Bi8940 Developmental Biology
Lesson 5
Vertebrate Organogenesis: Endo- and Mesodermal
Derivatives
Jan Hejátko
Laboratory of Molecular Plant Physiology,
Department of Functional Genomics and Proteomics,
and
Functional Genomics and Proteomics of Plants
CEITEC
Masaryk University,
Brno, Czech Republic
hejatko@sci.muni.cz, www.ceitec.eu
1
Outline of Lesson 5
Organogenesis in Vertebrates: Endo- and Mesodermal
Derivatives
 Mesoderm derivatives development
 somites formation and signalling
 formation of muscles
 endochordal ossification and signalling
 nephrogenesis
 formation of gonads
 hematopoesis and circular system development
 limbs formation
 Endoderm derivatives development
 alimentary canal and its derivatives formation
2
Outline of Lesson 5
Organogenesis in Vertebrates: Endo- and Mesodermal
Derivatives
 Mesoderm derivatives development
 somites formation and signalling
3
The developmental potential might be estimated (here during gastrulation) using
experiments, where donor cells are marked by staining with
tetramethylrhodaminisothyocyanate (TRITC) or recently, more specific via marker
gene expression.
The developmental potential in vertebrates is becoming narrower during
development.
Thus, though the cell is determined to form mesoderm, it is not determined at
this time to form a certain type of mesoderm. For example, the prospective
mesoderm cell may be capable of forming muscle, while its progeny may also be
able to form cartilage or body‐wall mesenchyme.
4
Bazální lamina
Zduřelé buňky
Pochva
Vacuoles with
proteoglycans
Direction of
Hensen’s
node
movement
Struna hřbetní
H2O
H2O
During gastrulation, as the Hensen’s node moves along the primitive streak, the
dorsal most mesoderm, adjacent to the forming neural tube, the midline
mesoderm, forms notochord (posteriorly from the level of prospecitve hidnbrain)
that is only a few cell layers in diameter in amniotes.
At the surface, notochord is covered by cellular sheath and basal lamina. The
notochord cells form vacuoles that accumulate proteoglycans. That causes that
the water is imported there and the notochord becomes stiffer, forming thus a
transient support to the developing embryo.
5
Induction of another ventral
pole via BMPs inhibition
As discussed previously, the notochord originates from the Hensen’s node cells
and has the ability to induce another neural tube formation (its ventral pole),
probably by blocking BMP signaling.
A gradient of BMPs is formed along the dorsoventral axis. High levels of BMP
favor not only dorsal ectoderm, but also mesoderm (see slides 15, 16) while low
BMP levels allow ventral ectoderm/mesoderm structures to develop.
6
Presomitický
(paraxiální)
mesoderm
Somit
(prvosegment,
myocél)
At both sides of the notochord, the so called presomitic (paraxial) mesoderm is
formed that will develop into somites, except the paraxial mesoderm in the head
region that does not form somites.
Beginning at the anterior end, the cells of presomitic mesoderm start to
aggregate, forming thus individual somites.
Process of cell aggregation is common to the beginning of development of most
of the mesodermal structures, as will be discussed further.
This process progresses towards the posterior end and it is accompanied by the
expression of C‐HAIRY, a mice homologue of Drosophila’s HAIRY gene. In
Drosophila, HAIRY expression coincides with segments formation (will be
discussed later).
C‐HAIRY is expressed in waves, every taking for about 90 minutes and allowing
thus individual somites formation. High levels of HAIRY expression remains at the
posterior portions of the newly formed somites.
Together with other genes, C‐HAIRY is a part of internal molecular clock (see the
next slide).
7
Palmeirim et al., Cell (1997)
In situ mRNA
localization of
C-HAIRY1 in
developing
chick embryo
Posterior poles
of new somites
basic helix-loop-helix
(b-HLH) TF caudal fissure appearing
The rhythm of somite production is characteristic of the species at a given temperature (90 min in the chick embryo
at 37°C and 20 min for the zebrafish embryo at 25°C). The total number of somites is constant within a given species.
It is usually about 50, although in some animals, such as snakes, it can reach up to 400 (Pourquie, Science, 2003).
In vertebrates, segmentation involves a molecular oscillator—the segmentation clock—which acts in the presomitic
mesoderm (PSM). Evidence for this oscillator was firstly provided by the observation of the regular pulses of
expression in PSM cells of the mRNA coding for the basic helix‐loop‐helix (b‐HLH) transcription factor c‐hairy1, a
vertebrate homolog of the protein encoded by the fly pair‐rule gene hairy (Palmeirim et al., Cell, 1997, see the
slide).
c‐hairy1 mRNA expression in the PSM defines a highly dynamic caudal‐to‐rostral (posterior/anterior) expression
sequence reiterated during formation of each somite.
(Top) In situ hybridization with c‐hairy1 probe showing different categories of c‐hairy1 expression patterns in
embryos aged of 15 (A, B, and C), 16 (D, E, and F), and 17 (G, H, and I) somites. Rostral to the top. Bar 5 200 mm.
(Bottom) Schematic representation of the correlation between c‐hairy1 expression in the PSM with the progression of
somite formation.
While a new somite is forming from the rostral‐most PSM (somite 0, S0), a narrow stripe of c‐hairy1 is observed in its
caudal aspect, and a large caudal expression domain extends rostrally from the tail bud region (stage I; A, D, and G).
As somite formation proceeds, as evidenced by the visualization of the appearing caudal fissure, the c‐hairy1
expression expands anteriorly, the caudal‐most domain disappears, and c‐hairy1 appears as a broad stripe in the
rostral PSM (stage II; B, E, and H).
When somite 0 is almost formed, the stripe has considerably narrowed, and c‐hairy1 is detected in the caudal part of
the prospective somite (stage III; C, F, and I) while a new caudal expression domain arises from the tail bud region (in
C can be seen the beginning of stage I of the next cycle).
This highly dynamic sequence of c‐hairy1 expression in the PSM was observed at all stages of somitogenesis
examined (from 1 to 25 somites), suggesting a cyclic expression of the c‐hairy1 mRNA correlated with somite
formation. Arrowheads point to the most recently completely formed somite (somite I, SI).
8
Pourquie., Science (2003)
II. III. III.II.I. I.
Several additional genes, referred to as cyclic genes, that exhibit a dynamic behavior similar to
that of c‐hairy1 have now been identified in fish, frog, chick, and mouse embryos, suggesting
that the segmentation clock is conserved among vertebrates.
The best characterized set of cyclic genes is involved in Notch signaling, suggesting that Notch
activation lies at the heart of the oscillator. Such genes encode several transcription factors of
the Hairy and Enhancer of Split (HES) family, acting downstream of Notch signaling, as well as
the glycosyl transferase Lunatic Fringe and the Notch ligand deltaC.
The NOTCH signalling pathway includes several negative feed‐back loops mediated e.g. by
Lunatic‐Fridge or transcriptional repressors from the HES family (see the slide).
A second group of cyclic genes linked to the Wnt signaling pathway has recently been
uncovered. Thus far, only one cycling gene in this class has been identified: the inhibitor of Wnt
signaling Axin2.
In the mouse, axin2 is expressed in a dynamic sequence similar to, but out of phase with, that of
the Notch‐related cyclic genes (Fig. A). axin2 is directly regulated by Wnt signaling and could
participate in the establishment of an autoregulatory negative feedback loop involved in its
periodic expression. axin2 oscillations persist in Notch pathway mutants, whereas both axin2
and lunatic fringe oscillations are disrupted in wnt3a mutants, indicating that Wnt signaling acts
upstream of the Notch‐regulated cyclic genes (Fig. B).
Therefore, in the mouse, the segmentation clock appears to be composed of a Wnt‐based
regulatory loop entraining a series of Notch‐based loops (Fig. B). The details of the interactions
between these different loops are presently not understood. Also, the conservation of the Wnt‐
based loop across vertebrates remains to be examined. The exact role of this oscillator in the
segmentation process remains unclear (Pourquie, Science, 2003).
9
Nusse, Nature (2005)
The name Wnt was coined as a catenation of Wg (wingless) and Int and is pronounced 'wint'. The wingless gene had
originally been identified as a recessive mutation affecting wing and haltere development in Drosophila
melanogaster. It was subsequently characterized as segment polarity gene in Drosophila melanogaster that functions
during embryogenesis and also during adult limb formation during metamorphosis.
The INT genes were originally identified as vertebrate genes near several integration sites of mouse mammary tumor
virus (MMTV) in mouse breast cancers. The Int‐1 gene and the wingless gene were found to be homologous, with a
common evolutionary origin evidenced by similar amino acid sequences of their encoded proteins.
Mutations of the wingless gene in the fruit fly were found in wingless flies, while tumors caused by MMTV were
found to have copies of the virus integrated into the genome forcing overproduction of one of several Wnt genes.
The ensuing effort to understand how similar genes produce such different effects has revealed that Wnts are a major
class of secreted morphogenic ligands of profound importance in establishing the pattern of development in the
bodies of all multicellular organisms studied (Wikipedia).
The canonical Wnt signalling pathway is shown on the slide. Here, the Wnt protein binds to its receptor, the
transmembrane protein Frizzled.
a. In cells not activated by Wnt, a complex between β‐catenin, Axin, APC and GSK3 causes phosphorylation of β‐
catenin and its consequent destruction. The Wnt receptors LRP6 and Frizzled are unoccupied. B. Without Axin, β‐
catenin is stabilized and it enters the nucleus to control gene expression.
Inset, binding of Wnt to cells results in phosphorylation (P) of LRP6 residues in its cytoplasmic tail. Zeng et al. and
Davidson et al. show that this is catalysed by the GSK3 and CK1 protein kinases. CK1 is attached to the membrane by a
lipid anchor domain. Several other sites on LRP6 that become phosphorylated are not shown here. The
phosphorylated LRP6 recruits Axin, removing it from the β‐catenin destruction complex and stabilizing β‐catenin.
Up‐to‐date and detailed information on these interactions and WNT signaling pathways will be presented during
advanced lecture of Dr. V. Bryja (Bi9903 Developmental Animal Physiology I and Bi9906 Developmental Animal
Physiology II).
10
Wikipedia
Notch signalling
In 1914, John S. Dexter noticed the appearance of a notch in the wings of the
fruit fly Drosophila melanogaster. The alleles of the gene were identified in 1917
by Thomas Hunt Morgan. Its molecular analysis and sequencing was
independently undertaken in the 1980s by Spyros Artavanis‐Tsakonas and
Michael W. Young.
The notch protein sits like a trigger spanning the cell membrane, with part of it
inside and part outside. Ligand proteins binding to the extracellular domain
induce proteolytic cleavage and release of the intracellular domain, which enters
the cell nucleus to alter gene expression.
Because most ligands are also transmembrane proteins, the receptor is normally
triggered only from direct cell‐to‐cell contact. In this way, groups of cells can
organize themselves, such that, if one cell expresses a given trait, this may be
switched off in neighbor cells by the intercellular notch signal. In this way, groups
of cells influence one another to make large structures (Wikipedia,
http://en.wikipedia.org/wiki/Notch_signaling_pathway).
11
Pourquie., Science (2003)
III.II.I. III.II.I. I.
TFs Mesp2/c‐meso1
↑  FGF8 = immature, posterior-located cells
↓ FGF8 = mature, anterior-located cells - ”determination front”
ac va on of segmenta on program, ↑ Paraxis, ↓posterior genes
(e.g.Brachyury), stopping oscillation genes
Notch-related
cyclic genes
FGF Signaling: Translation of the Clock Pulsation into Spatial Periodicity. Model for segment formation in
vertebrates based on mouse and chick data.
The FGF8/Wnt3A gradient, which regresses posteriorly during somitogenesis, is shown in black. The
anterior boundary of the gradient defines the determination front, which corresponds to the position of
the wavefront (thick black line). The phase I expression of Notch‐related cyclic genes is shown in red. The
expression of Mesp2/c‐meso1 is shown in blue.
Recent studies have shown that the secreted growth factor FGF8 (fibroblast growth factor 8) could be
implicated in converting the clock pulsation into the periodic arrangement of segment boundaries. FGF8
mRNA is strongly expressed in mesenchymal presomitic mesoderm (PSM) precursors in the primitive
streak and tail bud as well as in the posteriormost PSM, and its expression progressively decreases in
more anterior cells, thus establishing a gradient over two‐thirds of the PSM length (see the slide, black).
Overexpression of FGF8 in PSM cells can maintain their posterior identity and block segmentation,
suggesting that high concentrations of FGF8 are required to actively maintain newly formed posterior
PSM cells in an immature state. It was proposed that, because of the progressive decrease of FGF8
expression during maturation of the PSM, when cells become located in the anterior PSM, they reach a
threshold of FGF signaling allowing them to activate their segmentation program.
This threshold level, which was termed the “determination front” (see the slide), marks a transition in
genetic regulation in PSM cells, as shown by the activation of new sets of genes such as Paraxis, the down‐
regulation of posterior genes such as Brachyury, and the slowing down and stopping of the oscillations of
the clock genes.
Wnt3A was also recently proposed to assume a role similar to that of FGF8 by establishing a gradient‐
controlling segmentation in the PSM (20). However, because Wnt3A acts upstream of FGF8 in the PSM, it
could act together with or by way of the FGF8 gradient.
At the determination‐front level, gene coding for the transcription factor Mesp2/c‐meso1 becomes
periodically activated in a one‐somite wide domain, providing the earliest evidence for segmentation in
the PSM.
12
Kanálek pronefros
Vývod mesonefros
Tubulus mesonefros
Dermomyotom
Sklerotom
Neurální lišta
Myotom
Dermatom
Vývod mesonefros
Dorzální aorta
Somite is transient embryonal structure. Soon as the somites are formed, the
medioventral portion of the somite disagregates. The released cells, called
sclerotome abut the notochord and will differentiate into segmental cartilage, a
precursor of the vertebreae and proximal portion of the ribs.
The dorsal portion of the somites remains of epithelial character. It is called
dermamyotome, as it is comprised of dermatome and myotome.
Dermatome will give rise to dermis, the lower portion of the integuments that
contains some migrated ectoderm cells forming pigment cells, as discussed in
previous chapter.
Myotome will develop into muscles.
The relative positions of the forming kidney, pronephros and mesonephors, are
shown and will be discussed later.
The lateral portion of the dermamyotome will provide cells that will develop into
muscles of the body wall, limbs and distal portion of the ribs.
13
Importance of intercellular communications
•MYOD,MYF5
•Muscle cells specific TFs
•Mouse DELTA homologue
•Somites segmentation signalling
What is the molecular mechanism involved in the determination of the
narrowing developmental potential of mesoderm?
The explantation experiments have provided experimental evidence about
importance of the intercellular communications. As shown in the figure, the
nature of developing tissue is determined by the presence of surrounding tissue,
particularly neural tube (and its dorsal or ventral portion), lateral mesoderm and
overlying ectoderm.
Recently, some of the molecular determinants involved in the tissue
differentiation have been identified. These are e.g. muscle cells specific TFs
MYOD (see later slides) and MYF5. When ectopically expressed, these TFs induce
muscle cell differentiation and thus could be considered as muscle marker genes.
Also some of the signalling molecules involved in specific processes during
mesoderm structures differentiation were identified. This is e.g. homologue of
the DELTA gene from Drosophila that was identified in mouse. It is a signalling
molecule necessary for the somites segmentation.
14
Another important signalling molecule is SHH (SONIC HEDGHEOG) that we have
discussed previously (see Chapter 6 and the problem of the dorsoventral neural
tube patterning, inset).
SHH is secreted by the neural floor plate and by the notochord.
SHH secreted from the notochord alone is not able to induce sclerotome
formation.
However, notochord secreted SHH induces SHH production from the floor plate
of the neural tube (see previous slide), which allows to reach sufficient amount
of SHH to induce sclerotome differentiation and thus triggers the process of
cartilage formation.
15
Ectoderm and lateral
mesoderm derived
signals
Expression of muscle-specific
TFs
Dermamyotome differentiation
thanks to WNT and BMPs
signalling
Sclerotome formation thanks
to Shh signalling
Muscle-specific TFs
Complicate signalling networks are operating in the different developmental
processes during embryogenesis.
For the proper development of early dorsal mesoderm, interaction of specific
signals from different portions of neural tube, notochord, ectoderm and lateral
mesoderm and their proper timing and position are necessary.
WNT, secreted from the dorsal portion of neural tube together with ectoderm
and lateral mesoderm derived signals like BMPs is necessary for the
dermamyotome differentiation.
SHH triggers sclerotome formation.
Both dorsal and ventral signals are necessary for the later myotome formation
that is accompanied by the expression of muscle‐specific TFs, e.g. MyoD (see the
figure). SHH acts in the dorsal somite determination together with WNT either
directly or via unknown intermediate (X, see figure).
More about the Wnt signalling in the specialized lecture of Assoc. Prof. Bryja,
Developemental physiology of animals I and II (Bi9903 and Bi9906).
16
Outline of Lesson 5
Organogenesis in Vertebrates: Endo- and Mesodermal
Derivatives
 Mesoderm derivatives development
 somites formation and signalling
 formation of muscles
17
Unknown signal
(e.g. poor medium under in vitro conditions)
Muscle cells differentiate from myoblasts, the prospective muscle cells located in
myotome.
Myoblatsts stop proliferation that could be induced e.g. by the replacement of
cells from rich to poor medium and enter the G0 phase of the cell cycle. The
signal starting the development of muscle cells in embryos is still unkown.
18
Development of a given muscle differ and has its own timing and location and
undergo specific developmental pathways.
Myoblasts elongate, fuse and form syncitia in case of striated muscle cells. The
genes involved in the actin and myosin biosynthesis are activated.
There are differences of the developmental scheme based on the muscle type.
Heart muscles are distinctive but similar to striated cells. Smooth muscles do not
form syncitia and organization of actin and myosin is also different then in
striated muscle cells.
19
Outline of Lesson 5
Organogenesis in Vertebrates: Endo- and Mesodermal
Derivatives
 Mesoderm derivatives development
 somites formation and signalling
 formation of muscles
 endochordal ossification and signalling
20
IHH GLI,
PTC
PTHrP
PTH
Aggregation of
sclerotomes,
central cells
become
chondrocytes
Perichondrium
formation
Vertebrae and
proximal ribs
Long bones
Cell division
and swelling
Hypertrophic
chondrocytes
formation
Bone
formation
Blood vessels
invasion,
osteoblasts
movement and
hydroxyapatite
deposition Ochrustavice
(perichondrium)
Endochondral ossification
vs.
intramembraneous ossification
glioma-associated
oncogene homolog 1
patched
Sclerotome develops into cartilage that later allows vertebrae and proximal portion of ribs
formation. The cartilage for long bones of the limbs will develop from the lateral mesoderm.
Cartilage represents a template where the bone formation starts via a process called
endochondral ossification. Only the skull bones form without cartilage directly in mesenchymal
cells derived from the neural crest via a process called intramembranous ossification.
Sclerotome cells aggregate and the central cells become chondrocytes (cartilage cells), other
cells form a surrounding sheath, called perichondrium.
Chondrocytes divide and swell, forming thus hypertrophic chondrocytes.
The blood vessels invade the hypertrophic cartilage and bone forming osteoblasts move into the
area. Osteoblasts deposit hydroxyapatite, a crystaline form of calcium phosphate.
Growth and morphogenesis of the bones depends on a carefully regulated processes of
replacement of the cartilage by bone. If the process is too fast or vice versa too slow, it leads to
shortening or elongation of the forming bone, respectively.
Complicated feed back of signalling molecules, e.g. parathyroid hormone (PTH), parathyroid
hormone related peptide (PTHrP), INDIAN HEDGHEHOG (IHH) and other signalling molecules
(see the scheme above) takes place in that type of growth regulation.
GLI stands for glioma‐associated oncogene homolog 1 (zinc finger protein), PTC is PATCHED, a
receptor for IHH.
21
Outline of Lesson 5
Organogenesis in Vertebrates: Endo- and Mesodermal
Derivatives
 Mesoderm derivatives development
 somites formation and signalling
 formation of muscles
 endochordal ossification and signalling
 nephrogenesis
22
Splanchnic mesoderm
Splanchnický mesoderm
Somatic mesoderm
Somatický mesoderm
Intermediální mesoderm/nefrotom
Ledvinový
kanálek (odvodný
kanálek)Ledvinové tubuly
Filtrate
reabsorbtion
and cloaca
delivery
Liquid
filtration
The region between somite and splanchnic and somatic mesoderm is called
intremediate mesoderm (see the inset above, showing the cross‐section of the
chicken embryo), or also nephrotome, as it develops into tubules and ducts that
make up the kidney.
Some of these ducts will be co‐opted or shared with the developing reproductive
system.
The kidney consists of two major parts: tubules that filtrate liquid and small
molecules from the blood and ducts that reabsorb some of the filtered materials
and deliver the filtrate to the cloaca.
23
Tubuly pronefros
Vývod pronefros
(Wolfiiho vývod)
Tubuly
mesonefros
Vývod
mesonefros
Ureterové
divertikulum (pupen)
hagfishes (Agnatha) amphibians, fishes amniotes
sliznatky (bezčelistnatí)
Kulawik, 2005
Tubules differentiate in an anteriposterior axis, the more posterior, the more complex
structures develop.
In more simple organisms, e.g. in hagfishes (Agnatha) only the most primitive, i.e. most
anterior tubules develop, forming what is called pronephros.
During development of amphibians and fishes, the first developed pronephric‐type
anterior tubules degenerate and are replaced by more posterior and more complex
structures of mesonephros.
Finally, in amniotes, the anterior primimitive tubules act only during embryonic
development and the definitive kidney develop from a posterior located
metanephrogenic mesoderm forming metanephros.
Inset: Mesenchym of rabbit tongue during prenatal development. Image of median
cross‐section of the dorsum of the body of the tongue at day 15 of prenatal life in the
rabbit. Ep – epithelium, arrow – basement membrane, arrowhead – cell during mitosis,
Mz – mesenchyme. LM, x 40, Masson‐Goldner staining (Kulawik, 2005;
http://www.ejpau.media.pl/volume8/issue4/art‐17.html).
The image demonstrates the nature of mesenchyme as an undifferentiated loose
connective tissue that is derived mostly from mesoderm, although some is derived from
other germ layers; e.g. neural crest cells and thus originates from the ectoderm. Most
embryologists use the term mesenchyme only for those cells that develop from the
mesoderm (Wikipedia).
24
Gilbert, SF, Developmental biology
Intermediate mesoderm of a 13-day
mouse embryo
Pronephric duct stained
by fluorescently labelled
anti-cytokeratin Ab
General scheme of development in the vertebrate kidney. (A) The original
tubules, constituting the pronephros, are induced from the nephrogenic
mesenchyme by the pronephric duct as it migrates caudally. (B) As the
pronephros degenerates, the mesonephric tubules form. (C) The final
mammalian kidney, the metanephros, is induced by the ureteric bud, which
branches from the nephric duct. (D) The intermediate mesoderm of a 13‐day
mouse embryo showing the initiation of the metanephric kidney (bottom) while
the mesonephros is still apparent. The duct tissue is stained with a fluorescent
antibody to a cytokeratin found in the pronephric duct and its derivatives. (A‐C
after Saxén 1987; D courtesy of S. Vainio.)
25
Metanefrogenní
mesenchym
Metanepric
mesenchyme
Ureterové
divertikulum
Urereteric
diverticulum
Mesenhyme
aggregation
ectoderm-derived path cues
Wolffiho
vývod
The main duct formed during nephros formation is called Wolfian (pronephric)
duct. It elongates by migration mechanism via a path marked by cues from the
overlying ectoderm.
Near the connection of the duct to the cloaca, a bud diverts from the duct and
extends to the posterior metanephric mesenchyme, which is stimulated to
condense.
Wolfian duct persist only in males and degenerates in females (see further).
The developing kidney undergoes further development, see next slides.
26
Nefrogenní váček
Renal vesicle
A mesenchyme agregation
RV renal vesicle
U  uterus
CD collecting duct
27
Gilbert, SF, Developmental biology
Reciprocal induction in kidney development
Urereteric
diverticulum
induces
metaneprogenic
mesenchyme
aggregation
Metanephrogenic
mesechyme
induces bud to
branch
At the tip of
braches, the
epithelium of the
duct induces the
mesenchyme to
aggregate and
cavitate
Reciprocal induction in the development of the mammalian kidney. (A) As the 
ureteric bud enters the metanephrogenic mesenchyme, the mesenchyme 
induces the bud to branch. (B‐F) At the tips of the branches, the epithelium 
induces the mesenchyme to aggregate and cavitate to form the renal tubules and 
glomeruli (where the blood from the arteriole is filtered). When the mesenchyme 
has condensed into an epithelium, it digests the basement membrane of the 
ureteric bud cells that induced it and connects to the ureteric bud epithelium. 
The mesenchyme becomes the nephron (renal tubules and Bowman's capsule), 
while the ureteric bud becomes the collecting duct for the urine. (After Saxén 
1987.)
28
Reciprocal induction in kidney development
Gilbert, SF, Developmental biology
GDNF/GDNF GDNF/gdnf gdnf/gdnf
The role of glial-derived neurotrophic factor (GDNF)
WT1
Metanephrogenic mesenchyme
ability to respond to ureteric bud
The mechanisms of reciprocal induction
The induction of the metanephros can be viewed as a dialogue between the ureteric bud and the metanephrogenic mesenchyme. 
As the dialogue continues, both tissues are altered. There appear to be at least eight sets of signals operating in the reciprocal 
induction of the metanephros.
Step 1: Formation of the metanephrogenic mesenchyme
Only the metanephrogenic mesenchyme has the competence to respond to the ureteric bud to form kidney tubules, and if induced 
by other tissues (such as embryonic salivary gland or neural tube tissue), the metanephrogenic mesenchyme will respond by forming 
kidney tubules and no other structures (Saxén 1970; Sariola et al. 1982). Thus, the metanephrogenic mesenchyme cannot become 
any tissue other than nephrons. Its competence to respond to ureteric bud inducers is thought to be regulated by a transcription
factor called WT1, and if the metanephrogenic mesenchyme lacks this factor, the uninduced cells die (Kriedberg et al. 1993). In situ 
hybridization shows that WT1 is normally first expressed in the intermediate mesoderm prior to kidney formation and is then 
expressed in the developing kidney, gonad, and mesothelium (Pritchard‐Jones et al. 1990; van Heyningen et al. 1990; Armstrong et 
al. 1992). Although the metanephrogenic mesenchyme appears homogeneous, it may contain both mesodermally derived tissue 
and some cells of neural crest origin (Le Douarin and Tiellet 1974; Sariola et al. 1989; Sainio et al. 1994).
Step 2: The metanephrogenic mesenchyme secretes GDNF and HGF to induce and direct the ureteric bud
The second signal in kidney development is a set of diffusible molecules that cause the two ureteric buds to grow out from the 
nephric ducts. Recent research has shown that glial‐derived neurotrophic factor (GDNF) is a critical component of this signal. GDNF 
is synthesized in the metanephrogenic mesenchyme, and mice whose gdnf genes were knocked out died soon after birth from their 
lack of kidneys (Moore et al. 1996; Pichel et al. 1996; Sánchez et al. 1996). The GDNF receptor (the c‐Ret protein) is synthesized in 
the nephric ducts and later becomes concentrated in the growing ureteric buds (Figure 14.21; Schuchardt et al. 1996). Mice lacking 
the GDNF receptor also die of renal agenesis. Another protein synthesized by the metanephrogenic mesenchyme is hepatocyte 
growth factor (HGF; also known as scatter factor), and the receptor for HGF (the c‐met protein) is made by the ureteric buds. 
Antibodies to HGF will block ureteric bud outgrowth in cultured kidney rudiments (Santos et al. 1994; Woolf et al. 1995). The 
synthesis of GDNF and HGF by the mesenchyme is thought to be regulated by the WT1 protein.
The image: Ureteric bud growth is dependent on GDNF and its receptor. (A) The ureteric bud from a 11.5‐day wild‐type mouse 
embryonic kidney cultured for 72 hours has a characteristic branching pattern. (B) In embryonic mice heterozygous for the genes 
encoding GDNF, the size of the ureteric bud and the number and length of its branches are reduced. (C) In mouse embryos missing 
both copies of the GDNF gene, the ureteric bud does not form. (Scale bars = 100 μm.) (D) The receptors for GDNF are concentrated 
in the posterior portion of the nephric duct. GDNF secreted by the metanephrogenic mesenchyme stimulates the growth of the 
ureteric bud from this duct. At later stages, the GDNF receptor is found exclusively at the tips of the ureteric buds. (A‐C from Pichel
et al. 1996, photographs courtesy of J. G. Pichel and H. Sariola; D after Schuchardt et al. 1996.)
Further step follow, i.e. the ureteric bud secretes FGF2 and BMP7 to prevent mesenchymal apoptosis and LIF from the ureteric bud 
induces the mesenchyme cells to aggregate and to secrete wnt4 (for further reading see 
http://www.ncbi.nlm.nih.gov/books/NBK10089/). 
29
Primární sběrací kanálek
(vzniká větvením ureterického
divertikula)
Metanefrogenní mesenchym
Nefrogenní váček
Esovitý kanálek
Henle loop
Henleyova klička
Proximální tubulus
Bowmanův váček
Kapiláry glomerulu
Vacek, Embryologie (2006)
30
Outline of Lesson 5
Organogenesis in Vertebrates: Endo- and Mesodermal
Derivatives
 Mesoderm derivatives development
 somites formation and signalling
 formation of muscles
 endochordal ossification and signalling
 nephrogenesis
 formation of gonads
31
Můllerův vývod
Wolffiho vývod
Kloaka
Močovod
Nadvarle Nálevka vejcovodu
Děloha
Močová trubiceMočový měchýř
In paralel with the Wolfian duct, another duct, what is called Műlerian duct is
formed.
In males, the Műllerian duct degenerates due to anti‐Műllerian duct factor
production that induces apoptosis of its cells.
In females, the Műllerian duct persists to form the oviduct, the uterus and part of
the cervix.
32
Genitální lišta (plica genitalis)
Kanálek mesonefros
Medulární provazce
Gonads form from the so called genital ridge area.
The coelomic epithelium in the genital ridge region prolifereates to form sex
cords, which penetrate to the mesoderm and transiently associates with
mesonephros and both Mullerian and Wolfian ducts.
At that time, the primordial germ cells (PGCs) migrate from elsewhere (in
different vertebrates, the origin of the PGCs is different, e.g. in amphibians PGCs
arise in the endoderm). In humans, PGS originate from the yolk sac (see the next
slide).
Upon the moment of primitive sex cord formation, the further development of
males and females differs.
33
Yolk sac
Žloutkový váček
Perikard cavity
Perikadová dutina
Amnion cavity
Amniová dutina
Hindgut
Zaddní střevo
Body stalk
Zárodečný stvol
Allantois
Primordial germ cells
Prvopohlavní buňky
Allantois
Zaddní střevo
Genital ridge
Genitální lišta
Mesonephric
duct
Vývod
mesonephros
PGCs migrate along
the dorsal
mesenterium into
the hindgut and
from that further to
the genital ridge
Prvopohlavní buňky
migrující podél
dorzálního
mezentéria do
zadního střeva a
odtud do gen. lišty
Vacek, Embryologie (2006)
20-days-old human embryo 26-days-old human embryo
20 days old human embryo with PGC in the yolk sac (A, left).
Ventrolateral view into the trunk (coelom) cavity of the 3 mm large, 26 days old
human embryo (9B, right), showing the PGCs migrating to the genital ridge.
34
Continuous growth
and seminiferous
tubules formation
Testikulární
provazce
In males, the coelomic epithelium (sex cords) continues proliferation, becoming 
the seminiferous tubules.
35
Testikulární
provazce
Anti-Mullerian
duct factor
production by
differentiated
Serrtori cells
Produkce
Műllerovy inhibiční
látky
Ductuli
efferentes
Serrtori cells differentiate in the tubules and secrete the anti‐Műllerian duct
factor.
PGCs come to inhabit the tubules (cords), forming thus spermatozoa,
mesenchyme of the genital ridge becomes intersticial tissue of testis.
36
Ductuli
efferentes
Nadvarle
seminiferous tubules
The seminiferous tubules connect to the Wolfian duct, which becomes vas 
deferens of the male reproductive system.
37
Kortikální provazce
Degenerující
medulární provazce
In female genital ridge, the first epithelial cords degenerates and the secondary 
cords from the coelomic epithelium proliferate.
38
Primordiální folikuly
These secondary epithelial cords remain located near the surface and form the
prospective follicle cells, in which the PGCs reside and form oocytes.
The Wolfian ducts degenerates while Műllerian ducts develop into oviducts.
39
Vaječník
Vejcovod Děloha
Děložní čípek
40
Outline of Lesson 5
Organogenesis in Vertebrates: Endo- and Mesodermal
Derivatives
 Mesoderm derivatives development
 somites formation and signalling
 formation of muscles
 endochordal ossification and signalling
 nephrogenesis
 formation of gonads
 hematopoesis and circular system development
41
Somatický
mezoderm
Splanchnický
mezoderm
Střední
mezoderm
Krevní ostrůvky
Blood islands
inner cell layer of blood vessels
Hematopoesis (blood formation) is a stepwise process. The hematopoesis differs during development in terms of the location in the developing
embryo and differs also among animal groups.
In birds and mammals, the earliest blood vessels and erythrocytes form in the splanchnic mesoderm, surrounding the yolk sac, thus outside the
embryo proper. In the figure above (top left), there are shown blood islands forming in the splanchnopleura of chicken embryo.
Similarly to other mesodermal structures discussed previously, one of the first steps is mesodermal cell aggregation, leading to what is called blood
islands formation.
In the centres of the blood islands, the cells differentiate into erythrocytes while the surrounding cells form endothelium, i.e. the inner cell layer of
blood vessels. Note a certain parallel with osteocytes formation discussed before.
In embryonal mesoderm, similar process occurs leading to the formation of endothelial tubes arising from mesodermal agregations.
The larger tubes, prospective arterioles and venules‐will be surrounded by smooth muscle layers.
During early development most of the erytrocytes originates in the yolk sac while later on, the endothelial lining of the dorsal aorta and nearby blood
vessels will proliferate, to provide primitive additional erythrocytes and stem cells.
The location of hematopoesis changes further during development: The stem cells formed from the lining of dorsal aorta populate liver, which is the
place of blood formation of fetus until late in gestation. The final place of erythropoesis of adults becomes bone marrow populated by the stem cells.
Lymphocytes appear in the second half of the embryonic development of mammals.
The quality of the hemoglobin also changes during development (see next slide).
Important process of vessel system development represents the angiogenic sprouting, i.e. formation of new vessels.
During embryogenesis, this process occurs via so called vasculogenesis, where most of the blood vessels develop by in situ formation of endothelial
tubes.
Some of the vessels evolve thanks to invagination of existing endotehlium into surrounding tissue, a kind of invasion process called angiogenesis.
The process of angiogenesis is very important also in the adult age, where new vessels are formed e.g. after injury or tumors vascularization. Vascular
endothelial growth factor (VEGF) and angiopoetin are recently known factors involved in angiogenesis.
Interestingly, arteries, but not veins, in embryo often grow along nerves paths. Recent experiments show that nerves or surrounding Schwann cells
stimulate the growth of blood vessels and their differentiation into arteries.
42
Embryonic hemoglobin εεζζ
yolk sac
Fetal hemoglobin ααγγ
liver
Adult hemoglobin ααββ
bone marrow
The yolk sac erythrocytes produce so called embryonic form of α and β globin,
called ζ (zeta) and ε (epsilon), while later erythrocytes from the embryonic liver
contain adult α chains but fetal form of β globin, called γ (gamma).
Finally, the bone marrow produces ααββ tetramer of globins, forming the
peptide portion of the hemoglobin in adults.
43
Amniokardiakální
váček
Primordiální epimyocard
Primordiální endokardiální buňky
Epimyocard
(myoepikardový plášť)
Endokardiální trubice
Anteriorní intestinální portál
migration through
the Hensen’s node
Cells forming the heart migrate through the Hansen’s node and it is not clear up
to know, whether they aggregate in the future heart or if they just differentiate
there.
Heart forms as an endothelium in the splanchnic layer of the anterior ventral
mesoderm. Here, the islands of mesodermal cells form endothelium that
develops into a specialized muscular layer called myocardium.
Myocardium develops from the epimyocardium, an undifferentiated splanchnic
mesodermal layer of the embryonic heart that subsequently differentiates into
myocardium and epicardium.
The muscle fibres of myocardium undergo unique developmental pathway and
the tube forms heart.
In the figures, the ventral view is on the left‐hand side (A) while cross section is
on the right‐hand side (B).
44
Přední
střevo
Perikardiální dutina
Epimyokard
Endokardiální trubice
In amniotes, heart develops originally as a pair of endocardial tubes that fuse at 
the end, forming the single tube. 
45
Bulbus cordis
Right ventricle
Pravá komora
Left ventricle
Levá komora
Atrium
Síň
Sinus
venosus
migration of endocardial cells into the matrix between
the endocardium and outer myocardial cells chambers and
septa formation
Bulbus cordis
The heart starts functioning as a single tube, with blood entering posteriorly
through the sinus venosus, then moving through the atrium and ventricle and
finally passing anteriorly out from the bulbus cordis.
As the heart develops, it undergo a series of turning, forming a large loop to the
right side and then curls over itself, leading to the final shaping of the complex
heart structure.
Thus, what was originally anterior ventricular portions, turns to be posterior.
Internal heart structures (chambers and septa) develop via migration of
endocardial cells into the matrix that lies between the endocardium and outer
myocardial cells.
46
47
Aorta descendens
Pravá podklíčková tepna
Levá podklíčková tepna
Levá plícní tepna
Pravá plícní tepna
Oblouk krkavice Levý oblouk krkavice
Arteria carotis interna
(vnitřní krkavice)
Arteria carotis externa
(zevni krkavice)
entry from bulbus
cordis
Oblouk aorty
Hlavopažní kmen
pharyngeal (or branchial) arches
pharyngeal pouches
gills
aortic
arches
Circular system remodeling in mammals
Early development of the vertebrate vessel system is common, leading to the basic plumbing, which,
however, is remodeled during later development specifically in respective group of vertebrates.
In the region of prospective head and neck, the ridges form that are called pharyngeal (or branchial)
arches. Between the arches, the ectoderm and endoderm are closely apposed and ectoderm invaginates
forming thus pharyngeal pouches. In aquatic, nonamniote vertebrates these structures perforate and
develop into gills.
Within the mesoderm of pharyngeal arches will develop vessels forming six pairs of aortic arches. These
arches form in aquatic nonamniote vertebrates the capilary bed of the gills.
In the original anatomical arrangement, all blood is pumped out of the heart via bulbus cordis that
connects to the ventral aorta. From there, the blood goes via aortic arches back to the descending aorta
(see the figure A).
In amniotes, the above described anatomical arrangement gets extensively remodeled: Some of the aortic
arches atrophy while some develop into major arteries of the head and neck.
In mammals, the first two and the fifth arch atrophy.
The third arch together with anterior part of the ventral aorta develops into internal and external carotid
arteries, the major arteries of the head.
The fourth arch forms dorsal aorta, the major source of the blood in the posterior portion of the embryo.
From the dorsal aorta, several branches develop that supply kidney, digestive tract, yolk sac and so on.
Also the subclavian artery develops from the right fourth arch that supplies blood to the arterior limbs.
The sixth arch gives rise to pulmonary arteries to the lungs.
48
SíňKomora
1. Aortální oblouk
Aorta descendens
(sestupná aorta)
Anterior cardinal vein
Vena precardinalis
(přední kardinální žíla)
Vena postcardinalis
(zadní kardinální žíla)
Vena omphalomesenterica Arteria omphalomesenterica
V. cardinalis
communis
Žilný splav
Blood streaming in the initial circular system in amniote embryo
Yolk sac
Streaming of the blood in the initial system of stylized amniote vertebrates
embryo.
Blood is collected from capillary beds in all nascent tissues, empties into the
common cardinal vein and then into the entry chamber of the heart, the sinus
venosus.
Omphalomesenteric vein collects the blood from the yolk sac, while
omphalomesenteric artery brings the blood there.
Then the blood enters chambers of the atrium, passes to the ventricle and moves
out through the bulbus cordis once again.
49
V. cardinalis communis
Vena postcardinalis
(zadní kardinální žíla)
Vena precardinalis
(přední kardinální žíla)
Arteria carotis interna
Vena renalis
Vena portae
(vrátnicová žíla)
Ocasní tepna
a žílaVena omphalomesenterica
Arteria omphalomesenterica
4. Aortální oblouk
Blood streaming in the remodeled circular system in amniote embryo
In the later remodeled system, the venous blood gets diverted through the liver
on its way to the common cardial vein and back to the heart.
Extensive remodeling occurs including dramatic changes at the birth, when the
lungs start to respire and the yolk sac or placenta are out of function.
50
Outline of Lesson 5
Organogenesis in Vertebrates: Endo- and Mesodermal
Derivatives
 Mesoderm derivatives development
 somites formation and signalling
 formation of muscles
 endochordal ossification and signalling
 nephrogenesis
 formation of gonads
 hematopoesis and circular system development
 limbs formation
51
Střední mozek
Midbrain
Oko
Eye
Neurální trubice
Neural tube
Křídelní pupen
Wing bud
Nožní pupen
Leg bud
Somity
Somites
Ocas
Tail
Apikální ektodermová lišta
bud positioning via HOX genes
Mesenchymal aggregations
Limbs in vertebrates undergo specific developmental modifications, dependent
on the final limb type, e.g. wings (different in beds and birds), fins, flippers,
hooves etc..
Limb develop from the somatic mesoderm, where the what is called limb buds
develop, both anteriorly and posteriorly. The positioning of the limb buds is
determined by HOX genes (will be discussed later).
One of the most important structures that determines the proximodistal axis
patterning of the developing limb is so called apical ectodermal ridge (AER). The
role of AER will be discussed later.
As the bud grows, again, the mesenchymal aggregations that will be a precursors
of the cartilage appear. Ectoderm and mesoderm will differentiate into skin and
surface structures (e.g. feathers or hairs).
52
Limbs show complex organization in all (i.e. proximodistal, anteroposterior and
dorsoventral axes).
As an example, the major skeletal elements of the chick embryo is shown here.
53
Apoptosis
The tissue between the developing skeletal elements of digits might persist to
form webbing or undergoes apoptosis. The figure shows developmental stages
32‐35 (of chicken embryo, see slide 57), which correspond to 7.5‐8.5 days of
incubation.
54
Vertebrae and
proximal ribs
Long bones
The transplantation experiments using Japanese quail and chicken embryos have
provided important experimental evidence about the origin of cells involved in
the limb formation.
The nuclei of quail embryo might be easily distinguished by specific staining‐they
stain more intensely.
These experiments have shown that not all cells in the limb originate from lateral
(somatic) mesoderm. When the chicken somites 15‐20 were replaced with quail
ones, the striated muscles of developed limbs had muscle cells that originated
from quail somites.
55
AER
7 rounds of mitosis
Proximodistal limb patterning
As mentioned previously, AER plays a very important role in the proximodistal
patterning of the developing limbs. The proximodistal pattern arise in the
mitotically active region below the AER.
7 rounds of mitosis leading to the formation of successive cell layers is necessary
to complete the final proximodistal limb patterning. The later the AER is removed
before the process is completed, limbs that are defective in the more distal limb
structures will develop.
Numbers in the figure indicate the stage of development, when AER was
removed and the lines depict the level of limb truncation.
Besides AER, there are two additional important organizing centers. These are i)
the surrounding ectoderm and ii) the what is called zone of polarizing activity
(ZPA).
ZPA is located in the posterior mesoderm of the bud and its role will be discussed
later.
56
Hamburger & Hamilton Stage 4 (15 hours)
HH Stage 9 (31 hours 8 somite)
HH Stage 10 (33 hours)
HH Stage 16-17 (56 hours) HH Stage 38 (12 days)
Chicken embryo as a developmental model
Stages of chick development according to Hamburger & Hamilton, Series of
Embryonic Chicken Growth. J. Morphology, 88 49 ‐ 92 (1951).
The Hanburger and Hamilton dissected chicken development according to
different morphologically identifiable traits.
57
Outline of Lesson 5
Organogenesis in Vertebrates: Endo- and Mesodermal
Derivatives
 Mesoderm derivatives development
 somites formation and signalling
 formation of muscles
 endochordal ossification and signalling
 nephrogenesis
 formation of gonads
 hematopoesis and circular system development
 limbs formation
 Endoderm derivatives development
 alimentary canal and its derivatives formation
58
Slinné žlázy
Brzlík Příštitná tělíska
Štiíná žláza
Plíce
Játra
Žlučník
Řitní otvor
Slinivka břišní
Žaludek
Rathkeova výchlipka
Žlučník
Žlučník
Struna hřbetní
Jícen
Játra
Žaludek
Rektum
Alimentary canal
Amphibians
Amniotes
Rectal portion
communicates
Communication
only with yolk sac
Epithelial lining of the digestive system
Head mesoderm Splanchnic mesoderm
Endoderm forms what is called alimentary canal, i.e. the epithelial lining of the
digestive system.
Development of the connection of the digestive system to the surrounding
environment is specific for respective vertebrates groups.
E.g. in amphibians, the rectal portion of the digestive tube communicates with
the outside, while the oral opening forms later. In comparison, amniotes have
both ends blind with the midgut connected with the yolk sac and both cloacal
and oral opening form later.
The epithelium lining in concert with local surrounding of mesenchyme forms
outpocketings from the alimentary canal that develop into secretory and
endocrine glands of the head and neck, lungs, liver, gallbladder, and the
pancreas.
It is surrounded by the mesoderm – head mesoderm anteriorly, splanchnic
mesoderm in the rest of the body. Reciprocal interactions between the
surrounding mesoderm and endoderm participate in the final development of all
of the endodermal structures.
59
Rathkeova
výchlipka
Contact with diencephalon
Antherior pituitary
formation
Hypofýza
Pharyngeal arch-specific endoderm derivatives development
Eustachova trubice
Eustachian tube
Krční mandle
Palatine tonsil
Horní a dolní čelist
Maxillary and mandibular jaws
oral plate
As mentioned previously, all vertebrates form the so called arches in the wall of pharynx.
Endoderm will develop into many important structures here in an arch‐specific manner.
At the anterior end, the archenteron fuses with ectoderm and forms oral plate that later perforates and forms mouth.
Posterior to the oral plate is an inpocketing of ectoderm called Rathke’s pouch, which later contacts the
diencephalon. Diencephalon induces Rathke’s pouch to form the anterior pituitary.
In the region of individual arches, the endoderm cells associates with arches, prolifreates and invaginate in
association with the adjacent mesenchyme.
In aquatic vertebrates, gills develop from the arches and gill slits from the individual clefts of the pharynx.
However, in terrestrial vertebrates, the arches are only transient structures. However, the pouches between the
arches give rise to the set of important organs.
Pouch I forms the ustachian tube.
Pouch II the palastine tonsil.
The floor area of pouch I and II develop into the thyroid.
Pouches III and IV give rise to portions of the thymus and parathyroid gland.
Salivary glands develop also from pharyngeal endoderm. Interaction of the salivary endoderm and surrounding
salivary mesenchyme results into complicated branching of the salivary glands.
In a position of brancial archs I and II upper (maxillary) and lower (mandibular) jaws are formed. Teeth formation is a
result of mutual interactions between the mesenchyme and epithelium. Teeth are of ectodermal origin (progeny of
neural crest cells).
60
Hltan
Průdušnicová
rýha
Mesenchyme
interactions
Primární
průduškové
pupeny
Sekundární
průduškové
pupeny
Boční
průdušky Průdušnice
Lungs develop as an invagination that forms from the alimentary canal just
posteriorly from the pharynx.
Trachea develops from the pharynx evagination. Trachea then further branches
and forms bronchi and alveoli at their termini.
Again, similarly to the salinary glands branching, interaction of the endodermal
epithelium with the surrounding mesenchyme drives the branching and some of
the genes involved in the process were recently identified.
61
Střevo
Pojivová
tkáň
Acinus
(lalůček)
Krevní
cévy
Langerhansovy
ostrůvky
Pancreatic
diverticulum
•Glucagon
•Insulin
•Somatostatin
•Pancreatic peptide
•Zymogens production
•Precursors of peptidaeses, nucleases
and carbohydrate-hydrolyzing enzymes
Slinivka břišní
Posterior to the esophagus, the endodermal epitehlium develops into specialized cell
types that will form stomach.
Endodermal outpocketings of the epithelium in this region will result into liver,
gallbladder and pancreas formation.
These organs start to develop via evaginations into surrounding mesodermal
mesenchyme, called diverticula.
Liver start to develop as a hepatic diverticulum, that proliferates, branch and
differentiates as a glandular part of the future liver. Its base forms hepatic duct and
branch forms gallbladder.
Pancreas develops posteriorly from the dorsal and ventral pancreatic diverticula, which
introgress into the mesodermal mesencyme and again interact with it.
The pancreatic diverticula branch and form blind pockets, called pancreatic acini. Acini
will differentiate into exocrine cells that will produce zymogens, i.e. precursors of
peptidases, nucleases and carbohydrate‐hydrolyzing enzymes.
Some of the epithelial endodermal cells aggregate and develop into endocrine islets of
Langerhans cells. These islets contain at least four cell types and each of them will
produce different pancreatic hormones: glucagon, insulin, somatostatin or pancreatic
peptide.
62
Key Concepts
 Developmental potential is studied via e.g. transplantation experiments,,
while developmental fate is best studied using modern molecular techniques
in a normal embryo.
 Vertebrate embryos have a conservative axial organization proceeding from
the midline to the periphery: notochord, somite, nephrotome, or gonad and
lateral mesoderm (limb buds).
 Iterative oscillations of the gene expression in a combination with gradient of
gene expression allow proper body segmentation during embryonic
development. Underlying molecular mechanisms are at least partially
conserved in Drosophila and vertebrates.
 Normal tissue and organ formation depends not only on the presence of
signalling molecules, but also on the appropriate timing of ligand-receptor
interactions. Such communications can establish feed-back loops, as
demonstrated e.g. in case of cartilage and bone formation.
 Cell aggregation/disaggregation is one of the key developmental
mechanisms guiding new organ formation.
 Some structures are transient during development, e.g. pronephros, some,
e.g. circular system are extensively remodelled during development.
63