Successful pregnancy is considered a Th1–Th2 cooperation phenomenon (Th, T-helper), with a predominantly Th2-type lymphocytes response, together with the emerging role of interleukin (IL)-12, IL-15 and IL-18 and of other unidentified soluble factors dependent on natural killer (NK) cells. In the pathogenesis of recurrent spontaneous abortion (RSA), immunological factors have been involved such as decidual cells, complement system, cytokines and genes of the hystocompatibility complex that can determine the success or the failure of a pregnancy. A deeper insight into apparently unexplained RSA shows increasing evidences supporting both alloimmune and autoimmune mechanisms, with autoantibodies playing a major role. The best-characterised pathogenic autoantibodies are anti-phospholipid antibodies, and also other autoantibodies, such as anti-Ro/SSA and anti-La/SSB, have been found to be associated with an increased rate of abortion, poor pregnancy outcome and several other obstetric manifestations. This intriguing mixture has been unveiled only in the last few years with the discovery of novel pathogenic mechanisms that can be targeted in the prevention and treatment of obstetrical complications occurring in the course of an autoimmune disease.
Introduction
The relationships between pregnancy and autoimmunity are represented by a bidirectional model: autoimmune diseases (AIDs) can be affected by pregnancy, and, in turn, pregnancy can be affected by AIDs, mainly during the very early and very late stages of gestation. The coexistence of pregnancy and AID is far from rare . During early pregnancy subclinical and clinical autoimmune processes seem to be capable of causing pregnancy loss as well as foetal abnormalities such as neonatal lupus syndrome (NLS). Immune processes occurring acutely during late pregnancy can endanger both maternal and foetal life and need therefore to be clinically recognised as such. In normal pregnancy, the maternal immunological response against trophoblast antigens, concomitant with a transient depression of maternal cell-mediated immunity to protect semi-allogenic embryo from rejection, is the predominant mechanism for high live birth rate. Successful pregnancy is considered Th1–Th2 cooperation phenomenon (Th, T-helper), with a predominantly Th2-type lymphocytes response. Failure in the generation of Th2-type responses is associated with recurrent spontaneous abortion (RSA), obstetric complications and poor pregnancy outcome. A deeper insight into apparently unexplained infertility and RSA shows increasing evidences supporting both alloimmune and autoimmune mechanisms, in which natural killer (NK) cells and autoantibodies seem to play a relevant role. The autoimmune factors belong to the different branches of the immune system, including decidual cells, allo- and autoantibodies, the complement system, cytokines and antigen expression of major histocompatibility complex (MHC) hystocompatibility antigens (HLAs).
Anti-phospholipid syndrome (APS) and systemic lupus erythematosus (SLE) are the AIDs that most commonly jeopardise pregnancy, and potential adverse events consist of miscarriage, intrauterine foetal restriction, congenital heart block (CHB) and preterm delivery and flares of lupus activity. Indeed, the presence of anti-phospholipid antibodies (aPLs) and lupus activity are the most important predictors of poor obstetric outcome. Women with SLE have also increased rates of hypothyroidism and autoimmune thyroid disease defined as the presence of thyroid antibodies with or without thyroid dysfunction. Anti-thyroid antibodies and aPLs are associated with reduced fertility, miscarriage and preterm delivery, but the precise mechanisms by which thyroid antibodies, as well as those against other tissues, are suppressed during pregnancy, and often exacerbate after delivery, remain obscure. Presumably, the rapid reduction in immune suppressor functions following delivery leads to the reestablishment and/or exacerbation of these conditions.
In the pathogenesis of RSA, defined as three or more consecutive miscarriages, preexisting or underlying AIDs are often implied, although it may also occur in otherwise healthy women. RSA is manifested mainly by the tenth week of pregnancy, in pre-foetal period, and seems to be a distinct entity from foetal loss that occurs after the tenth week.
Wegmann first introduced the idea that the maternal–foetal relationship is bidirectional and that immunostimulation might be more important than immunosuppression . His group also suggested that allopregnancy is a Th2 phenomenon, the Th1 cytokines being abortifacient and the Th2 cytokines being protective. The main limit of the Th1/Th2 model in pregnancy is the exclusion of NK cells as having a central role and did not predict the NK-mediated immune control of vascularisation. Furthermore, the generic distinction within pro-inflammatory (IL-12, IL-15) and anti-inflammatory (IL-11, IL-13) cytokines cannot be considered as conclusive. Uterine NK cells (uNKs) comprise the largest population at implantation site, and their activity, characteristics and abundance suggest that they participate at the ‘decidualisation’ process, that, vice versa, induces NK activation and recruitment in each menstrual cycle.
This immunological response against trophoblast antigens, concomitant with a transient depression of maternal cell-mediated immunity to protect the semi-allogenic embryo from rejection, is the predominant mechanism for high live birth rate.
The interactions between the maternal immune system and the ‘foeto-placental unit’
Implantation is an event depending on at least several steps. At every step, there is a continuous embryo–uterus interaction, and, as the implantation window was discovered and the concept of uterine receptivity emerged, the idea that cytokines could be central in such a process dramatically raised . In human placentation, foetus-derived extravillous trophoblasts infiltrate the deciduas and replace the endothelium of the uterine spiral arteries to ensure the development of an adequate blood supply to the foetal–maternal unit .
During apposition/adhesion, the induction of adhesion molecules and the proper ligands are critical steps. Hence, it is very important that the expression of receptors/ligands at the cell surface of the embryo and uterus coincides. There is an involvement of interleukins, but also an important hormonal regulation, both in the uterus and on the embryo. Subsequently, invasion-penetration occurs, and, at this step several enzymes are involved, especially matrix metalloproteases (MMPs), counterbalanced by their inhibitors .
The interaction between immune system and maternal foetal placental unit is a role played by the distribution of helper T lymphocytes. The lymphocyte subpopulations change before and during pregnancy, with increased production of Th1 cytokines in non-pregnant endometrium .
In recent years, highly specialised cell subpopulations have been identified that represent a leap forward over previous models. The Th1/Th2 paradigm has been superseded by models that not only include other subpopulations, but overall do not limit the range of possible functions of Th1 and Th2 cells to a predetermined number . Within the CD4+ T cells, Th17 cells appear to constitute a new cell line, distinct from Th1 and Th2 cells, predominantly pro-inflammatory that is stimulated by IL-23 . Another cell type, the Foxp3 Treg, has been called a contributory factor in the pathology of pregnancy .
What has so far been demonstrated in women with RSA is a primarily Th1 pattern, whereas normal women show essentially a Th2 pattern, a condition that may be predictive of success of pregnancies.
Therefore, there is evidence that the foetal–placental unit may redirect maternal immunity towards a humoral response during pregnancy. It is known since long time that AIDs driven by cellular immunity, such as rheumatoid arthritis (RA), improve during pregnancy, while those driven by a humoral response, such as SLE, tend to deteriorate . Granulocyte-macrophage colony-stimulating factor (GM-CSF) plays a key role in this complicated balance as it can stimulate the production of prostaglandins, tumour necrosis factor (TNF), IL-1 and plasminogen activator, as well as the production of IL-6 . We showed that women with RSA have significantly lower levels of GM-CSF, and that one of the beneficial effects played by intravenous immunoglobulins (IVIgs) in the treatment of RSA, may include the induction of an increase in circulating GM-CSF levels .
The interactions between the maternal immune system and the ‘foeto-placental unit’
Implantation is an event depending on at least several steps. At every step, there is a continuous embryo–uterus interaction, and, as the implantation window was discovered and the concept of uterine receptivity emerged, the idea that cytokines could be central in such a process dramatically raised . In human placentation, foetus-derived extravillous trophoblasts infiltrate the deciduas and replace the endothelium of the uterine spiral arteries to ensure the development of an adequate blood supply to the foetal–maternal unit .
During apposition/adhesion, the induction of adhesion molecules and the proper ligands are critical steps. Hence, it is very important that the expression of receptors/ligands at the cell surface of the embryo and uterus coincides. There is an involvement of interleukins, but also an important hormonal regulation, both in the uterus and on the embryo. Subsequently, invasion-penetration occurs, and, at this step several enzymes are involved, especially matrix metalloproteases (MMPs), counterbalanced by their inhibitors .
The interaction between immune system and maternal foetal placental unit is a role played by the distribution of helper T lymphocytes. The lymphocyte subpopulations change before and during pregnancy, with increased production of Th1 cytokines in non-pregnant endometrium .
In recent years, highly specialised cell subpopulations have been identified that represent a leap forward over previous models. The Th1/Th2 paradigm has been superseded by models that not only include other subpopulations, but overall do not limit the range of possible functions of Th1 and Th2 cells to a predetermined number . Within the CD4+ T cells, Th17 cells appear to constitute a new cell line, distinct from Th1 and Th2 cells, predominantly pro-inflammatory that is stimulated by IL-23 . Another cell type, the Foxp3 Treg, has been called a contributory factor in the pathology of pregnancy .
What has so far been demonstrated in women with RSA is a primarily Th1 pattern, whereas normal women show essentially a Th2 pattern, a condition that may be predictive of success of pregnancies.
Therefore, there is evidence that the foetal–placental unit may redirect maternal immunity towards a humoral response during pregnancy. It is known since long time that AIDs driven by cellular immunity, such as rheumatoid arthritis (RA), improve during pregnancy, while those driven by a humoral response, such as SLE, tend to deteriorate . Granulocyte-macrophage colony-stimulating factor (GM-CSF) plays a key role in this complicated balance as it can stimulate the production of prostaglandins, tumour necrosis factor (TNF), IL-1 and plasminogen activator, as well as the production of IL-6 . We showed that women with RSA have significantly lower levels of GM-CSF, and that one of the beneficial effects played by intravenous immunoglobulins (IVIgs) in the treatment of RSA, may include the induction of an increase in circulating GM-CSF levels .
The restricted antigen presentation
To explain the loss of tolerance to foetal allografts, the phenomenon of antigen presentation restricted has been implicated that, if lost, can lead to the production of antibodies to the foetus .
The antigens that can lead in the production of antibodies can be processed in an endocytic manner by antigen-presenting cells (APCs) positive for MHC class II and then can be presented to CD4+ T cells as peptides bound to MHC.
The entry in the maternal circulation of foetal cells may trigger an antigenic response; nevertheless, the primary spring is the release of soluble MHC I and II antigens from placental foetal cells such as macrophages .
These antigens are then endocytosed by APCs present in the maternal lymphoid tissue with the production of maternal antibodies directed to foetal HLAs with the formation of soluble immune complexes that can easily undergo clearance without harm to the foetus. If antibodies directed towards the foetus escape this clearance mechanism, they may reach the placenta; the monomeric immunoglobulin-G’s (IgGs) are indeed selectively transported by the Fc receptors on the trophoblast . Antibodies to foetal cells, potentially harmful, are removed by placental macrophages after forming immune complexes with antigens present in the foetal milieu leading to the production of large amounts of antibodies direct to foetal cells that can cause damage to the foetus.
Production of alloantibodies
The production of maternal alloantibodies to paternal antigens is well documented, and it can be revealed since the early stages of pregnancy . This is a response direct towards few HLA antigens of the foetus, modulated by soluble HLA and by antidiotype antibodies .
The role of alloantibodies in the maintenance of pregnancy is still under debate. Under normal conditions, in early pregnancy there is the production of anti-MHC class II antibodies that have no pathogenic effect unless directed to antigens present on platelets .
By contrast, it seems to be clarified that their absence turns maternal immune response to a noxious Th1 pattern, and that women who shared HLA with the foetus with consequent lack of alloantibodies can abort repeatedly .
While it is clear why we have alloantibodies, there is great controversy concerning the mechanisms by which alloantibodies play these beneficial roles. Probably, they can suppress NK activity with the stimulation of the production of GM-CSF and CSF-1 .
Pregnancy and complement activation
The complement system consists of a complex group of serum proteins able to mediate several reactions, whether immune or not. Indeed, it participates in inflammatory processes through the release of biologically active products (the anaphilotoxins C3a, C4a, C5a), and it promotes cell lysis, chemotaxis, immune adherence, phagocytosis and anaphylaxis .
The trophoblast expresses non-maternal gene products; thus, it acts as a target for maternal alloantibodies and subsequent complement fixation. Deposit of complement components can be found on apical membranes of the syncytiotrophoblast; C3d and C9 (but not C4) can be seen at the level of basal membranes. C1 and C9 are located on the foetal vessel walls, C6 cytoplasmic granules as foetal vascular endothelium, and endothelial cells in culture can synthesise C3, C5, C6, C8, C9, H and have mRNA for B, D, I .
The activation of complement is a critical mechanism that can endanger pregnancy. The role played by complement is evident from several studies. It has been showed that targeted inhibition of complement activation may prevent features of preeclampsia in murine experimental models, for instance, by administering a single dose of the C3 inhibitor CR2-Crry to two strains of mice (DBA/2 and CBA/J) (expecting that the pregnancies might show features of preeclampsia and of immunologically mediated pregnancy loss) .
Other experimental observations suggest that increased complement activation causes and/or perpetuates inflammation during pregnancy. Complement activation is linked also with occurrence of preeclampsia . Indeed, excessive activation or insufficient regulation of complement recruits leucocytes and unleashes potent inflammatory and anti-angiogenic mediators associated with placental insufficiency and maternal endothelial dysfunction characteristic of preeclampsia .
Innate immunity
A deeper insight into apparently unexplained infertility and RSA shows increasing evidences supporting both alloimmune and autoimmune mechanisms, in which NK cells and autoantibodies seem to play a relevant role .
Peripheral NK cells
NK cells constitute the predominant leucocyte population present in the endometrium at the time of implantation and in early pregnancy while normally comprise about 10–15% of peripheral blood lymphocytes. During normal pregnancy, pNK cell activity and percentages tend to increase in the first trimester and then tend to decline from the second trimester with a second fall in the third trimester of pregnancy .
pNK cells are thymus independent cells that do not require pre-immunisation, thus reflecting their innate character . pNK cells are controlled by a highly complex system of inhibitory and activatory receptor–ligand interactions, sensing change in HLA expression. It has been suggested that pNK cells may either prevent/limit autoimmune responses or may have a permissive role in autoimmunity .
Uterine NK cells
Uterine NK (uNK) cells during menstrual cycle account for almost the 70% of total leucocytes prior to menstruation during late secretory phase. After implantation, the leucocytes increase from less than 10% of the stromal cells as they are in the proliferative and early secretory phase endometrium to over 30%, due to the increase in uNK .
uNK cells are classically CD56+/CD16− but can readily be activated by short-term exposure to cytokines such as IL-2 to become lymphokine-activated killer cells that express CD16 and release TNF and interferon gamma (IFN-γ), together with GM-CSF, IL-10, transfer growth factor beta (TGF-β), IL-8 and leukaemia inhibitory factor (LIF) . uNK cells are important for establishing spiral artery formation in the endometrium by releasing IFN-γ . It was demonstrated that IFN-γ-deficient mice exhibit preeclampsia-like symptoms and result in lower litter size . uNK cells support proper trophoblast and placental growth as well as vascularisation of the deciduas, and, through the production of immunomodulatory molecules, they participate in the creation of local immunosuppression at the maternal–foetal interface .
NK cells: harmful in pregnancy
Not only the status but also the number of NK cells appears to be crucial in the success of pregnancies . Increased circulating NK cell percentages are found exclusively during the secretory phase and not in endometrium. NK cells have been called to be implicated in RSA. Indeed, increased numbers and killing activity of NK cells in the peripheral blood predict the likelihood of another miscarriage and are considered as a causal and prognostic factor for sterility, infertility and miscarriage . High concentrations of NK cells of the conventional CD56+ 16+ type have been found in the uterus of women who have abortions, suggesting that the cytotoxic activity is probably present at the implantation site .
It is possible that the imbalance in the Th1/Th2 response could provoke an increase in NK cell levels. Indeed, the type 2 shift during pregnancy is predominantly in the NK cell (CD56bright and CD56dim) and NKT-cell (CD56+CD3+) populations than in the T-helper or cytotoxic T-cell populations .
It is not surprising that intravenous immunoglobulins are effective in several conditions of immune-mediated RSA. Indeed, we demonstrated that treatment by means of IVIG during pregnancy can revert and lower high levels of NK cells in women with RSA to values comparable with those of healthy women . The efficacy of IVIG treatment was further confirmed by the evidence of a negative modulation on NK levels even in a very short time, i.e. 30 min. after the single, very first infusion, an effect that never was reported in previous studies .
Among others, IVIG therapy was pursued as an approach of immunomodulation to down-regulate the maternal immune response to the embryo. According to the guidelines for IVIG treatment, we selected patients with severe clinical presentation of RSA or no responding to previous therapies as they were eligible for IVIG therapy. A downregulation of CD56+/16+ NK cell was observed in women treated by IVIG infusion and tested immediately after the first infusion, showing the acute immunomodulatory effect of IVIG. In treated pregnant RSA women, when studied at the second trimester, peripheral blood NK cells maintained at low levels, showing a long-acting beneficial effect of the therapy. In this study, increased peripheral blood NK cells are associated with pregnancy loss. Downregulation of CD56+/16+ NK cells by IVIG infusion results in a favourable pregnancy outcome in women with elevated NK cells. Therefore, women with RSA and elevated NK cells may benefit from IVIG treatment because IVIG not only reduce the number of NK cells – in the short time of the very first infusion and also during the whole pregnancy – but also produce reduction in the cytotoxic activity of these cells.
The autoantibodies
Anti-phospholipid antibodies
The APS is defined as the presence of at least one clinical manifestation in terms of arterial and/or venous thrombosis, or of obstetrical complications and the presence of anti-aPLs . Obstetrical complications are defined as one or more unexplained deaths of a morphologically normal foetus at or beyond the 10th week of gestation, with normal foetal morphology (demonstrated by ultrasound or direct examination of the foetus), one or more premature births of a morphologically normal neonate before the 34th week of gestation because of eclampsia or severe preeclampsia, or features of placental insufficiency (abnormal or non-reassuring foetal surveillance test(s), e.g., a non-reactive, non-stress test, suggestive of foetal hypoxaemia, abnormal Doppler flow velocimetry waveform analysis suggestive of foetal hypoxaemia, oligohydramnios or a postnatal birth weight less than the 10th percentile for the gestational age), or three or more unexplained consecutive spontaneous abortions before the 10th week of gestation, with maternal anatomic or hormonal abnormalities and paternal and maternal chromosomal causes excluded .
aPLs are a heterogeneous class of antibodies that show the common feature recognition of negatively charged plasma proteins, including precisely phospholipids . Wassermann et al. in 1906 identified in patients with syphilis antibodies directed towards negatively charged phospholipids, then isolated immunochemically by Pangborn . These autoantibodies are directed to a variety of protein compounds, being able to identify anti-cardiolipin (aCL), phosphatidylserine, phosphatidylethanolamine and anti-phospholipid-binding proteins (in particular anti-beta-2-glycoprotein-I (anti-β 2 GPI), prothrombin, annexin V, etc.). The aPLs interfere with normal conception in various ways, such as binding to phospholipids of the ovary, or by preventing the normal fertilisation of the ovum, or by binding the surface of the sperm. Of course, the reactivity of aPLs with cell membranes (cytoplasmic, mitochondrial, epithelial cells and endothelial cells, platelets and trophoblastic cells) is fundamental in the pathophysiology of APS.
The pathogenic role of aPLs has been clearly demonstrated in animal models. A brilliant experiment has shown that passive induction of aPLs in naive mice or active induction by immunisation with aPL or anti-β 2 GPI leads to reduction of fertility, increase in the re-absorption of the embryos, reduction in the number of embryos per pregnancy and finally reduction in weight of the embryos and placentas .
The pathogenic mechanisms by which aPLs contribute to the development of obstetrical disease are complex and involve not only the development of multiple micro-intra-placentar thrombi but also direct effects on the trophoblast. Thrombosis in decidual vessels is probably mediated by a network that provides an intense platelet activation accompanied by increased expression of adhesion molecules on the endometrial cells, an alteration of tissue factor, inhibition mediated by aPL factors and anticoagulants to endothelial cell dysfunction that causes the increase in circulating plasminogen activator inhibitor type 1 and antigen-like tissue plasminogen activation. Some aPLs can act specifically, such as the anti-annexin V, that interfere with this protein known to be essential in the placental coagulation system, and anti-β 2 GPI antibodies cause dysfunction of this glycoprotein, which is an important factor in the regulation of coagulation . Besides thrombosis, other factors serve as triggers of the condition.
The binding of aPLs to epithelial cells and platelets may reduce the nutrient supply to the foetus and cause thrombosis and placental infarction; nonetheless, there is the possibility that aPLs bind directly to the implant site or placental antigen. Other mechanisms have been implicated such as an alteration in the balance between thromboxane and prostacyclin with lack of inhibition of platelet aggregation. Then aPLs seem to be able to inhibit the secretion of human chorionic gonadotropin by trophoblasts, preventing the bond between the metal-proteinase urokinase to its receptor on the surface of the trophoblast (thus preventing the implant), or by inhibiting the synthesis of prostaglandins by decidual cells and therefore avoiding decidualisation, or by changing the cytokine pattern increasing the expression and secretion of pro-inflammatory cytokines such as TNF, IL-1 and IL-6, further causing thrombosis, placental apoptosis and activation of NK cells, and reducing the levels of IL-3, directly inhibiting cell growth and accelerating human placental cell apoptosis in culture and eventually causing the activation of complement ( Table 1 ) .
| Thrombosis in decidual vessels |
|
| Alteration of the thromboxane prostacycline balance |
|
| Anti-trophoblast actions |
|
Related posts:
Undifferentiated CTD: A wide spectrum of autoimmune diseases
Idiopathic inflammatory myopathies
Much more than thrombosis and pregnancy loss: The antiphospholipid syndrome as a ‘systemic disease’
Neurologic and other systemic manifestations in FMF: Published and own experience
Psoriatic arthritis
Tocilizumab – A novel therapy for non-organ-specific autoimmune diseases
Stay updated, free articles. Join our Telegram channel
Full access? Get Clinical Tree
