Anatomy & Physiology for Midwives 3: Third Edition

Chapter 8. The placenta

Learning objectives

• To describe the development of the placenta, membranes and umbilical cord.

• To describe the structure of the placenta, membranes and umbilical cord and recognize common structural variants.

• To identify the roles of the placenta.

• To discuss how abnormal placental development might affect fetal development including intrauterine growth retardation and other outcomes of the pregnancy.

• To describe methods for monitoring placental function.

• To outline the development of the placenta in twin pregnancies.

Introduction

The development of the placenta is critical for fetal survival because of the importance of the placenta in maternal–fetal transfer. It has a range of functional activities (Table 8.1), including complex synthetic capabilities, which are essential to the development of a normal term baby. The placenta flourishes in an immunologically foreign environment and has an important role in the immunological acceptance of the fetal allograft (see Chapter 10). Essentially, the placenta acts as a vascular parasite, depending on maternal blood for oxygen and nutrients and removal of waste products. The structure of the placenta means that, although optimal diffusion gradients are established, maternal and fetal blood never actually mixes.

Chapter case study

At Zara's 20-week scan, the ultrasonographer documented in her report that the placenta was situated on the anterior wall of the uterus with the majority of the placental body situated in the middle and lower pole of the uterine body with the lateral edge of the placenta approximately 2 cm from the internal cervical os.

• What are the possible complications that could arise from this situation?

• How do you think the midwife should discuss these possible complications with Zara and how they could be recognized?

• Why is it common for women with this situation to have vaginal bleeding around 34 weeks of pregnancy?

• Are there any conditions or factors related to low lying placentas and if so how can these be managed?

The placenta and the chorion (outer membrane) are derived from the trophoblast layer of blastocyst cells (see Fig. 6.6, p. 128). Other extraembryonic tissues develop from the inner cell mass. These include the amnion (inner membrane), the yolk sac, the allantois (a largely vestigial structure in humans) and the extraembryonic mesoderm. The umbilical cord and the blood vessels of the placenta are derived from the extraembryonic mesoderm.

The human placenta is haemochorial, which means that maternal blood comes into contact with the placental trophoblast cells. The uteroplacental unit is made up of both fetal and maternal components. The placenta as seen at delivery is just the fetal component or chorionic plate. The maternal component or basal plate is the placental bed which underlies the fetal component and the uteroplacental circulation that vascularizes the placental bed. Between the chorionic and basal plates is the intervillous space where the maternal–fetal exchange occurs. Conversion of the maternal spiral arteries to dilated and flaccid vessels is an essential step for successful pregnancy. Abnormal placental function is strongly associated with fetal complications, but study of the human placenta, particularly the maternal component, is not easy. Placentation in the human is unique, which means that observations from other species can be applied to humans only with caution. Placental reserve needs to exceed fetal requirements (otherwise the fetus could be compromised under conditions of hypoxia). It might be expected that placental size would increase in parallel with increased fetal size; however, the placental:fetal weight ratio actually decreases during gestation (Kingdom et al., 1993). Placental efficiency is best described as grams of fetus produced per gram of placenta developed (Wilson and Ford, 2001). Indeed, as there is a relationship between size at birth and life expectancy which is associated with the placental supply of nutrients; lighter placentas are more efficient placentas and associated with better long-term health outcome (Fowden et al., 2009). Placental efficiency is affected by the surface area for exchange, the thickness of the barriers between fetal and maternal circulations and the arrangement of the fetal and maternal blood vessels (‘vascular architecture’). It is the later variable that appears to account for species differences in placental efficiency. The most efficient human placentas are those which are small in diameter and thin; it is thought that these small placentas must functionally adapt to increase nutrient transporter abundance and become more efficient.

Instead, placental efficiency is enhanced by an increase in both the number of carrier proteins involved in the transport of substances across the placenta and the placental perfusion.

Uterine receptivity

The first phase of the development of uterine receptivity is regulated by oestrogen and progesterone which stimulate the presence of microvilli on the columnar epithelial cells of the endometrium. Smooth muscle myosin also increases and the stromal cells proliferate. The second phase is described as the blastocyst response phase (Banerjee and Fazleabas, 2010). The signals involved in the exquisitely sensitive dialogue between the embryo and the endometrium include human chorionic gonadotrophin (hCG), interleukin-1 and insulin-like growth factor (IGF) 2 (see Chapter 3). In the third phase following attachment and implantation, signals between the newly formed embryo and primed endometrium cause endometrial changes, or decidualization, whereby the stromal cells under the endometrial epithelium accumulate lipid and glycogen and become known as decidual cells. The stroma thickens and blood flow increases. Decidualization promotes changes in the endometrium that make it receptive to implantation. Synchronous development and communication between the maternal endometrium and the embryonic tissue are required for successful establishment of pregnancy. Whereas mammalian embryos have intrinsic invasive potential and can initiate implantation-type reactions in many different tissues, the endometrium protects itself from implantation except for the limited duration of the implantation window (Bischof et al., 2000).

During the implantation or nidation window, microvilli on the surface of the uterine endometrial cells fuse together to form single flower-like projections called pinopods or uterodomes (Murphy, 2000). These smooth bleb-like protrusions which lack the typical microvilli form under the influence of progesterone (during the mid-luteal phase) only in the preferred sites of embryo–endometrial interaction and thus act as markers of uterine receptivity (Usadi et al., 2003). The pinopods are only present for 2–3 days during which implantation must occur. The interaction between the endometrium and the developing trophoblast is facilitated by a number of cytokines, metalloproteinases, surface integrins and growth factors, including IGFs and their binding proteins, which create a specific microenvironment which modulates trophoblast function. It is evident that implantation is a rather inefficient process in the human; the probability of conception during a menstrual cycle (defined as fecundity) is about 30% and over three quarters of failed pregnancies are thought to be due to implantation defects (Wilcox et al., 1999).

Implantation

Implantation is the consequence of a well-organized sequence of events involving synchronized crosstalk between the receptive endometrium and a functional blastocyst (Achache and Revel, 2006). It occurs in three stages: apposition, adhesion and invasion. The trophoblastic cells overlying the inner cell mass are known as the polar trophoblast; it is these cells that initiate the adhesion and implantation processes. The morula enters the uterus about 4 days after fertilization. It may float freely in the uterus before it hatches out of the protective zona pellucida (Kingdom and Sibley, 1996). About 7 days after fertilization, the blastocyst hatches and comes into contact with the endometrium. The blastocyst rolls freely over the endometrium until it reaches a receptive area. This process is thought to be mediated by glycoproteins called selectins which are expressed on the polar trophoblast cells of the newly hatched blastocyst (Vitiello and Patrizio, 2007). The blastocyst then orientates itself so that the embryonic pole implants first. Inappropriate implantation of the blastocyst so there is reduced contact between the polar trophoblast and the uterine endothelium is thought to lead to abnormalities of umbilical cord insertion and even failure of pregnancy (Huppertz, 2008). The endometrium produces MUC1, a mucin-rich glycoprotein, to prevent the blastocyst adhering to areas of the endometrium with poor chances of implantation. The optimally receptive areas of the endometrium secrete chemokines and growth factors to attract the blastocyst to the pinopods. The apposition of the blastocyst to the endometrium triggers the production of adhesion molecules such as integrins and cadherins which firmly anchor the blastocyst to the endometrial pinopods (Fig. 8.1). This process is enhanced because the endometrial surface expresses receptors for selectins. The tethering of the blastocyst to the endometrium stimulates the polar trophoblastic cells to undergo rapid mitosis and proliferate as the invasion of the uterine wall commences. Implantation may be affected by maternal antiphospholipid syndrome hence the high incidence of pregnancy loss associated with this condition (Stone et al., 2006) (Case study 8.1).

Case study 8.1

Trudy is a 36-year-old, para 1, gravida 11. She attends the midwives clinic at 6 weeks gestation, very distressed as this was not a planned pregnancy. Trudy informs the midwife that her last baby was born at 32 weeks gestation, very small for dates and that Trudy had also developed fulminating pre-eclampsia which was the main reason for the early delivery. Three days after this delivery Trudy developed severe difficulty in breathing which was diagnosed as a pulmonary embolism. Trudy then informs the midwife that all her other pregnancies had been spontaneous miscarriages at around 10 weeks gestation.

The midwife reviews Trudy's post obstetric notes and discovers that Trudy has antiphospholipid syndrome and that at her last delivery it was documented that the placenta was small and infarcted. As a result of this, the midwife immediately refers Trudy to attend a consultant clinic as an emergency.

• Why did the midwife refer Trudy to the consultant as an emergency?

• What treatment would be offered to Trudy and how will the pregnancy be managed?

• What is the significance of the placental infarcts and what was the most likely cause?

Differentiation into cytotrophoblast and syncytiotrophoblast

There are two distinct cell layers in the blastocyst (see Chapter 6): the inner cell mass which is surrounded by an outer sphere of a single layer of mononucleated trophoblast cells. The trophoblast is the first cell type to differentiate; this outer layer rapidly proliferates and develops into the placental tissue and fetal membranes.

The trophoblast differentiates into two layers: the outer syncytiotrophoblast and the inner cytotrophoblastic layer. Some of the proliferative cytotrophoblast cells lose their cell membranes and coalesce to form a multinucleated syncytium (a united mass of fused cellular material): the syncytiotrophoblast (Fig. 8.2). This outermost layer of the placenta has little proliferative and transcriptional activity; the maintenance and growth of the syncytiotrophoblast throughout gestation is dependent on the incorporation of cytotrophoblast cells into the layer. Apoptosis (programmed cell death) of trophoblast tissue increases throughout pregnancy as a normal part of trophoblast turnover and syncytiotrophoblast formation. The nuclei of the cells newly incorporated into the syncytiotrophoblast are initially similar to the nuclei of the cytotrophoblast cells but then undergo morphological changes; the chromatin condenses so the nuclei become smaller and denser eventually resembling late apoptotic nuclei. Some of these nuclei aggregate and are packaged into syncytial knots which are shred from the apical surface of the syncytiotrophoblast into the maternal circulation. In a normal pregnancy, these syncytial knots can be identified in maternal blood in the uterine veins but are destroyed in the maternal pulmonary vessels. In pre-eclampsia, syncytiotrophoblast renewal is overactive; there is an increase in apoptosis often complicated by aponecrosis. The syncytial knots are more prominent and are smaller so they can survive the maternal pulmonary vasculature and trigger a maternal systemic inflammatory response including an activated endothelium and increase in proinflammatory markers (Hawfield and Freedman, 2009).

The surface of the syncytiotrophoblast is covered in microvilli which increase the surface area. The syncytiotrophoblast expresses transporters, enzymes and receptors on its surface. It also produces hCG and hPL which are crucial to the maintenance of the pregnancy. hCG enhances differentiation of cytotrophoblasts into the syncytiotrophoblast. Electron microscopy reveals the syncytiotrophoblast to be a mass of cytoplasm containing remnants of intercellular membrane, evenly dispersed nuclei and a few intermediate cells. This syncytial organization of cells is unusual; other than the trophoblast, multinucleated cells are seen only in some tumour cells and inflammatory giant cells (Chard, 1998). Interestingly, a range of tumour cells appears to secrete hCG (Iles and Chard, 1991) but at lower concentrations than those characteristic of trophoblast cells; these tumours usually affect individuals late in life.

The cytotrophoblasts, which form the inner monolayer of placental stem cells, are large clear discrete cuboidal cells each with a single nucleus, a few organelles and a well-defined cell membrane. These cells have marked mitotic activity and DNA synthesis. The syncytiotrophoblast increases in volume throughout the second week as cells detach from the proliferating layer of cytotrophoblast and fuse with the mass of syncytiotrophoblast. The syncytiotrophoblast has an invasive phenotype, secreting enzymes, which attack the endometrium, and hormones, which sustain the pregnancy. It is also involved in absorption of nutrients. The syncytiotrophoblast invasion is aggressive; between 6 and 9 days postfertilization the embryo becomes completely implanted into the endometrial stroma. The hydrolytic enzymes produced cause breakdown of the extracellular matrix between the cells of the endometrium thus eroding a pathway. The surface of the syncytiotrophoblast has tiny processes extending from it that penetrate between the endometrial cells, pulling the conceptus into the uterine wall. As implantation progresses, the expanding syncytiotrophoblast gradually envelops and encircles the blastocyst. The endometrial epithelium regenerates over the site of implantation, forming the decidua capsularis (Fig. 8.3). By 9 days, the embryo is completely embedded within the endometrial wall with the syncytiotrophoblast forming a complete mantle around the entire conceptus so it is the only embryonic tissue in direct contact with the maternal tissue; this is important in protecting the embryo from rejection, The syncytiotrophoblast has a developmental gradient (Huppertz, 2008); it is thicker and better developed over the embryonic pole. Implantation is complete by about 10–12 days after fertilization. A plug of a cellular material called the coagulation plug or operculum seals the small hole at the point of implantation (Fig. 8.4).

In the first week of development, as the free-floating embryo or conceptus moves towards the uterine cavity propelled by the cilia movement and muscular contraction of the uterine tube, the cells can obtain nutrients from secretions of the uterine tubes and endometrium and eliminate waste products by simple diffusion. It is thought that endometrial secretions of lactate and pyruvate and possibly some amino acids may be important in the nutrition of the embryo for up to 8 weeks (Burton et al., 2001). The nutrients and oxygen are taken up by the syncytiotrophoblast; this is known as histiotrophic nutrition. By the end of the first trimester, there is a transition to haemotrophic nutrition from the uteroplacental circulation which provides a system in which the maternal and fetal circulations come into close contact to facilitate transfer of substances from one system to the other.

As the syncytiotrophoblast penetrates the uterine wall, it comes into contact with the maternal endometrial capillaries and superficial veins. Fragments of these are engulfed within the syncytiotrophoblast forming fluid-filled trophoblastic lacunae (literally ‘little lakes’); these coalesce to form larger lacunae which are the precursors of the intervillous spaces. As maternal blood vessels are progressively invaded, the lacunae fill with maternal blood. Maternal capillaries near the syncytiotrophoblast expand to form maternal sinusoids which rapidly anastomose with the trophoblastic lacunae. As this development continues, the lacunae become separated by columns of syncytiotrophoblast, or trabeculae, which effectively form a framework on which the villous tree of the placenta develops. The trabecular columns project radially from the blastocyst. The cytotrophoblast at the core of the columns proliferates locally to form extensions, which grow into the columns of syncytiotrophoblast. The growth of these protrusions is induced by the newly formed extraembryonic mesoderm (Fig. 8.5). The result is the primary stem villus, an outgrowth of cytotrophoblast covered by syncytiotrophoblast, which penetrates into the blood-filled lacuna (Fig. 8.6).

Extravillous cytotrophoblast and remodelling of the uterine vessels

The cytotrophoblast layer has several distinct roles: (1) acting as proliferative progenitor or stem cell layer to generate and construct the developing syncytiotrophoblast which covers the villi; (2) forming the proliferative cell columns of the anchoring villi; (3) detaching from the cell columns and migrating into the maternal stroma to form interstitial cytotrophoblasts; and (4) migrating as non-proliferative extravillous cytotrophoblast cells to remodel the spiral arteries and replace the maternal endothelial cells (Knofler, 2010). These extravillous cytotrophoblast cells migrate from the villi beyond the leading edge of syncytiotrophoblast into the stroma. From about 12 days postfertilization, these cells invade the maternal capillaries and spiral arteries of the decidua. The extravillous cytotrophoblast cells initially plug the lumen of the maternal vessels that have been invaded and subsequently replace the maternal endothelium of these vessels. Plugging of the lumen of the invaded maternal blood vessels prevents bleeding and is achieved by day 14, which coincides with the expected date of the next menstrual period. If the maternal vessels are not plugged adequately during implantation and early development then vaginal bleeding may occur, which is associated with an increased risk of spontaneous miscarriage (sometimes haematomas can be seen on ultrasound investigation). The plugs prevent flow of maternal blood into the intervillous space in early pregnancy but allow a slow seepage of plasma (Jauniaux et al., 2003) so the lacunar spaces enclosed by the syncytiotrophoblast initially contain exudate from maternal vessels rather than blood. The developing placenta forms an effective barrier between the mother and developing embryo that persists up to 10 weeks' gestation when the trophoblastic plugs are dislodged and intervillous blood flow is established (Jauniaux et al., 1992). It is at this time that peak hCG secretion occurs (Meuris et al., 1995). The increasing oxygen level and concomitant oxidative stress also stimulate cytotrophoblast proliferation and differentiation and the increased expression of antioxidant enzymes. Doppler ultrasound shows there is no intervillous blood flow in normal pregnancies before this period and oxygen electrodes have demonstrated that an oxygen gradient exists across the placenta and decidua. This means that embryogenesis occurs in a relatively hypoxic environment. In fact, maternal–placental blood flow has been observed in the first trimester in a number of non-viable pregnancies but it is not clear whether this is a cause of the pregnancy failure or an effect (Kingdom and Sibley, 1996). The developing embryo is thought to be particularly vulnerable to damaging oxygen-free radicals during the sensitive period of organogenesis and the first-trimester placenta has limited antioxidant capacity; thus, limiting fetal exposure to oxygen may be protective. The differentiation of trophoblast cells is influenced by the local oxygen levels; hypoxia promotes trophoblast proliferation and normoxia inhibits proliferation and induces migration (Knofler, 2010). In most other mammalian species, organogenesis is complete and embryonic development is advanced before placental attachment. But in the human, implantation is highly invasive so the precocious conceptus is embedded in the uterine wall even before the primitive streak is evident (Jauniaux et al., 2006). Hence it is much more sensitive to oxidative stress. Embryonic and placental cells are particularly vulnerable because they are undergoing extensive cell division and DNA replication. The syncytiotrophoblast is exposed to the highest concentration of oxygen as it is closest to the maternal blood but it has low levels of antioxidant enzymes. Maternal metabolic disorders such as diabetes generate more oxidative free radicals and are associated with a higher incidence of miscarriage, vasculopathy and fetal structural defects which is thought to be due to oxidative stress (Jauniaux et al., 2006).

Between the 4th and 16th week of gestation, villus growth and considerable remodelling of the placenta occur, including remarkable changes in the maternal blood vessels underlying the fetal placenta ensuring that the spiral arteries are capable of delivering large volumes of blood to the placental intervillous spaces at an appropriate rate and pressure to protect the delicate fetal villi perfused by the low-pressure developing fetal circulation which are immersed in maternal blood which circulates at a much higher pressure and velocity. Failure of the transformation of the spiral arteries is associated with a number of common complications of pregnancy including pre-eclampsia, fetal growth restriction, recurrent first and second trimester losses, spontaneous preterm labour and premature rupture of the membranes (Burton et al., 2009a). In the early weeks, some of the cytotrophoblastic cells (described as extravillous cytotrophoblast) move from the tips of the anchoring villi to colonize the decidua and myometrium of the placental bed. It is this invasion of extravillous cytotrophoblast cells into the maternal blood vessels that promotes maternal recognition of the fetus and the subsequent production of blocking antibodies (see Chapter 10), which are important for the survival of the pregnancy. The extravillous cytotrophoblast cells are involved in physiologically remodelling the maternal spiral arteries which is completed by the end of the first trimester. After an apparent rest phase of a couple of weeks (weeks 14–16), there is a resurgence of the endovascular trophoblastic migration. The second wave of cytotrophoblast cells moves down the myometrial segments of the spiral arteries to their origin at the branching from the radial arteries. The cytotrophoblast cells are involved with the destruction of the maternal artery musculoelastic tissue and the replacement of the maternal endothelial wall with trophoblast resulting in a change in the vessel wall vasoresponsiveness. The result is conversion of the thick-walled muscular spiral arteries to compliant dilated sac-like uteroplacental vessels that have low impedance to blood flow (Fig. 8.7). The changes in the spiral arteries are augmented by interstitial trophoblast cells which migrate through the endometrial stroma and penetrate the spiral arteries from the outside; they continue to invade the myometrium where they transform into immotile giant cells (Burton et al., 2009a). Insufficient remodelling of the spiral arteries is associated with failure of the fetus to reach its genetic growth potential (intrauterine growth restriction (IUGR)) and pre-eclampsia (Box 8.1). Maternal spiral arteries maintaining a high vasculature resistance because of incomplete or failed remodelling are predisposed to hypoperfusion, hypoxia, reperfusion injury and oxidative stress. Although the trophoblast invasion and remodelling are limited to the spiral arteries, the radial, arcuate and uterine arteries also undergo profound dilation especially close to the site of implantation (Burton et al., 2009a and Burton et al., 2009b). These vessels unusually progressively increase rather than decrease in diameter as they reach their target organ.

Box 8.1

Pre-eclampsia

Pre-eclampsia is a placental condition; it can occur in the absence of a fetus in a molar pregnancy. One of the most convincing theories about the aetiology of pre-eclampsia and the associated condition of IUGR (which can occur independently or with pre-eclampsia) is that they are due to placental malperfusion secondary to deficient spiral artery conversion (Burton et al., 2009). In normal pregnancies, all spiral arteries in the placental bed are invaded by cytotrophoblast cells. In pre-eclampsia, it seems that only a proportion of the maternal vessels are invaded and that a significant number of vessels show complete absence of physiological changes. The second wave of arterial invasion may be the stage that is most compromised owing to the endovascular trophoblast failing to reach the intramyometrial portion of the vessels. This means that the spiral arteries are not completely transformed to uteroplacental vessels. Maternal uteroplacental blood flow is therefore restricted, which results in placental abnormalities and fetal complications such as IUGR. The effect is compounded by the persistence of vasoresponsiveness of the spiral arteries, which retain the ability to constrict and limit placental perfusion, like the spiral arteries of a non-pregnant uterus (see Chapter 4). Impaired placental perfusion increases the risk of ischaemia–reperfusion type insult which leads to the generation of reactive oxygen species (oxidative stress). Oxidative stress results in increased generation of oxygen-free radicals which lead to the formation of lipid peroxides which alter cell membranes. The incorporation of cholesterol, oxidized free fatty acids and LDLs into membranes is increased which leads to a biological cascade of leukocyte activation, platelet adhesion and aggregation and the release of vasoconstrictive agents (Jauniaux et al., 2006). Acute atherotic changes can lead to the development of intimal plaques which can project into the vessel lumen and restrict blood flow. In addition, endoplasmic reticulum (ER) stress is also triggered by ischaemia–perfusion and hypoxia. ER stress can lead to inhibition of protein synthesis and reduced expression of amino acid transporters (thus affecting growth) as well as activating apoptosis. There are serious maternal complications of pre-eclampsia which have been attributed to an as-yet unidentified placentally derived ‘factor X’ which is released into the maternal circulation; possible candidates include proinflammatory cytokines from the syncytiotrophoblast, products of placental oxidative stress, anti-angiogenic factors and trophoblastic apoptotic debris such as syncytiotrophoblast micro-fragments all of which could contribute to activation of maternal endothelial cells and cause the peripheral syndrome of pre-eclampsia (Fig. 8.8).

The remodelled vessels can passively dilate and accommodate a greatly increased blood flow (about 30% of maternal cardiac output) but they are not responsive to vasoactive agents. The effect of this interaction between the trophoblastic cells and the maternal blood vessels is that a low-pressure, high-conductance vascular system is established, which provides an adequate maternal blood flow to the placenta and thus a plentiful provision of oxygen and nutrients to the fetus. The maternal uteroplacental circulatory system is mostly complete by mid-gestation. In contrast, the fetal villous tree continues to branch and develop throughout the pregnancy, ensuring that the capacity of the placenta matches the growth of the fetus.

As the maternal cardiac output increases by about 40% (see Chapter 11), the net effect is to increase the uteroplacental blood flow by about 10-fold to over 500 mL/min (Kingdom and Sibley, 1996). Doppler ultrasound flow velocity waveforms provide diagnostic and prognostic information about maternal vessels, placental circulation and fetal vessels together with implications for both mother and fetus (Harman and Baschat, 2003). Flow in the uterine arteries gives a picture of the maternal vascular effects of the invading placenta, predicting likely pre-eclampsia and IUGR. Umbilical artery Doppler ultrasound depicts placental vascular resistance, which correlates with IUGR and effects of placental deficiency. Before pregnancy and in the first trimester, the uterine arterial waveform has a low end-diastolic flow velocity and early dichrotic notch during diastole. By 18–20 weeks' gestation, successful trophoblastic invasion alters this pattern to one showing a high diastolic flow velocity and loss of the dichrotic notch. If the dichrotic notch and low end-diastolic velocity persist, this indicates that the uterus still has high resistance to blood flow, which is predictive of intrauterine growth retardation (IUGR) and severe pre-eclampsia (Fig. 8.9).

A number of pathologies including placental infarction or abruption, pre-eclampsia and recurrent pregnancy loss are associated with defects within the placental vascular bed. Deficiencies of vitamin B12 and folate and hyperhomocysteinaemia are risk factors for these placenta-mediated diseases (Ray and Laskin, 1999). Smoking is associated with poorer outcomes of pregnancy including low birthweight, spontaneous abortion and placental abruption. Nicotine, cadmium and polyaromatic hydrocarbons in cigarette smoke adversely affect fertility, oocyte development and oestrogen synthesis (Shiverick and Salafia, 1999). Effects of these compounds on trophoblastic invasion and proliferation are thought to account for the increased miscarriage rate of women who smoke; however, smoking seems to reduce the incidence of pre-eclampsia, possibly because smoking reduces endothelium sensitivity. Severe placental dysfunction is more common with a male fetus (Edwards et al., 2000); this may be the cause of the male:female ratio decreasing from 164:100 soon after conception to 106:100 at birth. Placental pathologies may be due to defective trophoblast function and/or impaired maternal decidualization. The placental-related disorders of pregnancy, such as miscarriage and pre-eclampsia, are almost unique to humans (Jauniaux et al., 2006) and the incidence in other mammals is extremely low. It is thought that the rate of these disorders is increasing because of recent lifestyle changes such as hypercaloric diets and delayed childbirth; however, current populations of hunter-gatherers are also affected.

Vascularization of the placental villi

Fetal blood cells are derived from blood islands in the extraembryonic mesoderm surrounding the yolk sac (see Chapter 9). The blood vessels that perfuse the placenta also develop in this tissue. In the third week of postfertilization, the extraembryonic mesoderm associated with the cytotrophoblast penetrates into the core of the primary stem villi transforming them into secondary stem villi. This mesoderm develops into the blood vessels and connective tissue of the villi. It forms at the same time as the embryonic vasculature with which it will eventually connect. Haemangioblast cells (precursors of blood cells) appear and capillaries form. The linking of the blood vessels of the villi with the vessels of the embryo results in a circulating blood system so the villi begin to be perfused by the fetal circulation at about 28 days after fertilization. The fetal red blood cells containing embryonic haemoglobin allow oxygen transfer at low partial pressures of oxygen and low pH. The villi containing differentiated blood vessels are described as tertiary stem villi. By the end of the fourth week after fertilization, these villi cover the entire blastocyst surface forming a spherical shell of villi projecting outwards into the maternal tissue (Fig. 8.10). It is possible to remove a sample of the developing placental villi for genetic testing (Box 8.2). The placental barrier now effectively limits diffusion of gases, nutrients and waste materials. There are four layers: the endothelium lining the villus capillary, the connective tissue in the villus core, a layer of cytotrophoblast cells and a maternal-facing layer of syncytiotrophoblast (Fig. 8.11). By mid-gestation, most of the cytotrophoblast layer of many villi disappears and the placental barrier becomes very thin. The syncytiotrophoblast may even come into direct contact with the fetal capillary so the maternal and fetal blood may only be about 2–4 μm apart.

Box 8.2

Chorionic villus sampling

In the chorionic villus sampling (CVS) procedure, 20–40 mg of placental tissue can be obtained from a villus for genetic diagnosis of trisomy 21, for instance or of a single-gene abnormality such as cystic fibrosis or β-thalassaemia. After 10 weeks' gestation, the tissue can be extracted transabdominally by needle aspiration or transcervically using curved biopsy forceps. The collected trophoblast cells, which divide very rapidly, can be cultured for 24 h and then the chromosome number can be determined (see Chapter 7). Because of the problems associated with mosaicism (see Chapter 7), a more accurate determination of chromosome number and structure is obtained by using fibroblast cells taken from the vascular core of the villus. These cells grow more slowly so they have to be cultured for 2 weeks before being stained and examined (which means the results of the test take longer). As fibroblast cells are derived from the mesoderm, they originate from the inner cell mass and are embryonic rather than the trophoblast-derived cells from the outer layers of the villus. Placental mosaicism is associated with increased fetal loss and IUGR. There is a 1–2% procedure-related loss in CVS, although it should be remembered that the procedure is being performed because there is already a concern about the pregnancy. Pregnancies associated with genetic abnormality have a much higher risk of spontaneous failure.

Development of the discoid placenta and chorionic membrane

From the 4th week to the 16th week, the villus growth over the entire surface of the blastocyst is remodelled. Most of the villi orientated towards the uterine cavity degenerate and regress, leaving behind an area that develops into the typical placental structure and shape seen at delivery. As the embryo starts to enlarge, the uterine wall where it has implanted starts to protrude into the uterine cavity (Fig. 8.12). The protruding portion of the embryo is covered by the decidua capsularis, a thin layer or capsule of endometrium. The layer of decidua under the embryonic pole of the embryo is the decidua basalis. The remaining areas of the decidua are described as the decidua parietalis.

In the third month, as the fetus enlarges and grows to fill the uterus, the thin rim of decidua capsularis covering the bulge gradually thins and disappears so the chorion comes into contact with the decidua parietalis of the opposite wall of the uterus. Before the trophoblastic shell comes into contact with the uterine wall on the opposite side, cells of fetal origin can enter the uterine cavity and can be collected by flushing or aspiration from the endocervical canal (Kingdom et al., 1995). This is a route of non-invasive prenatal diagnosis, particularly for newer testing procedures that require fewer cells. The size of the chorionic, or embryonic, sac can be used to determine the gestational age of the embryo.

The uterine cavity is obliterated by 12 weeks' gestation. The enlarging blastocyst compresses the trophoblastic layer, distal to the entry pole, and limits nutrient supply and further growth by the villi in this region of the decidua capsularis. The underlying villi slowly degenerate and regress so by the fifth month this region becomes devoid of villi and smoother. This flattened surface of the decidua capsularis and atrophied chorion form the chorion laeve, the uteroplacental membrane, which is also known as the chorionic membrane or bald chorion. Effectively, the chorion is extraplacental trophoblast with similar immunological properties; it may also be an important source of hCG, particularly in early pregnancy. The portion of the trophoblastic tissue associated with the decidua basalis implants further and receives a plentiful supply of nutrients so it continues growing. This area of the chorion, therefore, retains villi that proliferate and progressively arborize forming the chorion frondosum (‘frondosus’ is Latin for leaf), which ultimately develops into the definitive discoid fetal placenta. The placenta is a union between the chorion frondosum derived from the fertilized ovum and the decidua basalis (basal plate) formed from the maternal uterine wall. It is anatomically complete by the end of the first trimester but continues to grow throughout the pregnancy.

Development of the amnion (inner membrane)

The amniotic cavity first appears at about day 7. The primitive ectoderm cells enclosing the cavity become flattened forming amnioblasts, cells which become the fetal-facing amniotic membrane. These cells secrete amniotic fluid, thus the embryo is enclosed in the fluid-filled amniotic sac. The outer surface of the amnioblast cell layer becomes covered with mesoderm. As the embryo expands, the amnion comes into contact with the chorion. The chorionic cells are lined with mesoderm cells on the inner side. When the amnion and chorion meet, the two layers of mesoderm loosely fuse. The fetal membranes protect the fetus and secrete factors into the amniotic fluid which affects amniotic fluid activity and also can influence the maternal uterine physiology. An important aspect of examining the placenta and membranes is to ensure that both membranes are present following birth. The amnion and chorion should be easily separated; the chorion is attached to the edge of the placenta, whereas the amnion can be separated from the surface of the placenta with attachment around the base of the cord.

Amniotic fluid has an important role in protecting the fetus, cushioning it from external impact and stresses preventing fetal injury. It also allows symmetrical fetal growth and movement, preventing fetal parts from adhering together or to the amnion, allows practice breathing and swallowing exercises and increases placental surface area. Amniotic fluid has bacteriostatic properties and is also important in maintaining a constant body temperature; it is also involved in maintaining amnion integrity, discouraging myometrial contractions and maintaining cervical length and consistency (Harman, 2008). In the first half of gestation, before skin keratinization takes place, fluid and electrolytes can diffuse freely across the skin (Jauniaux and Gulbis, 2000). Although the amnioblasts actively secrete amniotic fluid, the composition of the fluid at this time is similar to that of fetal tissue fluid. After 20 weeks, the skin becomes keratinized and transudation from maternal and fetal blood vessels contributes less to the amniotic fluid. Fetal urine and lung secretions are also important. Fetal swallowing and exchange across the amnion mean that turnover of fluid is rapid, particularly close to term. Lung fluid may contribute abut 100 mL per day and fetal urine 7–10 mL per hour. The fetus may swallow up to 1 L of fluid per day; the extra water crosses the gut, enters the fetal circulation and can then cross the placenta. By term, the normal volume of amniotic fluid is 500–1000 mL. Polyhydramnios is an excess amount of fluid (over 2000 mL), which is usually associated with multiple pregnancies or fetal swallowing problems such as oesophageal atresia. A deficiency of amniotic fluid (<500 mL) is classified as oligohydramnios, a condition often associated with impaired fetal renal function.

Amniotic fluid provides a useful tool to monitor fetal development and well-being. A small amount of amniotic fluid can be removed in amniocentesis for measurement and testing. Amniotic fluid contains many maternal and fetal proteins and fetal cells, which can be used for genetic testing (see Chapter 7). If the fetus has a neural tube defect (see Chapter 9), concentrations of AFP (alpha-fetoprotein, derived from spinal fluid) in the amniotic fluid are very high. Levels of AFP are low in Down's syndrome (trisomy 21) and are measured as part of the triple test (see Chapter 7). It has been proposed that this could be due to the persistence of extraembryonic coelom or related to interferon receptor levels (Chard, 1998). Components of amniotic fluid may also be used to predict preterm labour, premature cervical effacement and fetal infection (Harman, 2008). Recent research has investigated the role of amniotic fluid in inhibiting bacterial growth in burns victims and in promoting healing of burns and other skin injuries. Amniotic fluid is a potential source of pluripotent (stem) cells and novel anti-inflammatory compounds (Harman, 2008).

Growth and maturation of the placental villi

The placental villi continue to grow for most of the pregnancy. There is a widely held belief that the placenta ages during the pregnancy and that at term it is about to decline into functional senescence. Instead, the continuous morphological changes should perhaps be viewed as an increase in functional efficiency rather than ageing. Thus in early pregnancy, the placenta is a highly invasive and proliferative tissue and in later pregnancy, although its growth rate slows down, it continues to mature and increase in efficiency. Placental efficiency is favoured by the attenuated maternal–fetal barrier and reduced diffusion distance rather than by an increase in weight. Although the rate of placental growth does decline in the later part of gestation, this decrease in growth rate is not irreversible or inevitable. If the maternal environment becomes unfavourable, for instance because of maternal anaemia or increased altitude, fresh villus growth will ensue and the placenta will expand its surface area and continue branching past term. In all placentas, total placental DNA levels continue to increase linearly beyond the 40th week of gestation.

Growth of the placenta (see Fig. 8.6, p. 178) can be divided into three stages. Earlier in pregnancy, the trabeculae develop side-branches of syncytiotrophoblast protrusions (syncytial spouts) which may be filled with a core of cytotrophoblast. These primary villi protrude into the intervillous spaces. Later on, more lateral branches develop and the layers forming the placental barrier become more refined. In the ninth week, the tertiary stem villi lengthen to form mesenchymal villi as extraembryonic mesodermal cells penetrate the cytotrophoblast; the presence of this mesenchymal core transforms the villi into secondary or mesenchymal villi. Haematopoietic stem or progenitor cells develop within the mesoderm of the secondary villi forming the first placental blood cells and endothelial cells. Maternal and embryonic vascular systems do not connect; their development is similar and coordinated but independent and separate. The formation of placental blood vessels and cells transforms the villi into tertiary villi. Placenta blood vessels are formed in two processes: vasculogenesis is the formation of the first blood vessels from cells differentiated from the mesenchymal core and angiogenesis is the development of new vessels from existing vessels (Demir et al., 2007). These processes of blood vessel development are controlled by oxygenation, and the vascular endothelial growth factor (VEGF) family of growth factors and their receptors (Arroyo and Winn, 2008).

By the 16th week, the terminal extensions of the tertiary stem villi reach their maximum length. At this stage, the villi are described as immature intermediate villi. The cells of the cytotrophoblast layer become more dispersed within the villi creating gaps in the cytotrophoblast layer of the villus wall. Near the end of the second trimester, the tertiary stem villi form numerous side-branches and are described as mature intermediate villi. The earliest mature intermediate villi finish forming by about week 32 and then begin to produce small nodule-like secondary branches characteristic of the terminal villi. This is the final structure of the placental villous tree. The terminal villi are not formed by active outgrowth of the syncytiotrophoblast but by coiled and folded villus capillaries that bulge against the villus wall and expand by unfurling. Two types of chorionic villi can be identified: deep villi anchor the placenta to the decidua basalis; shorter villi extend into the intervillous spaces and have a nutritive role.

The blood-filled intervillous space into which the villi project is formed from the trophoblastic lacunae that grow and coalesce. Therefore, the intervillous space is lined on both sides with syncytiotrophoblast. The maternal face of the placenta is the basal plate, which consists of syncytiotrophoblast lining plus a supporting layer of decidua basalis. The fetal side is formed of the layers of chorion of the chorion plate.

The functional unit within the placenta is the placentome (or placental lobe), a villous tree arising from the chorionic plate within the intervillous space, which is perfused by a spiral artery. There are about 50–100 such units within the placenta. The villous tree has rami (major branches) and smaller ramuli. The terminal villi have little impedance to flow and therefore an increased fetoplacental flow; they are probably the major sites of nutrient and gaseous exchange in late gestation. The progressive development and branching of the placental tree structure is important for fetal growth and development. For instance, in IUGR pregnancies requiring elective preterm delivery, there are fewer terminal villi, which seem to have an abnormal extravillous cytotrophoblast structure. By term, the surface area of the placental villi is estimated to be 12–14 m² (Jauniaux et al., 2006).

On the basal (maternal) surface, the placenta is subdivided into cotyledons by wedge-like placental septa, which appear in the third month. The placental (decidual) septa grow into the intervillous space from the maternal side of the placenta, separating the villi into 10–40 cotyledons. The placental septae do not fuse with the chorionic plate, so maternal blood can flow freely from one cotyledon to another. This means that the villi are bathed in a lake of maternal blood which is constantly exchanging; this organization of placental perfusion is described as haemochorial. Haemochorial placentation is efficient because the trophoblast is in contact with maternal blood optimizing maternal–fetal transport of gases, nutrients, water and ions. The trophoblast can also endocytose the immunoglobulin IgG. In addition, hormones produced by the fetoplacental unit can easily access the maternal circulation. There are, however, some costs to the haemochorial arrangement; bleeding may be extensive at parturition and cells can be transferred between mother and fetus, for instance, resulting in microchimerisms (see Chapter 10) or erythroblastosis fetalis (haemolytic disease of the newborn due to Rhesus incompatibility).

Placental blood flow

The fetal blood reaches the placental blood system via the two umbilical arteries which spiral around the umbilical vein (Fig. 8.13). On reaching the chorion, the vessels usually each supply half of the placenta. The arteries (which are vessels carrying blood away from the fetal heart and therefore carry deoxygenated blood) divide repeatedly to form a branching network of smaller arteries and capillaries running through the intervillous space. The fetal blood flow through the placenta is about 500 mL/min, propelled by the fetal heart. Smooth muscle fibres contracting in the villi may help to pump blood back from the placenta to the fetus.

The maternal blood enters the intervillous space via about 50–100 of the remodelled spiral arteries. There is a pressure gradient from the maternal arteries to the intervillous space to the maternal veins. The blood leaves the intervillous space via the endometrial veins. Most organs have a progressive decrease in arterial diameter as the blood nears its target tissue. In the uteroplacental vessels, the remodelled spiral arteries increase in diameter as the vessels approach the intervillous space. Therefore, the intervillous space is a low-pressure system; the blood gently flows through and washes over the fetal placental tissue. The placenta has little resistance to maternal blood flow and a high vascular conductance so there is little fall in pressure across the intervillous space. The main determinant of the rate of maternal blood flow is the vascular resistance in the myometrial arteries. Myometrial contractions can decrease or stop afferent blood flow to the intervillous space. This effect is probably due to the compression or occlusion of the veins draining this space. During a contraction, the space distends so the fetus is not totally deprived of oxygen.

Fetal hypoxia, IUGR (see Box 8.3) or fetal death is often attributed to ‘placental insufficiency’. A proportion of those babies with a low birthweight (<2.5 kg) probably failed to achieve their growth potential because placental transfer of oxygen and nutrients was inadequate. However, the fetal placenta is rarely insufficient. Like all essential organs, it has a considerable physiological reserve. It has been estimated that the placenta could lose 30–40% of its villi (and therefore surface area) without affecting its function. However, the placentas of growth-restricted fetuses may exhibit pathological changes such as reduced syncytiotrophoblast area, increased placental apoptosis or increased thickness of the exchange barrier (Arroyo and Winn., 2008).

Box 8.3

Intrauterine growth restriction

UUGR increases the risk of disability or death for the fetus and neonate in the perinatal period and predisposes the individual born following IUGR to later adult disease (see Chapter 12). Although there is no internationally agreed definition for IUGR, it is usually defined small for gestation age (SGA) and as a birth weight below a certain percentile (10th, 5th or 3rd percentile). It may be further qualified as also including a longitudinal decrease in the growth of the abdominal circumference, increased fetal head:abdomen circumference ratio or oligohydramnios. Risk factors for IUGR include acquired blood borne infections (such as malaria, rubella and cytomegalovirus), pre-existing maternal disease (cardiovascular, endocrine, autoimmune), aneuploidies, metabolic factors and placental disorders including abnormal placental position. However, for most infants with IUGR, there is no known cause; idiopathic IUGR is often described as ‘placental insufficiency’. In IUGR, there are marked reductions in the placental delivery of amino acids to the fetus (Cetin and Alvino, 2009) which is reflected in decreased deposition of tissue. This is probably due to both blood flow and arteriovenous differences in nutrient concentrations being compromised as well as reduced activity in the placental amino acid transport systems. Placental transport of glucose in IUGR seems no different to a normal placenta but the placental expression of the lipoprotein receptors and lipoprotein lipases involved in transfer of maternal fatty acids is altered. In IUGR, there also appears to be increased placental permeability and increased placental oxygen utilization. Where there is IUGR with a normal umbilical blood flow, amino acid and LCPUFA concentrations are significantly reduced and non-esterified fatty acids levels are increased. However, when umbilical blood flow is also impaired in severe IUGR, fetal blood flow to the brain, liver and heart is altered leading to the fetus becoming hypoxic and lactacidaemic with subsequent effects on the fetal growth trajectory. Associated with these changes, the fetus adapts to nutrient intake not meeting demand by increasing protein catabolism, reducing metabolic rate and making endocrine adaptations; these adaptation mechanisms are not without cost and may have lifelong health consequence (see Chapter 12). The most serious complications of IUGR occur in fetuses weighing less than 500 g. One approach to correcting IUGR might be placental gene therapy, particularly as growth factors (such as insulin growth factors, placental growth factor and VEGF) are all implicated in failure of trophoblast remodelling of the uterine spiral arteries which appears to be the underlying abnormality in both IUGR and pre-eclampsia. Although development of such treatment is theoretical at the moment, it does raise interesting ethical questions about whether the patient in such a scenario is the woman or her fetus and whether gene therapy could be justified if it was safe and effective, where there was reasonable certainty that the fetus would suffer irreversible and substantial harm without the intervention and where the risk to the mother was negligible.

Placental insufficiency really describes inadequate maternal uteroplacental blood flow, which is probably due to incomplete conversion of the spiral arteries during the early stages of pregnancy. Studies suggest the uteroplacental perfusion is greatly reduced because of a failure of the trophoblast invasion into the myometrium and subsequent remodelling of the spiral arteries (Brosens et al., 2002) which results in fewer terminal villi and other placental abnormalities affecting blood vessels and membranes involved in diffusion. It is possible that some small-stem arterioles may be occluded by fetal platelets (Wilcox and Trudinger, 1991). Measurement of abnormal oxygen and amino acid levels in the umbilical vein blood suggests a defect in placental transport mechanisms (Cetin et al., 1990). However, it is not established whether these changes are causative or adaptive. Compensatory mechanisms exist in the fetus, which result in redistribution of blood to the fetal brain at the expense of the lower body. This is supported by the finding that amniotic fluid volume is decreased presumably because blood flow to the kidneys is reduced (Kingdom and Sibley, 1996). Substances that cause vasoconstriction, such as cocaine and alcohol, are implicated in preterm labour, possibly because they cause a decrease in blood flow to the placenta affecting uterine contractility and sensitivity.

Case study 8.2 details an example of a small baby.

Case study 8.2

Polly was diagnosed as carrying a small-for-dates baby. She spontaneously delivered Thomas at 39 weeks, and although he weighed only 2.4 kg, he appeared healthy and vigorous. The midwife noted that the third stage appeared complete but failed to identify that the placenta appeared relatively large.

• Do you think that there is any need to weigh placentas and to compare the placental and birth weights?

• Are there any situations where the weight and condition of the placenta may be used as a possible indicator for disease states in later life?

Blood leaving the right atrium is diverted into the ductus arteriosus, into the aorta and down to the lower body (see Chapter 15). At term, about 40–50% of the fetal cardiac output goes to the placenta via the umbilical arteries. Blood flow from the aorta to the umbilical arteries is high because the resistance to flow in these vessels is low compared with the systemic circulation of the lower body. The vessels of the fetoplacental circulation lack autonomic innervation but a variety of substances can affect the smooth muscle of the stem villous arteries. Of particular importance are paracrine agents, which have a local effect on the fetoplacental circulation. Both prostacyclin and nitric oxide (NO), which have vasodilatory and anticoagulant effects, are produced from the vessel endothelium. It is suggested that flow-mediated release of NO may have an important role (Learmont et al., 1994). Diffusion of NO into the intervillous space affecting maternal uteroplacental vessels may also be important (Myatt et al., 1993). The heterogeneous cells of the placental vessel endothelium also produce endothelin-1 (a potent vasoconstrictor), substance P, serotonin, ATP, atrial natriuretic peptide (ANP) and neuropeptide Y (NPY) (Cai et al., 1993 and Myatt et al., 1992); whether these substances have a physiological role is yet to be established.

Optimal placental exchange requires adequate vascularization of the placental bed by the maternal arteries matched by circulation of fetal blood to the placenta. The fetus does not appear to have a mechanism to increase umbilical flow in response to hypoxia or volume depletion. It has a limited ability to increase cardiac output. Therefore, the fetus adapts to hypoxia or decreased nutrient availability by decreasing oxygen consumption and growth rate. The cardiac output is redistributed to the heart, brain and adrenal glands at the expense of the flow to the body and gut. Hypoxia and acidosis cause cerebral vasodilatation and constriction of the pulmonary and femoral vessels. Blood flow to the liver is high when oxygen and nutrients are plentiful but the hepatic circulation is bypassed if placental exchange is compromised.

It is hypothesized that perfusion of the placental vessels is controlled to match the maternal perfusion of the uteroplacental vessel in a similar way to the perfusion–ventilation matching in the neonatal or adult pulmonary system (see Chapter 1). If an area of the placenta is underperfused by the maternal blood flow, hypoxia ensues. The endothelium of the placental vessels responds by vasoconstricting (by decreasing NO synthesis and increasing endothelin-1 production) so fetoplacental blood flow is diverted to a better-perfused villous tree.

Placental transport mechanisms

Many substances are transported from the maternal blood in the intervillous space to the fetal blood in the capillaries of the villi and vice versa. By term, most exchange occurs in the terminal villi, which have a high surface area and small diffusion distance, perhaps of only a few micrometres in some areas. The surface area of the placenta is calculated to be 5 m² at 28 weeks, increasing to about 11 m² at term (Carlson, 1994). The precise mechanisms of placental transport for many substances are not clear. Transport mechanisms include simple diffusion and transporter-mediated processes; these can be active or facilitated. The effectiveness of the transport mechanism depends on the morphological characteristics of the placenta (like surface area and barrier thickness) and on the abundance and distribution of specific transporters. Simple and facilitated diffusion depends on the concentration gradient, the placental permeability and the surface area; however, passive diffusion alone is not likely to be adequate for fetal requirements for nutrients.

Lipophilic substances (soluble in lipid such as respiratory gases) are soluble in cell membranes so their transport depends on the concentration gradient and the relative rates of maternal and fetal blood flow. Because the placenta provides a large surface area, the transfer of respiratory gases depends on the maternofetal concentration difference which depends on the flow rates of the uterine and umbilical circulations (Desforges and Sibley, 2010). Diffusion of hydrophilic substances (soluble in aqueous solutions but poor solubility in lipid bilayers) is limited by the diffusion distance (placenta thickness) and the surface area of the membranes of the placental barrier. The fetal capillary endothelium probably limits transport of large proteins (such as albumin, IgG and AFP). Transport studies of the syncytiotrophoblast suggest that it is much more permeable than was previously believed and probably has channels or pores which offer a route continuous with, and containing, extracellular fluid (Kingdom and Sibley, 1996), which allows the diffusion of large proteins. The transfer of substances across the placenta occurs in both directions, to and from the fetus.

There are specific transport proteins on the placental plasma membrane involved in the efficient transfer of metabolically important substances. Some of these proteins form channels and others act as shuttles or carriers. Glucose demands are high and are estimated to be 4–8 mg/kg/min (Aldoretta and Hay, 1995). Glucose transport is predominantly carried out by the facilitative-diffusion glucose transporter GLUT1 and also by GLUT3 (Knipp et al., 1999). Glucose transport by GLUT1 is concentration-dependent, bidirectional and independent of insulin. GLUT1 expression is higher on the maternal side of the syncytiotrophoblast which favours mother-to-fetus transfer and protects glucose being transported from the fetus during maternal hypoglycaemia. GLUT1 expression is downregulated by hyperglycaemia so fetal development is partially protected if there is maternal hyperglycaemia. GLUT3 is not the major facilitator of glucose transport to the developing fetus but may be important during periods of maternal hypo- or hyperglycaemia. Fetal glucose levels are usually slightly lower and directly related to maternal levels. The difference between fetal and maternal glucose levels increases with the severity of IUGR.

Some substances, such as certain amino acids and calcium, are transported by active (energy-dependent) transport against their electrochemical gradients. There are several amino acid transfer systems in the uterine and placental tissues. The system A amino acid transporter is sodium-dependent and transports neutral amino acids with short or linear side chains such as alanine, glutamine, methionine, serine, proline and glycine (Desforges and Sibley, 2010). System A activity is affected by substrate availability and hormones; it is also inhibited by hypoxia and oxidative stress. The sodium-independent amino acid transport systems L and γ+ transport branched chain amino acids and lysine. Placental transfer of amino acids is reduced in IUGR probably because there are fewer transporters expressed and thus a reduced maximum rate of transfer. Amino acid levels are usually higher in the fetus than in the mother as expected with active transport systems but where fetus is growth-restricted, maternal amino acid levels are not as low as in normal pregnancies.

Fatty acids, particularly the long-chain polyunsaturated fatty acids (LCPUFA), are important for synthesis of phospholipids and cell membranes, and growth and development of the brain and nervous system. The fetus is dependent on placental transfer which is mediated by specific fatty acid binding and transfer proteins and preferentially transports LCPUFA (Cetin and Alvino, 2009). The placental transfer of maternally derived fatty acids requires placental lipases which hydrolyse lipoprotein-borne triacylglycerides and phospholipids. The placenta nucleoside transporters enable the fetus to meet its high demands for nucleosides to synthesize nucleotides; transport of adenosine is decreased by ethanol. There are additional mechanisms for the transport of some very large molecules such as receptor-mediated pinocytosis for IgG (see Chapter 10). There is a net flux of water to the fetus, mostly across the placenta.

Steroid hormones cross the placenta but peptide hormones seem to be poorly transferred. Gas transfer occurs by diffusion and is probably limited by blood flow. As well as oxygen and carbon dioxide, the placenta permits diffusion of other gases such as carbon monoxide and inhalation anaesthetics. The placenta itself has a high rate of oxygen consumption; much of this oxygen is used for oxidative phosphorylation of glucose. The consequent production of ATP is used mainly for synthesis of peptide and steroid hormone and for transport of nutrients.

The placenta has a protective function and limits transfer of some xenobiotic substances to the fetus. The placenta expresses cytochrome P450 enzymes which metabolize and detoxify a number of drugs. There are also export pumps in the syncytiotrophoblast which reduce placental transfer of potentially toxic substances. However, some bacteria (such as the one that causes syphilis), some protozoa (such as the parasite that causes toxoplasmosis) and a number of viruses (including HIV, cytomegalovirus, rubella, polio and varicella) can cross the placenta and affect outcome.

The placenta acts as a nutrient sensor and regulates nutrient transfer depending on the ability of the maternal circulation to supply the nutrients. Furthermore, the placenta has its own nutrient demands; it extracts a fixed proportion of maternal nutrients (70% of the glucose and 40% of the oxygen) (Miller et al., 2008) and can also take nutrients, such as amino acids, from the fetal circulation for its own nutrient needs (Cetin and Alvino, 2009). This means the fetus is vulnerable to nutrient deprivation as it is restricted to the surplus nutrients that remain after placental demands are met. So even slight placental dysfunction can restrict nutrient transfer and blood flow to the fetus whilst maintaining the high level of placental nutrition. Likewise, placental oxygen uptake also seems to remain constant even when there are acute reductions in uterine oxygen supply, so it is the fetal level of oxygenation which is compromised in such conditions. If uterine perfusion is reduced, delivery of glucose and amino acids to the fetus can be compromised. Such a reduction in substrate availability can affect growth and metabolism of the fetus. The fetal compensatory responses include down-regulation of the insulin and IGF1 axis and hepatic glucose metabolism. This results in glycogenolysis and endogenous protein breakdown which increase fetal glucose and amino acid levels but potentially compromise growth. Endocrine response includes hypothyroidism, bone demineralization and up-regulation of the adrenocortical axis. Fetal red blood cell mass increase which not only may exacerbate placental dysfunction but also results in increased risk of thrombocytopenia, increased blood viscosity, and platelet aggregation.

Placental hormone production

Placental hormones have a role in adjusting maternal physiology to provide the optimal environment for fetal development (see Chapter 11); however, roles for all of the placental products have not yet been elucidated. Concentrations of placental protein hormones are higher in the maternal blood than in the fetus because the fetal circulation limits the transfer of large molecules (Firth and Leach, 1996). Conversely, levels of steroid hormones are about 10 times higher in the fetal circulation (Chard, 1998). Although the levels of placental protein hormone fluctuate randomly, a true circadian rhythm of placental secretion has never been demonstrated (Chard, 1998). This includes secretion of hCG, which is not higher in urine specimens collected in the morning (Kent et al., 1991). Hormonal concentrations also change in response to environmental challenges such as changes in maternal blood flow, compromised maternal diet and hypoxia. In the fetal compartment, environmental conditions that favour fetal growth increase concentrations of anabolic hormones such as thyroid hormones, insulin and IGFs and decrease concentrations of catabolic hormones such as catecholamines and cortisol (Fowden and Forhead, 2009).

The placenta has a broad endocrine capacity and diversity, producing many hormones that other endocrine organs also produce. The syncytiotrophoblast is probably the source of most placental products including hormones, growth factors and cytokines, although the cytotrophoblast may also produce hCG, human placental lactogen (hPL), inhibin, relaxin and placental releasing hormones (Chard, 1998). The major steroids produced are progesterone and oestrogens (oestriol). The production of oestrogens requires both maternal and fetal precursors, so monitoring maternal oestrogen levels during the pregnancy is a useful indicator of fetal well-being. Cholesterol from maternal low-density lipoprotein (LDL) is mostly used as the precursor for steroid hormone production. Oestriol synthesis requires 16α-hydroxydehydroandrosterone sulphate derived from the fetal liver and adrenal gland. Most of the steroid hormones produced enter the mother's circulation, affecting her physiology (see Chapter 11). The placenta also produces neuropeptides (although it has no nerves), which may regulate placental hormone production, leptin, growth factors and cytokines.

Embryo development requires progesterone to maintain uterine quiescence. Initially hCG from the trophoblast rescues the corpus luteum from atresia (see Chapters 4 and 6) thus maintaining the production of oestrogen and progesterone. Release of hCG from the trophoblast seems to begin about 7 days after fertilization (Chard, 1998). However, levels of hCG and luteal steroid hormones are not directly related in normal pregnancies (Hamilton-Fairley and Johnson, 1998). Hormone levels fall following in vitro fertilization (IVF) despite increasing levels of hCG (Johnson et al., 1993a). The relationship between hCG and steroid hormone production is stronger in anembryonic pregnancies (Johnson et al., 1993b), where embryonic development has failed, suggesting that the embryo itself takes over the control of steroid hormone production by the corpus luteum. The corpus luteum becomes redundant at about 7 weeks after fertilization when steroid hormone production is taken over by the placenta. The change in site of production is described as the luteoplacental shift; a relative progesterone and oestradiol deficiency can develop and lead to abortion (Schindler, 2004). Inadequate hormone production by the corpus luteum early in pregnancy before this shift, or insufficient placental development, is also thought to be responsible for early miscarriages.

hCG can be detected in maternal serum and urine once implantation has occurred about 9–10 days postfertilization when secretions from the trophoblast can enter the maternal vessels. hCG is composed of two subunits; the α-subunit is common to all glycoprotein hormones such as LH (luteinizing hormone), FSH (follicle-stimulating hormone) and TSH (thyroid-stimulating hormone). Dissociated α- and β-subunits of hCG, as well as the intact dimer (the complete hCG molecule formed of two subunits), are measurable in pregnancy (Kingdom and Sibley, 1996). The concentration of α-subunits progressively increases throughout the pregnancy reaching maximal levels at about 36 weeks. The concentration of free β-subunits parallels the concentration of intact dimer, reaching a peak about 10 weeks after fertilization, and then declines to a plateau. It is thought that the cytotrophoblast produces α-subunits and the more differentiated syncytiotrophoblast produces both α- and β-subunits (Kingdom and Sibley, 1996). Raised levels of β-subunits of hCG are associated with Down's syndrome, reflecting abnormal formation of the syncytiotrophoblast (Spencer et al., 1992). Levels of the hormone are also significantly higher in cases of severe pre-eclampsia (see Box 8.1, p. 181) and IUGR (Wenstrom et al., 1994). hCG promotes cytotrophoblast differentiation and migration of extravillous trophoblasts (Banerjee and Fazleabas, 2010). The role of hCG in maintaining hormone production by the corpus luteum is clearly established. However, the peak of hCG production is reached after the function of the corpus luteum has already started to decline, suggesting other roles for the hormone. hCG may have immunoregulatory roles and also be involved with fetal testosterone production and male sexual differentiation (see Chapter 5). Because of the structural similarity between hCG and LH, hCG is used in ART for controlled ovarian stimulation (see p. 133) because it reduces the risk of ovarian hyperstimulation syndrome. It has been suggested that higher levels of hCG in early pregnancy are associated with a long-term protection against breast cancer (Banerjee and Fazleabas, 2010). In the myometrium, hCG inhibits smooth muscle contraction via several mechanisms and so may be important in preventing premature delivery (Ticconi, 2007).

hPL (also known as human chorionic somatomammotropin, hCS) is also a product of the syncytiotrophoblast; it is structurally and functionally similar to human growth hormone. Levels of hPL increase throughout gestation and correlate well with placental syncytiotrophoblast mass so it can be used clinically to evaluate placental function. By term, 1–3 g of hPL is produced per day (equivalent to 5–7 mg/mL in maternal blood). hPL affects maternal metabolism, erythropoietin activity, fetal growth, mammary gland development and ovarian function; it induces insulin resistance and carbohydrate intolerance (see p. 281) and is important in the provision of fatty acids for the fetus. Lower hPL levels are associated with pre-eclampsia, aborting molar pregnancy (hydatidiform mole), choriocarcinoma and placental insufficiency. Higher than normal hPL levels are associated with multiple pregnancies, placental tumours, intact molar pregnancy, diabetes and Rhesus incompatibility. However, there are reports of women with abnormally low or absent levels of hPL having completely normal pregnancies (Kingdom and Sibley, 1996).

PGH is synthesized by the syncytiotrophoblast. It has high somatogenic (growth) activity and low lactogenic activity. PGH is structurally similar to hPL and pituitary growth hormone. It gradually replaces pituitary growth hormone in the maternal circulation (Evain-Brion and Malassine, 2003). It is secreted continuously only into the maternal circulation (whereas pituitary growth hormone has a pulsatile secretory pattern) and is important in facilitating maternal metabolic adaptation to pregnancy and regulating the amount of glucose and amino acids available for placental extraction from the maternal circulation. Together with hPL, PGH regulates the serum levels of insulin-IGFs; levels of PGH correlate with the birthweight of the newborns.

The allantois and yolk sac

The allantois and yolk sac are semivestigial structures that have a more important role in other species, such as birds and reptiles, where the yolk sac is important in nutrition of the maternally isolated eggs and the allantois has a respiratory and excretory role. The allantois forms from a pocket of the hindgut embedded within the umbilical cord, which is incorporated into the developing urinary system. Blood cells develop in the wall of the allantois during weeks 3–5 and its blood vessels become the vessels of the umbilical cord which forms in the region of the body stalk and is covered by the developing amnion. The embryonic structures and the right umbilical vein disappear, leaving two arteries and one vein. The yolk sac develops on the ventral side of the embryonic disc and is important in nutrition of the embryo while the uteroplacental circulation is forming. The primordial germ cells (see Chapter 5) and the blood islands develop in the tissue of the yolk sac. The yolk sac becomes thin and elongated and is incorporated into the umbilical cord and primitive gut. Its role in haematopoiesis is taken over by the liver in the sixth week of development. The remnant of the yolk sac lies between the chorion and amnion; it is a calcified yellow nodule about 4 mm long (Kaplan, 2008).

The placenta at term

The mature placenta is an oval/round disc with a diameter of about 18–22 cm and 2–3 cm thick in the middle, petering out towards the edges. At the placental margins, the basal and chorionic surfaces of the placenta unite to form the fetal chorionic membrane. On average, a placenta weighs about one-sixth of the weight of the fetus, about 470–500 g. The amniotic membrane is smooth so the fetal aspect of the placenta, the chorionic plate, appears shiny and grey. The amnion is a single layer of epithelial cells and avascular connective tissue which is weakly attached and can be easily removed from the delivered placenta. The basal plate is maternal surface of the placenta which appears grooved and lobed with a dull red coloration. This basal surface is a mixture of extravillous trophoblasts, maternal decidua cells and immune cells such as macrophages and natural killer cells, extracellular matrix, fibrinoid and blood clots. The chorionic membrane retains the ridged appearance owing to the regression of the early villi. The umbilical cord is usually inserted slightly eccentrically into the chorionic plate. The umbilical cord gets progressively longer with the duration of the pregnancy as fetal activity and traction on the cord increases its length. At term, the umbilical cord is normally between 50 and 60 cm long. If the cord is abnormally short, it can cause bleeding problems. If it is long (>70 cm), it may prolapse through the cervix or entangle with the fetus, possibly forming knots that could obstruct fetal circulation during delivery, causing potentially fetal distress and death. Abnormal cord length is used clinically as a marker of undetected intrauterine events (Calvano et al., 2000); although it is not clear whether cord length is long because of traction associated with entanglement or whether entanglements occur because the cord is long. There is thought to be a genetic component to abnormally long cords (Kaplan, 2008). For a normal vaginal delivery, a cord length of at least 32 cm is thought to be necessary to avoid fetal distress, cord tearing and possible abruption. Short cords are associated with poor fetal movement (such as in oligohydramnios) and prenatal exposure to alcohol and drugs such as cocaine; short cords have been associated with compromised neurological development (Kaplan, 2008). Most umbilical cords are twisted; usually the twists are counterclockwise ‘left twists’ every few centimetres. Cords without any twists are associated with single umbilical artery (SUA) and an increased risk of perinatal mortality. Excessive twisting, which can compromise blood flow, is also associated with fetal morbidity and mortality. True knots in the umbilical cord occur in about 1% of births (Moore and Persaud, 1998). The vessels of the cord, two arteries carrying blood from the fetus and one vein carrying blood to the fetus, are embedded in Wharton's jelly. This jelly is a connective tissue that protects the vessels of the cord.

Examination of the placenta

Examination of the placenta, membranes and umbilical cord at delivery is an important responsibility of the attendant midwives. A quick visual inspection of the maternal and fetal surfaces can pick up the occasional abnormal specimen; unusual odour which could indicate a bacterial infection and substantial amounts of fresh clot could indicate premature placental separation. Cord length and diameter are assessed and the distance of insertion to the nearest placental margin. The cord is checked for vessel number, true knots, twisting, discolouration, congestion and thrombosis. There are usually two arteries and the persisting left umbilical vein. The SUA occurs in about 1% of births and is associated with an increased frequency of fetal and chromosomal abnormalities, particularly of the renal or cardiovascular systems (Benirschke, 1994); however, it is normal for the arteries to fuse together just above the fetal surface. Sometimes more vessels are present because the right umbilical vein has not regressed. Although an abnormality in the number of vessels is associated with congenital abnormalities, it is not clear whether the wrong number of vessels is a cause or a result of the abnormality. About 20% of infants with SUA will have other major congenital abnormalities; the remainder are often slightly small and have an increased risk of perinatal mortality (Kaplan, 2008). Macrosomic babies of diabetic mothers tend to have thick oedematous cords, whereas thin delicate cords are associated with IUGR. The cord can be inserted into the placental bed in different ways. Insertion of the cord is usually approximately central but may be lateral. Abnormal insertion of the cord can create problems at delivery (Table 8.2). If the cord is ruptured, it indicates that there may have been some fetal blood loss during labour. The chorionic plate opacity and colour including the amount of fibrin and thrombosis in the subchorionic region can also be assessed.

The type of membrane insertion and their completeness are assessed. The membranes are easier to examine if the placenta is held up by the cord. Usually, the two membranes hang down in a neat uniform way. The placenta is continuous with the chorion but the amnion should be able to be separated from the chorion up to the base of the cord. If the membranes are ragged and torn, some parts of the membrane may be retained in the uterus, which can impede uterine involution and staunching of blood loss. Meconium can discolour the membranes in late gestation; fresh meconium may also be present.

A healthy placenta is normally rounded and uniform but shape is quite variable. Unusual shapes may be a result of uterine cavity abnormalities. Placental weight is usually between 350 and 750 g; excessively light or heavy placentas are associated with pathological conditions. An excessively large or oedematous (soft) placenta is associated with maternal diabetes, hydrops or cardiac abnormalities. Maternal diabetes tends to result in placenta with a deep red colour. The placenta is also examined for abnormal numbers of lobes (Table 8.2) or missing areas of the maternal surface, which could indicate that a lobe has been retained, potentially causing serious postpartum bleeding. Depending on whether twins are monozygotic or dizygotic, the placenta may be shared or regions fused (Box 8.4). The maternal surface is examined for completeness, adherent blood clot, lesions (such as infarcts and thrombi) and degree of calcification. Cysts are common on the surface and are associated with fibrin deposition but are usually not significant. True placental infarcts are also common, they tend to be small (<1 cm) and located at the placental margins. Haemorrhages on the maternal surface are usually due to premature separation and are more common with hypertensive disorders, ascending infection, smoking and cocaine use (Kaplan, 2008).

Box 8.4

The placenta in multiple pregnancies

Dizygotic (non-identical) twins and monozygotic (identical) twins resulting from early splitting of the blastocyst prior to implantation can have separate placentas and membranes. However, if the two blastocysts implant in close proximity, the placentas and chorion may fuse. If monozygotic twins arise from division of the inner cell mass, they usually have separate amnions but share the placenta and chorion. The vascular systems within the placenta may remain separate but can fuse. If the vascular systems fuse within the placenta, twin-to-twin transfusion syndrome may occur where the twins have an unequal blood supply. This condition, which occurs in 10–15% of monozygotic twins (thus affecting 1 in 400 pregnancies, 1 in 1600 babies) with a shared placenta, can threaten the survival of both twins because the donor twin is anaemic and the recipient twin is polycythaemic and prone to cardiac hypertrophy and heart failure. Modern treatments have increased the survival of twins but the survivors are at increased risk of brain injury and neurodevelopment consequences.

Sometimes a dense raised white rim may be observed on the periphery of the fetal placental bed; this is called a circumvallate placenta. It is thought to be caused by deep implantation of the placenta into the maternal decidua and subsequent partial separation of the placental from the uterine wall. This results in the folding back of the membrane towards the chorionic surface. The rim is a double fold of fetal membranes with degenerated decidua and fibrin between them. The clinical significance of the condition is uncertain but it may be a risk factor for antenatal haemorrhage and/or severe intermittent uterine contractions. It is more common in multigravidae and in women who have previously had a circumvallate placenta.

Case study 8.3 details an example of placental abnormality revealed by inspection.

Key points

• The placenta derives largely from the trophoblast layer of the embryo, which differentiates into two layers: the cytotrophoblast and the syncytiotrophoblast.

• The cytotrophoblast undergoes rapid mitosis and the syncytiotrophoblast aggressively digests and invades the maternal endometrial wall.

• Fragments of maternal blood vessels are engulfed forming lacunae and a framework for villi development.

• Extravillous cytotrophoblast invades the maternal circulation resulting in remodelling of the spiral arteries.

• Extraembryonic mesoderm, originating from the inner cell mass, invades the core of the villi and establishes the vasculature of the villi.

• The villi continue to grow and remodel throughout the pregnancy; the barrier to diffusion is reduced as fetal requirements increase.

• The placenta has specific transport mechanisms and a range of endocrine activities. It also has an important immunological role.

• Amniotic fluid, produced by the amniotic membrane, cushions and protects the fetus. It is also important in the development of the respiratory system.

• Inadequate maternal uteroplacental blood flow, described as placental insufficiency, is associated with the aetiology of pre-eclampsia and IUGR.

• Examination of the placenta is important in detecting any abnormality or retention of placental tissue.

Application to practice

The placenta has an important physiological role in supporting and maintaining pregnancy. Dysfunctioning of the placenta and its development results in abnormal conditions, which may be observed in pregnancy which may affect the health of the mother and baby before and after birth.

Knowledge of the gross anatomy and the variants in the placental structure is essential in the postnatal examination of the placenta and membranes.

Case study 8.3

Following what appeared to be a normal delivery, the midwife inspected the placenta and membranes. She discovered a hole in the membranes that had blood vessels leading to it, radiating out from the main body of the placenta.

• What do you think the midwife concluded from these findings?

• What care and observation will the woman require?

• How will this be explained to the woman and what information might she require?

Annotated further reading

Benirschke, K.; Kaufmann, P., Pathology of the human placenta. ed 5 (2006) Springer, New York .

A comprehensive reference text, which covers the structure of the placenta at birth, types of placenta, early development and cellular details.

Cudihy, D.; Lee, R.V., The pathophysiology of pre-eclampsia: current clinical concepts, J Obstet Gynaecol 29 (2009) 576–582.

This is an overview of existing knowledge about pre-eclampsia, which can still be described as a ‘medical mystery’, which identifies and attempts to unify findings of past and current scientific investigation providing examination of the known risk factors, the epidemiologic trends, and recent research about the pathogenesis of the disease.

Desforges, M.; Sibley, C.P., Placental nutrient supply and fetal growth, Int J Dev Biol 54 (2010) 377–390.

Detailed review of the mechanisms involved in placental transfer and how the capability of the placenta to supply nutrients affects fetal growth.

Fisk, N.M.; Duncombe, G.J.; Sullivan, M.H., The basic and clinical science of twin-twin transfusion syndrome, Placenta 30 (2009) 379–390.

Recent review about twin-to-twin transfusion syndrome which includes discussion on the underlying placental pathophysiology, the resulting fetal pathophysiology, fetal surveillance and recent therapeutic advances.

Harman, C.R., Amniotic fluid abnormalities, Semin Perinatol 32 (2008) 288–294.

A clear and well-written description of the functions and dynamics of normal amniotic fluid volume and composition with detailed sections on oligohydramnios and polyhydramnios.

A review of placental development which also covers placental functions and transport mechanisms.

Huppertz, B., The anatomy of the normal placenta, J Clin Pathol 61 (2008) 1296–1302.

An excellent review describing the development of the placenta including the gross and microscopic anatomy and histology of the delivered placenta.

Arroyo, J.A.; Winn, V.D., Vasculogenesis and angiogenesis in the IUGR placenta, Semin Perinatol 32 (2008) 172–177.

A clearly written and well-illustrated description of placenta blood vessel formation in normal placental development and in pregnancies complicated by intrauterine growth restriction.

Kaplan, C.G., Gross pathology of the placenta: weight, shape, size, colour, J Clin Pathol 61 (2008) 1285–1295.

A comprehensive review of placental abnormalities, illustrated with photographic examples, which includes guidance on placental examination.

Kingdom, J.; Jauniaux, E.; O'Brien, S., The placenta: basic science and clinical practice. (2000) Royal College of Obstetricians and Gynaecologists, London .

A comprehensive handbook covering scientific and clinical aspects of placental structure and function, placental pathology including growth restriction and pre-eclampsia), placental infection, preterm labour, placental malignancy, clinical assessment of the placenta and placental complications in labour.

Tyson, R.W.; Staat, B.C., The intrauterine growth-restricted fetus and placenta evaluation, Semin Perinatol 32 (2008) 166–171.

An excellent review which relates evaluation of the placenta to the pathophysiology underlying intrauterine growth restriction; includes an excellent table of the common gross and microscopic characteristics of the placenta in growth restriction.

References

Achache, H.; Revel, A., Endometrial receptivity markers, the journey to successful embryo implantation, Hum Reprod Update 12 (2006) 731–746.

Aldoretta, P.W.; Hay Jr, W.W., Metabolic substrates for fetal energy metabolism and growth, Clin Perinatol 22 (1) (1995) 15–36.

Arroyo, J.A.; Winn, V.D., Vasculogenesis and angiogenesis in the IUGR placenta, Semin Perinatol 32 (2008) 172–177.

Benirschke, K., Obstetrically important lesions of the umbilical cord, J Reprod Med 39 (1994) 226.

Banerjee, P.; Fazleabas, A.T., Endometrial responses to embryonic signals in the primate, Int J Dev Biol 54 (2010) 295–302.

Bischof, P.; Meisser, A.; Campana, A., Paracrine and autocrine regulators of trophoblast invasion: a review, Placenta 21 (Suppl A) (2000) S55–S60.

Brosens, J.J.; Pijnenborg, R.; Brosens, I.A., The myometrial junctional zone spiral arteries in normal and abnormal pregnancies: a review of the literature, Am J Obstet Gynecol 187 (5) (2002) 1416–1423.

Burton, G.J.; Hempstock, J.; Jauniaux, E., Nutrition of the human fetus during the first trimester: a review, Placenta 22 (Suppl A) (2001) S70–S77.

Burton, G.J.; Woods, A.W.; Jauniaux, E.; et al., Rheological and physiological consequences of conversion of the maternal spiral arteries for uteroplacental blood flow during human pregnancy, Placenta 30(2009) 473–482.

Burton, G.J.; Yung, H.W.; Cindrova-Davies, T.; et al., Placental endoplasmic reticulum stress and oxidative stress in the pathophysiology of unexplained intrauterine growth restriction and early onset preeclampsia, Placenta 30(Suppl A) (2009) S43–S48.

Cai, W.Q.; Bodin, P.; Sexton, A.; et al., Localization of neuropeptide Y and atrial natriuretic peptide in the endothelial cells of human umbilical blood vessels, Cell Tissue Res 272 (1993) 175–181.

Calvano, C.J.; Hoar, R.M.; Mankes, R.F.; et al., Experimental study of umbilical cord length as a marker of fetal alcohol syndrome, Teratology 61 (3) (2000) 184–188.

Carlson, B.M., Human embryology and developmental biology. (1994) Mosby, St. Louis .

Cetin, I.; Corbetta, C.; Sereni, L.P.; et al., Umbilical amino acid concentrations in normal and growth-retarded fetuses sampled in utero by cordocentesis, Am J Obstet Gynecol 162 (1990) 253–261.

Cetin, I.; Alvino, G., Intrauterine growth restriction: implications for placental metabolism and transport. A review, Placenta 30 (Suppl A) (2009) S77–S82.

Chard, T., Placental metabolism, In: (Editors: Chamberlain, G.; Broughton Pipkin, F.) Clinical physiology in obstetrics ed 3 (1998) Blackwell, Oxford, pp. 419–436.

Demir, R.; Seval, Y.; Huppertz, B., Vasculogenesis and angiogenesis in the early human placenta, Acta Histochem 109 (2007) 257–265.

Desforges, M.; Sibley, C.P., Placental nutrient supply and fetal growth, Int J Dev Biol 54 (2010) 377–390.

Edwards, A.; Megens, A.; Peek, M.; et al., Sexual origins of placental dysfunction, Lancet 355 (2000) 203–204.

Evain-Brion, D.; Malassine, A., Human placenta as an endocrine organ, Growth Horm IGF Res 13 (Suppl A) (2003) S34–S37.

Firth, J.A.; Leach, L., Not trophoblast alone: a review of the contribution to the fetal microvasculature to transplacental exchange, Placenta 17 (1996) 89.

Fowden, A.L.; Forhead, A.J., Endocrine regulation of feto-placental growth, Horm Res 72 (2009) 257–265.

Fowden, A.L.; Sferruzzi-Perri, A.N.; Coan, P.M.; et al., Placental efficiency and adaptation: endocrine regulation, J Physiol 587 (2009) 3459–3472.

Hamilton-Fairley, D.; Johnson, M.R., The ovary, In: (Editors: Chamberlain, G.; Broughton Pipkin, F.) Clinical physiology in obstetrics ed 3 (1998) Blackwell, Oxford, pp. 396–416.

Harman, C.R.; Baschat, A.A., Comprehensive assessment of fetal wellbeing: which Doppler tests should be performed? Curr Opin Obstet Gynecol 15 (2) (2003) 147–157.

Harman, C.R., Amniotic fluid abnormalities, Semin Perinatol 32 (2008) 288–294.

Hawfield, A.; Freedman, B.I., Pre-eclampsia: the pivotal role of the placenta in its pathophysiology and markers for early detection, Ther Adv Cardiovasc Dis 3 (2009) 65–73.

Huppertz, B., The anatomy of the normal placenta, J Clin Pathol 61 (2008) 1296–1302.

Iles, R.K.; Chard, T., Human chorionic gonadotrophin expression by bladder cancers: biology and clinical potential, J. Urol. 145 (1991) 453.

Jauniaux, E.; Gulbis, B., Fluid compartments of the embryonic environment, Hum Reprod Update 6 (3) (2000) 268–278.

Jauniaux, E.; Jurkovic, D.; Campbell, S.; et al., Doppler ultrasonographic features of the developing placental circulation: correlation with anatomic findings, Am J Obstet Gynecol 166 (1992) 585.

Jauniaux, E.; Gulbis, B.; Burton, G.J., The human first trimester gestational sac limits rather than facilitates oxygen transfer to the foetus: a review, Placenta 24 (Suppl A) (2003) S86–S93.

Jauniaux, E.; Poston, L.; Burton, G.J., Placental-related diseases of pregnancy: involvement of oxidative stress and implications in human evolution, Hum Reprod Update 12 (2006) 747–755.

Johnson, M.R.; Bolton, V.N.; Riddle, A.F.; et al., Interactions between the embryo and corpus luteum, Hum Reprod 8 (1993) 1496–1501.

Johnson, M.R.; Riddle, A.F.; Irvine, R.; et al., Corpus luteum failure in ectopic pregnancy, Hum Reprod 8 (1993) 1491–1495.

Kaplan, C.G., Gross pathology of the placenta: weight, shape, size, colour, J Clin Pathol 61 (2008) 1285–1295.

Kent, A.; Kitau, M.J.; Chard, T., Absence of diurnal variation in urinary chorionic gonadotrophin excretion at 8–13 weeks gestation, Br J Obstet Gynaecol 98 (1991) 1180.

Kingdom, J.; Sibley, C., The placenta, In: (Editors: Hillier, S.G.; Kitchener, H.C.; Neilson, J.P.) Scientific essentials of reproductive medicine (1996) Saunders, Philadelphia, pp. 312–318.

Kingdom, J.C.P.; Awad, H.; Fleming, J.E.E.; et al., Obstetrical determination of relative placental size, Placenta 14 (1993) A36.

Kingdom, J.C.P.; Sherlock, J.; Rodeck, C.H.; et al., Detection of trophoblast cells in transcervical samples collected by lavage and cytobrush, Obstet Gynecol 86 (1995) 283–288.

Knipp, G.T.; Audus, K.L.; Soares, M.J., Nutrient transport across the placenta, Adv Drug Delivery Rev 38 (1) (1999) 41–58.

Knofler, M., Critical growth factors and signalling pathways controlling human trophoblast invasion, Int J Dev Biol 54 (2010) 269–280.

Lala, P.K.; Chakraborty, C., Factors regulating trophoblast migration and invasiveness, Placenta 24 (1) (2003) 575–587.

Learmont, J.G.; Braude, P.R.; Poston, L., Flow induced dilation is modulated by nitric oxide in isolated human small fetoplacental arteries, J Vasc Res 31 (Suppl 1) (1994) 26.

Meuris, S.; Nagy, A.M.; Delogne-Desnoeck, J.; et al., Temporal relationship between the human chorionic gonadotrophin peak and the establishment of the inter-villus blood flow in early pregnancy, Hum Reprod 10 (1995) 947.

Miller, A.W.F.; Hanretty, K.P., In: Obstetrics illustrated ed 5 (1998) Churchill Livingstone, New York, p. 12, 99.

Miller, J.; Turan, S.; Baschat, A.A., Fetal growth restriction, Semin Perinatol 32 (2008) 274–280.

Moore, K.L.; Persaud, T.V.N., Before we are born: essentials of embryology and birth defects. ed 5 (1998) Saunders, Philadelphia .

Murphy, C.R., Understanding the apical surface markers of uterine receptivity: pinopods-or uterodomes? Hum Reprod 15 (2000) 2451–2454.

Myatt, L.; Brewer, A.S.; Brockman, D.E., The comparative effects of big endothelin-1, endothelin-1, and endothelin-3 in the human fetal–placental circulation, Am J Obstet Gynecol 167 (1992) 1651–1656.

Myatt, L.; Brockman, D.E.; Eis, A.L.W.; et al., Immunohistochemical localization of nitric oxide synthase in the human placenta, Placenta 14 (1993) 487–495.

Ray, J.G.; Laskin, C.A., Folic acid and homocyst(e)ine metabolic defects and the risk of placental abruption, pre-eclampsia and spontaneous pregnancy loss: a systematic review, Placenta 20 (7) (1999) 519–529.

Schindler, A.E., First trimester endocrinology: consequences for diagnosis and treatment of pregnancy failure, Gynecol Endocrinol 18 (1) (2004) 51–57.

Shiverick, K.T.; Salafia, C., Cigarette smoking and pregnancy I: ovarian, uterine and placental effects, Placenta 20 (4) (1999) 265–272.

Spencer, K.; Coombes, J.; Mallard, S.A.; et al., Free beta human chorionic gonadotrophin in Down's syndrome screening: a multicentre study of its role compared to other biochemical markers, Ann Clin Biochem 29 (1992) 506–518.

Stone, S.; Pijnenborg, R.; Vercruysse, L.; et al., The placental bed in pregnancies complicated by primary antiphospholipid syndrome, Placenta 27 (2006) 457–467.

Ticconi, C.; Zicari, A.; Belmonte, A.; et al., Pregnancy-promoting actions of HCG in human myometrium and fetal membranes, Placenta 28 (Suppl A) (2007) S137–S143.

Usadi, R.S.; Murray, M.J.; Bagnell, R.C.; et al., Temporal and morphologic characteristics of pinopod expression across the secretory phase of the endometrial cycle in normally cycling women with proven fertility, Fertil Steril 79(2003) 970–974.

Vitiello, D.; Patrizio, P., Implantation and early embryonic development: implications for pregnancy, Semin Perinatol 31 (2007) 204–207.

Wenstrom, K.D.; Owen, J.; Boots, L.R.; et al., Elevated second trimester human chorionic gonadotropin levels in association with poor pregnancy outcome, Am J Obstet Gynecol 171 (1994) 1038–1041.

Wilcox, G.R.; Trudinger, B.J., Fetal platelet consumption: a feature of placental insufficiency, Obstet Gynecol 77 (1991) 616–621.

Wilcox, A.J.; Baird, D.D.; Weinberg, C.R., Time of implantation of the conceptus and loss of pregnancy, N Engl J Med 340 (1999) 1796–1799.

Wilson, M.E.; Ford, S.P., Comparative aspects of placental efficiency, Reprod Suppl 58 (2001) 223–232.