UNSW Embryology
Musculoskeletal - Limb Development
© Dr Mark Hill (2008)
Introduction
Somites form and then begin to differentiate forming sclerotome,
dermomyotome and then dermatome and myotome. The lateral portion of the
myotome edge migrates at level of limbs (upper limb first then lower) and mixes
with somatic mesoderm. Meanwhile the dermotome continues to contribute cells to myotome.
Limb development occurs at different
times for forelimbs and hindlimbs. In the
mid-4th week, human upper limb buds first
form and lower limbs about 2 days later.
The limbs form at vertebra segmental levels C5-C8 (upper limbs) L3-L5 (lower limbs).

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Upper Limb (Stage 23) |
Changes in External Appearance of Upper Limb (embryonic stages, not to scale. More? Upper Limb Movie) |
Page Links: Introduction | Some Recent Findings |
Appendicular Skeleton | Limb Axes |
Hands and Feet | Limb Ossification |
Molecular Development | Carnegie Stage - Upper Limb |
Limb Movies | Somite Development |
Limb Development Abnormalities | References
| Glossary
Other Pages: Stage 13/14 Limbbuds | Stage 22 | Stage 22 Upper Limb | Stage 22 Lower Limb
Some Recent Findings
Sun M, Ma F, Zeng X, Liu Q, Zhao XL, Wu FX, Wu GP, Zhang ZF, Gu B, Zhao YF, Tian SH, Lin B, Kong XY, Zhang XL, Yang W, Lo W, Zhang X.
Triphalangeal thumb-polysyndactyly syndrome and syndactyly type IV are caused by genomic duplications involving the long-range, limb-specific SHH enhancer.
J Med Genet. 2008 Apr 16;
Nissim S, Allard P, Bandyopadhyay A, Harfe BD, Tabin CJ.
Characterization of a novel ectodermal signaling center regulating Tbx2 and Shh in the vertebrate limb. Dev Biol. 2007 Apr 1;304(1):9-21
"The data presented here identify the non-AER border of dorsal-ventral ectoderm as a new signaling center in limb development that localizes the ZPA to the limb margin.
This finding explains the tight restriction of Shh expression to the posterior margin throughout limb outgrowth as well as the tight restriction of Shh expression to the anterior margin in
many mutants exhibiting preaxial polydactyly."
Nissim S, Hasso SM, Fallon JF, Tabin CJ.
Regulation of Gremlin expression in the posterior limb bud. Dev Biol. 2006 Nov 1;299(1):12-21. (More? OMIM - Gremlin)
"Proper outgrowth of the limb bud requires a positive feedback loop between Sonic hedgehog (Shh) in the zone of polarizing activity (ZPA) and Fgfs in the overlying apical ectodermal ridge.
The Bmp antagonist Gremlin is expressed in a domain anterior to the ZPA and is thought to act as a signaling intermediate between Shh and Fgf."
Appendicular Skeleton
The appendicular skeleton consists of: Shoulder girdle, Upper limb (arm, hand), Pelvic girdle, Lower limb (leg, foot).
Limb Axes
The limb is initially mesenchyme with an ectodermal covering. Within this mesenchyme blood vessels begin forming, the largest (marginal vein) is adjacent to the tip of the limbbud.
At the tip of the limbbud lies a
thickened ridge of ectoderm, the apical
ectodermal ridge (AER), which has a role
in patterning the structures that form
within the limb. The majority of cell
division (mitosis) occurs just deep to AER
and this region is known as the progress
zone. Another region at the base of the
limbbud beside the body, the zone of
polarizing activity (ZPA) has a similar
patterning role to the AER, but in
determining another axis of the limb.
(More? Molecular Development).
Hands and Feet
In the 5th week hand and footplates
appear at the ends of limb buds and ridges
form digital rays. Cells between the
digital rays are removed by programmed
cell death (apoptosis).
Late in Carnegie stage embryogenesis
(Stage 20-23, 8th week) limb rotation
occurs. Forelimbs and hindlimbs rotate in
different directions, upper limb rotates
dorsally, lower limb rotates ventrally,
thumb and toe rostral, knee and elbow face
outward.
Limb Ossification
Bones within the limb form by
endochondrial ossification (begins
Carnegie stage 18) throughout embryo. This
process is the replacement of cartilage
with bone (week 5-12).
For more information see Bone Development
Molecular Development
There is another page with an overview of Molecular Development of Musculoskeletal structures and for more
specific information about factors involved in limb patterning look at the Molecular Development Page or use the links below.
Limb Initiation
- FGF beads can induce additional limb
formation (More?
see factors- FGF)
- FGF10 , FGF8 (lateral plate intermediate
mesoderm)
- prior to bud formation
- FGF8 (limb ectoderm) FGFR2
- FGF can respecify Hox gene expression (Hox9-
limb position)
- Hox could activate FGF expression
Limb Specification (Fore- Hind-)
Limb Patterning- Axes
- Wing has been used as Model of limb
development as chick wing easy to manipulate:
removal, grafting, additional ARER, ZPA etc
Limb Patterning- Axes
- Proximodistal Axis
- Dorsoventral Axis
- somite provides dorsal signal to
mesenchyme
- which dorsalizes ectoderm
- ectoderm then signals back (Wnt7a) to
mesenchyme to pattern limb
- Anteroposterior Axis
- ZPA zone of polarizing
activity
- mesenchymal posterior region of limb
- addition of extra ZPA duplicated
digits
- signal is Shh (More?
see factors- SHH)
- Limb Axes
- Limb Patterning- Axes (More?
see Limb Axes)
- Signals give positional information which
is interpreted by Hox gene expression
establishing programs of differentiation.
(More?
see factors- Hox)
- Proximodistal Axis
- Dorsoventral Axis
- Anteroposterior Axis
Carnegie Stage
Carnegie stage 12 to 23 Human Forelimb Development
Limb Movies
Below are links to movies generated from the above Carnegie images. Note that the series of movie images are not to scale as
the early images would be far too small to see any detail.
Arm Movie Page or
Movie on new page (179Kb)
Somite Development
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Mesoderm beside the notochord (axial
mesoderm) thickens, forming the paraxial mesoderm
as a pair of strips along the rostro-caudal axis.
Paraxial mesoderm towards the rostral end,
begins to segment forming the first somite.
Somites are then sequentially added
caudally. The somitocoel, is a cavity
forming in early somites, which is lost as the
somite matures.
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Cells in the somite differentiate medially to
form the sclerotome (forms vertebral column) and
laterally to form the dermomyotome. The
dermomyotome then forms the dermotome (forms
dermis) and myotome (forms muscle).
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Pig Embryo Serial Sections
(st13/14)
Serial section D3 (rotated 90 degrees)
Mouseover to identify structures
see also links below
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The myotome differentiates to form 2 components
dorsally the epimere and ventrally the hypomere,
which in turn form epaxial and hypaxial muscles
respectively. The bulk of the trunk and limb muscle
coming from the Hypaxial mesoderm. Different
structures will be contributed depending upon the
somite level.
Note these are very simplified figures.
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Limb Abnormalities
Limb Abnormalities both genetic and environmentally derived and now covered in more detail on a page Limb Abnormalities).
References
Nissim S, Allard P, Bandyopadhyay A, Harfe BD, Tabin CJ.
Characterization of a novel ectodermal signaling center regulating Tbx2 and Shh in the vertebrate limb. Dev Biol. 2006 Dec 9;
McKee PR, et al. [See
Related Articles] Skeletal relationships in the human embryonic foot based on three-dimensional reconstructions. Acta Anat (Basel). 1987;129(1):34-42.
Search PubMed: Feb 2007 "limb bud development" 1,264 reference articles of which 132 were reviews.
Search PubMed Now:
limb bud development |
limb development |
musculoskeletal development
(More? PubMed- Medline)
Selected Lists of References from PubMed March 1999 search results are available for Department of Anatomy computers without internet
access: Somite Reviews | Somitogenesis Abstracts | Mesoderm Review List
Computers with internet access can search from either Below or directly from PubMed Internet Access
Glossary
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Terms
- anlage- (Ger. ) primordium,
structure or cells which will form a future
structure.
- apical ectodermal ridge-
(=AER) specialized region of ectoderm at the tip of
the growing limbbuds that specifies proximo/distal axss
of limb development
- aponeurosis- thin flat
sheet tendon.
- apoptosis- the process
of programmed cell death. In development of the limbs
occurs in the "paddle" if both the hand and foot,
generating the separated digits. Occurs in many tissues
of the embryo and adult.
- axillary fossa-
the future "armpit" region
- brachial plexus-
mixed spinal nerves innervating the upper limb form a
complex meshwork (crossing).
- cartilage- connective
tissue from mesoderm in the embryo forms initial skeleton
replaced by bone. In adult, found on surface of bone
joints.
- dermatome-
- dermomyotome- dorsolateral half of each somite that
forms the dermis and muscle.
- dorsal root ganglia- (=spinal
ganglia) sensory ganglia derived from the neural crest
lying laterally paired and dorsally to the spinal cord
(in the embryo found ventral to the spinal cord).
Connects centrally with the dorsal horn of the spinal
cord.
- dura mater-
- dystrophy-
- ectoderm- the layer (of
the 3 germ cell layers) which form the nervous system
from the neural tube and neural crest and also generates
the epithelia covering the embryo.
- ectodermal ring-
the thickened ring of ectoderm seen dorsally in the early
(stage13/14) embryo adjacent to the dermatome. Ectoderm
ventrally is relatively thin, gaining its dermatome
component at a later stage.
- endochondrial
ossification- the process of replacement of the
cartilagenous framework by osteoblasts with bone.
- epaxial myotome- the
dorsal portion of the myotome that generates dorsal
skeletal muscles (epaxial muscles).
- extracellular matrix- material
secreted by and surrounding cells. Consists if fibers and
ground substance.
- extensor-
- fascia-
- fascicle- (=bundle)
- femur-
- fibroblast growth factors-
(FGF) a family of at least 10 secreted proteins that bind
membrane tyrosine kinase receptors. A patterning switch
with many different roles in different tissues. (FGF8 =
androgen-induced growth factor (AIGF)
- fibroblast growth factor
receptor- receptors comprise a family of at least 4
related but individually distinct tyrosine kinase
receptors (FGFR1- 4). They have a similar protein
structure, with 3 immunoglobulin-like domains in the
extracellular region, a single membrane spanning segment,
and a cytoplasmic tyrosine kinase domain.
- flexor-
- fossa-
- growth factor-
usually a protein or peptide that will bind a cell
membrane receptor and then activates an intracellular
signaling pathway. The function of the pathway will be to
alter the cell directly or indirectly by changing gene
expression. (eg shh)
- hox- (=homeobox)
family of transcription factors that bind DNA and
activate gene expression. Expression of different Hox
genes along neural tube defines rostral-caudal axis and
segmental levels.
- humerus-
- hypaxial myotome- the
ventral portion of the myotome that generates ventral
skeletal muscles (hypaxial muscles).
- inguinal fossa-
the region of the lower limb ajacent to flexor surface
(exuivilant to the axillary fossa of the upper
limb).
- lumbar plexus-
mixed spinal nerves innervating the lower limb form a
complex meshwork (crossing).
- mesenchyme-
- mesoderm- the middle
layer of the 3 germ cell layers of the embryo. Mesoderm
outside the embryo and covering the amnion, yolk and
chorion sacs is extraembryonic mesoderm.
- metacarpal
cartilage-
- muscle- 3 main types of
muscle (smooth, cardiac and skeletal) all derived from
mesoderm but different regions.
- myotome-
- myoblast- the
undifferentiated mononucleated muscle cells that will
fuse together to form a multinucleated myotube, then
mature into a muscle fibre.
- MyoD- transcription factor
involved in the determination of muscle cells in the
somite. A basic helix-loop-helix factor which binds
DNA.
- myotome- the portion of
the dermamyotome that generates skeletal muscle. Has 2
components epaxial (dorsal muscles ) hypaxial (ventral
muscles).
- neural crest- cell
region at edge of neural plate, then atop the neural
folds, that remains outside and initially dorsal to the
neural tube when it forms. These paired dorsal lateral
streaks of cells migrate throughout the embryo and can
differentiate into many different cell
types(=pluripotential). Those that remain on the dorsal
neural tube form the sensory spinal ganglia (DRG). Neural
crest cells migrate into the somites.
- patella-
- Pax- name derived from
Drosophila gene 'paired' (prd) the 'paired box' is a
amino end 124 amino-acid conserved domain (signature aa
35-51: P-C-x(11)-C-V-S). Transcription factor of
the helix-turn-helix structural family, DNA binding, and
activating gene expression. In human, nine member
proteins from Pax-1 to Pax-9. Regulate differentiation of
many different tissues. Some members of the family (PAX3,
PAX4, PAX6, PAX7) also contain a functional homeobox
domain.
- sclerotome-
ventromedial half of each somite that forms the vertebral
body and intervertebral disc.
- segmentation- to
break a solid structure into a number of usually equal
size pieces.
- spinal nerve- mixed
nerve (motor and sensory) arising as lateral pairs at
each vertebral segmental level.
- somatic
mesoderm- derived from lateral mesoderm closest to
the ectoderm and separated from other component of
lateral mesoderm (splanchnic, near endoderm) by the
intraembryonic coelom.
- somite- segmental block
(ball) of mesoderm formed from paraxial mesoderm adjacent
to notochord (axial mesoderm). Differentiates to form
initially sclerotome and dermamyotome (then dermotome and
myotome).
- somitic
mesoderm-
- somitocoel- a transient
cavity that appears within each of the the early forming
somites then is lost.
- somitogenesis- the
process of segmentation of the paraxial mesoderm to form
"mesoderm balls" beginning cranially (humans day20) and
extending caudally at 1 somite/90 minutes until approx.
44 pairs have been formed.
- sonic hedgehog- (=shh)
secreted growth factor that binds patched (ptc) receptor
on cell membrane. SHH function is different for different
tissues in the embryo. In the nervous system, it is
secreted by the notochord, ventralizes the neural tube,
inducing the floor plate and motor neurons. In the Limb
it is secreted by the zone of polarizing activity (ZPA)
organizing limb axis formation.
- syndactyly- fusion of
digits.
- tarsal-
- Tbx- T-box genes
(transcription factor) involved in mouse forelimb (Tbx4)
and hindlimb (Tbx5) specification.
- tibia-
- transcription
factor- a factor (protein or protein with steroid)
that binds to DNA to alter gene expression, usually to
activate. (eg steroid hormone+receptor, Retinoic
acid+Receptor, Hox, Pax, Lim, Nkx-2.2).
- Wnt7a- The designation 'Wnt'
was derived from 'wingless' and 'int'. The Wnt gene was
first defined as a protooncogene, int1. Humans have at
least 4 Wnt genes: Wnt7a gene is at 3p25 encoding a 349aa
secreted glycoprotein. A patterning switch with different
roles in different tissues. The mechanism of Wnt
distribution (free diffusion, restricted diffusion and
active transport) and all its possible cell receptors are
still being determined. At least one WNT receptor is
Frizzled (FZD). The Frizzled gene family encodes a
seven-transmembrane receptor.
- zone of polarizing activity-
(zpa) dorsal region with forming limbbud mesenchyme that
secretes shh and regulates limb axis formation.
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Comments

This page is an overview of Limb development covering a few key issues on pattern formation and mesoderm development with links to additional resources.
Limb development requires a study of several systems, a coordinated establishment of pattern (molecular) on mainly
mesodermal components which also have a covering ectoderm (skin) and invading neural (sensory, motor, neural crest) components.
There is additional page covering Limb Development Abnormalities
Please email Dr Mark Hill if you wish to make a comment about this current project.