Implantation
Ji Yan
10-12-2012
• Infertility has been a serious worldwide problem,
especially in developed countries. It has been
reported that developed countries have much
high infertility rate and much lower pregnancy
rate (Cohen JE, 2010).
• The World Health Organization estimates that 812% of all couples world-wide experience
infertility, at least 7.3 million people in the United
States, 3.5 million people in the United Kingdom
and 40 million couples in China suffer infertility.
• The World Health Organization concluded that
in 47 developing countries (excluding China)
186 million women between the ages of 15-49
are infertile.
• Of the pregnancies that are lost, 75%
represent a failure of implantation, and
therefore are not clinically recognized as
pregnancies.
• Implantation involves an intricate
discourse between the embryo and uterus
and is a gateway to further embryonic
development. (NATURE REVIEWS | GENETICS, 2006 )
• Implantation is the process in which the
blastocyst physically and physiologically
comes into intimate contact with the uterus.
(KevinYLee etal. 2004)
• Although implantation involves the interplay
of numerous signaling molecules, the
hierarchical instructions that coordinate the
embryo-uterine dialogue are not well
understood.
• A better understanding of peri-implantation
biology could alleviate female infertility and
help to develop novel contraceptives.
NATURE REVIEWS | GENETICS, 2006
• The successful implantation of an embryo is
contigent upon cellular and molecular crosstalk between the uterus and embryo.
• Thus, deficiencies in uterine receptivity,
embryo development, or the embryo-utrerine
dialogue will compromise fertility.
Embryo
• The differentiated and expanded
blastocyst is composed of three cell types:
the outer polarized epithelial
trophectoderm, the primitive endoderm,
and the pluripotent ICM.
• The ICM provides the future cell lineages
for the embryo
• The trophectoderm, the very first epithelial
cell type in the developmental process,
makes the initial physical and
physiological connection with the uterine
luminal epithelium.
• Trophoblast cells produce a variety of
growth factors, cytokines, and hormones
that influence the conceptus and maternal
physiology in an autocrine, paracrine,
and/or juxtacrine manner.
• Implantation is the process in which the
blastocyst physically and physiologically
comes into intimate contact with the uterus.
(KevinYLee etal. 2004)
• The successful implantation of an embryo is
contigent upon cellular and molecular crosstalk between the uterus and embryo.
• Thus, deficiencies in uterine receptivity,
embryo development, or the embryo-utrerine
dialogue wil compromise fertility.
Ovary
Oviduct
Perimetrium
Cervix
Myometrium
GE
Endometrium LE
• Fertilization occurs in the fallopian tube within
24 to 48 hours after ovulation.
• The initial stages of development, from
fertilized ovum (zygote) to a mass of 12 to 16
cells (morula), occur as the embryo, passes
through the fallopian tube.
• The morula enters the uterine cavity
approximately two to three days after
fertilization.
• The appearance of a fluid-filled inner cavity
within the mass of cells marks the transition
from morula to blastocyst and is
accompanied by cellular differentiation.
The fertilization of an egg by a sperm,
normally occurs in the fallopian tubes.
The next step for the fertilized egg is to
implant into the walls of the uterus,
beginning the initial stages of pregnancy.
If fertilization and/or implantation does not
take place, the system is designed to
menstruate (the monthly shedding of the
uterine lining).
The events of implantation include:
apposition of the blastocyst to the uterine
luminal epithelium
adhesion to the epithelium
penetration through the epithelium basal
lamina and invasion into the stromal
vasculature
Figure 1. Blastocyst
Apposition and Adhesion.
The diagram shows a
preimplantation-stage
blastocyst (approximately
six to seven days after
conception) and the
processes
thought to be necessary for
uterine receptivity and
blastocyst apposition and
adhesion.
Figure 2. Blastocyst Implantation
The diagram shows an invading
blastocyst (about 9 to 10 days
after conception) and the
processes necessary for
trophoblast invasion.
• Although various cellular aspects and
molecular pathways of implantation
process have been identified, a
comprehensive understanding of the
implantation process is still missing.
• A better understanding of the molecular
signals that regulate uterine receptivity
and implantation may lead to strategies
to correct implantation failure and
improve pregnancy rates.
• Recent advances in molecular and genetic
approaches have led to the discovery of
numerous molecules involved in embryouterine interactions; however, the precise
sequence and details of the signaling
cascades for many of these molecules
have not yet been defined.
• We attempts to focus on a select number
of signaling molecules and pathways that
are implicated in embryo-uterine
interactions in relation to implantation.
Steroid hormone signaling
• Ovarian estrogen (E2) and progesterone
(P4) are essential for blastocyst
implantation in the progesterone-primed
uterus in mice and rats.
 Estrogen action is normally considered to
involve its interaction with a nuclear receptor
(ER), a ligand-dependent transcription factor .
 Progesterone receptor (PR)
adhesion molecules: cell-cell
interactions
•
•
•
•
•
•
selectins
galectins
Muc-1
integrins
cadherins
trophinin
vasoactive factors
• It has long been speculated that vasoactive
agents, such as histamine and PGs, are
involved in many aspects of reproduction
including ovulation, fertilization, implantation,
and decidualization.
Histamine is a well-known neurotransmitter in
the brain, but it is also involved in other
physiological responses including gastric acid
secretion, regulation of allergic reactions, and
vascular permeability.
Because the process of implantation is
considered a proinflammatory reaction and
because increased vascular permeability at the
site of blastocyst implantation is common to
many species, it was suggested that histamine
plays a role in implantation and decidualization .
• PGs possess vasoactive, mitogenic, and
differentiating properties and are implicated
in various female reproductive functions.
growth factors
• The expression of various growth factors
and their receptors in the uterus in a
temporal and cell-specific manner during
the periimplantation period suggests that
these factors are important for implantation.
growth factors
• EGF family
EGF, TGFα, HB-EGF, amphiregulin,
betacellulin, epiregulin, neuregulin
• TGFβs
• FGFs
• IGFs
• PDGF
cytokines
TNFα
IL-1
M-CSF
LIF
Homeobox genes
• Hoxa-10 and Hoxa-11 are highly
expressed in developing genitourinary
tracts and the adult female reproductive
tract, suggesting roles in reproductive
events.
• Hmx3 is primarily expressed in the
myometrium during early pregnancy.
• In brief, implantation in mammals
absolutely depends upon synchronized
development of the blastocyst to the stage
when it is competent to implant and the
uterus to the stage when it is receptive to
blastocyst growth and implantation.
• Ovarian estrogen (E2) and progesterone
(P4) are the primary effectors that direct
the prereceptive uterus to a receptive
state via a number of locally expressed
growth factors, cytokines, transcription
factors,and vasoactive mediators in the
uterus.
FIG. 1. A scheme of
signaling networks in
embryo-uterine
communication during
implantation in mice.
Le, Luminal epithelium;
Ge, glandular epithelium;
S, stroma;
CE, catechol estrogen.
• Tissue remodeling and angiogenesis are two
hallmark events during implantation and
decidualization.
• Increased vascular permeability and
angiogenesis are crucial to successful
implantation, decidualization, and placentation.
Vascular epithelial growth factor
(VEGF)
• VEGF, originally discovered as a vascular
permeability factor, is also a potent mitogen
for endothelial cells and a key regulatory
growth factor for vasculogenesis and
angiogenesis.
• Targeted disruption of even one allele of the
Vegf gene results in embryonic death in uteri
during midgestation with aberrant blood
vessel formation.
Angiogenic signaling in the uterus during implantation
• The proangiogenic factor VEGF and its receptor
Flt1 (VEGFR1) and Flk1 (VEGFR2) are
primarily important for uterine vascular
permeability and angiogenesis before and
during the attachment phase of the implantation
process.
• Among the factors regulating uterus
development, vascular remodeling
promoters are critical for uterus function
and fertility. VEGF as a major factor
promotes endothelial cell growth and
blood vessel development.
• In addition, VEGF has been found playing
major roles in non-endothelial cells.
• Gene expression studies and genetically engineered
mouse models have provided valuable clues to the
implantation process with respect to specific growth
factors, cytokines, lipid mediators, adhesion
molecules, and transcription factors.
• A staggering amount of information from microarray
experiments is also being generated at a rapid pace.
If properly annotated and explored, this information
will expand our knowledge regarding yet-to-beidentified unique, complementary, and/or redundant
molecular pathways in implantation.
(Endocrine Reviews 25: 341–373, 2004)
• High-throughput sequencing technology has been
used for transcriptone analysis .
• Taking advantage of the Illumina Genome
Analyzer platform, we performed transcriptional
profiling on the VEGF-repressed mouse uterus.
• The Digital Gene Expression (DGE) tag profiling
allows us to quantify gene expression levels of
relative samples.
• In this study, a genetically repressed VEGF
mouse model is used to profile uterus
transcriptome at post coitus 2.5.
• Go analysis of uterus expressed genes,
uterus specific genes and VEGF-reguated
genes, and their related signaling pathways
were analyzed.
Gene ontology (GO)
•
The Gene Ontology project provides an ontology of defined
terms representing gene product properties. The ontology
covers three domains:
– cellular component, the parts of a cell or its
extracellular environment;
– molecular function, the elemental activities of a gene
product at the molecular level, such as binding or
catalysis;
– biological process, operations or sets of molecular
events with a defined beginning and end, pertinent to the
functioning of integrated living units: cells, tissues,
organs, and organisms.
KEGG pathway
•
•
KEGG (Kyoto Encyclopedia of Genes and
Genomes) is a collection of databases dealing
with genomes, enzymatic pathways, and
biological chemicals.
The PATHWAY database records networks of
molecular interactions in the cells, and variants
of them specific to particular organisms.
Up-regulated genes
Down-regulated genes
A: Up-regulated genes
B: Down-regulated genes
• To determine whether VEGF regulate uterus
development through antisense genes
expression, the antisense transcript levels were
analyzed.
• Antisense expression levels showed correlation
with sense expression which may indicate nonspecific transcription near promoters and
enhancers.
Genes relatived to muscle
devolpment
• Enzymes: Sult1d1, Ctla2a, Hdc, Pkdcc, Pcx,
Gamt, Usp2, Mylk, Car2, Mif, Rnf187
• Receptors: P2ry14, Usmg5, Ramp3, Oxtr,
Mt2, F2r, Sfrp2, Ly6e,Fgfl1
• Structure proteins: Acta2, Actg2, Eln, Gjb2,
Utrn, Myh11
• Glycoprotein: Inhbb, CD24a, Tnc, Ihh
• Transcript factors: Hopx, Tcf23, Lef1, Elf3,
Skil
• Regulatory proteins: Cnn1, Cabl1, Fxyd4
GE
LE
V
Dox+
Dox-