Preimplantation development
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Fertilization
Chick (days from onset
of incubation)
Mouse (days from time
Incubation
of mating)
Rat (days from time
of mating)
Human (days post ovulation)
Implantation
Primitive
streak
Neural
plate
Pharyngula arches
15–20 somites,
rostral neuropore
closed
Upper
limb bud
Lower
limb bud
Days 2 4 6 8 10 12 14 16 18 20 22 24 26 28 30
Fig . 8 .2 Within developmental biology, evidence concerning the nature of developmental processes has come mainly from studies in vertebrate
embryos, most commonly amniote embryos of the chick, mouse and rat . This chart illustrates the comparative timescale of development in these
animals and in humans .
All cleavage divisions after fertilization are dependent on continuing
30 Stage protein synthesis. In contrast, passage through the earliest cycles, up to
23 eight cells, is independent of mRNA synthesis. Thereafter, experimental
inhibition of transcription blocks further division and development,
indicating that activation of the embryonic genome is required. There
22 is also direct evidence for the synthesis of embryonically encoded pro-
25
teins at this time. As the genes of the embryo first become both active
21 and essential, the previously functional maternally derived mRNA is
destroyed. However, protein made on these maternal templates does
persist at least during blastocyst growth. Spontaneous developmental
20 20
arrest of embryos cultured in vitro seems to occur during the cell cycle
of gene activation, but it is not caused by total failure of that activation
19 process. Early cleavage, up to the formation of eight cells, requires
pyruvate or lactate as metabolic substrates, but thereafter more glucose
15 18 is metabolized and may be required.
The earliest time at which different types of cells can be identified
17 within the cleaving embryo is when 8–16 cells are present. Up to the
formation of eight cells, cells are essentially spherical, touch each other
10 loosely, and have no specialized intercellular junctions or significant
16
extracellular matrix; the cytoplasm in each cell is organized in a radially
15
symmetric manner around a centrally located nucleus. Once eight cells


14 have formed, a process of compaction occurs. Cells flatten on each other
12 13
5 to maximize intercellular contact, initiate the formation of gap and
11 focal tight junctions, and radically reorganize their cytoplasmic confor-
10 mation from a radially symmetric to a highly asymmetric phenotype.
8 9
7 This latter process includes the migration of nuclei towards the centre
6
0 of the embryo, the redistribution of surface microvilli and an underly-
14 21 28 35 42 49 56 ing mesh of microfilaments and microtubules to the exposed surface,
Days post fertilization and the localization of endosomes beneath the apical cytoskeletal
Fig . 8 .3 Human developmental stages 10–23 . The greatest embryonic mesh. As a result of the process of compaction, the embryo forms a
length in mm (ordinate) is plotted against age in postfertilizational primitive protoepithelial cyst, which consists of eight polarized cells, in
weeks (abscissa), with the stages superimposed according to current which the apices face outwards and basolateral surfaces face internally.
information . (Data provided by courtesy of Professor R O’Rahilly . See also The focal tight junctions, which align to become increasingly linear, are
O’Rahilly and Müller (2010) .) localized to the boundary between the apical and basolateral surfaces.
)mm(
eziS


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Outer acrosomal membrane Fusion of plasma membrane
and outer acrosomal membrane Vesiculation Acrosome-reacted sperm
Acrosomal contents Plasma membrane
Inner acrosomal membrane
Nucleus
Head
Neck
Midpiece
Initiation of acrosome reaction
Binding
Cortical granule
Tail
Plasma membrane
Perivitelline space
Continuation of reaction
Zona pellucida
Acrosome-reacted
sperm
Zona reaction Penetration
Excluded sperm Fusion
Cortical reaction Calcium wave starting at sperm entry point
Fig . 8 .4 The fertilization pathway: a succession of steps . After a sperm binds to the zona pellucida, the acrosome reaction takes place (see detail at top) .
The outer acrosomal membrane (blue), an enzyme-rich organelle in the anterior of the sperm head, fuses at many points with the plasma membrane
surrounding the sperm head . Then those fused membranes form vesicles, which are eventually sloughed off from the head, exposing the acrosomal
enzymes (red) . The enzymes digest a path through the zona pellucida, enabling the sperm to advance . Eventually, the sperm meets and fuses with the
secondary oocyte plasma membrane and this triggers cortical and zona reactions . First, enzyme-rich cortical granules in the oocyte cytoplasm release
their contents (yellow) into the zona pellucida, starting at the point of fusion and progressing right and left . Next, in the zona reaction, the enzymes
modify the zona pellucida, transforming it into an impenetrable barrier to sperm as a guard against polyspermy (multiple fertilization) .
elements of compaction. The entire process can function in the absence
of both mRNA and protein synthesis. Post-translational controls are
sufficient and seem to involve regulation through protein phosphoryl-
ation. Significantly, although E-cadherin is not synthesized and present
on the surface of cleaving blastomeres, it first becomes phosphorylated
when eight cells are visible, at the initiation of compaction.
The process of compaction is important for the generation of cell
diversity in the early embryo. As each polarized cell divides, it retains
significant elements of its polar organization, so that its daughter cells


inherit cytocortical domains, the nature of which reflects their origin
and organization in the original parent cell in the eight-celled embryo.
A B Thus, if the axis of division is aligned approximately at right angles to
the axis of cell polarity, the more superficially placed daughter cell
Fig . 8 .5 A, An unfertilized human secondary oocyte surrounded by the inherits all the apical cytocortex and some of the basolateral cytocortex
zona pellucida; the first polar body can be seen . Spermatozoa are visible and is polar, whereas the more centrally placed cell inherits only baso-
outside the zona pellucida . B, A fertilized human ootid before fusion of lateral cytocortex and is apolar. In contrast, if the axis of division is
the pronuclei . Two polar bodies can be seen beneath the zona pellucida . aligned approximately along the axis of the cell polarity, two polar
daughter cells are formed. In this way, two-cell populations are formed
in the 16-cell embryo that differ in phenotype (polar, apolar) and posi-
Gap junctions form between apposed basolateral surfaces and become tion (superficial, deep). The number of cells in each population in any
functional. one embryo will be determined by the ratio of divisions along, and at
The process of compaction involves the cell surface and the calcium right angles to, the axis of eight-cell polarity. The theoretical and
dependent cell–cell adhesion glycoprotein, E-cadherin (also called observed limits of the polar to apolar ratio are 16 : 0 and 8 : 8. The outer
L-CAM or uvomorulin). Neutralization of its function disturbs all three polar cells contribute largely to the trophectoderm, whereas the inner


Preimplantation development
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8
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A B
A B
C D
Fig . 8 .6 Successive stages of cleavage of a human ootid . A, The two-cell
stage . B, The three-cell stage . C, The five-cell stage . D, The eight-cell
stage .
apolar cells contribute almost exclusively to the inner cell mass in most
C
embryos.
In cleavage, the generation of cell diversity, to either trophectoderm
or inner cell mass, occurs in the 16-cell morula and precedes the forma- Fig . 8 .7 Human embryos . Formation of a morula and blastocyst within
tion of the blastocyst. During the 16-cell cycle, the outer polar cells the zona pellucida and blastocyst hatching from the zona pellucida . A, A
continue to differentiate an epithelial phenotype, and display further ball of cells, the morula, with the cells undergoing compaction . B, The
aspects of polarity and intercellular adhesion typical of epithelial cells, blastocyst cavity is developing and the inner cell mass can be seen on
one side of the cavity . C, The blastocyst is beginning to hatch from the
while the inner apolar cells remain symmetrically organized. During
zona pellucida .
the next cell division (16–32 cells), a proportion of polar cells again
divide differentiatively, as in the previous cycle, each yielding one polar
and one apolar progeny, which enter the trophectoderm and inner cell
mass lineages, respectively. Although differentiative division at this time
is less common than at the 8- to 16-cell transition, it has the important
function of regulating an appropriate number of cells in the two tissues
of the blastocyst. Thus, if differentiative divisions were relatively infre-
quent at the 8- to 16-cell transition, they will be more frequent at the
16- to 32-cell transition, and vice versa.
After division to 32 cells, the outer polar cells complete their dif-
ferentiation into a functional epithelium, display structurally complete
zonular tight junctions and begin to form desmosomes. The nascent
trophectoderm engages in vectorial fluid transport in an apical to basal
direction to generate a cavity that expands in size during the 32- to


64-cell cycles and converts the ball of cells, the morula, to a sphere, the
blastocyst (Fig. 8.7). Once the blastocyst forms, the diversification of
the trophectoderm and inner cell mass lineages is complete, and troph-
Fig . 8 .8 A human blastocyst nearly completely hatched from the zona
ectoderm differentiative divisions no longer occur. In the late blastocyst,
pellucida . The blastocyst can now expand to its full size .
the trophectoderm is referred to as the trophoblast, which can be
divided into polar trophoblast, lying in direct contact with the inner
cell mass, and mural trophoblast, surrounding the blastocyst cavity
(Fig. 8.8). development. Even at this early stage, cells of the inner cell mass are
already arranged into an upper layer (i.e. closest to the polar tropho-
Blastocyst blast), the epiblast, which will give rise to the embryonic cells, and a
lower layer, the hypoblast, which has an extraembryonic fate. Thus, the
The blastocyst ‘hatches’ from its zona pellucida at 6–7 days, possibly dorsoventral axis of the developing embryo and a bilaminar arrange-
assisted by an enzyme similar to trypsin (see Figs 8.7C, 8.8). Tropho- ment of the inner cell mass are both established at or before implanta-
blast oozes out of a small slit; many embryos form a figure-of-eight tion. (The earliest primordial germ cells may also be defined at this
shape bisected by the zona pellucida, especially if it has been hardened stage.)
during oocyte maturation and cleavage. Such half-hatching could result
Attachment to the uterine wall
in the formation of identical twins. Hatched blastocysts expand and
differentiation of the inner cell mass proceeds (see Fig. 8.8).
The outer cells of the blastocyst – the trophoblast or trophectoderm On the sixth postovulatory day, the blastocyst adheres to the uterine
– are flattened polyhedral cells, which possess ultrastructural features mucosa and the events leading to the specialized, intimate contact of


typical of a transporting epithelium. The trophoblast covering the inner trophoblast and endometrium begin. Implantation, which is the term
cell mass is the polar trophoblast and that surrounding the blastocyst used for this complicated process, includes the following stages: dissolu-
cavity is the mural trophoblast. The free, unattached blastocyst is tion of the zona pellucida; orientation and adhesion of the blastocyst on
assigned to stage 3 of development at approximately 4 days post ovula- to the endometrium; trophoblastic penetration into the endometrium;
tion, whereas implantation (before villus development) occurs within migration of the blastocyst into the endometrium; and spread and pro-
a period of 7–12 days post ovulation and over the next two stages of liferation of the trophoblast, which envelops and specifically disrupts


PreimPlantation develoPment
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and invades the maternal tissues (Ch. 9). The interactions between the A B
hatched blastocyst and the maternal endometrium are not well under-
stood. Developmentally competent embryos appear to enhance the
uterine environment and promote their own implantation (Brosens
et al 2014, Macklon and Brosens 2014).
The site of implantation is normally in the endometrium of the
posterior wall of the uterus, nearer to the fundus than to the cervix; it
may be in the median plane or to one or other side, but may occur
elsewhere in the uterus, or in an extrauterine or ectopic site.
Ectopic implantation
The conceptus may be arrested at any point during its migration through C D
the uterine tube and implant in its wall. Previous pelvic inflammation
damages the tubal epithelium and may predispose to such delay in
tubal transport. The presence of an intrauterine contraceptive device or
the use of progesterone-based oral contraceptives may also predispose
to ectopic pregnancy, probably because of alteration in the normal tubal
transport mechanisms.
Nidation of the embryo as an ectopic pregnancy most frequently
occurs in the wider ampullary portion of the uterine tube, but may also
occur in the narrow intramural part or even in the ovary itself. Most
ectopic pregnancies are anembryonic, although the continuing growth
of the trophoblast will produce a positive pregnancy test, and may cause
rupture of the uterine tube and significant intraperitoneal haemorrhage. E F
Ectopic pregnancies with a live embryo are the most dangerous because
they grow rapidly and may be detected only when they have eroded the
uterine tube wall and surrounding blood vessels, as early as 8 weeks of
pregnancy. Similarly, cornual ectopics (in the intramural part of the
tube) may present with catastrophic haemorrhage because there is a
substantial blood supply in the surrounding muscularis.
Ovarian or abdominal pregnancies are exceptionally rare. Although
some are presumed to have been caused by fertilization occurring in
the vicinity of the ovary (primary), most are probably caused secondar-
ily and result from an extrusion of the conceptus through the abdomi-


nal ostium of the tube.
Fig . 8 .9 Relationships of the extraembryonic membranes in different
Apart from their important clinical implications, these conditions
types of twinning . A, Diamnionic, dichorionic separated; i .e . separation
emphasize the fact that the conceptus can implant successfully into of the first two blastomeres results in separate implantation sites .
tissues other than a normal progestational endometrium. Prolonged B, Diamnionic, dichorionic fused; here the chorionic membranes are fused
development can occur in such sites and is usually terminated by a but the fetuses occupy separate choria . C, Diamnionic, monochorionic;
mechanical or vascular accident and not by a fundamental nutritive or reduplication of the inner cell mass can result in a single placenta and
endocrine insufficiency or by an immune maternal response. Abdomi- chorionic sacs but separate amniotic cavities . D, Monoamnionic,
nal implantation may occur on any organ, e.g. bowel, liver or omentum. monochorionic; duplication of the embryonic axis results in two embryos
If such a pregnancy continues, this makes removal of the placenta at sharing a single placenta, chorion and amnion . E, Incomplete separation
delivery or abortion hazardous as a result of haemorrhage; conse- of the embryonic axis results in conjoined twins . F, Unequal division of
quently, the placenta is usually left in situ to degenerate spontaneously. the embryonic axis or unequal division of the blood supply may result in
an acardiac monster .
Twinning
Spontaneous twinning occurs once in about every 80 births. Monozy-
single ovum, or both. It is most likely to be seen in women treated with
gotic twins arise from a single ovum fertilized by a single sperm. At
drugs to stimulate ovulation.
some stage up to the establishment of the axis of the embryonic area
The range of separation of twin embryos is reflected in the separation
and the development of the primitive streak, the embryonic cells sepa-
of the extraembryonic membranes. The types of placentation that can
rate into two parts, each of which gives rise to a complete embryo. The
occur are shown in Figure 8.9. Monoamniotic, monochorionic placen-


process of hatching of the blastocyst from the zona pellucida may result
tae are associated with the greatest perinatal mortality (50%), caused
in constriction of the emerging cells and separation into two discrete
both by entanglement of the umbilical cords impeding the blood
entities. There is a gradual decrease in the average thickness of the zona
supply and by various vascular shunts between the placentae, which
pellucida with increasing maternal age, which may be causally related
may divert blood from one fetus to the other. Artery–artery anastomoses
to the increase in frequency of monozygotic twinning with increased
are the most common, followed by artery–vein anastomoses. If the
maternal age. The resultant twins have the same genotype but the
shunting of blood across the placentae from one twin to the other is
description ‘identical twins’ is best avoided, since most monozygotic
balanced by more than one vascular connection, development may
twins have differences in phenotypes. Late separation of twins from a
proceed unimpaired. However, if this is not the case, one twin may
single conceptus may result in conjoined twins; these may be equal or
receive blood from the other, leading to cardiac enlargement, increased
unequal, as in acardia. After twinning, monozygotic embryos enter a
urination and polyhydramnios in the recipient, and anaemia, oligo-
period of intense catch-up growth. Despite starting out at half the size,
hydramnios and atrophy in the donor.
each twin embryo or fetus is of a size comparable to a singleton fetus
Dizygotic twins have either completely separate chorionic sacs or
in the second trimester of pregnancy, but declines in relative size in the
sacs that have fused. Such placentae are separated by four membranes,
last 10 weeks of pregnancy. The sex of monozygotic twins will be the
two amnia and two choria; in addition, these placentae have a ridge of
same. Monoamniotic, monochorionic, monozygotic twins are most
firmer tissue at the base of the dividing membranes, caused by the abut-
likely to be female, as are acardiac twins. The male to female ratio for
ting of two expanding placental tissues against each other.


all monozygotic twins is 0.487, and for monoamniotic, monochorionic
twins it is 0.231.
Dizygotic twins represent the most frequent form of twinning. They
FORMATION OF EXTRAEMBRYONIC TISSUES
result from multiple ovulations, which can be induced by gonado-
trophins or drugs commonly used in patients with infertility. Dizygotic
twins may be different sexes; like-sex pairs are more common. The male The earliest developmental processes in mammalian embryos involve
to female ratio is 0.518. Multiple births greater than twinning, such as the production of those extraembryonic structures that will support and
triplets or quadruplets, can arise from multiple ovulations, or from a nourish the embryo during development. Production of these layers


Formation of extraembryonic tissues
169
8
retPaHC
begins before implantation is complete. At present, it is unclear where mural trophoblast and, rarely, may also share gap junctions. The visceral
the extraembryonic cell lines arise. The trophoblast was considered to hypoblast cells are cuboidal; they have a uniform apical surface towards
be a source but evidence now points to the inner cell mass as the site the blastocyst cavity but irregular basal and lateral regions, with flanges
of origin. Figure 8.1 shows the sequence of development of various and projections underlying one another and extending into intercel-
tissues in the early embryo. lular spaces. There is no basal lamina subjacent to the visceral hypo-
blast, and the distance between the hypoblast cells and the epiblast
Epiblast and amniotic cavity basal lamina is variable.
A series of modifications of the original blastocystic cavity develops
beneath the hypoblast later than those developing above the epiblast.
Epiblast cells are closest to the implanting face of the trophoblast and
While the amniotic cavity is enlarging within the sphere of epiblast
have a definite polarity; they are arranged in a radial manner with
cells, the parietal hypoblast cells are proliferating and spreading along
extensive junctions near the centre of the mass of cells, supported by
the mural trophoblast until they extend most of the way around the
supranuclear organelles. A few epiblast cells are contiguous with
circumference of the blastocyst, converging towards the abembryonic
cytotrophoblast cells; apart from this contact, a basal lamina surrounds
pole. At the same time, a space appears between the parietal hypoblast
what is initially a spherical cluster of epiblast cells, and isolates them
and the mural trophoblast that limits the circumference of the hypo-
from all other cells. Those epiblast cells adjacent to the hypoblast
blastic cavity. A variety of terms have been applied to the parietal
become taller and more columnar than those adjacent to the tropho-
hypoblast layer: extraembryonic hypoblast, extraembryonic endoderm
blast, and this causes the epiblast sphere to become flattened and the


and exocoelomic (Heuser’s) membrane. The cavity that the layer ini-
centre of the sphere to be shifted towards the polar trophoblast. Amni-
tially surrounds is the primary yolk sac (primary umbilical vesicle), and
otic fluid accumulates at the eccentric centre of the now lenticular
the resultant smaller cavity lined by hypoblast is the secondary yolk sac.
epiblast mass, which is bordered by apical junctional complexes and
It has been suggested that the secondary yolk sac forms in a variety of
microvilli. As further fluid accumulates, an amniotic cavity forms,
ways, including cavitation of visceral hypoblast (a method similar to
roofed by low cuboidal cells that possess irregular microvilli. The cells
formation of the amnion), rearrangement of proliferating visceral
share short apical junctional complexes and associated desmosomes,
hypoblast and folding of the parietal layer of the primary yolk sac into
and rest on an underlying basal lamina. The demarcation between true
the secondary yolk sac. Further development of the yolk sac is described
amnion cells and those of the remaining definitive epiblast is clear. The
on page 177.
columnar epiblast cells are arranged as a pseudostratified layer with
The visceral hypoblast cells are thought to be important in many
microvilli, frequently a single cilium, clefted nuclei and large nucleoli;
aspects of the early specification of cell lines and control of the timing
the cells have a distinct, continuous basal lamina. Cell division in the
of early development. They induce the formation and position of the
epiblast tends to occur near the apical surface, causing this region to
primitive streak in the midline of the embryo, thus establishing the first
become more crowded than the basal region. At the margins of the
axis of the embryonic disc, and they influence the timing of epiblast
embryonic disc, the amnion cells are contiguous with the epiblast; there
epithelial-to-mesenchyme transition at the primitive streak. Hypoblast
is a gradation in cell size from columnar to low cuboidal within a two-
cells remain beneath the primitive streak; their experimental removal
to three-cell span (Fig. 8.10; see Fig. 9.1). Further development of the


in avian embryos causes multiple embryonic axes to form. Hypoblast
amnion and amniotic fluid is described on page 178.
cells are also believed to be necessary for successful induction of the
Hypoblast and yolk sac head region and for the successful specification of the primordial germ
cells. With the later formation of the embryonic cell layers from the
epiblast, the visceral hypoblast appears to be sequestered into the sec-
Hypoblast is the term used to delineate the lower layer of cells of the
ondary yolk sac wall by the expansion of the newly formed embryonic
early bilaminar disc, most commonly in avian embryos. This layer is
endoderm beneath the epiblast, although there is now evidence that
also termed anterior, or distal, visceral endoderm in the mouse embryo.
some hypoblast cells may remain in the final endoderm layer and con-
Just before implantation, the hypoblast consists of a layer of squamous
tribute to the gut, and that they may also play a role in the induction
cells that is only slightly larger in extent than the epiblast. The cells
of cardiogenesis. For a review of the roles of hypoblast cells, see Stern
exhibit polarity, in that apical microvilli face the cavity of the blastocyst
and Downs (2012).
and apical junctional complexes, but they lack a basal lamina. During
After the formation of the secondary yolk sac, a diverticulum of
early implantation, the hypoblast extends beyond the edges of the
the visceral hypoblast, the allantois, forms towards one end of the
epiblast and can now be subdivided into those cells in contact with the
embryonic region and extends into the local extraembryonic mesoblast.
epiblast basal lamina, the visceral hypoblast, and those cells in contact
It passes from the roof of the secondary yolk sac to the same plane
with the mural trophoblast, the parietal hypoblast. The parietal hypo-
as the amnion. Further development of the allantois is described on
blast cells are squamous; they may share adhesion junctions with the
page 178.
Extraembryonic mesoblast
By definition, extraembryonic tissues encompass all tissues that do not
contribute directly to the future body of the definitive embryo and, later,


the fetus. At stage 5, blastocysts are implanted but do not yet display
trophoblastic villi (see Fig. 8.10); they range from 7 to 12 days in age.
A feature of this stage is the first formation of extraembryonic meso-
blast, which will come to cover the amnion, secondary yolk sac and the
internal wall of the mural trophoblast, and will form the connecting
stalk of the embryo with its contained allantoenteric diverticulum. The
origin of this first mesoblastic extraembryonic layer is by no means
clear; it may arise from several sources, including the caudal region of
the epiblast, the parietal hypoblast and subhypoblastic cells. The tro-
phoblastic origin of extraembryonic mesoblast is questioned because
there is always a complete basal lamina underlying the trophoblast; the
migration of cells out of an epithelium is usually associated with previ-
ous disruption of the basal lamina. Certainly, the origin of extraembry-
onic cells will change over time as new germinal populations are
established.
Hypoblast
The first mesoblastic extraembryonic layer gives rise to the layer
Cytotrophoblast
known as extraembryonic mesoblast, arranged as a mesothelium with
Syncytiotrophoblast underlying extraembryonic mesenchymal cells; this also appears to
Fig . 8 .10 The implanting conceptus at stage 5b . The embryo at this stage form an extracellular structure corresponding to the magma reticulare,
is composed only of the abutting epiblast and hypoblast layers . It is between the mural trophoblast and the primary yolk sac in the stage 5
suspended within the blastocyst cavity (chorionic cavity) and surrounded embryo. Later extraembryonic mesoblast populations mushroom
by a layer of cytotrophoblast . A large mass of syncytiotrophoblast, with beneath the cytotrophoblastic cells at the embryonic pole, forming
interconnecting lacunae, is penetrating the maternal endometrium . the cores of the developing villus stems, and villi and the angioblastic


PreimPlantation develoPment
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cells that will give rise to the capillaries within them and the earliest
Bonus e-book video
blood cells.
Initially, the extraembryonic mesoblast connects the amnion to the
chorion over a wide area. Continued development and expansion of
the extraembryonic coelom means that this attachment becomes Video 8 .1 Human in vitro fertilization and early development .
increasingly circumvented to a connecting stalk, which is a permanent
connection between the future caudal end of the embryonic disc and
the chorion. The connecting stalk forms a pathway along which vascular
anastomoses around the allantois establish communication with those
of the chorion.
KEY REFERENCES
Brosens JJ, Walker MS, Teklenburg G 2014 Uterine selection of human O’Rahilly R, Müller F 2010 Developmental stages in human embryos: revised
embryos at implantation. Sci Reports 4:3894. and new measurements. Cells Tissues Organs 192:73–84.
Macklon NS Brosens JJ 2014 The human endometrium as a sensor of Stern CD, Downs KM 2012 The hypoblast (visceral endoderm): an evo-devo
embryo quality. Biol Reprod 91:98. perspective. Development 139:1059–69.
O’Rahilly R, Müller F 1987 Developmental Stages in Human Embryos. Streeter GL 1942 Developmental horizons in human embryos. Descriptions
Washington: Carnegie Institution. of age group XI, 13 to 20 somites, and age group XII, 21 to 29 somites.
O’Rahilly R, Müller F 1999 The Embryonic Human Brain. An Atlas of Contrib Embryol Carnegie Inst Washington 30:211–45.
Developmental Stages, 2nd ed. New York: Wiley-Liss.
O’Rahilly R, Müller F 2000 Minireview: Prenatal ages and stages – measures
and errors. Teratology 61:382–4.


Preimplantation development
170.e1
8
retPaHC
REFERENCES
Brosens JJ, Walker MS, Teklenburg G 2014 Uterine selection of human O’Rahilly R, Müller F 2010 Developmental stages in human embryos: revised
embryos at implantation. Sci Reports 4:3894. and new measurements. Cells Tissues Organs 192:73–84.
Macklon NS Brosens JJ 2014 The human endometrium as a sensor of Stern CD, Downs KM 2012 The hypoblast (visceral endoderm): an evo-devo
embryo quality. Biol Reprod 91:98. perspective. Development 139:1059–69.
O’Rahilly R, Müller F 1987 Developmental Stages in Human Embryos. Streeter GL 1942 Developmental horizons in human embryos. Descriptions
Washington: Carnegie Institution. of age group XI, 13 to 20 somites, and age group XII, 21 to 29 somites.
O’Rahilly R, Müller F 1999 The Embryonic Human Brain. An Atlas of Contrib Embryol Carnegie Inst Washington 30:211–45.
Developmental Stages, 2nd ed. New York: Wiley-Liss.
O’Rahilly R, Müller F 2000 Minireview: Prenatal ages and stages – measures
and errors. Teratology 61:382–4.


CHAPTER
9
Implantation and placentation
and during leukocyte emigration from the blood into the tissues.
IMPLANTATION
Flanges of syncytial trophoblast grow between the cells of the uterine
luminal epithelium towards the underlying basal lamina without
Implantation involves the initial attachment of the trophoblastic wall apparent damage to the maternal cell membranes or disruption of the
of the blastocyst to the endometrial luminal epithelium and its decidual intercellular junctions. Instead, shared junctions, including tight junc
response. It is now apparent that, during this process, there is an inter tions, are formed with many of the maternal uterine epithelial cells.
active dialogue between the implanting embryo and endometrial Implantation continues with erosion of maternal vascular endothe
decidu al stromal cells. Competent preimplantation human embryos lium and glandular epithelium, and phagocytosis of secretory products,
actively enhance the uterine environment for their implantation, until the blastocyst occupies an uneven implantation cavity in the
whereas developmentally impaired embryos induce endoplasmic retic stroma (interstitial implantation) (Fig. 9.1). In the early postimplant
ulum stress responses in decidual cells that inhibit implantation ation phase, the maternal surface is resealed by reepithelialization and
(Brosens et al 2014). the formation of a plug, which may contain fibrin. As the blastocyst
The blastocyst/trophoblast lineage gives rise to three main cell types burrows more deeply into the endometrium, syncytial trophoblast
in the human placenta: syncytiotrophoblast cells form the epithelial forms over the mural cytotrophoblast, but never achieves the thickness
covering of the villous tree and are the main endocrine component of of the syncytial trophoblast over the embryonic pole.
the placenta; villous cytotrophoblast cells represent a germinative popu The syncytiotrophoblast, which expresses neither class I nor class II
lation that proliferates throughout pregnancy, fusing to generate syn major histocompatibility complex (MHC) antigens, secretes numerous


cytiotrophoblast; and extravillous trophoblast cells are nonproliferative hormones; human chorionic gonadotrophin (hCG) can be detected in
and invade the maternal endometrium. The first two cell lines can be maternal urine from as early as 10 days after fertilization and forms the
seen from stages 4 and 5 onwards. The cytotrophoblast cells that form basis for tests for early pregnancy. This hCG prolongs the life of the
the mural and polar trophoblast are cuboidal and covered externally corpus luteum, which continues to secrete progesterone and oestrogens
with syncytial trophoblast (syncytiotrophoblast), a multinucleated during approximately the first 2 months of pregnancy, until these essen
mass of cytoplasm that forms initially in areas near the inner cell mass tial hormones are produced by the placenta.
after apposition of the blastocyst to the uterine mucosa (see Fig. 8.10). Menstruation ceases on successful implantation. The endometrium,
Preimplantation embryos produce matrix metalloproteinases now known as the decidua in pregnancy, thickens to form a suitable
(MMPs) that mediate penetration of the maternal subepithelial basal nidus for the conceptus. Decidualization of the endometrial stroma
lamina by the syncytiotrophoblast. Trophoblast cells express Lselectin may occur without an intrauterine pregnancy, e.g. in the presence of an
(usually seen as a mediator of neutrophil rolling and tethering in ectopic pregnancy, after prolonged treatment with progesterone, and in
inflamed endothelium), and the maternal epithelium upregulates the late secretory phase of a nonconception cycle.
selectinoligosaccharidebased ligands. Thus, differentiating cytotro Decidual differentiation is not evident in the stroma at the earliest
phoblast cells appear to use processes that also occur in vasculogenesis stages of implantation, and it may not be until a week later that fully
Uterine glands
Uterine blood vessel
Villous stems
Amniotic cavity
Amnion
Secondary yolk sac
Yolk sac wall
Lacuna
Extraembryonic coelom
(chorionic cavity) Chorion
Epithelium of
Decidua
endometrial surface


Extraembryonic mesoblast
Cytotrophoblast
Syncytiotrophoblast
Coagulum over
implanting blastocyst
Fig. 9.1 The implanting conceptus at stage 6. The embryo is composed of epiblast, with the amniotic cavity above it, and hypoblast, with the secondary
yolk sac below it (see also Figs 8.10, 10.1). Both cavities are covered externally with extraembryonic mesoblast, which also lines the larger chorionic
cavity. Primary villi cover the outer aspect of the conceptus and extend into the maternal endometrium, and in places, maternal blood fills the lacunae. 171


ImplantatIon and placentatIon
172
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noItceS
differentiated cells are present. During decidualization, the interglan As the blastocyst implants, the syncytiotrophoblast invades the
dular tissue increases in quantity (Burton et al 2010). It contains a uterine tissues, including the glands and walls of maternal blood vessels
substantial population of leukocytes (large granular lymphocytes, (see Fig. 9.1; Fig. 9.3), and increases rapidly in thickness over the
macrophages and T cells) distributed amongst large decidual cells. The embryonic pole (Fig. 9.4). A progressively thinner layer covers the rest
most numerous are uterine natural killer (NK) cells, which accumulate of the wall towards the abembryonic pole. Microvilluslined clefts and
in the endometrium during the secretory phase of the cycle and persist lacunar spaces develop in the syncytiotrophoblastic envelope (days
until midpregnancy; they interact with invading extravillous tropho 9–11 of pregnancy) and establish communications with one another.
blast cells expressing human leukocyte antigen G and C (HLAG, Initially, many of these spaces contain maternal blood derived from
HLAC). Decidual cells are stromal cells that contain varying amounts dilated uterine capillaries and veins, as the walls of the vessels are par
of glycogen, lipid and vimentintype intermediate filaments in their tially destroyed. As the conceptus grows, the lacunar spaces enlarge, and
cytoplasm. They are generally rounded but their shape may vary, become confluent to form intervillous spaces.
depending on the local packing density. They may contain one, two or The projections of syncytiotrophoblast into the maternal decidua are
sometimes three nuclei, frequently display rows of clublike cytoplas called primary villi. They are invaded initially with cytotrophoblast and
mic protrusions enclosing granules, and are associated with a charac
teristic capsular basal lamina. Decidual cells produce a range of secretory
products, including insulinlike growth factor binding protein 1 (IGF
Decidua basalis
BP1) and prolactin, which may be taken up by the trophoblast. These
Intervillous space


secretions probably play a role in the maintenance and growth of the
conceptus in the early part of postimplantational development, and can
be detected in amniotic fluid in the first trimester of pregnancy. Uterine
Extracellular matrix, growth factors and protease inhibitors pro tube
duced by the decidua all probably modulate the degradative activity of Villous
the trophoblast and support placental morphogenesis and placental Connecting stems
accession to the maternal blood supply. Once implantation is complete, stalk of chorion
distinctive names are applied to different regions of the decidua (Fig. frondosum
9.2). The part covering the conceptus is the decidua capsularis; that Umbilical
between the conceptus and the uterine muscular wall is the decidua cord Yolk sac
basalis (where the placenta subsequently develops); and that which
Decidua Vitelline
lines the remainder of the body of the uterus is the decidua parietalis. parietalis duct
There is no evidence that their respective resident maternal cell popula
tions exhibit sitespecific properties. Chorion Amniotic
cavity
Chorionic
DEVELOPMENT OF THE PLACENTA cavity Embryo
within
Decidua amnion
Formation of the human placenta requires a developmental progression
capsularis
that proceeds in a specific chronological order: the development of
primary, secondary and anchoring villi, and the local differentiation of
haemangioblast cells within them, occur concurrently with the modifi Myometrium Plug of mucus
cation of maternal blood vessels to ensure their patency. With the onset Fig. 9.2 The gravid uterus in the second month. A placental site precisely
of the embryonic heart beat, a primitive circulation exists between the in the uterine fundus, as indicated on the figure, is rather unusual; the
embryo and the secondary yolk sac. An embryonic–placental circula dorsal, ventral or lateral wall of the corpus uteri is more usual. The
tion starts around week 8 of gestation; over the course of 40 weeks, the maternal endometrium is now termed decidua; different regions are
placenta develops into a highly vascular organ. distinguished.
Syncytiotrophoblast Lacunae Syncytiotrophoblast Endometrial gland


Cytotrophoblast Endometrial gland Cytotrophoblast Spiral artery
Mesenchyme Spiral artery Interstitial extravillous trophoblast Decidua
Amnion Decidua Endovascular extravillous trophoblast
Glandular secretions in intervillous space
A B Plasma in intervillous space
Fig. 9.3 A, The growing syncytiotrophoblast erodes decidual glands and spiral arteries, and their contents form lacunae within the syncytiotrophoblast.
Cytotrophoblastic cells extend as villi into the syncytiotrophoblast. B, Cells originating from the cytotrophoblast move into the decidua, forming interstitial
extravillous trophoblast cells, the third line of cytotrophoblastic cells. These surround the opened glands and spiral arteries in the decidua and inner
myometrium. Cells that move through the tunicae adventitia, media and interna to enter the lumen of the vessels become endovascular extravillous
trophoblast. These cells remodel the walls and plug the lumen of the spiral arteries, permitting only plasma to enter the forming intervillous space.
Glandular secretions in the intervillous space provide histiotrophic nutrition to the embryo.


development of the placenta
173
9
RetpaHc
then with extraembryonic mesenchyme (days 13–15) to form second maternal blood, along the spiral arteries as far as the inner myometrial
ary placental villi. Capillaries develop in the mesenchymal core of the region (Huppertz et al 2014).
villi during the third week post conception. The cytotrophoblast within The definitive placenta is composed of a chorionic plate on its fetal
the villi continues to grow through the invading syncytiotrophoblast aspect and a basal plate on its maternal aspect, separated by an interven
and makes direct contact with the decidua basalis, forming anchoring ing intervillous space containing villous stems with branches in contact
villi. Further cytotrophoblast proliferation occurs laterally so that neigh with maternal blood (see Fig. 9.5). During the first trimester, develop
bouring outgrowths meet to form a spherical cytotrophoblastic shell ment takes place in a lowoxygen environment supported by histio
around the conceptus (Fig. 9.5; see Fig. 9.3B). Lateral projections from trophic nutrition from the endometrial glands, which discharge into
the main stem villus form true villi. Elongation of growing capillaries the intervillous space until at least 10 weeks (see Fig. 9.3A) (Burton et al
outstrips that of the containing villi, leading to looping of vessels. This 2002, Burton et al 2007, Burton and Fowden 2012). When maternal
obtrusion of both capillary loops and new sprouts results in the forma blood bathes the surfaces of the chorion that bound the intervillous
tion of terminal villi. space, the human placenta is defined as haemochorial. Different grades
As secondary villi form, single mononuclear cells become detached of fusion exist between the maternal and fetal tissues in many other
from the distal (anchoring) cytotrophoblast and infiltrate the maternal mammals (e.g. epitheliochorial, syndesmochorial, endotheliochorial).


decidua (Fig. 9.6; see Fig. 9.3B). These extravillous cells are the third The chorion is vascularized by the allantoic blood vessels of the body
line of the original trophoblastic cells. Interstitial extravillous trophob stalk, and so the human placenta is also termed chorioallantoic
last cells invade the maternal spiral arteries from their adventitia. The (whereas, in some mammals, a choriovitelline placenta either exists
smooth muscle and internal elastic lamina are replaced with extracel alone or supplements the chorioallantoic variety). The human placenta
lular fibrinoid deposits (Harris 2010). Those cells that pass through the is also defined as discoidal, in contrast to other shapes in other
endothelial basement membrane and gain access to the vessel lumen mammals, and deciduate because maternal tissue is shed with the pla
are termed endovascular extravillous trophoblast (see Figs 9.3B, 9.6). centa and membranes at parturition as part of the afterbirth.
These cells plug the maternal spiral arteries until the end of the first
trimester; the cells also migrate antidromically, against the flow of Growth of the placenta
Expansion of the entire conceptus is accompanied by radial growth of
the villi and, simultaneously, an integrated tangential growth and
Fig. 9.4 The conceptus
expansion of the trophoblastic shell. Eventually, each villous stem
at about stage 14. The
forms a complex that consists of a single trunk attached by its base to
embryonic pole shows
the chorion, from which second and thirdorder branches (intermedi
extensive villous
ate and terminal villi) arise distally. Terminal villi are specialized for
formation at the chorion
frondosum, whereas exchange between fetal and maternal circulations; each one starts as a
the abembryonic pole syncytial outgrowth and is invaded by cytotrophoblastic cells, which
is smooth and then develop a core of fetal mesenchyme as the villus continues to grow.
villous-free at the The core is vascularized by fetal capillaries (i.e. each villus passes
chorion laeve. through primary, secondary and tertiary grades of histological differen


(Photograph courtesy tiation). The germinal cytotrophoblast continues to add cells that fuse
of P Collins.) with the overlying syncytium and so contribute to the expansion of the
haemochorial interface. Terminal villi continue to form and branch
within the confines of the definitive placenta throughout gestation,
projecting in all directions into the intervillous space (see Fig. 9.5).
From the third week until about the second month of pregnancy,
the entire chorion is covered with villous stems. They are thus continu
ous peripherally with the trophoblastic shell, which is in close appos
ition with both the decidua capsularis and the decidua basalis. The villi
adjacent to the decidua basalis are stouter and longer, and show a
greater profusion of terminal villi. As the conceptus continues to
expand, the decidua capsularis is progressively compressed and thinned,
the circulation through it is gradually reduced, and adjacent villi slowly
atrophy and disappear. This process starts at the abembryonic pole; by
the end of the third month, the abembryonic hemisphere of the
Cotyledonary septum
DEVELOPMENT
Syncytiotrophoblast Basal plate
Lacunae Cytotrophoblastic
enlarging as cell column
intervillous
spaces
Lacunae in syncytium Lacunar
circulation
Maternal vessels
in decidua
Intervillous
space
Cytotrophoblast
Fetal vessels Stem villus
Chorionic
Maternal blood True villus Secondary syncytial fusion
plate
Ingrowth of cytotrophoblast and Terminal villus Connective tissue
mesoblast bearing fetal vessels Amnion
Fig. 9.5 Placental development.


ImplantatIon and placentatIon
174
2
noItceS
Myometrium
Spiral artery
Uterine gland
Decidua
Cotyledonary septum
Syncytiotrophoblast
Cytotrophoblast
Interstitial extravillous trophoblast
Endovascular extravillous trophoblast
Glandular secretions in intervillous space
Plasma in intervillous space
Fig. 9.6 As the placenta grows, the interstitial and endovascular extravillous trophoblast cells continue to remodel the spiral arteries into large-bore,
low-resistance uteroplacental vessels. These arteries remain occluded by endovascular extravillous trophoblast until the end of the first trimester,
promoting a low oxygen environment.
conceptus is largely denuded. Eventually, the whole chorion apposed Chorionic plate
to the decidua capsularis is smooth and is now termed the chorion
The chorionic plate is covered on its fetal aspect by amniotic epithe
laeve; this will later become apposed to the amnion, forming chorio
lium; on the stromal side of the epithelium, a connective tissue layer
amnion (see Figs 9.2, 9.9). In contrast, the villous stems of the disc
carries the main branches of the umbilical vessels (see Fig. 9.5; Fig. 9.7).
shaped region of chorion apposed to the decidua basalis increase
Subjacent to this are a diminishing layer of cytotrophoblast and then
greatly in size and complexity, and the region is now termed the chorion
the inner syncytial wall of the intervillous space. The connective tissue
frondosum (see Fig. 9.4). The chorion frondosum and the decidua
layer is formed by fusion between the mesenchymecovered surfaces of
basalis constitute the definitive placental site (see Fig. 9.2). Abnormali
amnion and chorion, and is more fibrous and less cellular than Whar
ties in this process may account for the persistence of villi at abnormal
ton’s jelly (of the umbilical cord), except near the larger vessels. The
sites in the chorion laeve of the gestational sac and hence the presence


umbilical vessels radiate and branch from the cord attachment, with
of accessory or succenturiate lobes within the membranes of the defini
variations in the branching pattern, until they reach the bases of the
tive placenta. The presence of the entire villous ring of the primitive
trunks of the villous stems; they then arborize within the intermediate
placenta beyond the second month postmenstruation leads to the
and terminal villi. There are no anastomoses between vascular trees of
development of placenta membranacea, a giant definitive placental
adjacent stems. The two umbilical arteries are normally joined at, or
structure surrounding the whole gestational sac during the remainder
just before they enter, the chorionic plate, by some form of substantial
of pregnancy.
transverse anastomosis (Hyrtl’s anastomosis).
Coincidentally with the growth of the embryo and the expansion of
the amnion, the decidua capsularis is thinned and distended, and the
Basal plate
space between it and the decidua parietalis is gradually obliterated. By
the second month of pregnancy, the three endometrial strata recogniz The basal plate, from fetal to maternal aspect, forms the outer wall of
able in the premenstrual phase, i.e. compactum, spongiosum and the intervillous space. The trophoblast and adjacent decidua are
basale, are better differentiated and easily distinguished. The glands in enmeshed in layers of fibrinoid and basement membranelike extracel
the spongiosum are compressed and appear as oblique slitlike fissures lular matrix to form a complex junctional zone. In different places, the
lined by low cuboidal cells. By the beginning of the third month of basal plate may contain syncytium, cytotrophoblast or fibrinoid matrix,
pregnancy, the decidua capsularis and decidua parietalis are in contact. remnants of the cytotrophoblastic shell, and, at the site of implantation,
By the fifth month, the decidua capsularis is greatly thinned, and it areas of necrotic maternal decidua (the socalled Nitabuch’s stria) (see


virtually disappears during the succeeding months. Figs 9.5 and 9.7). Nitabuch’s stria and the decidua basalis contain
Focal bleeding often occurs in the periphery of the developing pla cytotrophoblast and multinucleate trophoblast giant cells derived from
centa at the time of the formation of the membranes (8–12 weeks). This the mononuclear extravillous interstitial cytotrophoblast population
complication, which is termed threatened miscarriage, is a common that infiltrate the decidua basalis during the first 18 weeks of pregnancy.
clinical complication of pregnancy and can lead to a complete miscar These cells penetrate as far as the inner third of the myometrium but
riage if the haematoma extends to the definitive placenta (Jauniaux et al can often be observed at or near the decidual–myometrial junction.
2006). They are not found in the decidua parietalis or the adjacent myo
At term, the placental diameter varies from 200 to 220 mm, the metrium, suggesting that the placental bed giant cell represents a dif
mean placental weight is 470 g, its mean thickness is 25 mm and the ferentiative end stage in the extravillous trophoblast lineage. The striae
total villous surface area is 12–14 m2, providing an extensive and inti of fibrinoid are irregularly interconnected and variable in prominence.
mate interface for materno–fetal exchange; the uteroplacental circula Strands pass from Nitabuch’s stria into the adjacent decidua, which
tion carries approximately 600 ml of maternal blood per minute. There contains basal remnants of the endometrial glands and large and small
are no lymph vessels in the placenta and, equally, no aggregates of decidual cells scattered in a connective tissue framework that supports
lymphoid cells. Systematic evaluation and careful description of the an extensive venous plexus.
placenta after delivery have been recommended for correlation with Initially, only the central area of a placenta contains extravillous
later neonatal neurodevelopmental outcomes (Roescher et al 2014). trophoblast cells, both within and around the spiral arteries. These cells


development of the placenta
175
9
RetpaHc
Myometrium
Maternal blood vessels
Basal plate
Cytotrophoblastic cell column Fibrinoid deposit
Cytotrophoblast
Syncytiotrophoblast
Anchoring villus
Orifices of maternal vessels
Syncytial sprout
Terminal villi
Syncytial fusion
Intervillous space
Hofbauer cell
Intermediate villus
Stem villus
Syncytiotrophoblast
Cytotrophoblast
Fetal mesenchyme Fetal blood vessels Chorionic plate
Amniotic epithelium
Fig. 9.7 The arrangement of the placental tissues from the chorionic plate (fetal side) to the basal plate or decidua basalis (maternal side).
gradually extend laterally, reaching the periphery of the placenta around feature occupying the peripheral margin of the placenta and commu
midgestation; the extent of invasion is progressively shallower towards nicating freely with the intervillous space, has not been confirmed.
the periphery. Invaded maternal vessels show a 5–10fold dilation of Recent anatomic and in vivo studies have shown that human placen
the vessel mouth and altered responsiveness to circulating vasoactive tation is, in fact, not truly haemochorial in early pregnancy (Video 9.1)
compounds (Hung et al 2001, Burton et al 2009). (Jauniaux et al 2003). From the time of implantation, the extravillous
The conversion of maternal musculoelastic, spiral arteries to large trophoblast not only invades the uterine tissues but also forms a con
bore, lowresistance uteroplacental vessels is considered key for a suc tinuous shell at the level of the decidua. The cells of this shell anchor
cessful human pregnancy. Failure to achieve these changes is a feature the placenta to the maternal tissue and also form plugs in the tips of
of common complications of pregnancy, such as earlyonset pre the uteroplacental arteries (see Figs 9.3B, 9.6) (Burton et al 1999). The
eclampsia, miscarriage and fetal growth restriction. shell and the plugs act like a labyrinthine interface that filters maternal


From the third month onwards, the basal plate develops placental blood, permitting a slow seepage of plasma but no true blood flow into
or cotyledonary septa, which are ingrowths of the cytotrophoblast the intervillous space (Burton et al 2003). This creates a physiological
covered with syncytium that grow towards, but do not fuse with, the placental hypoxia, which may protect the developing embryo against
chorionic plate (see Fig. 9.5). The septa circumscribe the maternal the deleterious and teratogenic effects of oxygen free radicals. As a
surface of the placenta into 15–30 lobes, often termed cotyledons. Each result, there is a uteroplacental O gradient that exerts a regulatory effect
2
cotyledon surrounds a limited portion of the intervillous space associ on placental tissue development and function (Jauniaux et al 2003). In
ated with a villous trunk from the chorionic plate. From the fourth particular, it influences cytotrophoblast proliferation and differenti
month, these septa are supported by tissue from the decidua basalis. ation along the invasive pathway, villous vasculogenesis and the form
Throughout the second half of pregnancy, the basal plate becomes ation of the chorion laeve. An O gradient persists within lobules of the
2
thinned and progressively modified: there is a relative diminution of placenta throughout pregnancy, with the central region well oxygenated
the decidual elements, increasing deposition of fibrinoid, and admix compared to the periphery, owing to the direction of maternal blood
ture of fetal and maternal derivatives. flow (Burton and Jauniaux 2011).
At the end of the first trimester, the endovascular extravillous tro
Intervillous space phoblastic plugs are progressively dislocated, allowing maternal blood
The intervillous space contains the main trunks of the villous stems and to flow progressively more freely and continuously within the intervil
their arborizations into intermediate and terminal villi (see Figs 9.5, lous space (see Figs 9.6, 9.7, Video 9.1). During the transitional phase


9.7). A villous trunk and its branches may be regarded as the essential of 10–14 weeks’ gestation, twothirds of the primitive placenta disap
structural, functional and growth unit of the developing placenta. pears, the chorionic cavity is obliterated by the growth of the amniotic
At term, from the inner myometrium to the intervillous space, the sac, and maternal blood flows progressively throughout the entire pla
walls of most spiral arteries consist of fibrinoid matrix, within which centa (Jauniaux et al 2003). These events bring the maternal blood
cytotrophoblast is embedded. This arrangement allows expansion of closer to the fetal tissues, facilitating nutrient and gaseous exchange
the arterial diameter (and so slows the rate of arterial inflow and reduces between the maternal and fetal circulations (GutierrezMarcos et al
the perfusion pressure), independent of the local action of vasoconstric 2012).
tive agents. Endothelial cells, where present, are often hypertrophic. The
veins that drain the blood away from the intervillous space pierce the Structure of a placental villus
basal plate and join tributaries of the uterine veins. The presence of a Chorionic villi are the essential structures involved in exchanges
marginal venous sinus, which hitherto has been described as a constant between mother and fetus. The villous tissues separating fetal and


ImplantatIon and placentatIon
176
2
noItceS
A B C
Anchoring villus
Arterio–capillary– Endothelium of Fetal blood
venous network fetal capillary
Placental
Connective membrane
tissue core
Syncytiotrophoblast
Syncytiotrophoblast
Fetal capillary
Cytotrophoblast
Hofbauer cell
Fibrinoid material
Fetal capillaries
Persisting cytotrophoblast cells
Nuclear aggregation or syncytial knot
Vein
Artery
Stem villus
Fig. 9.8 A, A chorionic villus and its arterio–capillary–venous system carrying fetal blood. The artery carries deoxygenated blood and waste products
from the fetus, and the vein carries oxygenated blood and nutrients to the fetus. B–C, Transverse sections through individual chorionic villi at 10 weeks
(B) and at full term (C). The villi would be within the intervillous space, bathed externally in maternal blood. The placental membrane, composed of fetal
tissues, separates the maternal blood from the fetal blood.
maternal blood are therefore of crucial functional importance. From the plasm is more strongly basophilic than that of the cytotrophoblastic
chorionic plate, progressive branching occurs into the villous tree, as cells and is packed with organelles consistent with its secretory pheno
stem villi give way to intermediate and terminal villi. Each villus has a type. Where the plasma membrane adjoins basal lamina, it is often
core of connective tissue containing collagen types I, III, V and VI, as infolded into the cytoplasm, whereas the surface bordering the intervil
well as fibronectin. Crossbanded fibres (30–35 nm) of type I collagen lous space is set with numerous long microvilli, which constitute the
often occur in bundles, whereas type III collagen is present as thinner brush border seen by light microscopy.
(10–15 nm) beaded fibres, which form a meshwork that often encases Glycogen is thought to be present in both layers of the trophoblast
the larger fibres. Collagens V and VI are present as 6–10 nm fibres at all stages, although it is not always possible to demonstrate its pres


closely associated with collagens I and III. Laminin and collagen type ence histochemically. Lipid droplets occur in both layers and are free
IV are present in the stroma associated with the basal laminae that in the core of the villus. In the trophoblast, they are found principally
surround fetal vessels and also in the trophoblast basal lamina. Overly within the cytoplasm, but they also occur in the extracellular space
ing this matrix are ensheathing cyto and syncytiotrophoblast cells between cytotrophoblast and syncytium, and between the individual
bathed by the maternal blood in the intervillous space (see Figs 9.5, cells of the cytotrophoblast. The droplets diminish in number with
9.7; Fig. 9.8). Cohesion between the cells of the cytotrophoblast, and advancing age and may represent fat in transit from mother to fetus,
also between the cytotrophoblast and the syncytium, is provided by and/or a pool of precursors for steroid synthesis. Membranebound
numerous desmosomes between the apposed plasma membranes. granular bodies of moderate electron density occur in the cytoplasm,
In earlier stages, the cytotrophoblast forms an almost continuous particularly in the syncytium, some of which are probably secretion
layer on the basal lamina. After the fourth month, it gradually expends granules. Other membranebound bodies, lysosomes and phagosomes,
itself producing syncytium, which comes to lie on the basal lamina over are involved in the degradation of materials engulfed from the intervil
an increasingly large area (56% at term), and becomes progressively lous space.
thinner. Cytotrophoblastic cells persist until term but, because the Although many nuclei are dispersed within the syncytioplasm, other
increase in villous surface area outstrips their proliferation, they are are aggregated into specializations referred to as syncytial knots and
usually disposed singly. In the first and second trimesters, cytotropho syncytial sprouts (Burton et al 2003). Knots must be distinguished from
blastic sprouts, covered in syncytium, are present and represent a stage syncytial sprouts, which are markers of trophoblast proliferation. In the


in the development of new villi. Cytotrophoblast columns at the tips immature placenta, syncytial sprouts represent the first stages in the
of anchoring villi extend from the villous basal lamina to the maternal development of new terminal villi, which later become invaded by
decidual stroma. cytotrophoblast and villous mesenchyme (Fogarty et al 2013). Occa
The cells of the villous cytotrophoblast (Langhans cells) are pale sionally, adjacent syncytial sprouts make contact and fuse to form
staining with a slight basophilia. Ultrastructurally, they have a rather slender syncytial bridges. The sprouts may become detached, forming
electrontranslucent cytoplasm and relatively few organelles. They maternal syncytial emboli, which pass to the lungs. It has been com
contain intermediate filaments, particularly in association with desmo puted that some 100,000 sprouts pass daily into the maternal circula
somes. Between the desmosomes, the membranes of adjacent cells are tion; deported trophoblast may be a mechanism by which the maternal
separated by approximately 20 nm. Sometimes, the intercellular gap immune system is maintained in a state of tolerance towards paternal
widens to accommodate microvillous projections from the cell surfaces; antigens (Chamley et al 2011). In the lungs, syncytial sprouts provoke
occasionally, it contains patches of fibrinoid. A smaller population of little local reaction and apparently disappear by lysis. However, they
intermediate cytotrophoblast may also be found in the chorionic villi. may occasionally form foci for neoplastic growth. Syncytial sprouts are
This postmitotic population represents a state of partial differentiation present in the term placenta. Syncytial knots are aggregates of nuclei
between the cytotrophoblast stem cell and the overlying syncytium. with particularly condensed heterochromatin, and may represent a
The syncytiotrophoblast is the multinucleated epithelium of the sequestration phenomenon by which senescent nuclear material is
placenta and is an intensely active tissue layer, across which most trans removed from adjacent metabolically active areas of syncytium.
placental transport must occur (Ellery et al 2009). It is a selectively Fibrinoid deposits are frequently found on the villous surface in


permeable barrier that allows water, oxygen and other nutritive sub areas lacking syncytiotrophoblast. They may constitute a repair mecha
stances and hormones to pass from mother to fetus, and some of the nism in which the fibrinoid forms a wound surface that is subsequently
products of excretion to pass from fetus to mother. It secretes a range reepithelialized by trophoblast. The extracellular matrix glycoprotein
of placental hormones into the maternal circulation. Syncytial cyto tenascin has been localized in the stroma adjacent to these sites.


Fetal membranes
177
9
RetpaHc
The core of a villus contains small and large reticulum cells, fibro acute phase of a newly acquired maternal infection. As the maternal
blasts and macrophages (Hofbauer cells). Early mesenchymal cells circulation inside the placenta only starts to function at the end of the
probably differentiate into small reticulum cells, which, in turn, produce first trimester, transmission of Toxoplasma to the embryos is rare during
fibroblasts or large reticulum cells. The small reticulum cells appear to that period, but highly damaging to the developing fetus. Plasmodium
delimit a collagenfree stromal channel system through which Hofbauer falciparum is transferred by villous sequestration of maternal infected
cells migrate. Mesenchymal collagen increases from a network of fine erythrocytes or a breakdown of the villous barrier during labour. The
fibres in early mesenchymal villi to a densely fibrous stroma within stem presence of maternal rubella in the early months of pregnancy is of
villi in the second and third trimesters. After approximately 14 weeks, especial importance in relation to the production of congenital anoma
the stromal channels found in immature intermediate villi are infilled lies. (For further reading about TORCH perinatal infections, caused by
by collagen to give the fibrous stroma characteristic of the stem villus. Toxoplasma gondii, rubella virus, CMV and herpes simplex virus (HSV),
Fetal placental vessels include arterioles and capillaries; lymphatic see Stegmann and Carey (2002).)
vessels are not present. Pericytes may be found in close association with
the capillary endothelium, and from late first trimester, the vessels are
surrounded externally by a basal lamina. From the second trimester FETAL MEMBRANES
(and a little later in terminal villi), dilated thinwalled capillaries are
found immediately adjacent to the villous trophoblast; their respective
The implanting conceptus consists initially of three cavities and their
basal laminae apparently fuse to produce a vasculosyncytial interface;
surrounding epithelia. The original blastocyst cavity, surrounded by
the distance separating the maternal and fetal circulations may be
trophoblast, is now termed the chorionic cavity (synonymous with


reduced to as little as 2–3 µm. The endothelial cells are nonfenestrated,
extraembryonic coelom). It is a large cavity containing the much smaller
display numerous caveolae, and are linked by conspicuous junctional
amniotic cavity and secondary yolk sac (see Fig. 9.1). The apposition
complexes incorporating tight and adherens junctions.
of the latter two cavities delineates the extent of the early embryo. The
chorionic cavity becomes lined with extraembryonic mesoblast, which
transport across placental villi
is also reflected over the outer surface of the amnion and yolk sac. A
The mechanism of transfer of substances across the placental barrier
fourth cavity, the allantois, develops later as a caudal hypoblastic diver
(membrane) is complex. The volume of maternal blood circulating
ticulum that becomes embedded within the extraembryonic mesen
through the intervillous space has been assessed at 500 ml per minute.
chyme, forming the connecting stalk of the embryo. It does not have a
Simple diffusion suffices to explain gaseous exchange. Transfer of ions
direct mesothelial covering.
and other watersoluble solutes is by paracellular and transcellular dif
fusion and transport; the relative importance of each of these for most Chorion
individual solutes is unknown, and the paracellular pathway is mor
phologically undefined. Glucose transfer involves facilitated diffusion,
The chorion consists of developing trophoblast and extraembryonic
while active transport mechanisms carry calcium and at least some
mesothelium. It varies in thickness during development, both tempor
amino acids. The fatsoluble and watersoluble vitamins are likely to
ally and spatially. It is thickest at the implantation site throughout gesta
pass the placental barrier with different degrees of facility (Jauniaux and
tion as the chorion frondosum and then the placenta, and thinner as
Gulbis 2000a, Jauniaux et al 2004, Jauniaux et al 2005). The water
gestation progresses over the remainder of its surface as the chorion
soluble vitamins B and C pass readily. Water is interchanged between
laeve (see Fig. 9.4). At term, the chorion consists of an inner cellular


fetus and mother (in both directions) at approximately 3.5 litres per
layer containing fibroblasts and a reticular layer of fibroblasts and Hof
hour. The transfer of substances of high molecular weight, such as
bauer cells, which resembles the mesenchyme of an intermediate villus.
complex sugars, some lipids and hormonal and nonhormonal pro
The outer layer consists of cytotrophoblast 3–10 cells deep, resting on
teins, varies greatly in rate and degree and is not so well understood;
a pseudobasement membrane, which extends beneath and between
energydependent selective transport mechanisms, including receptor
the cells. Occasional obliterated villi within the trophoblast layer are
mediated transcytosis, are likely to be involved.
the remnants of villi present in the chorion frondosum of the first tri
Lipids may be transported unchanged through and between the cells
mester. Although the interface between the trophoblast and decidua
of the trophoblast to the core of the villus. The passage of maternal
parietalis is uneven, no trophoblast infiltration of the decidua parietalis
antibodies (immunoglobulins) across the placental barrier confers
occurs.
some degree of passive immunity on the fetus; it is widely accepted that
transfer is by micropinocytosis. Investigation of transplacental mechan
Yolk sac
isms is complicated by the fact that the trophoblast itself is the site of
synthesis and storage of certain substances, e.g. glycogen.
The placenta is an important endocrine organ. Some steroid hor As the secondary yolk sac forms, it delineates a cavity lined with par
mones, various oestrogens, βendorphins, progesterone, hCG and ietal, and perhaps visceral, hypoblast, continuous with the developing
human chorionic somatomammotropin (hCS), also known as placental endoderm from the primitive streak (Ch. 10; see Fig. 10.10). The sec
lactogen (hPL), are synthesized and secreted by the syncytium. The tro ondary yolk sac is the first structure that can be detected ultrasono
phoblast contains enzyme systems that are associated with the synthesis graphically within the chorionic cavity (Jauniaux et al 1991). Its


of steroid hormones, as well as enzymes that inactivate maternal hor diameter increases slightly between 6 and 10 weeks of gestation, reach
mones, allowing the fetus to develop and mature in a protected environ ing a maximum of 6–7 mm, after which its size decreases.
ment; for example, the fetus is protected from 10 times higher maternal The inner cells of the yolk sac (denoted endoderm in many studies,
glucocorticoid levels by the placental enzyme 11betahydroxysteroid although this layer is restricted to the embryo itself) display a few short
dehydrogenase 2 (11betaHSD2), which converts biologically active microvilli and are linked by juxtaluminal tight junctions (Jones and
maternal cortisol to inactive cortisone (Murphy et al 1974). Jauniaux 1995). Their cytoplasm contains numerous mitochondria,
It has been suggested that leukocytes may migrate from the maternal whorls of rough endoplasmic reticulum, Golgi bodies and secretory
blood through the placental barrier into the fetal capillaries. It has also droplets, giving them the appearance of being highly active synthetic
been shown that some fetal and maternal red blood cells may cross the cells. With further development, the epithelium becomes folded to
barrier. The former may have important consequences, e.g. in Rhesus form a series of cystlike structures or tubules, only some of which com
incompatibility. municate with the central cavity. The cells synthesize several serum
The majority of drugs are small molecules and are sufficiently proteins in common with the fetal liver, such as αfetoprotein (AFP),
lipophilic to pass the placental barrier from 7 weeks of gestation in α1antitrypsin, albumin, prealbumin and transferrin (Gulbis et al
quantities that yield measurable concentrations in both coelomic and 1998). With rare exceptions, the secretion of most of these proteins is
amniotic fluid. Many are tolerated by the fetus but some may exert grave confined to the embryonic compartments (Jauniaux and Gulbis 2000b).
teratogenic effects on the developing embryo/fetus, depending on the The yolk sac becomes coated with extraembryonic mesenchyme,


gestational age and the doses used by the mother (e.g. thalidomide, which forms mesenchymal and mesothelial layers. A diffuse capillary
cocaine) (Ross et al 2015). A welldocumented association exists plexus develops between the mesothelial layer and the underlying sec
between maternal alcohol ingestion and fetal abnormalities. Addiction ondary yolk sac wall, and subsequently drains through vitelline veins
of the fetus can occur to substances of maternal abuse such as heroin. to the developing liver. The mesothelial layer bears a dense covering of
A wide variety of bacteria (Listeria), protozoa (Toxoplasma gondii, microvilli; the presence of numerous coated pits and pinocytotic vesi
Plasmodium falciparum) and viruses, including cytomegalovirus (CMV), cles gives it the appearance of an absorptive epithelium (Jones and
human immunodeficiency virus (HIV) and rubella, are known to pass Jauniaux 1995, Burke et al 2013).
the placental barrier from mother to fetus. Toxoplasma and CMV are The secondary yolk sac plays a major role in the early embryonic
transferred by cellular invasion of the villous trophoblast during the development of all mammals. In laboratory rodents, it has been


ImplantatIon and placentatIon
178
2
noItceS
demonstrated as one of the initial sites of haemopoiesis. Human data intraabdominal urachus; this, in turn, continues into the apex of the
indicate that it has an absorptive role for molecules of maternal and bladder.
placental origin found in the chorionic cavity (Gulbis et al 1998), and
mediates the main movement of molecules passing from the chorionic Amnion (chorio-amnion)
cavity to the yolk sac and, subsequently, to the embryonic gut and
circulation.
The original amniotic cells develop from the edges of the epiblast of After week 9, the cellular components of the wall of the secondary
the embryonic disc, which ultimately form the interface with the skin yolk sac start to degenerate, and their function is subsumed into
at the umbilical region. Between the tenth and twelfth weeks of preg
exchanges at the placental chorionic plate. With embryonic develop
nancy, the amniotic cavity expands until it makes contact with the
ment of the midgut, the connection of the yolk sac to the embryo
chorion to form the chorioamnion, an avascular membrane that per
becomes attenuated to a slender and elongated vitelline intestinal duct.
sists to term. The amniotic membrane extends along the connecting
Both the yolk sac and its duct remain within the extraembryonic coelom
stalk and forms the outer covering of the umbilical cord. After birth,
(chorionic cavity) throughout gestation, located between the amnion
the site of this embryonic/extraembryonic junction is important because
and chorion as they fuse, near the placental attachment of the umbilical
the extraembryonic cell lines will die, causing the umbilical cord to
cord.
degenerate and detach from the body wall. In cases of anomalous
development of the ventral body wall, e.g. gastroschisis and exompha
Allantois
los, the reflections of the amnion along the forming umbilical cord may
be incomplete (see below).
The allantoenteric diverticulum (see Fig. 10.10) arises early in the third


The inner surface of the amnion consists of a simple cuboidal epi
week as a solid, endodermal outgrowth from the dorsocaudal part of
thelium. It has a microvillous apical surface, beneath which is a cortical
the yolk sac into the mesenchyme of the connecting stalk. It soon
web of intermediate filaments and microfilaments. There are no tight
becomes canalized. When the hindgut is developed, the proximal
junctional complexes between adjacent cells and cationic dyes penetrate
(enteric) part of the diverticulum is incorporated in its ventral wall. The
between the cells as far as the basal lamina. The intercellular clefts
distal (allantoic) part remains as the allantoic duct and is carried ven
present scattered desmosomes, but elsewhere the clefts widen and
trally to open into the ventral aspect of the cloaca or terminal part of
contain interlacing microvilli. These features are consistent with selec
the hindgut (Fig. 9.9A). The allantois is a site of angiogenesis, giving
tive permeability properties. The epithelium synthesizes and deposits
rise to the umbilical vessels that connect to the placental circulation.
extracellular matrix into the compact layer of acellular stroma located
The extraembryonic mesenchyme around the allantois forms the con
beneath the basal lamina, as well as the basal lamina itself.
necting stalk, which is later incorporated into the umbilical cord.
Towards the end of gestation, increasing numbers of amniotic
In the fetus, the allantoic duct, which is confined to the proximal
cells undergo apoptosis. Apoptotic cells become detached from the
end of the umbilical cord, elongates and thins. However, it may persist
amnion and are found in the amniotic cavity at term. The highest inci
as an interrupted series of epithelial strands at term, in which case the
dence is in weeks 40–41, independent of the onset of labour. Apoptosis
proximal strand is often continuous at the umbilicus with the median
may play a role in the fragility and rupture of the fetal membranes
at term.
Human amniotic epithelial cells are thought to be pluripotent


Placental area (chorion frondosum) because they arise so early from the conceptus. They can be distin
A guished from the epiblast cells from day 8. Amniotic cells lack the MHC
Connecting stalk antigen and so the amnion can be exposed to the maternal immune
attachment system without eliciting a maternal immune response. Cultured human
becomes more amniotic epithelial cells express a range of neural and glial markers,
Allantoic canal ventrally placed
including glial fibrillary acidic protein, myelin basic protein, vimentin
Amniotic cavity Chorionic cavity/ and neurofilament proteins, suggesting that these cells may supply
extraembryonic neurotrophic factors to the amniotic fluid. They also appear to have a
coelom hepatocyte gene expression profile, showing albumin production, gly
Chorion laeve
cogen storage and albumin secretion in culture. In organ culture, they
Yolk sac have been shown to secrete 30fold larger amounts of albumin than in
monolayer culture, and to secrete α1antitrypsin (Takashina et al
Pericardial coelom 2004). Amnion is used in the repair of corneas after trauma and as a
graft material for reconstructing vaginas in women with cloacal
abnormalities.
Prenatal changes to the chorio-amnion
At term, the surface area of the chorioamnion is 1000–1200 cm2, with
B 30% overlying the placenta and the remaining 70% in contact with the
Yolk sac decidua (Myatt and Sun 2010). There is a diminution of the decidual
Umbilical cord
component of the chorion and an increase in chorionic apoptosis
Line of fusion
Obliterated of amnion with towards term at the supracervical site, overlying the cervical os, which
yolk duct chorion, i.e. line encompasses the site of rupture (Chai et al 2013). The amnion and
of obliterated choriodecidua show varying degrees of separation over the putative
Umbilical coelom
extraembryonic rupture site; the mean rupture strength of the zone is 60% of the


coelom remaining membranes, associated with changes in collagen organiza
tion and increase in MMP9 (Strauss 2013). Term vaginal delivery is
Amniotic cavity
Allantoic duct associated with a threefold increase in apoptosis in the chorion close
to the rupture site, compared to fetal membranes from nonlabour,
elective caesarean delivery (Harirah et al 2012).
In preterm, premature rupture of membranes (PPROM), the chorion
is reported to be thinner at all sites (Fortner et al 2014). The presence
of bacteria in the chorion at term, ascending via the vagina, has been
reported without concomitant chorioamnionitis. However, chorio
amnionitis rates are higher in preterm labour and this may be a con
Fig. 9.9 A, A Longitudinal section of a conceptus showing the cavities
tributing cause (Harirah et al 2012, Fortner et al 2014). Loss of
associated with development. The amniotic cavity and yolk sac are both
chorioamnion intracellular cytokeratins, which become downregu
covered with extraembryonic mesoblast and are contained within the
larger chorionic cavity, which is lined with extraembryonic mesoblast. The lated in infection, is thought to make the amnion vulnerable to apop
embryo is attached to the chorion frondosum by the connecting stalk into tosis, shear stress and rupture (Vanderhoeven et al 2014).
which the allantois projects. B, A longitudinal section of a conceptus at a There is increasing evidence that infection and inflammation of the
later stage, showing the diminution of the chorionic cavity, expansion of chorioamnion is associated with prenatal changes to the developing
the amniotic cavity, relative attenuation of the yolk sac and the structures brain, especially the cortex, periventricular white matter and the devel
that give rise to the umbilical cord. oping cerebral blood vessels (Harteman et al 2013, Roescher et al


Umbilical cord
179
9
RetpaHc
2014). This association also extends to the neonatal period and post of cytokines, have also been detected in amniotic fluid; the concentra
natal gut maturation (Wynn and Neu 2012). Histologically confirmed tions of most cytokines differ from those in maternal blood and reflect
chorioamnionitis is associated with changes in cerebral blood flow the physiological status of the fetus (Tong 2013). Proteomic analysis of
soon after birth; male infants appear to be more affected than female human amniotic fluid at 16–18 weeks has identified more than 800
(Koch et al 2014). proteins and peptides (Cho et al 2007). It appears that amniotic fluid
may be acting as a pathway for the transport of signalling molecules to
target cells. In animal studies, substances injected into the amniotic
AMNIOTIC FLUID cavity were detected within the embryos within 2 minutes, demonstrat
ing a rapid exchange with all fetal tissues (Tong 2013).
The amniotic fluid, or liquor amnii, provides a buoyant medium that A number of clinical markers in amniotic fluid are used as predic
supports the delicate tissues of the young embryo and allows free move tors of fetal health, including interleukin (IL)10, IL6, cell free fetal
ment of the fetus up to the later stages of pregnancy. It also diminishes DNA and cell free fetal mRNA. Genes and proteins specific to the fetal
the risk to the fetus of injury from outside. Amniotic fluid is derived brain, lung, kidney, blood and heart have been identified in amniotic
from multiple sources throughout gestation. These include secretions fluid, differential gene expression investigated in a range of trisomic
from amniotic epithelium, filtration of fluid from maternal vessels via fetuses and lung maturity predicted by measurement of lecithin, sphin
the parietal decidua and amniochorion, filtration from the fetal vessels gomyelin and phosphatidylglycerol (Edlow and Bianchi 2012, Ross
via the chorionic plate or the umbilical cord, and fetal urine and fetal and Beall 2014). Bacterial species have been isolated from amniotic


lung secretions. In early pregnancy, diffusion from intracorporeal fluid, and microbial invasion of the amniotic cavity is considered a
vessels via fetal skin provides another source. Once the gut is formed, leading cause of preterm premature delivery (DiGiulio 2012). It is
fetal swallowing of amniotic fluid is a normal occurrence; the fluid is thought such bacteria access the amniotic cavity via the vagina and
absorbed into the fetal circulation. cervix, although many oral bacteria have also been identified, suggest
Amniotic fluid volume increases logarithmically during the first half ing that a haematogenous route is also likely. The establishment of
of pregnancy, from less than 10 ml at 8 weeks’ gestation to 30 ml at 22 pathogenesis is influenced by microbial adhesion, biofilm formation,
weeks and 770 ml at 28 weeks. After 30 weeks, the volume may remain and immune evasion and the maternal and fetal immune systems
unchanged to 36 weeks. It decreases towards the expected delivery date, (DiGiulio 2012).
and volumes decrease sharply in postterm gestation, averaging 515 ml
at 41 weeks (Beall et al 2007, Ross and Beall 2014).
UMBILICAL CORD
Human fetal urine output from the metanephric kidney increases
from 110 ml/kg/day at 25 weeks to almost 200 ml/kg/day at term; this
corresponds to almost 1000 ml/day. The rate of human fetal lung secre The formation of the connecting stalk and the early formation of the
tion has not been measured but is presumed to be in the range of umbilical cord are described in Chapter 10. The umbilical cord ulti
60–100 ml/kg/day near term (Callen 2008). Animal studies indicate mately consists of an outer covering of flattened amniotic epithelial
that lung fluid production is about onethird that of urine production, cells and an interior mass of mesenchyme of diverse origins (see Fig.
and that half of the fluid leaving the lungs enters the amniotic fluid and 9.9). It contains two tubes of hypoblastic origin, the vitellineintestinal


half is swallowed (Beall et al 2007, Ross and Beall 2014). and allantoic ducts, and their associated vitelline and allantoic (umbili
It is estimated that human fetuses swallow up to 760 ml/day of cal) blood vessels. The yolk stalk and continuing duct extend the length
amniotic fluid near term, although this decreases in the days before of the cord, whereas the allantoic duct extends only into its proximal
delivery. The remainder of the amniotic fluid is thought to be absorbed part.
via an intramembranous pathway directly across the amniotic cavity The mesenchymal core is derived from the somatopleuric extra
and fetal surface of the placenta into fetal blood vessels (Beall et al embryonic mesenchyme covering the amniotic folds, splanchnopleuric
2007, Ross and Beall 2014). Intramembranous flow is thought to reach extraembryonic mesenchyme of the yolk stalk (which carries the vitel
400 ml/day at term (Callen 2008). line vessels and clothes the yolk duct), and similar allantoic mesen
Amniotic fluid volume is estimated at routine antenatal ultrasound chyme of the connecting stalk (which clothes the allantoic duct and
scans, despite some concerns about its objectivity (Beattie and Rich initially carries two umbilical arteries and two umbilical veins). These
2007, Callen 2008, Gilbert 2012); it may be expressed as an amniotic various mesenchymal compartments fuse and are gradually trans
fluid index. A deficiency of amniotic fluid is termed oligohydramnios formed into the loose connective tissue (Wharton’s jelly) that character
and absent amniotic fluid is anhydramnios. Oligohydramnios in the izes the more mature cord. The tissue consists of widely spaced elongated
second or third trimester is usually the result of urinary tract malforma fibroblasts separated by a delicate threedimensional meshwork of fine
tions, e.g. bilateral renal agenesis or obstruction of the lower urinary collagen fibres, which contains a variety of hydrated glycosaminogly
tract, uteroplacental insufficiency or premature rupture of the mem cans and is particularly rich in hyaluronic acid.


branes. The urachus may play a critical role in the resolution of oligo The vitelline and allantoic (umbilical) vessels, which are initially
hydramnios in lower urinary tract obstruction by acting as a fistula symmetrical, become modified as a result of changes in the circulation.
between the bladder and the amniotic space. The major concern with The vitelline vessels involute, whereas most of the allantoic (umbilical)
oligohydramnios at less than 20 weeks is the significant risk of pulmo vessels persist. The right umbilical vein disappears but the two umbilical
nary hypoplasia and neonatal death. A volume of amniotic fluid in arteries normally remain. Occasionally, one umbilical artery may disap
excess of 2 litres is generally considered to be abnormal and constitutes pear; there is some correlation within structural anomalies, most often
polyhydramnios (see Fig. 14.5A). Maternal causes include cardiac and cardiac, in such cases. The vessels of the umbilical cord are rarely
renal problems and diabetes mellitus, which causes fetal hyperglycae straight, and are usually twisted into either a right or lefthanded cylin
mia and polyuria. Fetal causes include reduced fetal swallowing due to drical helix. The number of turns involved ranges from a few to over
congenital malformations, e.g. anencephaly; upper intestinal tract 300. This conformation may be produced by unequal growth of the
obstruction (oesophageal and duodenal atresia); compressive pulmo vessels, or by torsional forces imposed by fetal movements. Its func
nary disorders (congenital diaphragmatic hernia); and neuro muscular tional significance is obscure; perhaps the pulsations and contractions
impairment of swallowing. of the helical vessels assist the venous return to the fetus in the umbili
Amniotic fluid has, for many years, been regarded as a medium that cal vein.
physically supports the developing embryo and fetus. More recently, it Anomalies of the fetal anterior abdominal wall, such as exomphalos
has also been conceptualized as part of the embryonic and early fetal and gastroschisis, may affect the arrangement of the outer covering of


extracellular matrix, and its composition throughout pregnancy has amnion cells along the proximal end of the umbilical cord. Exomphalos
been investigated (Tong et al 2009). Techniques for biochemical analy arises from a failure of the lateral folds along the ventral surface of the
sis have revealed similarities and differences between amniotic fluid and embryo, resulting in failure of the normal embryonic regression of the
fetal umbilical cord blood and maternal serum. Although, early in midgut from the umbilical stalk into the abdominal cavity. The abdomi
development, amniotic fluid resembles blood plasma, it also demon nal contents, including intestines and liver or spleen, covered by a sac
strates extremely low oxygen tension, reflecting the physiological pla of parietal peritoneum and amnion, protrude into the base of the
cental hypoxia noted in the first trimester (Ross and Beall 2014, Jauniaux umbilical cord. In gastroschisis, the insertion of the umbilical cord is
et al 2003). In the second and third trimesters, primary electrolytes are intact and there is evisceration of the intestine through a small abdomi
similar in amniotic fluid, umbilical cord blood and maternal serum, nal wall defect that is usually located to the right of the umbilical cord;
whereas glucose, cholesterol and some enzymes are markedly lower in this results in free loops of bowel in the amniotic cavity. Theories con
amniotic fluid. Umbilical cord blood and maternal serum contain 8 cerning the aetiology of this defect include abnormal involution of the
and 12.5 times more protein than amniotic fluid, respectively (Tong right umbilical vein or disruption of the omphalomesenteric artery by
et al 2009). More than 100 metabolites, including cortisol and a range ischaemia.


ImplantatIon and placentatIon
180
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noItceS
Mature umbilical vessels, particularly the arteries, have a strong mus with vulnerability to injury and fetal haemorrhage. This is especially
cular coat that contracts readily in response to mechanical stimuli. The important if the placenta is lowlying, and may be associated with vasa
outermost bundles pursue an interlacing spiral course, and when they praevia, in which case fetal blood vessels run across the internal os.
contract, they produce shortening of the vessel and thickening of the Inadvertent rupture of the fetal vessels in spontaneous labour or at the
media, with folding of the interna and considerable narrowing of the time of amniotomy (artificial rupture of membranes to induce labour)
lumen. This action may account for the periodic sharp constrictions of will cause fetal haemorrhage and may prove fatal. Infection of the
contour, the socalled valves of Hoboken, which often characterize umbilical cord, funisitis, is associated with chorioamnionitis and later
these vessels. neurodevelopmental outcomes (Roescher et al 2014, Buhimschi and
The fully developed umbilical cord is, on average, some 50 cm long Buhimschi 2012).
and 1–2 cm in diameter. Its length varies from 20 to 120 cm; exception Recent studies have examined the optimal time for cutting the
ally short or long cords are associated with fetal problems and compli umbilical cord after delivery and found that early clamping is associated
cations during labour. A long umbilical cord may prolapse through the with neonatal anaemia and iron deficiency. Delayed clamping, defined
cervix into the vagina once the fetal membranes rupture and this may as ligation of the umbilical cord 2–3 minutes after birth or when cord
be exacerbated by conditions that prevent the fetal head from fully pulsation stops, reduces the prevalence of iron deficiency at 4 months
occupying the maternal pelvis, e.g. pelvic tumours (fibroids), ovarian of age and results in improved ferritin levels (Andersson et al 2011,
cysts, placenta praevia and prematurity. Compression of the cord by the McDonald et al 2013).
presenting part of the fetus, or an umbilical artery spasm, will lead to


fetal hypoxia and death, if untreated. The risk of perinatal death rises
as the interval from diagnosis to delivery increases. The treatment is
either funic replacement (pushing the cord back above the fetal head)
Bonus e-book video
or, more commonly, immediate caesarean section, depending on factors
such as fetal viability.
The distal end of the umbilical cord usually attaches in the central
portion of the placenta, but in 0.2% of pregnancies, velamentous inser Video 9.1 Ultrasound features of the maternal placental blood flow.
tion is observed (i.e. into the membranes) and this may be associated
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