Download Development from Neural Crest Cells

Development from Neural Crest Cells
Neural Crest Cells as Precursor Cells for Multiple Lineages
• Neural crest cells migrate extensively to
generate multiple differentiated cell types.
• neurons and glial cells of the sensory,
sympathetic and parasympathetic nervous
system
• epinephrine-producing (medulla) cells of
the adrenal gland
• pigment-containing cells of the epidermis
• majority of the skeletal and connective
tissue components in the head
(cf. for trunk & posterior  somites)
• The fate of the neural crest cells depends,
to a large degree, on where they migrate
to and settle.
Birth of Neural Crest Cells
(NCCs)
• Quail neural plate graft into chick non-neural ectoderm
induces a second neural crest cell population.
(contributions from both neural and epidermal ectoderms)
• High expression of BMPs initially in the neural ectoderm
(neural plate)  later in dorsal neural tube.
• Ventral NT : NC/FP  Shh  Noggin  BMP inhibition
• High expression of Wnt6 initially in epidermal ectoderm
 later in surface ectoderm
• Cells under the influences from both high levels of BMPs and
Wnt6 give rise to neural crest cells.
• These presumptive neural crest cells can be identified by
their characteristic expression of two marker genes such as
Slug and FoxD3.
• FoxD3 : NCC fate specification; FoxD3 inhibition  No NCC;
Forced FoxD3 expression  Ectopic NCC formation
• Slug : NCC migration out of epithelium
Four Main Domains of Neural Crest
1) Cranial (Cephalic) Neural Crest
A. migrate dorsolaterally to produce craniofacial mesenchyme
 cartilage, bone, neurons, glia, and connective tissues in the face
B. enter anterior pharyngeal arches and pouches
 thymic cells, odontoblasts (tooth primordia), middle ear & jaw bones
2) Trunk Neural Crest (S8-27)
A. migrate ventrolaterally and remain in the anterior half of
each sclerotome  dorsal root ganglia (sensory neurons)
B. migrate more ventrally  sympathetic ganglia, adrenal medulla,
and nerve clusters around aorta
C. migrate dorsolaterally into the ectoderm
 melanocytes (pigment synthesizing cells)
3) Vagal (S1-7) and Sacral (S28 to posterior) Neural Crest
 parasympathetic enteric ganglia of the gut (colon)
(cf. neurons related to the peristaltic bowel movement)
4) Cardiac (somite 1-3) Neural Crest (posterior to cranial NC)
A. migrate into the third, fourth and sixth pharyngeal arches
 melanocytes, neurons, cartilage, and connective tissue
B. entire muscular-connective tissue wall of the large arteries
(arise from the heart)
C. septum separating the pulmonary circulation from the aorta
Migration Pathways of Trunk Neural Crest Cells
1) Ventral Pathway
• Sensory neurons (dorsal root ganglia)
• Sympathetic neurons
• Adrenomedullary cells
• Schwann cells
• In birds and mammals, this migration is going
through only anterior section of each sclerotome.
2) Dorsolateral Pathway
• melanocytes : melanin-forming cells
• migrate between epidermis and dermis layers
• enter the ectoderm through small holes in the basal lamina
HNK-1 Staining
Neural Crest Cell Emigration from the Neural tube
• The neural crest cells are originally part of an epithelium.
• The NCCs undergo EMT (Epithelial-to-Mesenchymal
Transformation), thus the NCCs can be detached from
other cells. Then they migrate away from the epithelial
sheet.
• Wnts, FGFs, and BMPs together induce the expression of
NCC specific genes such as Slug and RhoB.
• RhoB : Cytoskeletal changes to promote migration
(microfilament polymerization)
• Slug Dissociates the intercellular tight-junctions to render
cell movements.
• Downregulation of N-cadherin expression at the time of
migration
• Reappearance of N-cadherin expression upon NCC
aggregation after reaching to the final destination
(e.g. to form dorsal root and sympathetic ganglia)
HNK-1 : Red, RhoB : Green
Overlapping Staining : Yellow
Recognition of Surrounding Extracellular Matrices
• ECMs such as fibronectin, laminin, tenascin, various collagens and proteoglycans promote
migration of neural crest cells.
• Integrin expression in NCCs upon migration (e.g. integrin α4β1)
• When these integrin proteins are lacking, NCCs released from the neural tube become
disoriented and often time undergo apoptosis.
• Thrombospondin, expressed only in the anterior portion of the sclerotome, promotes NCC
adhesion and migration by cooperating with fibronectin and laminin.
• NCC migration impeding molecules
- Ephrine proteins (Eph) expressed in the posterior section of each sclerotome
- Ephrine receptor (Eph-R) expressed in NCCs
- Eph binding to Eph-R activates tyrosine kinase activity of Eph-R in NCCs.
- Then, Eph-R signaling interferes microfilament polymerization and more ….
• This patterning also correlates with the actual structures of the neural crest derived peripheral
nervous system such as dorsal root ganglia segmental positioning.
• SCF (Stem Cell Factor) supports NCC proliferation for future melanocytes, protects apoptosis,
and also functions as a chemotactic factor. (e.g. Forced expression of SCF in the chick footpad
can unusually recruit melanocytes to this region.)
The migration of neural crest cells can be regulated
both by the ECMs and soluble signaling factors secreted at their potential destinations.
Ephrine (+) region
HNK-1 (+) cells
negative correlation
Motor neurons are
emerged only from
the anterior portion
of each sclerotome
FN Coating & Ephrin Strip
Injection of Fluorescence-Labeled Neural Crest Cells  Diverse Destinations
Pluripotency of Neural Crest Cells
• The vagal (neck) NCCs produce cholinergic neurons in parasympathetic ganglia.
• The thoracic (trunk) NCCs produce adrenergic neurons in sympathetic ganglia.
• But, when those NCCs are reciprocally transplanted, a new differentiation fate is determined
based on their new locations.
• NCCs express enzymes synthesizing for both acetylcholine and norepinephrine at pre-migratory
stages. After migration, one of either enzyme is downregulated.
• NCCs from the hindbrain region normally migrate into the eye and interact with the pigmented
retina to become scleral cartilage. (i.e. no putative destinations to give rise to neural tissues)
• But, when transplanted into the trunk region, they can be differentiated into sensory ganglion
neurons, adrenomedullary cells, glia and Schwann cells.
• Individual NCCs are pluripotent as they leave the crest.
• Further differentiation of a neural crest cell depends on its eventual location.
• BUT, some recent efforts have demonstrated that the initial NCC populations can be a mixture of
different precursor cells.
Gradual Determination of Neural Crest Cell fates
Pluripotency of NCCs
changes upon
interactions with
surrounding tissues
during migration
Signal Transduction Pathways Regulate the NCC Fate Specification
The Fates of Neural Crest Cells Are
Directed by the Tissue Environment Where They Settle
• Ephrin signals inhibit early ventral migration, but later facilitate dorsolateral migration.
Thus it seems critical for controlling the timing or onset of lateral migration of NCCs.
• LIF (leukemia inhibition factor) from heart converts adrenergic neurons into cholinergic neurons.
• BMP2 secreted by the heart, lungs, and dorsal aorta  cholinergic neuron differentiation
• GGF (glial growth factor; neuregulin)  glial fates (cf. suppresses neuronal fates)
• Wnt1  sensory neurons
• Endothelin-3  adrenergic neuron
• Endothelin-3 + Wnts  melanocyte differentiation
• Glucocorticoids  adrenomedullary cell differentiation (chromaffin cells)
(cf. suppresses neuronal fates)
• NGF and FGF2  sympathetic neurons
Cranial Neural Crest
• Trunk NCCs  melanocytes, neurons and glia
• Cranial NCCs  cartilage, bone, melanocytes,
neurons and glial cells
• Trunk NCCs can not give rise to cartilage tissues.
• But, if trunk NCCs are placed into the head region,
these NCC can normally differentiate into head
cartilages and bones, which suggests that tissues
around cranial NCCs provide competence factors
for cranial NCCs to become cartilage or bone.
Rhombomeres  NCCs  Pharyngeal Arches  Various Cranial Structures
• Rhombomere 1 and 2  first pharyngeal (mandibular) arch  jaw, ear, frontonasal process
• Rhombomere 4  second pharyngeal arch  neck, middle ear
• Rhombomere 6  third and fourth pharyngeal arches  thymus, thyroid, parathyroid glands
 clavicle (collar bone) ; neck muscle attachment site
• Rhombomere 3 and 5  no direct migration out of these rhombomeres
NCC Specific β-Galactosidase Transgenic Mice
NC Originated Precursor Cells  Majority of Cranial Structures
Vertebrate Skeletons
Intramembranous
Ossification
Endochondral
Ossification
Mesenchymal Cells
Mesenchymal Cells
Chondrogenesis
Skull
Facial Bones
Teeth
Cartilage
(Chondrocyte & ECMs)
Somite (Sclerotome)
Axial : Vertebrae, Ribs
Lateral Plate Mesoderm
Appendicular : Limbs
Mesenchyme is made of loosely scattered cells that make up
the majority of the undifferentiated tissue in the embryo.
Intramembraneous Ossification
periosteum
NCC  Mesenchyme  Condensation  Osteoblasts
 Osteocytes (Bone Cells)
 Capillaries
CBFA1 Is Essential for Osteogenesis
BMP 2, 4, 7
Mesenchymal
Cells
CBFA1 (Runx2)
Expression
Induction of
Bone Specific Matrix Proteins
such as Osteocalcin & Osteopontin
Wildtype
CBFA1 K.O.
Cartilage Tissues Prefigure Entire Axial and Appendicular Skeletons
Alcian Blue : Cartilage Specific Staining Reagent
Endochondral Ossification & Longitudinal Bone Growth
Proliferating Chondrocytes
Highly Proliferative & Resistant to Apoptosis
Responsible for Longitudinal Bone Growth
Hypertrophic Chondrocytes
Growth Arrested & Sensitive to Apoptosis
Target Site for Osteogenesis & Vascularization
Tooth Development
Ectoderm
Epithelium  Enamel Knot  Cusp Formation  (Pre)Odontoblast  Dentin Secreting Cells
NCC  Ectomesenchyme  FGF8  Pax9  Mesenchyme Condensation  Dental Papilla
Mesoderm
Enamel Knot  Shh, FGF4, BMP 2, 4, 7
Jaw Epithelium  Ameloblast  Enamel Secreting Cells
Cardiac Neural Crest
• The heart originally forms in the neck region directly beneath the pharyngeal arches.
• Cardiac neural crest : caudal region of the cranial neural crest
• Cardiac neural crest cells give rise to the endothelium of the aortic arch arteries and the septum
between the aorta and the pulmonary artery.
• In mice, pax3 can be a cardiac neural crest marker.
• Mutation in pax3 results in fewer cardiac neural crest cells and this condition causes persistent
truncus arteriosus (the failure of the aorta and pulmonary artery to separate).
• This mutation is also related to the congenital defects in thymus, thyroid, parathyroid glands.
Cranial Placode
• The anterior borders between the epidermal and
neural ectoderm; local and transient thickenings
of the ectoderm in the head and neck; Sensory
apparatus (e.g. nose, ear, taste receptors, lens)
• The otic placode, which develops into the
sensory cells of the inner ear, is induced in the
region of the ectoderm where the presumptive
neural plate meets the presumptive epidermis.
• FGF19 and Wnt8c synergistically induce the otic
placode.
• The epibranchial placodes form dorsally to where the pharyngeal
pouches contact the epidermis to give rise to facial neurons and
vagal nerves, etc.
• Secondary NCCs from the placodes do not travel ventrally, rather
they migrate dorsally to form glial cells and these glia form the
tracks that guide neurons from the epibranchial placodes to the
hindbrain. Therefore, it is very critical for hindbrain innervation.