations (Fig. 2).



View larger version (18K):
[in this window]
[in a new window]
  		FIGURE 1. Stress exposure increases abortion in mice, which is abrogated by dydrogesterone treatment via CD8-dependent pathways. A, Effect of stress exposure on gestation day 5.5 and dydrogesterone application during pregnancy on abortion rate, as determined on day 13.5 of gestation. The abortion rate was calculated as follows: % abortion rate = A/(A + V) x 100, whereby A means the number of dead placental units and V means viable placental units. The bars depict the mean percentage ± SEM; *, represents p < 0.05, as analyzed by the nonparametric Mann-Whitney U test. B, Effect of stress exposure and dydrogesterone application during pregnancy on the abortion rate in CD8-depleted mice.

 


View larger version (32K):
[in this window]
[in a new window]
  		FIGURE 2. Stress exposure, dydrogesterone treatment, or CD8 depletion did not affect number of implantations. Effect of stress exposure, dydrogesterone application, and CD8 depletion during pregnancy on the number of implantations on the various experimental = gestational days. This figure depicts the total number of implantations, which comprise viable placental units (pointed out by the gray arrows in the photo of a uterus taken on gestation day 13.5) in addition to hemorrhagic units as an indicator for dead placental units (white arrows), thus the photo depicts an abortion rate of 50%. The data in the diagram are depicted as the mean ± SEM; no significant differences between the groups and days, respectively, were analyzed by the nonparametric Mann-Whitney U test.

 
Differential expression of CD8+ uterus cells in stressed mice

Now, we wished to determine the relative number of uterine CD8+ cells, because this population has been suggested to have a crucial role in pregnancy maintenance (8, 29, 33). As shown in Fig. 3A, stressed mice presented a significant decrease of the percentage of CD8+ uterus cells, as analyzed by flow cytometry, on day 7.5 of gestation compared with control mice. Strikingly, in stressed mice which received dydrogesterone, the percentage of CD8+ uterus cells had been restored back to levels seen in control mice. Interestingly, on day 9.5, mice exposed to stress presented a significantly higher percentage of CD8+ uterus cells, compared with control or stressed and dydrogesterone-treated mice. However, the percentages of CD8+ cells where at a lower level, compared with day 7.5 of gestation. On gestation day 10.5, we observed that the significant differences between the percentages of CD8+ uterine cells in the groups tested were no longer detectable.



View larger version (22K):
[in this window]
[in a new window]
  		FIGURE 3. CD8-positive decidual lymphocytes are decreased after stress exposure, dydrogesterone treatment restores them. A, The percentage of CD8+ cells within the uterine lymphocyte population in the various experimental groups on gestation days 7.5, 9.5, and 10.5. The data were obtained by flow cytometry and are presented as mean ± SEM. *, p < 0.05, **, p < 0.01, ***, p < 0.001, as analyzed by the nonparametric Mann-Whitney U test. B, The percentage of CD8+ cells within the uterine lymphocyte population in stressed, dydrogesterone-treated mice with and without CD8 depletion on gestation day 7.5.

 
We further looked at the expression of CD8 in the peripheral blood cells by flow cytometry and found no significant effect of stress or dydrogesterone treatment with respect to CD8+ cells (Table I).


View this table:
[in this window]
[in a new window]
  		Table I. Percentage of CD8+ blood cells analyzed by flow cytometry

 
Influence of stress and dydrogesterone treatment on cytokine profile in uterus cells

As mentioned earlier, Th2-like cytokines appear to contribute to the maintenance of pregnancy, whereas Th1 cytokines have been shown to be deleterious for pregnancy. In the present study, we observed that on gestation day 7.5 and 9.5, stressed mice showed a significantly increased percentage TNF-{alpha}+ uterus cells (Fig. 4). On gestation day 10.5, 5 days after stress exposure, levels of TNF-{alpha} were back to the percentage seen in control mice. Interestingly, injection of dydrogesterone significantly lowered the relative number of TNF-{alpha}-producing uterine cells on days 7.5, 9.5, and even on 10.5. The absolute numbers of CD8+/TNF-{alpha}+ uterus cells are presented in Table II.



View larger version (17K):
[in this window]
[in a new window]
  		FIGURE 4. Stress exposure up-regulates the percentage of TNF-{alpha}-positive decidual cells; dydrogesterone abolishes this proinflammatory response. Percentage of TNF-{alpha}-positive cells within the uterine lymphocyte population of the various experimental groups on gestation days 7.5, 9.5, and 10.5. TNF-{alpha}-positive cells were identified by flow cytometry for intracellular proteins. The data are presented as mean ± SEM. *, p < 0.05, **, p < 0.01, as evaluated by nonparametric Mann-Whitney U test.

 

View this table:
[in this window]
[in a new window]
  		Table II. The absolute numbers of cells isolated from the uterus and analyzed by flow cytometry

 
Further, we investigated the presence of the Th1 cytokines IFN-{gamma} and IL-12 in uterus cells. As depicted in Fig. 5, nonstressed mice presented a significantly greater percentage of IFN-{gamma}+ cells immediately after stress exposure, on gestation day 7.5, compared with stressed as well as stressed dydrogesterone-treated mice. However, on day 9.5, when the percentages in controls significantly decreased compared with day 7.5, this Th1 down-regulation was not present in stressed mice. When the stressed mice were treated with dydrogesterone, IFN-{gamma} down-regulation did occur in uterine cells. On day 10.5, significant differences with respect to IFN-{gamma}+ uterus cells were no longer detectable as the relative number was rather low compared with day 7.5. The absolute numbers of CD8+/IFN-{alpha}+ and IL-12+uterus cells are presented in Table II.



View larger version (16K):
[in this window]
[in a new window]
  		FIGURE 5. Flow cytometry to detect IFN-{gamma}-positive decidual lymphocytes. Intracellular detection of IFN-{gamma} in the uterine lymphocyte population on gestation days 7.5, 9.5, and 10.5 of the various experimental groups, again determined by flow cytometry. **, p < 0.01, as analyzed by the nonparametric Mann-Whitney U test.

 
When looking at IL-12 expression in uterus cells, we observed no significant differences between the three groups and the different days of gestation (Fig. 6).



View larger version (16K):
[in this window]
[in a new window]
  		FIGURE 6. Stress or dydrogesterone do not significantly alter levels of uterine IL-12. Percentage of IL-12-positive uterine lymphocyte populations from nontreated, stressed, and dydrogesterone-treated, stressed mice on gestation days 7.5, 9.5, and 10.5. Bars shows mean ± SEM. Data were analyzed by nonparametric Mann-Whitney U test.

 
As presented in Fig. 7, stressed mice treated with dydrogesterone showed significantly higher percentages of pregnancy protective IL-4+ uterus cells beginning on gestation day 7.5 and, even more strikingly, maintained until day 10.5. The absolute numbers of CD8+/IL-4+ uterus cells are presented in Table II.



View larger version (16K):
[in this window]
[in a new window]
  		FIGURE 7. Dydrogesterone treatment up-regulates IL-4 in uterus cells. Percentage of IL-4-positive uterine lymphocyte population of the various experimental groups on days 7.5, 9.5, and 10.5 of gestation. Data are represented as the mean ± SEM. *, p < 0.05, **, p < 0.01, as evaluated by nonparametric Mann-Whitney U test.

 
Depletion of CD8+ cells abolished the protective effect of dydrogesterone on the abortion rate in stressed mice by affecting the Th1/Th2 cytokine profile

To investigate whether the protective effect of progesterone substitution with dydrogesterone may be mediated by CD8+ cells, we specifically depleted this cell population using a mAb (33). As presented in Fig. 1B, we observed that the decrease of stress-triggered abortion in dydrogesterone-treated mice could be significantly abrogated if these mice have been CD8-depleted. Further, the dydrogesterone-induced down-regulation of the Th1 cytokines TNF-{alpha}, IFN-{gamma}, and IL-12 present in uterus cells has been abolished in the CD8-depleted mice (Fig. 8A). Depletion of uterine CD8+ cells by injection of Lyt 2.1 has been confirmed by flow cytometry, as presented in Fig. 3B. Interestingly, the ratio between Th1/Th2 cytokines in stressed mice changed upon depletion of CD8 cells; Fig. 8B depicts the predominance of Th1 cytokine levels in stressed, dydrogesterone-treated and CD8-depleted mice with a ratio of 1, compared with a ratio of 0.4 in stressed, dydrogesterone-treated mice with no CD8 depletion.



View larger version (31K):
[in this window]
[in a new window]
  		FIGURE 8. Depletion of CD8 abrogates the pregnancy protective effect of dydrogesterone treatment via an increase of proinflammatory cytokines and the Th1/Th2 ratio. A, Percentage of uterine lymphocyte population positive for the cytokines TNF-{alpha}, IFN-{gamma}, IL-12, IL-4, and IL-10 in stressed mice treated with dydrogesterone, compared with mice that were CD8-depleted in addition to stress and dydrogesterone treatment. Data are represented as the mean ± SEM. *, p < 0.05, **, p < 0.01, as evaluated by nonparametric Mann-Whitney U test. B, Th1/Th2 ratio has been calculated as follows: sum of percentages for TNF-{alpha}+, IFN-{gamma}+, and IL-12+ uterine cells, divided by 3 (to obtain the mean Th1 value). The sum of percentages for IL-4+ and IL-10+ uterine cells was calculated and divided by 2 (to obtain the mean Th2 value). The Th1/Th ratio has been obtained by dividing the mean Th1 value by the mean Th2 value.

 

   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
References
 
It is well known that progesterone mediates effects which are beneficial for the onset, development, and maintenance of pregnancy (10, 11, 12, 13, 14). In contrast, stress interferes with the course of pregnancy and may lead to abortions, putatively via decrease of progesterone (15, 16). Previous studies in our laboratory have demonstrated that stress-triggered abortion can be therapeutically approached by application of a progesterone derivative (31).

In this study, we now deliver evidence that stress exposure induces a decrease of uterine CD8+ cells within 24 h. This decrease can be restored by application of the progesterone derivative dydrogesterone, an observation which is clearly in agreement with the observations of a pregnancy protective CD8+ T cell subpopulation preventing abortion, which are present between days 4.5 and 8.5 of gestation (8, 33). These CD8+ cells, which have been suggested to coexpress the {gamma}{delta} TCR, appear to play a crucial role in Th1/Th2 balance (8, 34), which determines success and failure of pregnancy. Because we were able to restore this population by dydrogesterone, it appears that these cells are highly regulated by adequate levels of progesterone and express progesterone receptors. In our CD8 depletion experiments, we observed that the pregnancy protective effect of dydrogesterone treatment with respect to the abortion rate is almost abolished, thus, we conclude that the protective effect of progesterone and dydrogesterone is predominantly mediated by uterine CD8+ cells (Fig. 9).



View larger version (27K):
[in this window]
[in a new window]
  		FIGURE 9. Synopsis of endocrine-immune bidirectionality in stress-triggered abortion. Hypothetical scenario of the mechanism involved in endocrine-immune bidirectionality during pregnancy. A, In normally progressing pregnancies, adequate levels of progesterone led to a predominance of pregnancy protective cytokines, possibly via PIBF secreted by uterine CD8+ cells, which results in fetal tolerance and pregnancy maintenance. B, In stressed pregnant individuals, levels of progesterone are inadequate and consequently, PIBF is diminished and Th1 cytokines prevail, which results in fetal rejection and pregnancy failure. C, Substitution of progesterone in stressed individuals, e.g., through dydrogesterone application, restores the pregnancy protective PIBF and Th2 predominance and consequently, stress-triggered abortions are abrogated and fetal tolerance ensured.

 
Interestingly, the population of uterine CD8+ cells changes subsequently (gestation days 9.5 and 10.5) from pregnancy protective to abortogenic, suggesting the presence of different functional subpopulations of uterine CD8+ cells throughout gestation. Functional CD8 is a dimeric protein made of either two {alpha}-chains (CD8{alpha}{alpha}) or an {alpha}-chain and a {beta}-chain (CD8{alpha}{beta}) (35, 36, 37). Importantly, CD8{alpha}{alpha} and CD8{alpha}{beta} are differentially expressed on a variety of lymphocytes. CD8{alpha}{beta} is expressed predominantly on the surface of {alpha}{beta}TCR+ T cells and thymocytes (38, 39, 40, 41). Expression of CD8{alpha}{alpha} has been identified on a subset of {gamma}{delta} TCR+ intestinal intraepithelial lymphocytes, NK cells, dendritic cells, and a small fraction of CD4+ T cells, but {alpha}{alpha}-chains are essentially absent on cells of the circulation (41, 42, 43, 44). The differential distributions of CD8{alpha}{alpha} and CD8{alpha}{beta} suggest that these two forms of CD8 are likely to mediate distinct functions. Based on the experimental set-up in our present study, we cannot differentiate whether the CD8 cells express an {alpha}{alpha} or {alpha}{beta}-chain. However, from preliminary results, we know that ~25% of uterine CD8 cells also express CD11c, thus belonging to the family of uterine dendritic cells. The percentage of CD8+/CD11c+ cells slightly diminished with stress and could be restored by dydrogesterone application (data not shown). Gorczynski et al. (45) suggested that CD8+ {gamma}{delta} T suppressor cells and CD8{alpha}{alpha} indoleamine 2, 3 dioxygenase-producing suppressor cells could be stimulated by CD200+CD8– dendritic cells. These CD8+ cells represent an important subset for the induction of cytokines such as IL-4, IL-10, and TGF-{beta} that inhibit macrophages, TCR {alpha}{beta}+ cells and NK or NK {gamma}{delta}T cells, which produce IFN-{gamma}, TNF-{alpha}, and IL-12 and induce placental hemorrhage by up-regulation of fgl2– prothrombinase (25). However, CD8 depletion did not entirely abrogate the protective effect of progesterone and mimicked an abortion rate as seen in the stressed mice. This could be due to cellular infiltration of the fetomaternal interface by progesterone receptor-negative macrophages and NK cells, which are also capable of producing high levels of abortogenic cytokines. Another possible reason may be the existence of a T regulatory cell subpopulation, e.g., Johansson et al. (46) recently described a novel population of small CD3+B220lowCD4–CD8– T cells in the mouse female genital tract with regulatory functions.

The evaluation of IL-4-producing cells in blood and uterus revealed that the percentage of cells producing IL-4 was increased in dydrogesterone-treated stressed mice compared with control or stressed mice. This dramatic increase may be due to the mediating effect of increased PIBF or a direct effect of dydrogesterone on cytokine production, similar to the effect of progesterone itself as demonstrated by Piccinni (47). It has been highly accepted that progesterone induces the production of Th2 cytokines in vivo and vitro (24, 27, 28). Szekeres-Bartho and colleagues and the Margni group (48, 49) report that progesterone induces the secretion of PIBF through lymphocytes and may be involved in the synthesis of asymmetric Abs. PIBF has an antiabortive effect by suppressing NK activity and increases the secretion of Th2 cytokines by immunosuppressor cells (8, 22, 24). In our previous experiment, we further observed that PIBF levels were decreased after stress exposure and could be restored by treatment of dydrogesterone (31).

In our present study using flow cytometry, we observed that the population of TNF-{alpha}+ uterus cells increases after stress exposure, compared with the control group. This stress effect can be counteracted by dydrogesterone treatment. This result confirms the notion of Th1 cytokines deleterious for pregnancy maintenance (21). Indeed, other Th1 cytokines, e.g., IFN-{gamma}, increased after stress exposure in murine matings (23, 50). Interestingly, Tangri and Raghupathy (51) demonstrated that the placenta of an abortion-prone mating such as CBA/J x DBA/2J produced high levels of TNF-{alpha} and IFN-{gamma}, whereas in low abortion mating, e.g., CBA/J x BALB/c, IL-10 is dominant. However, based on recent publications, the Th1/Th2 paradigm has been frequently challenged of late (52, 53), pointing toward the importance for future research in reproductive immunology to deliver additional experimental evidence.

In our present experiments, the percentage of IL-12-producing uterus cells did not change over the days investigated in the various groups, bringing into question the function of IL-12 as a Th1 inducer on the uterine level. We suggest that the physiologic function of IL-12 in pregnancy may rather be around the time of blastocyst adhesion, and additional studies are necessary to elucidate the role of this cytokine in the regulation of the immune system during pregnancy maintenance.

In conclusion, our data imply that stress leads to increased abortions by an alteration of the hormonal system with a consecutive dysregulation of the Th1/Th2 ration via CD8+ cell-dependent pathways. Progesterone substitution by dydrogesterone can abrogate this effect by inducing a pregnancy protective Th2-biased immune response in the decidua.


   Acknowledgments
 
We thank R. Pliet, P. Moschansky, E. Hagen, and P. Busse for their technical support.


   Footnotes
 
1 This work was supported by grants from Solvay Pharmaceuticals Germany and the Charité. S.M.B. received a scholarship from the German Student Exchange Program (Deutscher Akademischer Austauschdienst). Back

2 Address correspondence and reprint requests to Dr. Petra C. Arck, Biomedizinisches Forschungszentrum, Raum 2.0549, Augustenburger Platz 1, 13353 Berlin, Germany. E-mail address: petra.arck{at}charite.de Back

3 Abbreviations used in this paper: PIBF, progesterone-induced blocking factor; RT, room temperature. Back

Received for publication November 3, 2003. Accepted for publication March 1, 2004.


   References
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 References
 

  1. Medawar, P. B.. 1953. Some immunological and endocrine problems raised by evolution of viviparity in vertebrates. Symp. Soc. Exp. Biol. 7:320.
  2. Arck, P., J. Dietl, D. A. Clark. 1999. From the decidual cell internet: trophoblast-recognizing {gamma}{delta} T cells. Biol. Reprod. 60:227.[Abstract/Free Full Text]
  3. Norwitz, E. R., D. J. Schust, S. J. Fisher. 2001. Implantation and the survival of early pregnancy. N. Engl. J. Med. 345:1400.[Free Full Text]
  4. Raghupathy, R.. 2001. Pregnancy: success and failure within the Th1/Th2/Th3 paradigm. Semin. Immunol. 13:219.[Medline]
  5. Clark, D. A., G. Chaouat, R. M. Gorczynski. 2002. Thinking outside the box: mechanisms of environmental selective pressures on the outcome of the materno-fetal relationship. Am. J. Reprod. Immunol. 47:275.
  6. Clark, D. A.. 1999. Signaling at the fetomaternal interface. Am. J. Reprod. Immunol. 41:169.
  7. Coulam, C. B., D. Wagenknecht, J. A. McIntyre, W. P. Faulk, J. F. Annegers. 1991. Occurrence of other reproductive failures among women with recurrent spontaneous abortion. Am. J. Reprod. Immunol. 25:96.
  8. Clark, D. A., P. C. Arck, G. Chaouat. 1999. Why did your mother reject you: immunogenetic determinants of the response to environmental selective pressure expressed at the uterine level. Am. J. Reprod. Immunol. 41:5.
  9. Stray-Pedersen, B., S. Stray-Pedersen. 1984. Etiologic factors and subsequent reproductive performance in 195 couples with a prior history of habitual abortion. Am. J. Obstet. Gynecol. 148:140.[Medline]
  10. Das, C., K. J. Catt. 1987. Antifertility actions of the progesterone antagonist RU 486 include direct inhibition of placental hormone secretion. Lancet 2:599.[Medline]
  11. Wang, M. W., R. B. Heap, M. J. Taussig. 1992. Abnormal maternal behaviour in mice previously immunized against progesterone. J. Endocrinol. 134:257.[Abstract/Free Full Text]
  12. Graham, J. D., C. L. Clarke. 1997. Physiological action of progesterone in target tissues. Endocr. Rev. 18:502.[Abstract/Free Full Text]
  13. Deanesly, R.. 1973. Termination of early pregnancy in rats after ovariectomy is due to immediate collapse of the progesterone-dependent decidua. J. Reprod. Fertil. 35:183.[Abstract/Free Full Text]
  14. Elger, W., M. Fahnrich, S. Beier, S. S. Qing, K. Chwalisz. 1987. Endometrial and myometrial effects of progesterone antagonists in pregnant guinea pigs. Am. J. Obstet. Gynecol. 157:1065.[Medline]
  15. Wiebold, J. L., P. H. Stanfield, W. C. Becker, J. K. Hillers. 1986. The effect of restraint stress in early pregnancy in mice. J. Reprod. Fertil. 78:185.[Abstract/Free Full Text]
  16. Chrousos, G. P., D. J. Torpy, P. W. Gold. 1998. Interactions between the hypothalamic-pituitary-adrenal axis and the female reproductive system: clinical implications. Ann. Intern. Med. 129:229.[Abstract/Free Full Text]
  17. Kovats, S., E. K. Main, C. Librach, M. Stubblebine, S. J. Fisher, R. DeMars. 1990. A class I antigen, HLA-G, expressed in human trophoblasts. Science 248:220.[Abstract/Free Full Text]
  18. King, A., D. S. Allan, M. Bowen, S. J. Powis, S. Joseph, S. Verma, S. E. Hiby, A. J. McMichael, Y. W. Loke, V. M. Braud. 2000. HLA-E is expressed on trophoblast and interacts with CD94/NKG2 receptors on decidual NK cells. Eur. J. Immunol. 30:1623.[Medline]
  19. Mellor, A. L., J. Sivakumar, P. Chandler, K. Smith, H. Molina, D. Mao, D. H. Munn. 2001. Prevention of T cell-driven complement activation and inflammation by tryptophan catabolism during pregnancy. Nat. Immunol. 2:64.[Medline]
  20. Frumento, G., R. Rotondo, M. Tonetti, G. Da Monte, U. Benfatti, G. B. Ferrara. 2002. Tryptophan-derived catabolites are responsible for inhibition of T and natural killer cell proliferation induced by indoleamine 2,3-dioxygenase. J. Exp. Med. 196:459.[Abstract/Free Full Text]
  21. Raghupathy, R.. 1997. Th1-type immunity is incompatible with successful pregnancy. Immunol. Today 18:478.[Medline]
  22. Raghupathy, R., M. Makhseed, F. Azizieh, N. Hassan, M. Al-Azemi, E. Al-Shamali. 1999. Maternal Th1- and Th2-type reactivity to placental antigens in normal human pregnancy and unexplained recurrent spontaneous abortions. Cell. Immunol. 196:122.[Medline]
  23. Arck, P. C., D. A. Ferrick, D. Steele-Norwood, P. J. Egan, K. Croitoru, S. R. Carding, J. Dietl, D. A. Clark. 1999. Murine T cell determination of pregnancy outcome. Cell. Immunol. 196:71.[Medline]
  24. Szekeres-Bartho, J., T. G. Wegmann. 1996. A progesterone-dependent immunomodulatory protein alters the Th1/Th2 balance. J. Reprod. Immunol. 31:81.[Medline]
  25. Clark, D. A., G. Chaouat, P. C. Arck, H. W. Mittrücker, G. A. Levy. 1998. Cytokine-dependent abortion in CBA/J x DBA/2J mice is mediated by the procoagulant fgl2 prothrombinase. J. Immunol. 160:545.[Abstract/Free Full Text]
  26. Ogando, D. G., D. Paz, M. Cella, A. M. Franchi. 2003. The fundamental role of increased production of nitric oxide in lipopolysaccharide-induced embryonic resorption in mice. Reproduction 125:95.[Abstract]
  27. Szekeres-Bartho, J., G. Chaouat, R. Kinsky. 1990. A progesterone-induced blocking factor corrects high resorption rates in mice treated with antiprogesterone. Am. J. Obstet. Gynecol. 163:1320.[Medline]
  28. Szekeres-Bartho, J., D. Philibert, G. Chaouat. 1990. Progesterone suppression of pregnancy lymphocytes is not mediated by glucocorticoid effect. Am. J. Reprod. Immunol. 23:42.
  29. Check., J. H., M. Arwitz, J. Gross, M. Peymer, J. Szekeres-Bartho. 1997. Lymphocyte immunotherapy (LI) increases serum levels of progesterone induced blocking factor (PIBF). Am. J. Reprod. Immunol. 37:17.
  30. Schoeler, H.. 1962. Maintenance of pregnancy in spayed animals with 6-dehydro-retroprogesterone, and masculinization of the female foetus as a test or androgenic activity of progestational compounds. J. Endocrinol. 24:15.
  31. Joachim, R. A., A. C. Zenclussen, B. Polgar, A. Douglas, S. Fest, M. Knackstedt, B. F. Klapp, P. C. Arck. 2003. The progesterone derivate dydrogesterone abrogates murine stress triggered abortion by inducing a Th2 biased local immune response. Steroids 68:931.[Medline]
  32. Marquez, M. G., A. Galeano, S. Olmos, M. E. Roux. 2000. Flow cytometry analysis of intestinal intraepithelial lymphocytes in a model of immunodeficiency in Wistar rats. Cytometry 41:115.[Medline]
  33. Arck, P. C., F. Merali, G. Chaouat, D. A. Clark. 1996. Inhibition of immunoprotective CD8+ T cells as a basis for stress-triggered substance P-mediated abortion in mice. Cell. Immunol. 171:226.[Medline]
  34. Szekeres-Bartho, J., R. Kinsky, M. Kapovic, G. Chaouat. 1991. Complete Freund’s adjuvant treatment of pregnancy females influences resorption rates in CBA/J x DBA/2 matings via progesterone-mediated immunomodulation. Am. J. Reprod. Immunol. 26:82.
  35. Terry, L. A., J. P. DiSanto, T. N. Small, N. Flomenberg. 1990. Differential expression and regulation of the human CD8{alpha} and CD8{beta} chains. Tissue Antigens 35:82.[Medline]
  36. Moebius, U., G. Kober, M. L. Griscelli, T. Hercend, S. C. Meuer. 1991. Expression of different CD8 isoforms on distinct human lymphocyte subpopulations. Eur. J. Immunol. 21:1793.[Medline]
  37. Torero, D., P. L. Tazzari, J. P. DiSanto, P. F. di Celle, D. Raspadori, R. Conte, M. Gobbi, G. B. Ferrara, N. Flomenberg, F. Lauria. 1992. Heterogeneous immunophenotype of granular lymphocyte expansions: differential expression of the CD8{alpha} and CD8{beta} chains. Blood 80:1765.[Abstract/Free Full Text]
  38. Panaccio, M., M. T. Gillespie, I. D. Walker, L. Kirszbaum, J. A. Sharpe, G. H. Tobias, I. F McKenzie, N. J Deacon. 1987. Molecular characterization of the murine cytotoxic T-cell membrane glycoprotein Ly-3 (CD8). Proc. Natl. Acad. Sci. USA 84:6874.[Abstract/Free Full Text]
  39. Norment, A. M., D. R. Littman. 1988. A second subunit of CD8 is expressed in human T cells. EMBO J. 7:3433.[Medline]
  40. Shiue, L., S. D. Gorman, J. R. Parnes. 1988. A second chain of human CD8 is expressed on peripheral blood lymphocytes. J. Exp. Med. 168:1993.[Abstract/Free Full Text]
  41. Torres-Nagel, N., E. Kraus, M. H. Brown, G. Tiefenthaler, R. Mitnacht, A. F. Williams, T. Hunig. 1992. Differential thymus dependence of rat CD8 isoform expression. Eur. J. Immunol. 22:2841.[Medline]
  42. Gapin, L., H. Cheroutre, M. Kronenberg. 1999. Cutting edge: TCR{alpha}{beta}+ CD8{alpha}{alpha}+ T cells are found in intestinal intraepithelial lymphocytes of mice that lack classical MHC class I molecules. J. Immunol. 163:4100.[Abstract/Free Full Text]
  43. Jarry, A., N. Cerf-Bensussan, N. Brousse, F. Seiz, D. Guy-Grand. 1990. Subsets of CD3+ and CD3– lymphocytes isolated from normal human gut epithelium display phenotypic features different from their counterparts in peripheral blood. Eur. J. Immunol. 20:1097.[Medline]
  44. Hayday, A., E. Theodoridis, E. Ramsburg, J. Shires. 2001. Intraepithelial lymphocytes: exploring the third way in immunology. Nat. Immunol. 2:997.[Medline]
  45. Gorczynski, R. M., S. Hadidi, G. Yu, D. A. Clark. 2002. The same immunoregulatory molecules contribute to successful pregnancy and transplantation. Am. J. Reprod. Immunol. 48:18.
  46. Johansson, M., N. Lycke. 2003. A unique population of extrathymically derived {alpha}{beta} TCR+CD4–CD8– T cells with regulatory functions dominates the mouse female genital tract. J. Immunol. 170:1659.[Abstract/Free Full Text]
  47. Piccinni, M. P., M. G. Giudizi, R. Biagiotti, L. Beloni, L. Giannarini, S. Sampognaro, P. Parronchi, R. Manetti, F. Annunziato, C. Livi. 1995. Progesterone favors the development of human T helper cells producing Th2-type cytokines and promotes both IL-4 production and membrane CD30 expression in established Th1 cell clones. J. Immunol. 155:128.[Abstract]
  48. Kelemen, K., I. Bognar, M. Paal, J. Szekeres-Bartho. 1996. A progesterone-induced protein increases the synthesis of asymmetric antibodies. Cell. Immunol. 167:129.[Medline]
  49. Malan Borel, I., S. M. Freire, E. Rivera, A. Canellada, R. A. Binaghi, R. A. Margni. 1999. Modulation of the immune response by progesterone-induced lymphocyte factors. Scand. J. Immunol. 49:244.[Medline]
  50. Hildebrandt, M., P. C. Arck, S. Kruber, H. U. Demuth, W. Reutter, B. F. Klapp. 2001. Inhibition of dipeptidyl peptidase IV (DP IV, CD26) activity abrogates stress-induced, cytokine-mediated murine abortions. Scand. J. Immunol. 53:449.[Medline]
  51. Tangri, S., R. Raghupathy. 1993. Expression of cytokines in placentas of mice undergoing immunologically mediated spontaneous fetal resorptions. Biol. Reprod. 49:850.[Abstract]
  52. Svensson, L., M. Arvola, M. A. Sallstrom, R. Holmdahl, R. Mattsson. 2001. The Th2 cytokines IL-4 and IL-10 are not crucial for the completion of allogeneic pregnancy in mice. J. Reprod. Immunol. 51:3.[Medline]
  53. Zenclussen, A. C., S. Fest, P. Busse, R. Joachim, B. F. Klapp, P. C. Arck. 2002. Questioning the Th1/Th2 paradigm in reproduction: peripheral levels IL-12 are down-regulated in miscarriage patients. Am. J. Reprod. Immunol. 48:245.



This article has been cited by other articles:


Home page
 Hum Reprod UpdateHome page
L. R. Guerin, J. R. Prins, and S. A. Robertson
Regulatory T-cells and immune tolerance in pregnancy: a new target for infertility treatment?
Hum. Reprod. Update, September 1, 2009; 15(5): 517 - 535.
[Abstract] [Full Text] [PDF]

Home page
 J. Immunol.Home page
L. M. Moldenhauer, K. R. Diener, D. M. Thring, M. P. Brown, J. D. Hayball, and S. A. Robertson
Cross-Presentation of Male Seminal Fluid Antigens Elicits T Cell Activation to Initiate the Female Immune Response to Pregnancy
J. Immunol., June 15, 2009; 182(12): 8080 - 8093.
[Abstract] [Full Text] [PDF]

Home page
 Hum ReprodHome page
E. Kondoh, T. Okamoto, T. Higuchi, K. Tatsumi, T. Baba, S. K. Murphy, K. Takakura, I. Konishi, and S. Fujii
Stress affects uterine receptivity through an ovarian-independent pathway
Hum. Reprod., April 1, 2009; 24(4): 945 - 953.
[Abstract] [Full Text] [PDF]

Home page
 Am. J. Pathol.Home page
R. A. Joachim, B. Handjiski, S. M. Blois, E. Hagen, R. Paus, and P. C. Arck
Stress-Induced Neurogenic Inflammation in Murine Skin Skews Dendritic Cells Towards Maturation and Migration: Key Role of Intercellular Adhesion Molecule-1/Leukocyte Function-Associated Antigen Interactions
Am. J. Pathol., November 1, 2008; 173(5): 1379 - 1388.
[Abstract] [Full Text] [PDF]

Home page
 Reproductive SciencesHome page
N. S. Macklon, M. H. van der Gaast, A. Hamilton, B. C. J. M. Fauser, and L. C. Giudice
The Impact of Ovarian Stimulation With Recombinant FSH in Combination With GnRH Antagonist on the Endometrial Transcriptome in the Window of Implantation
Reproductive Sciences, April 1, 2008; 15(4): 357 - 365.
[Abstract] [PDF]

Home page
 J EndocrinolHome page
C Dosiou, A E Hamilton, Y Pang, M T Overgaard, S Tulac, J Dong, P Thomas, and L C Giudice
Expression of membrane progesterone receptors on human T lymphocytes and Jurkat cells and activation of G-proteins by progesterone
J. Endocrinol., January 1, 2008; 196(1): 67 - 77.
[Abstract] [Full Text] [PDF]

Home page
 Biol. Reprod.Home page
S. M Blois, U. Kammerer, C. A. Soto, M. C Tometten, V. Shaikly, G. Barrientos, R. Jurd, D. Rukavina, A. W Thomson, B. F Klapp, et al.
Dendritic Cells: Key to Fetal Tolerance?
Biol Reprod, October 1, 2007; 77(4): 590 - 598.
[Abstract] [Full Text] [PDF]

Home page
 Biol. Reprod.Home page
M. Tometten, S. Blois, A. Kuhlmei, A. Stretz, B. F. Klapp, and P. C. Arck
Nerve Growth Factor Translates Stress Response and Subsequent Murine Abortion via Adhesion Molecule-Dependent Pathways
Biol Reprod, April 1, 2006; 74(4): 674 - 683.
[Abstract] [Full Text] [PDF]

Home page
 Proc. Natl. Acad. Sci. USAHome page
P. A. Nepomnaschy, K. B. Welch, D. S. McConnell, B. S. Low, B. I. Strassmann, and B. G. England
Cortisol levels and very early pregnancy loss in humans
PNAS, March 7, 2006; 103(10): 3938 - 3942.
[Abstract] [Full Text] [PDF]

Home page
 J. Immunol.Home page
L. Shao, A. R. Jacobs, V. V. Johnson, and L. Mayer
Activation of CD8+ Regulatory T Cells by Human Placental Trophoblasts
J. Immunol., June 15, 2005; 174(12): 7539 - 7547.
[Abstract] [Full Text] [PDF]

Home page
 Ann Rheum DisHome page
R H Straub, F Buttgereit, and M Cutolo
Benefit of pregnancy in inflammatory arthritis
Ann Rheum Dis, June 1, 2005; 64(6): 801 - 803.
[Full Text] [PDF]

Home page
 J. Immunol.Home page
S. Blois, M. Tometten, J. Kandil, E. Hagen, B. F. Klapp, R. A. Margni, and P. C. Arck
Intercellular Adhesion Molecule-1/LFA-1 Cross Talk Is a Proximate Mediator Capable of Disrupting Immune Integration and Tolerance Mechanism at the Feto-Maternal Interface in Murine Pregnancies
J. Immunol., February 15, 2005; 174(4): 1820 - 1829.
[Abstract] [Full Text] [PDF]

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow Request Permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Blois, S. M.
Right arrow Articles by Arck, P. C.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Blois, S. M.
Right arrow Articles by Arck, P. C.

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS