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Bio 127 - Section IIIOrganogenesis Part 1
I. The Stem Cell ConceptII. The Emergence of the EctodermIII. Neural Crest Cells and Axonal SpecificityIV. Paraxial and Intermediate Mesoderm
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I. Stem Cells Role in the Development of Tissues and Organs
• Gastrulation produces the three germ layers
• Germ layer interactions induce organogenesis
• More and more we see that this requires the development of stem cells and their ‘niches’– Places that these cells can remain relatively
undifferentiated and yet provide differentiated progeny
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A. The Stem Cell Concept
• Division of stem cells produces one new stem cell and one differentiated daughter– Sometimes potential is unrealized and you get two
new stem cells
• In some organs: frequent replenishing divisions– gut, epidermis, bone marrow– example: billions of blood cells are destroyed by the
spleen every hour
• In others, they only divide in response to stress or the need to repair the organ– heart, prostate
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b. Stem Cell Terminology
Totipotent = zygote and 4-8 blastomeres
Pluripotent = inner cell mass, “ESC”
COMMITTED STEM CELLS:
Multipotent = adult stem cellshematopoietic, mammary, gut
Unipotent = adult stem cellsspermatogonia, melanocyte
HSC
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Maturational series of neuronal stem cells
VOCABULARY
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c. Types: Embryonic Stem Cells
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c. Types: Adult Stem Cells
• Committed stem cells with limited potential– hematopietic stem cells - hair stem cells– mesenchymal stem cells - melanocyte stem cells– epidermal stem cells - muscle stem cells– neural stem cells - tooth stem cells– gut stem cells - germline stem cells– mammary stem cells
• Hard to extract and culture– HSC are less than 1 in 15,000 bone marrow cells– Transplants work very well, however– Mammary, neural, muscle, others all being worked on
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c. Types: Mesenchymal Stem Cells
• Surprising degree of differentiation plasticity- muscle, fat, bone, cartilage- PDGF, TGF-B, FGF combinations determine fate
• Found in lots of niches in both embryo and adult– umbilical cord blood, baby teeth– marrow, fat muscle, thymus, dental pulp
• Paramedic response to injury– Migrate from niche to provide paracrine stimulus to
repair injured tissues w/wo differentiating on-site
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2. The Stem Cell Niche
• Part of organogenesis in many tissues requires developing special sites for stem cells to live
• Microenvironments wherein the cells that stay don’t differentiate but those that leave do
• Unique combinations of local paracrine signaling, cell-ECM and cell-cell interactions
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Hematopoietic stem cells and the bone marrow niche
-Both are Committed Stem Cells-Progenitor cells can’t self-renew
This is what allows us to do bone marrow transplants
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So, what’s going on in the bone marrow niche?
Hematopoietic stem cellscan form all blood cells.
Mesenchymal stem cellscan migrate to injury sites.
Controls on differentiation:-bone cell matrix-stromal paracrine factors-pericyte paracrine factors-systemic hormones-neuronal signals
So far.....Wntangiopoietinstem cell factorDelta-NotchIntegrin-ECM
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The mouse tooth stem cell niche (we don’t have one)
A balance of “positive – negative”FGF3 – BMP4 and activin -- follistatin
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Stem cell niche in Drosophila testesThe ‘hub’ consists of ~12 somatic cells: the cells in direct contact with themremain stem cells, while the daughters without contact become sperm progenitors
Hub cells Unpaired JAK-STAT Stem Cell Division
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Stem cell niche in Drosophila testes
Cadherinsappear tohold firstcentrosomeclose to the ‘hub’
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Niche Break-Down May be Part of Aging
• Too much cell differentiation– Can deplete the capacity for renewal– Graying hair may result from too many
melanocyte differentiations
• Too much cell division– Cancers may result from excess division– Myeoloproliferative disease is too much
marrow division without differentiation
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Neurulation is a developmental process that takes the organism from the gastrula stage through development of a functional central nervous system
Structure Process Structure
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The first organ system to begin development in vertebrates is the central nervous system
Two Major Steps:1. Formation of the neural tube2. Differentiation of neurons
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REMEMBER: Hensen’s Node (chick) and Spemann’s Organizer (frog) pass organizing power to the notochord
Secreted factors from the notochordcause neurulation in ectoderm above
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Interestingly, the primary mechanism is by means of inhibition....
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Figure 9.1 Major derivatives of the ectoderm germ layer
EctodermalCompetencies
DifferentiatedPhenotypes
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So, where are we starting?
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Establishing Neural Cells from the Ectoderm
• Competence: multipotent cells with the ability to form neurons with the right signals
• Specification: the right signals are there but cell change could still be repressed by other signals
• Determination: the cells have entered the neuronal pathway and cannot be repressed
• Differentiation: the cells leave the mitotic cycle and express the genes characteristic of neurons
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As the node regresses, it leaves the notochord behind anterior to posterior and the overlying neural plate starts to form neural tube in the same pattern
PrimaryNeurulation
SecondaryNeurulation
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Combining Primary and Secondary Neurulation to form the Neural Tube
• Primary = Folding of the Neural Plate into a tube structure directly
• Secondary = Mesenchymal Coalescence followed by hollowing out into a tube
• The Neural Tube proper results from the joining of the two
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• In Birds: everything anterior to the hind limbs is Primary Neurulation
• In Mammals: the sacral vertebrae back through the tail is Secondary Neurulation
• In Amphibians and Fish: only the tail is Secondary Neurulation
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Primary Neurulation in the Chick
Neural plate cells elongateinto columnar epithelium
As much as half of the ectodermcan be induced to form neural plate!
neural convergent extensioncombined with epidermal epiboly
medial hinge point cellsare anchored to notochord
MHP cells flatten and become wedge-shapedto facilitate bending
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Primary Neurulation in the Chick
dorsolateral hinge pointsform between neural and epidermal cells, not crest
as the tube nears closure,neural crest cells undergoEMT and migrate away
*remember: closureresults from neural cells
switching from E- to N-cadherin
Birds close at mid-brain and “zip” in 2 directionsMammals have three primary points of closure
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Works the same on the dorsal surface of amphibian “sphere”
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Human Neural Closure
-spina bifida: posterior neuropore-anencephaly: anterior neuropore-craniorachischisis: the whole tube
Neural tube defects are common: 1 in 1000 live births
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Folate Supplementation Reduces Rate of Defects
Folate-binding protein in the neural folds as neural tube closure occurs
A fungal contamination of corn produces the teratogenfumonisin that appears to disrupt the function of FBP
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Secondary Neurulation
The coalescence of the two neural tubes is not well understood and may be important in some defects
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Differentiation of the Neural Tube
• Three simultaneous levels of development
– Gross anatomy: bulges and constrictions form the chambers of the brain and spinal cord
– Tissue anatomy: the cell populations in the wall rearrange to form functional domains
– Cell biology: the neuroepithelial cells differentiate into neurons and glia
• Two simultaneous axes of development
– Anterior-Posterior: the forebrain back toward the spinal column
– Dorsal-Ventral: the axis from the roof plate of the tube, near the epidermis, and the floor plate, near the notochord
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Figure 9.9 Early human brain development (Part 1)
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Figure 9.9 Early human brain development (Part 2)
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Rhombomeres of the chick hindbrain
r1
r2
r3
r4
r5
r6r7
Neural crest cells from above specific rhombomeres form the cranial nerve ganglia
5th trigeminal
7th facial and8th vestibuloacoustic
9th glossopharyngeal
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The size of the vertebrate brain increases very rapidly in early neurulationdue to an osmotic Na+ gradient dumped into the presumptive ventricle: for example, the chick’s brain volume increases 30-fold from day 3-5
The increase in size determines how manyneurons are able to ultimately divide and form
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Occlusion of the neural tube allows expansion of the future brain
Relaxesafterexpansion
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Anterior-Posterior Specification of Neurons: Evolutionary conservation of homeotic gene organization and transcriptional expression in fruit flies and mice
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Dorsal-ventral specification of the spinal neural tube
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Dorsal-ventral specification of the spinal neural tube
Sensory Input
Motor Output
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Concentration Gradient-Dependent Transcription Factor Expression
growth factors transcription factors
Pax7
Pax6
Nkx6.1
TGF-B
Shh
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Differentiation of Neurons in the Brain
• Neuroepithelium of neural tube starts as one layer of stem cells
• Humans have 100 billion neurons and 1 trillion glial cells
• Neuroepithelium gives rise to:– Ependymal cells: line the ventricles, secrete CSF– Neurons: electrical, regulation, thought, senses– Glia: brain construction, neuron support, insulation
and maybe memory storage?
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Diagram of a neuron
We have very few dendrites at birth,up to 100,000 connections in 1st year!
can be 2-3 feet long
microtubules followssignal
gradient
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Figure 9.16 Axon growth cones
Actin microspikes provide migratory traction and signal sensing
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Figure 9.17 Myelination in the central and peripheral nervous systems
Multiple sclerosis is ademyelination disease
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Neural stem cells in the germinal epithelium
Neural tubestart as onelayer ofstem cells,all in the cell cycle
Position of nucleusdepends on cell cycle
Stem cell divisionsare all horizontal
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Neuron Birthdays
• Differentiating cells are born from vertical divisions
• Stem cell stays attached, distal sister migrates away and leaves the cell cycle
• Early birthdays form closer layers, later birthdays form more distal layers
• Neuronal function, neurotransmitter type and connections formed depends on Anterior-Posterior, Dorsal-Ventral position (eg. Hox, TGF-B v. Shh )
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Complexity Increases the Further Anterior You Go
Initially three basiclayers are formed
stemcells
cellbodies“graymatter”
myelinaxons“whitematter”
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Figure 9.20 Development of the human spinal cord
Original formation of the germinal neuroepithelial layer
Differentiated three adult layers:1. ventricular zone = ependyma2. Intermediate zone = mantle3. Marginal zone = myelin layer
Becomes encased in connective tissue
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Figure 9.19 Differentiation of the walls of the neural tube (Part 1)
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Differentiation of the Cerebellum
Adds three additional layers:- The Purkinje cell layer- The inner and outer granular layers
Cerebellum coordinates complex movements- Purkinje’s have ~100,000 synapses on their dendrites- Axons control all cerebellar output- Not too sure about the role of granular neurons
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Cerebellar neurons have typical brain migration mechanism
They crawl along glial processes from layer to layer
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Figure 9.21 Cerebellar organization
Bergman gliaprovide themigratoryprocesses inthe cerebellum
phenomenaldual-photonconfocalmicroscopy!
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Cerebral cortex is the most complex of all
The major addition is the formation of the neocortex- Stratifies into 6 layers, each with unique inputs and outputs- Adult form not completed until middle of childhood
Also organized Anterior-Posterior and Dorsal-Ventral- Hox genes and TGF-B v. Shh- eg. layer 6 inputs and outputs in visual cortex
differ from layer 6 connections in auditory cortex
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Figure 9.25 Evidence of adult neural stem cells
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Development of the Sensory Systems
• They form from the cranial ectodermal placodes which are made competent by endo, meso signals
– We’ll focus on the lens placode
– The olfactory placode forms nasal epithelium, nerves
– The otic placode forms inner ear, acoustic ganglia
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Reciprocal induction between neural tube and overlying ectoderm
1. Optic vesicle evaginates fromdiencephalon, contacts ectoderm
2. Optic vesicle Delta binds to ectoderm Notch, induces placode
The induction of lens causes the cells to elongate, invaginate and grab holdof the optic vesicle cells with adhesive filopodia to ride its movement inward
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Reciprocal induction between neural tube and overlying ectoderm
3. Lens signals cause two layers of opticcup to form pigmented and neural retina
4. Lens tissue is pulled under the surface, induced to make Crystallin
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Key Cell Differentiation Events
Without theexpression of the retinalhomeoboxgene (Rx) nooccular tissuesdevelop at all
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Key Cell Differentiation EventsThe neural retinaforms 7 majorlayers of neurons
The epithelium ofthe posterior layerof optic cup arecompetent to makeall of them
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Key Cell Differentiation Events
The tips of the optic cup form a ring of pigmented muscle, the iris, which controls the pupil dilation and gives eye color
The junction between the iris and the neuralretina form the ciliary body, which secretesthe vitreous humor tocontrol pressure andthe curvature of eye.
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Key Cell Differentiation Events
surfaceectoderm
Neural crest
“Lens fibers” are elongated cells from the lens placode
Neural crest mesenchymemigrate in to form cornea
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Key Cell Differentiation Events
Anterior chamber fillswith vitreous humor
Neural crest cell MET, dehydrate andform tight junctions to become cornea.Stem cell population at corneal edge.
Lens fibers extrudetheir nuclei and formmassive amounts ofCrystallin proteins.Stem cells in epithelium.
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Figure 9.31 Sonic hedgehog separates the eye field into bilateral fields
Too little Shh and the eyefields don’t separate on midline
“Cyclopism”
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Figure 9.32 Surface-dwelling (A) and cave-dwelling (B) Mexican tetras (Astyanax mexicanus)
Too much Shh andeye fields don’t form.
Vision sacrificed incave dwellers in favorof better olfaction andbigger jaws!
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The Epidermis and Cutaneous Appendages
• Remember, the ectoderm forms:
– Neural tube
– Neural crest
– Epidermis
• The epidermis is the outer layer of the skin with mesoderm-derived dermis underneath
– Largest organ, tough, impermeable, renewable
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The Epidermis and Cutaneous Appendages
• The cells of the skin include
– Basal layer stem cells, keratinocyte daughters
– Dermal fibroblasts
– Neural crest-derived melanocytes
• The same three cell types form hair follicles
– Each must become specialized to do so
– Similar interactions form feathers and scales
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Layers of the human epidermis
The same basic design asthe neuroepithelium: stemcell divisions are horizontal,differentiation divisions arevertical.
Melanocytes transfer melanin granulesdirectly to the cells of the Malpighian layer.
Daughters migrate away,withdraw from cell cycle,differentiate to keratinocytes
Outerlayeris deadcells
Innerlayerdividesthroughlifetime
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Layers of the human epidermis
Keratinocytes are the epitomyof “taking one for the team”!
They then arrest their own metabolismand die, leaving a tough membrane andfiber protective layer over the basal layer.
As they differentiate, theyexpress heavy keratin fibersin their cytoplasm, hook themto cadherins and integrins intheir plasma membrane andto collagen and neighbor cells.
Outerlayeris deadcells
Innerlayerdividesthroughlifetime
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• Basal cells express Delta-family member Jagged
• When it binds to distal sister Notch it starts keratinocyte differentiation
• Time from basal layer to sloughed is 2 weeks in mouse, a little longer in us
• As part of their differentiation they push their nucleus to the side of the cell
• We lose 1.5 grams of them per day with enough DNA in each (or perhaps a few) to identify us!
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Reciprocal induction in development of hair follicles
1. Dermal fibroblasts induceplacode formation in basal cells
2. Placode cells induce thosefibroblasts to form dermal papilla
placodesinksinto
dermis
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Reciprocal induction in development of hair follicles
3. dermal papilla induces stemcells to differentiate daughters
differentiated daughters include:hair shaft, sebocytes, root sheath
Part of differentiation includes migration to absorb papilla.
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Reciprocal induction in development of hair follicles
- Hair shaft is keratinocytes with melanin just like skin itself.- Sebaceous gland secretes lubricant onto hair and skin.- “Bulge” is the stem cell niche for basal cells and melanocytes.
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Figure 9.42 Model of follicle stem cell migration and differentiation
Stem cells migrate from the bulge: 1. down the outer root sheath to thefollicle root, 2. up to the sebaceous gland, and 3. up to the basal layer.
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Figure 9.39 Patterning of hair follicle placodes by Wnt10 and Dickkopf
Evenly spaced hair follicles result fromthe epithelial cells releasing both Wnt10AND its inhibitor Dickkopf. Wnt causesautocrine formation of placodes close by,while Dickkopf blocks nearby neighborsfrom being able to form them.
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