Female patients in reproductive age with systemic lupus erythematosus and fertility complications together are observed by rheumatologists, gynecologists, and reproductive immunologists. The paper notes the presence of autoantibodies to zona pellucida, to phospholipids (phosphatidyl serine, phosphatidyl ethanolamine, phosphatidyl inositol, phosphatidyl glycerol, phosphatidic acid, annexin V, beta-2 glycoprotein I, and cardiolipin) and of isoantibodies to sperm cells. Isoantibodies to sperm cells are not significantly predominant, but autoimmunity is well expressed in IgG positivity against phosphatidyl inositol, phosphatidyl ethanolamine, phosphatidyl serine, cardiolipin, and beta-2 glycoprotein I, as well as antizona pellucida antibodies in IgG isotype. According to the levels of autoantibodies we have to choose preventive treatment to protect mother and her foetus.
Although autoimmune diseases (AIDs) are not in general considered a major cause of impaired reproductive capacity some AIDs are associated with infertility or pregnancy wastage. Autoimmune conditions may affect all stages of fertility, from ovarian to implantation failure and pregnancy loss. Systemic lupus erythematosus (SLE) is one of the most common autoimmune disorders that affect women during their childbearing age. Prevalence of SLE in Europe is similar as in USA, ranging from 3.3 to 4.8 per 100.000 person-years [
Typical clinical symptoms of SLE include fatigue, fever, arthritis, a photosensitive rash serositis, Raynaud phenomenon, glomerulonephritis, vasculitis, and hematologic abnormalities. Although patients with SLE are as fertile as women in the general population, their pregnancies could be associated with complications [
High rate of fetal losses up to 45 percent in SLE women has been described in some studies [
There is a wide variety of autoantibodies associated with SLE. Some of the antibodies are helpful in the diagnostics of the illness, while others are more useful in detecting and monitoring disease activity or potential complications. Antibodies to native double-strand DNA (dsDNA) are relatively specific for the diagnosis of SLE. Serum antinuclear antibodies (ANAs) are found in nearly all individuals with active SLE. Significant for SLE diagnosis is assessment of spectrum extractable nuclear antibodies (Sm, La, Ro), antibodies to ribonucleoprotein (RNP), complement and N-methyl-D-aspartate receptor (ENA panel).
Antiphospholipid antibodies (aPLs) form a large group of antibodies that are detected in patients with SLE as well as with other autoimmune conditions. These antibodies are associated with a wide range of potential complications during pregnancy, including miscarriage, fetal death, intrauterine growth restriction, prematurity, and preeclampsia-especially in the primary antiphospholipid syndrome (APS).
In SLE women, pregnancy should be best planned during periods of disease stabilization and nephritis remission lasting at least six months. Closed collaboration of rheumatologist, obstetrician, and neonatologist is necessary for successful pregnancy and delivery. Fulfilling these prerequisites, there is still around 5% of women with SLE that have fertility or pregnancy problems [
A complication in reproduction is very often a reason for thorough immunological examination. We present here our experience with SLE patients from the perspective of reproductive immunology.
The general aim of our paper is to evaluate the results of screening tests in reproductive immunology in women of childbearing age with SLE. The primary aim is to investigate the occurrence of autoantibodies to zona pellucida and eight various phospholipids and occurrence of isoantibodies to sperm cells in patients with SLE in remission planning pregnancy.
The study group consists of 52 woman with SLE (mean time from diagnosis
The control group consisted of 25 age-matched healthy fertile women (age 24–43 years, mean
Detection of autoantibodies to zona pellucida, eight phospholipids (phosphatidyl-serine, phosphatidyl-ethanolamine, phosphatidyl-inositol, DL-glycerol, phosphatidic acid, annexin V, beta-2-glycoprotein I, and cardiolipin), and isoantibodies to sperm cells was performed as described below.
Both groups were tested with
We have performed tray agglutination test (TAT) [
The i-MAR test was performed to analyze the antisperm response in IgG and IgA class. One microliter of native sperm suspension, 1
We use ovulatory cervical mucus taken by a special syringe 5 days after condom protected intercourse that is planned to happen during ovulation time. Sperm capillary penetration test serves as screening test, and i-MAR test detects local sperm antibodies of IgG and/or IgA [
Classic ELISA tests (Laboserv) for detection of zona pellucida antibodies in isotype of IgG and IgM were used.
ELISA was also used for detection of aPL against phosphatidyl serine, phosphatidyl ethanolamine, phosphatidyl inositol, DL-glycerol, and phosphatidic acid. For ELISA, polystyrene microtiter plates were coated with 50
Serum from patients and controls, diluted to a ratio of 1 : 50 in TBS containing 10% FCS were added to the wells after the wash. The plates were incubated for 2 hours at room temperature and then washed five times with TBS containing 0.05% Tween 20. Fifty microliters of peroxidase-conjugated antihuman immunoglobulin (IgG, IgA, or IgM) was added and incubated for 1 hour at room temperature. The plates were then washed five times in TBS Tween. Fifty microliters of substrate solution were added to each well and incubated in the darkness for 30 min at room temperature. The reaction was stopped by adding 50
For detection and quantification of serum antiannexin V (Szabo-Scandic Handels GMBH & Co KG, Vienna, Austria) commercial ELISA kits were used, as well as for detection of IgG and IgA antibodies against beta2-glycoprotein I (beta2GPI) (Immunotech, Prague Division, Czech Republic), cardiolipin levels in IgG and IgM (Millenia, London, UK) (Table
Prevalence of positive results for each performed antibody analysis with comparison between study and control groups.
Antibody | Number of positive subjects | Odds ratio (95% confidence interval) | |
---|---|---|---|
Study group ( |
Control group ( |
||
Antisperm antibodies (ASAs) | |||
Serum ASA | 5/52 (9.6%) | 1/25 (4.0%) | 2.55 (0.28–23.10) |
Cervical ovulatory mucus ASA | 6/52 (11.5%) | 1/25 (4.0%) | 3.13 (0.36–27.52) |
Antizona pellucida antibodies (aZP) | |||
aZP IgG |
|
|
N/A |
aZP IgM | 1/52 (1.9%) | 0/25 (0.0%) | N/A |
Antiphospholipid antibodies (aPL) against | |||
phosphatic acid IgG | 1/52 (1.9%) | 0/25 (0.0%) | N/A |
phosphatic acid IgM | 0/52 (0.0%) | 0/25 (0.0%) | N/A |
phosphatidyl glycerol IgG | 2/52 (3.8%) | 0/25 (0.0%) | N/A |
phosphatidyl glycerol IgM | 0/52 (0.0%) | 0/25 (0.0%) | N/A |
phosphatidyl inositol IgG |
|
|
N/A |
phosphatidyl inositol IgM | 2/52 (3.8%) | 1/25 (4.0%) | 0.96 (0.08–11.12) |
phosphatidyl ethanolamine IgG |
|
|
N/A |
phosphatidyl ethanolamine IgM | 3/52 (5.8%) | 1/25 (4.0%) | 1.47 (0.15–14.88) |
phosphatidyl serine IgG |
|
|
N/A |
phosphatidyl serine IgM | 2/52 (3.8%) | 2/25 (8.0%) | 0.46 (0.06–3.47) |
cardiolipin IgG |
|
|
|
cardiolipin IgM | 4/52 (7.7%) | 0/25 (0.0%) | N/A |
beta-2-glycoprotein IgG |
|
|
N/A |
beta-2-glycoprotein IgA | 9/52 (17.3%) | 0/25 (0.0%) | N/A |
annexin-V | 7/52 (13.5%) | 0/25 (0.0%) | N/A |
Note: statistically significant values are marked in bold script.
Because equality of variance test proved that both groups were similar in age (as an independent baseline characteristic), we were able to assess individual antibody prevalence percentages as follows: nonzero prevalences of tested antibodies were all higher in the study group compared to controls with the exception of antiphosphatidyl inositol IgM and antiphosphatidyl serine IgM (3.8% versus 4.0% and 3.8% versus 8.0%, resp.). Difference between study group and control group in antisperm antibodies (ASAs) predominance was only nominal and statistically insignificant in both serum ASA (9.6% versus 4.0%, odds ratio 2.55, 95% confidence interval 0.28–23.10) and cervical ovulatory mucus ASA (11.5% versus 4.0%, OR 3.13, 95% CI 0.36–27.52). Concerning antizona pellucida (aZP) and antiphospholipid (aPL) antibodies, no significant difference was observed in IgM subtypes across all tests. However, in aZP IgG subclass a difference was found to be significant in favor of study group (12 out of 52 subjects, 23.0%, versus zero of 25 subjects). The same was true for almost all individual aPL antibodies in IgG isotype, the exception being antiphosphatic acid IgG and antiphosphatidyl glycerol IgG (detailed results are shown in Table
Pregnancy is the most significant exception to the immunological rules where the mothers body not only tolerates, but also broadly supports semiallogenic blastocyst, embryo, and fetus. Remarkable tolerance is provided by complex immunoregulatory mechanisms between the mother and fetal trophoblast. Failure of this tolerance is one of the most common causes of fetal loss. As mentioned in many papers, reproductive failure including recurrent pregnancy loss (RPL) and infertility has been linked to presence of various autoantibodies [
In SLE as well as in other autoimmune diseases an overproduction of autoantibodies is described, especially against organ-nonspecific antigens. This feature is based on the loss of B-cell tolerance to antigens, nucleic acids, and their binding proteins. Antinuclear antibodies can trigger proliferation of autoreactive lymphocytes in genetically susceptible individuals under the influence of environmental factors (infections, drugs, toxins, smoking, and hormonal factors) leading to expansion of their specificity towards more nuclear antigens or other structures (elements of blood, plasma components, coagulation factors, and complement cascade) [
The aim of our study was to evaluate the occurrence of variety of antibodies possibly responsible for immunological reproduction failure in women with SLE and compare these results with findings in healthy controls.
Many retrospective and prospective studies have proven effect of antisperm antibodies in various human body fluids (serum, semen, cervical ovulatory mucus, follicular, or peritoneal fluid) on fertility of women and men [
It seems clear that repeated sperm exposition is not sufficient for most women to start production of antisperm antibodies. Other factors such as mechanical, chemical, or inflammatory lesions of the mucosa in female genital system play an important role [
Antisperm antibodies (ASAs) in both of our groups were determined to evaluate the difference. In contrast to the above mentioned facts we have found no significant difference between antisperm antibody prevalence in study group and control group. Our prevalence data from control group are in consent with evidence about the presence of these antibodies in healthy women with undisturbed reproductive capability (several authors reported that these antibodies occur in approximately 1 to 2.5% of fertile men and in 4% of fertile women) [
Zona pellucida (ZP), ten micrometers “strong” oogenetic glycoprotein matrix, is formed during oogenesis and its thickness increases with the growth of the oocyte. It serves basically as a protective layer surrounding the oocyte as it matures and is composed of three major glycoproteins ZP1, ZP2, and ZP3. These glycoproteins seem to be the important ovarian antigens participating in the etiology of some infertility disorders, including autoimmune premature ovarian failure (POF). POF can be seen in approximately 1-2% of healthy women, in 30% is connected with some autoimmune causes and is often presented in women with SLE [
In the present study we have found borderline significant prevalence of aZP IgG antibodies in the study group in comparison with the control group.
Antiphospholipid antibodies (aPL) form a group of antibodies that is probably associated with compromised fertility not only in SLE women. Although some retrospective studies reported nonsignificant association between antiphospholipid antibodies and pregnancy loss in patients with SLE other studies have confirmed statistically significant interrelation between them [
A review of 10 studies of 554 women with SLE found that fetal demise was more common in those with aPL (38 to 59 percent versus 16 to 20 percent in those without such antibodies), LAs (36 versus 13 percent), or aCL (39 versus 18 percent). Fetal loss typically occurs after 10 weeks gestation [
The relationship between autoimmunity and reproduction has long been recognized. Although in one critical review about the impact of abnormal autoimmunity on female fertility doubts about influence of aPL to POF in SLE were expressed SLE remains the autoimmune disease mostly compromised by humoral changes. It seems possible that another unexplained mechanism (e.g. presence of other antibodies as an IgG antilaminin-I antibodies, anticorpus luteum glycoprotein and antilymphocytotoxic autoantibodies, or antimitochondrial M5 type antibodies) could be involved [
Based on our results and long-term experience, we believe that it is appropriate in women with SLE with recurrent and otherwise unexplained fetal loss to recommend a detailed evaluation by a specialist in reproductive immunology. In the case of positive findings of autoantibodies, preventive treatment is to be considered (e.g. corticosteroids, low molecular weight heparin, low dose of aspirin, intravenous immunoglobulin therapy). Because the evidence supporting association of various individual antibodies with reproductive failure is inconclusive a new meta-analysis of antibody profiles in women at risk may be of benefit.
The authors state no conflict of interests.
The authors would like to thank all west Bohemian rheumatologists for close collaboration. The study was supported by the Grant GA UK no. 696212 from the Grant Agency of the Charles University in Prague, and from Charles University Research Fund (Project Number P36).