Introduction

The traditional way to study pregnancy immunology follows the classical transplantation model, which views the fetus as an allograft. A more recent approach focuses on the unique, local uterine immune response to the implanting placenta. This requires a detailed knowledge of implantation and placental structure because this impacts greatly on the type of immune response produced by the mother. At the implantation site, cells from the mother and the fetus intermingle during pregnancy. Unravelling what happens here is crucial to our understanding of why some human pregnancies are successful but others are not.

Nidation

The invasive implantation undertaken by the human embryo brings fetally derived trophoblast cells into direct contact with maternal cells in the uterine mucosa. Initial contact is followed by adhesion between the embryonic trophectoderm of the blastocyst and the uterine surface epithelium. As the blastocyst penetrates through the surface epithelium into the uterine mucosa, this trophectoderm layer differentiates into an outer multinucleated syncytiotrophoblast (primitive syncytium) and an inner layer of primitive mononuclear cytotrophoblast. Lacunae soon appear in the syncytium, and these rapidly enlarge by fusing with each other. The uteroplacental circulation is potentially established when this lacuna system erodes through the uterine capillaries. The intervillous space of the definitive placenta is a derivation of these lacunae.

The subsequent differentiation of trophoblast occurs along two main pathways, villous and extravillous ( Fig. 6.1 ). Villous trophoblast is in contact with maternal blood in the intervillous space, and its main functions are transport of nutrients and oxygen to the fetus and secretion of hormones. In contrast, extravillous trophoblast is involved in the establishment of the placental blood supply and intermingles with maternal uterine tissues. At the tips of some chorionic villi, cytotrophoblast cells proliferate into cytotrophoblast columns that anchor these villi to the underlying decidua. From these columns, individual trophoblast cells break off to invade the decidua. These interstitial extravillous trophoblast cells appear to move towards the decidual spiral arteries, encircling these vessels, which then show endothelial swelling and a characteristic ‘fibrinoid’ destruction of the smooth muscle of the media. How trophoblast cells induce these changes in the vessel wall is unknown. When migrating trophoblast cells reach the decidual–myometrial junction, many become multinucleated placental bed giant cells. These can be regarded as the endpoint of the extravillous pathway of trophoblast differentiation.

Schematic representation of the implantation site. A, Placental villi (top) are shown with anchoring cytotrophoblast cell columns and trophoblast invasion into the maternal decidua (bottom) . Maternal blood from decidual spiral arteries (A) fills the intervillous space in direct contact with syncytiotrophoblast (ST). Distinct trophoblast populations are shown. Villous trophoblast comprises: cytotrophoblast (CT) syncytiotrophoblast form the two layers covering placental villi and do not stain for human leukocyte antigen (HLA)-G. Extravillous trophoblast includes cytotrophoblast cell columns (COL), interstitial trophoblast (IT), endovascular trophoblast (ET) and placental bed giant cells (GCs); all stain strongly for HLA-G. Anchoring cell columns coalesce to form a continuous trophoblast shell (TS). From this shell, interstitial trophoblasts (ITs) invade through the decidual stroma to encircle and destroy the arterial media, which is replaced by fibrinoid material (F). ETs move in retrograde fashion down spiral arteries, displacing endothelial cells. On reaching the inner layer of the myometrium, trophoblast cells differentiate to multinuclear giant cells (GCs). The inset shows a representation of cellular interactions within the decidua. ITs are seen between large decidual stromal cells (S). Maternal leukocytes present are mainly uterine natural killer NK (uNK) cells with a few macrophages (Ms) and occasional T cells (Ts).

B, Pathways of trophoblast differentiation and trophoblast subtypes at the implantation site.

Panel A adapted from Moffett-King A. Natural killer cells and pregnancy. Nat Rev Immunol 2 (9):656–663, 2002.

Cytotrophoblast columns that lie over the openings of the decidual spiral arteries form a plug of cells that are known as endovascular trophoblast. Early in gestation, these plugs occlude the lumen of the vessels (see Fig. 6.1A ). This limits the influx of blood in the first trimester so that there is only seepage of serum into the intervillous space. This means that early in pregnancy, the embryo in the first trimester exists in a low-oxygen environment. From these plugs, some endovascular trophoblasts move down the inside of the artery, replacing the endothelium, and become incorporated into the vessel wall. At around 10 weeks of gestation, the endovascular plugs disperse, and maternal blood flow to the intervillous space is established.

Transformation of the spiral arteries by trophoblast is crucial to successful implantation because these changes convert the arteries from muscular vessels into flaccid sacs capable of transmitting the increased blood flow required for the developing fetoplacental unit. Failure of this arterial transformation will result in reduced conductance and poor perfusion of the placenta, which will affect the development of the villous tree. This in turn will lead to clinical conditions such as miscarriage, stillbirth, fetal growth restriction and pre-eclampsia ( Fig. 6.2 ).

Disorders of human pregnancy resulting from abnormal placentation. A, The blood supply to a human pregnant uterus. B, Normal pregnancy. Maternal blood flow to the intervillous space begins at around 10 weeks’ gestation. The spiral arteries of the placental bed are converted to uteroplacental arteries by the action of migratory extravillous trophoblast cells. Both the arterial media and the endothelium are disrupted by trophoblast cells, converting the artery into a wide-calibre vessel that can deliver blood to the intervillous space at low pressure. The small basal arteries are not involved and remain as nutritive vessels to the inner myometrium and decidua basalis. C, Pre-eclampsia and fetal growth restriction. When trophoblast cell invasion is inadequate, there is deficient transformation of the spiral arteries. The disturbed pattern of blood flow leads to reduced growth of the branches of the placental villous tree, which results in poor fetal growth.

Decidualisation

The uterine endometrial mucosa into which trophoblast invades is transformed into decidua during pregnancy. Morphologically, the most obvious changes occur in the stromal cells, which become rounded and glycogen rich. There is also infiltration by large numbers of bone marrow–derived cells. These changes begin during the luteal phase of the menstrual cycle (predecidual change), but if pregnancy occurs, the decidualisation process continues. This is unlike the situation in most other species in which decidualisation only begins at implantation. Decidualisation is under the control of sex hormones oestrogen and progesterone. Both glandular and stromal cells of the endometrium increasingly express oestrogen and progesterone receptors until the time of ovulation, and expression then declines soon after in the glands. Expression of progesterone receptors continues in the stroma throughout the secretory phase and in early decidua. Prolonged exposure to progesterone results in large, rounded cells that secrete high levels of prolactin and insulin growth factor binding protein-1. Other changes include secretion of interleukin-15 (IL-15), metalloproteinases and chemokines and the laying down of a pericellular rim of matrix proteins, particularly fibronectin.

Current opinion favours the view that decidualisation facilitates implantation by providing an appropriate substrate for trophoblast migration and a fertile soil for nourishment of the developing fetus throughout gestation. However, it is also possible that decidua provides a restraining influence against overinvasion by trophoblast. This accords with observations that, in situations in which decidualisation is inadequate, such as in ectopic pregnancies or implantation over a previous caesarean section scar, trophoblast invasion is unrestrained, leading to conditions such as placenta accreta. It is likely that decidua provides a balance, allowing migration of trophoblast but only to a certain depth. Thus mammalian reproduction may be considered as a parental tug-of-war between the requirements of the fetus to derive as much nourishment as possible from the mother and the defence of the mother to reduce this nutritional burden for the sake of her own health and for future pregnancies.

Trophoblast Interaction with Extracellular Matrix

Cell migration depends on the expression of adhesion molecules which bind to extracellular matrix (ECM) proteins. For example, the physiological migration of epithelial cells in wound healing and the pathological invasion of cancer cells require cell–matrix interactions. Trophoblast migration into decidua appears to use similar mechanisms. There are four families of adhesion molecules, of which the most important for adhesion to the ECM are the integrins. These are transmembrane glycoproteins consisting of noncovalently associated α and β subunits. Different α and β subunits exist and the way they combine determines the ligand specificity of the integrin. For example, the heterodimers α1β1 and α6β4 are receptors for the ECM protein, laminin, while α5β1, α4β1 and α4β7 bind fibronectin.

Using monoclonal antibodies specific for various subunits, the pattern of expression of integrins by different trophoblast populations at the implantation site is now well documented. The α6β4 integrin is expressed on the villous cytotrophoblast layer and the cytotrophoblast cells of the cell columns nearest the villous core. This integrin disappears further out in the cell columns to be replaced by the heterodimers α5β1 that continue to be expressed by the interstitial trophoblast invading into decidua. Thus the α6β4 laminin receptor is downregulated with a concomitant upregulation of the α5β1 fibronectin receptor as trophoblast invades the decidua. This observation is similar to that seen during the healing of a skin wound where the sessile keratinocytes that form the normal epidermis express α6β4, but keratinocytes which migrate to close over the wound express α5β1. Binding of trophoblast to fibronectin results in signalling through integrins to the trophoblast cell with changes in gene expression that will affect trophoblast function. In pre-eclampsia, trophoblast fails to downregulate β4 as seen in normal pregnancy, indicating that dysregulation of these integrins could contribute to the inadequate trophoblast invasion of decidua associated with this pathological condition.

Matrix Degradation by Trophoblast

Besides adhesion to ECM proteins, cellular migration also requires degradation of the matrix. This involves the production of proteolytic enzymes by the migrating cell. The two main groups of enzymes are members of the plasminogen activator (PA) system of serine proteases and the family of matrix metalloproteinases (MMPs). The MMP family comprises three main classes based on their substrate specificities: the collagenases, the gelatinases and the stromelysins. There is an intricate interaction between the PA and MMP systems, and together they can break down the major components of ECM. The activity of these proteases is subjected to close control by specific inhibitors. There are two inhibitors for PA (PAI), designated as PAI-1 and PAI-2, and two tissue inhibitors for MMP (TIMP), designated as TIMP-1 and TIMP-2.

Trophoblast cells possess proteolytic activity which can be demonstrated in vitro by their digestion of the surrounding matrix on which the cells are seeded. Zymogram studies have shown that trophoblast cells produce a wide array of proteases, this production being greater in first-trimester trophoblast compared with trophoblast later in gestation, which therefore mirrors the invasive capacity of early trophoblast. These observations have led to the conclusion that PA, MMP, PAI and TIMP together provide an intricate network that controls matrix degradation during trophoblast invasion. Although it is clear that changes in integrin and protease expression play an important role in regulating trophoblast differentiation and invasion, the factors that control these changes in normal and pathological pregnancies are poorly understood.