2009 Lecture 14

From Embryology
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Limb Development

Appendicular skeleton.jpg

Introduction

This lecture is an introduction to the events in limb development. Initially somites develop and then begin to differentiate forming sclerotome, dermomyotome and then dermatome and myotome. The lateral portion of the hypaxial 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.

The appendicular skeleton consists of: Shoulder girdle, Upper limb (arm, hand), Pelvic girdle, Lower limb (leg, foot).

UNSW Embryology Limb Development | Limb Abnormalities 2008 Lecture 2008 Lecture | 1 slide/page | 4 slide/page | 6 slide/page

Carnegie stage 1-23

Lectopia Audio Lecture Date: 15-09-2009 Lecture Time: 12:00 Venue: BioMed E Speaker: Mark Hill Limb

Lecture Objectives

Human Embryo stage 14 SEM
  • Understanding of limb positioning
  • Understanding of differences in developmental timing of upper and lower limbs
  • Understanding of regions and factors determining limb axes
  • Understanding of limb rotation
  • Understanding of limb muscle, blood vessel, bone and nerve formation
  • Brief understanding of limb molecular factors and cell death
  • Brief understanding of limb abnormalities

Textbook References

  • The Developing Human: Clinically Oriented Embryology (8th Edition) by Keith L. Moore and T.V.N Persaud - Moore & Persaud Chapter 15 the skeletal system
  • Larsen’s Human Embryology by GC. Schoenwolf, SB. Bleyl, PR. Brauer and PH. Francis-West - Chapter 11 Limb Development


  • Before we Are Born (5th ed.) Moore and Persaud Ch16,17: p379-397, 399-405
  • Essentials of Human Embryology Larson Ch11 p207-228

Limb Buds

  • Limbs are initially undifferentiated mesenchyme (mesoderm) with an epithelial (ectoderm) covering.
  • Blood vessels then begin forming, the largest (marginal vein) is adjacent to tip of the limbbud.


Upper and Lower Limb

Stage20-23 limbs a.jpg

Stage14 somites limbbuds.png

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).

Limb Axis Formation

Four Concepts - much of the work has been carried out using the chicken and more recently the mouse model of development.

  1. Limb Initiation
  2. Proximodistal Axis
  3. Dorsoventral Axis
  4. Anteroposterior Axis

Limb Initiation

  • Fibroblast growth factor (FGF) coated beads can induce additional limb
  • 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 then activate FGF expression

Note that during the embryonic period there is a rostrocaudal (anterior posterior) timing difference between the upper and lower limb development

  • this means that developmental changes in the upper limb can precede similar changes in the lower limb (2-5 day difference in timing)

Limb Identity

Forelimb and hindlimb (mouse) identity appears to be regulated by T-box (Tbx) genes, which are a family of transcription factors.

  • hindlimb Tbx4 is expressed.
  • forelimb Tbx5 is expressed.
  • Tbx2 and Tbx3 are expressed in both limbs.

Related Research - PMID: 12490567 | Development 2003 Figures | Scanning electron micrographs of E9 Limb bud wild-type and Tbx5del/del A model for early stages of limb bud growth | PMID: 12736217 | Development 2003 Figures

Body Axes

  • Anteroposterior - (Rostrocaudal, Craniocaudal, Cephalocaudal) from the head end to opposite end of body or tail.
  • Dorsoventral - from the spinal column (back) to belly (front).
  • Proximodistal - from the tip of an appendage (distal) to where it joins the body (proximal).

Proximodistal Axis

  • Apical Ectodermal Ridge (AER) formed by Wnt7a
  • then AER secretes FGF2, 4, 8
  • stimulates proliferation and outgrowth

apical ectodermal ridge | AER and vascular channel

Dorsoventral Axis

  • Somites - provides dorsal signal to mesenchyme which dorsalizes ectoderm
  • Ectoderm - then in turn signals back (Wnt7a) to mesenchyme to pattern limb

Wnt7a

  • name was derived from 'wingless' and 'int’
  • Wnt gene first defined as a protooncogene, int1
  • Humans have at least 4 Wnt genes
  • Wnt7a gene is at 3p25 encoding a 349aa secreted glycoprotein
  • patterning switch with different roles in different tissues
  • mechanism of Wnt and receptor distribution still being determined (free diffusion, restricted diffusion and active transport)

One WNT receptor is Frizzled (FZD)

  • Frizzled gene family encodes a 7 transmembrane receptor

Fibroblast growth factors (FGF)

  • Family of at least 17 secreted proteins
  • bind membrane tyrosine kinase receptors
  • Patterning switch with many different roles in different tissues
  • FGF8 = androgen-induced growth factor, AIGF

FGF receptors

  • comprise a family of at least 4 related but individually distinct tyrosine kinase receptors (FGFR1- 4) similar protein structure
    • 3 immunoglobulin-like domains in extracellular region
    • single membrane spanning segment
    • cytoplasmic tyrosine kinase domain

Anteroposterior Axis

  • Zone of polarizing activity (ZPA)
  • a mesenchymal posterior region of limb
  • secretes sonic hedgehog (SHH)
  • apical ectodermal ridge (AER), which has a role in patterning the structures that form within the limb
  • majority of cell division (mitosis) occurs just deep to AER in a region known as the progress zone
  • A second 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


Wing as Limb Model

  • chicken wing easy to manipulate
    • removal, addition and rotation of limb regions
    • grafting additional AER, ZPA
    • implanting growth factor secreting structures

UNSW Embryology - Axes Formation - Limb | Signal Factors - Wnt

Limb Muscle Mass

(a) Skeletal muscle derived from somites, blocks of mesodermal cells (b) Myoblasts form at each edge of a dermomyotome. Axial myoblasts form the myotome Lateral myoblasts migrate to the limb bud (c) Dermotome skin elements (dermis, hypodermis). Myotome to axial muscle.

Origin of limb muscle cells - Migrations traced by grafting cells from a quail embryo into a chick embryo

  • two species very similar in development
  • quail cells recognizable by distinctive nucleoli
  • Quail somite cells substituted for somite cells of 2 day chick embryo
  • wing of chick sectioned a week later
  • found muscle cells in chick wing derive from transplanted quail somites

Dorsal/Ventral Muscle Mass

Forelimb Muscles

Limb Muscle - Differentiation, Skeletal muscle differentiates the same

  1. Muscle precursor cells migrate to the muscle location
  2. Form beds of proliferating myoblasts
  3. Myoblasts fuse together to form myotubes
  4. Myotubes begin to express contractile proteins, form sarcomeres
  5. mature into myofibers, Innervation determines final muscle maturation

Dermomyotome MyoD

Limb Tissues- Bones (Bone development covered in detail in previous lecture)

  • cartilage template, endochondrial ossification
  • begins Carnegie stage 18 throughout embryo
  • process replaces cartilage with bone (week 5-12), except at future joint sites

Hand and Footplates

Depletion of BMP Signaling Causes Interdigital Syndactyly
  • 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)
  • 3-5 day difference between hand and foot development


hand growth

Apoptosis

Cell Biology - Cell Death Lecture | Cell Biology - Apoptosis Lecture

Limb Rotation

Stage20-23 limbs a.jpg
  • 8th week limbs rotate in different directions (Humans Stage 20-23)
  • thumb and toe rostral
  • knee and elbow face outward
  • upper limb rotates dorsally
  • lower limb rotates ventrally

Limb Innervation

brachial plexus
Adult Dermatomes
  • spinal cord segmental nerves form a plexus adjacent to each limb
  • Brachial (upper) lumbar (lower)
  • Plexus forms as nerves invade the limb bud mesechyme
  • Fetal period - touch pads become visible on hands and feet

brachial plexus origin

Limb Abnormalities

Genetic

Trisomy21 hand.jpg
  • Human Gene Mutations - mutation of any of the patterning genes will result in limb abnormalities (Will put Table on Web page mutations and terminology)

Type II syndactyly- HoxD13

Maternal

  • thalidomide Phocomelia
  • short ill-formed upper or lower limbs
  • hyperthermia

Muscle Development

Duchenne Muscular Dystrophy

  • X-linked dystrophy
  • large gene encoding cytoskeletal protein- Dystrophin
  • progressive wasting of muscle, die late teens

Becker Muscular Dystrophy

  • milder form, adult onset

Congenital Hip Dislocation

Congenital Hip Dislocation
  • Instability: 1:60 at birth; 1:240 at 1 wk: Dislocation untreated; 1:700
  • congenital instability of hip, later dislocates by muscle pulls or gravity
  • familial predisposition female predominance
  • Growth of femoral head, acetabulum and innominate bone are delayed until the femoral head fits firmly into the acetabulum

Online Links

References

Textbooks

  • The Developing Human: Clinically Oriented Embryology (8th Edition) by Keith L. Moore and T.V.N Persaud - Moore & Persaud
  • Larsen’s Human Embryology by GC. Schoenwolf, SB. Bleyl, PR. Brauer and PH. Francis-West -

Online Textbooks

  • Madame Curie Bioscience Database Chapters taken from the Madame Curie Bioscience Database (formerly, Eurekah Bioscience Database)

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Images

Stage13

Stage13 bf1c.jpg Stage13 sem1c.jpg

Stage14

Stage14 bf2cl.jpg Stage14 sem1c.jpg


Glossary Links

Glossary: A | B | C | D | E | F | G | H | I | J | K | L | M | N | O | P | Q | R | S | T | U | V | W | X | Y | Z | Numbers | Symbols | Term Link

Course Content 2009

Embryology Introduction | Cell Division/Fertilization | Cell Division/Fertilization | Week 1&2 Development | Week 3 Development | Lab 2 | Mesoderm Development | Ectoderm, Early Neural, Neural Crest | Lab 3 | Early Vascular Development | Placenta | Lab 4 | Endoderm, Early Gastrointestinal | Respiratory Development | Lab 5 | Head Development | Neural Crest Development | Lab 6 | Musculoskeletal Development | Limb Development | Lab 7 | Kidney | Genital | Lab 8 | Sensory - Ear | Integumentary | Lab 9 | Sensory - Eye | Endocrine | Lab 10 | Late Vascular Development | Fetal | Lab 11 | Birth, Postnatal | Revision | Lab 12 | Lecture Audio | Course Timetable


Cite this page: Hill, M.A. (2019, October 16) Embryology 2009 Lecture 14. Retrieved from https://embryology.med.unsw.edu.au/embryology/index.php/2009_Lecture_14

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© Dr Mark Hill 2019, UNSW Embryology ISBN: 978 0 7334 2609 4 - UNSW CRICOS Provider Code No. 00098G