Study Questions 6 Flashcards
- What parts do each somite become divided into (5 parts)?
· Sclerotome (vertebrae, ribs, cartilage)
· Myotome (musculature of the back, ribs and limbs)
· Dermatome (dermis of the back)
· Syndetome (tendons)
o Endothelial cells (Cells that generate vascular cells in the dorsal aorta)
- Draw the relative geometric shapes and positions of the dermatome, sclerotome and two subdivisions of the myotome in respect to the notochord, neural tube, lateral plate mesoderm and overlaying epidermis. Why is this particular arrangement important?
This particular arrangement is important because the placement of morphogen generating structures determines what types of somites get exposed to what and at what concentration. Those factors determine the developmental
- Which proteins control somite boundaries?
Notch – receptor protein, has signal pathway associated with gene expression for boundary formation
· Stimulate gene expression of Hairy1
Hairy1 - transcription factor that is expressed in a wave like fashion throughout the unsegmented paraxial mesoderm
· Leaves a thin band that defines the posterior of next somite
- Explain the results of the following experiment involving protein Notch
Quail presumptive boundary cells transplanted into unsegmented paraxial mesoderm of chick
a. Result = boundary formed
b) Quail non-boundary cells transplanted into unsegmented paraxial mesoderm of chick
a. Result = no boundary induced
c) Quail non-boundary cells transplanted into unsegmented paraxial mesoderm of chick followed by local ectopic (unusual place) expression of notch via induction of graft by electroporation to activate Notch
a. Result = somite boundary induced
- How does protein Notch influence gene expression? (This question is about regulation of gene expression by Notch, NOT about somite clock and wave mechanism.)
The protein Notch is a transcription factor because it enters the nucleus to change gene expression.
· Notch is a receptor located at cell membrane. It interacts with membrane bound ligand on adjacent cell (ex. Delta). Upon notch/ligand binding, intracellular domain of notch is cleaved and the cleaved polypeptide goes to nucleus to influence gene expression.
- What is the current theory explaining somite formation involving Notch and Hairy 1 (among other factors)?
The theory is clock and wave mechanism that follows Hensen node retraction
FGF expression trigger oscillating ‘clock’ signal that causes
· Notch’s (as transcription factor) gene expression of Hairy1 (transcription factor) and Notch inhibitor
o Hairy1 = posteriorizes somite, leaves a band at the anterior point of the unsegmented paraxial mesoderm which is then the posterior of the next somite
o As more notch is created from Wnt pathway, more notch inhibitor made = decreasing levels of notch = decreasing levels of inhibitor = eventual re-increase of notch = wave
§ Simple negative feedback loop (sinusoidal/wave expression)
· Expression of Hairy1 allow for determination of the posterior of each somite
• an oscillating signal (the “clock”): Notch, which stimulates gene expression of Hairy 1 (one of “effectors”), as well as its own regulatory protein, inhibitor (“clock inhibitor”); this creates repeating “waves”
- What is the role of ephrin and ephrin receptor in somite formation?
The ephrin ligand (ephrin-b2) and tyrosine kinase receptor (EphA4) allows somites to separate from unsegmented paraxial mesoderm. This allows for somite repulsion and separation from unsegmented paraxial mesoderm. Ephrin prevents NCC movement in certain areas because expression modifications can cause cytoskeleton arrangements.
- Describe the process of somite epithelization. What are the roles of fibronectin and N-cadherin?
● After somite formation Ectodermal signals initiates mesenchymal-toepithelial transition (MET)
● Form basal membrane, sub-apical surface
● Fibronectin – ECM organizing protein
● N-cadherin – adhesion protein
○ Work with fibronectin to organize outer layers into epithelium posterior to anterior (for chickens)
● Epithelialization occur in posterior part of anterior somite (top of gap to bottom of gap
○ Central cells remain mesenchymal while outer cells become epithelial layer
● Somite forms ANTERIOR TO POSTERIOR
● Epithelialization forms POSTERIOR TO ANTERIOR
● Gap forms within anterior mesenchymal Presomitic mesoderm (PSM)
Cells posterior to the gap undergo epithelialization at later stage than cells at the gap anterior
Cells at anterior part of gap will complete epithelialization first and become the POSTERIOR edge of somite
- Do somites continue to exist in the body, or are they only temporary present in the embryo?
● Somites are only temporarily present in the embryo (transient structures)
● Important for subsequent development
○ Vertebrae, ribs, dorsal skin dermis, back skeletal muscle, body walls, limbs (bones, muscle, skin, limbs) Determine migration path of NCC and spinal nerve axons (function)
- Somites first subdivide into sclerotome, dermatome and myotome. What signals are involved in these subdivisions and where are they coming from?
Sclerotome: High conc. Sonic Hedgehog (shh) from notochord, floor plate
Induces sclerotome cells to secrete Pax1 transcription factor -Cartilage, vertebrae (Pax1)
Dermatome: Neurotropin3 (NT3) and Wnt1 (Shh antagonist) from roof plate-adipocyte
Myotome: Primaxial – low concentration of Wnt Abaxial – Wnt (epidermal), BMP4, Fgf5 (lateral plate mesoderm)
lateral Dermamyotome – pax3 induces MyoD
medial Dermamyotome – Wnt &SHH induce Myf5 induces MyoD -Dermatome, myotome, sclerotome
muscle
- Where are the cells that form the vertebrae derived from? What forms the anterior part of each vertebrae? What forms the posterior part of each vertebrae? Why does this happen?
Somites of paraxial mesoderm generate the vertebrae
● Sclerotome (somite derivatives) form the anterior part of each vertebra
● 2/3 of sclerotome form posterior part of each vertebra
● This occurs because the outgrowth of spinal neurons occur in order to reach and innervate the myotome
○ To do this, the spinal nerve goes through the sclerotome, cutting each segment in 2 pieces (1/3 anterior, 2/3 posterior)
■ 2/3 posterior piece combine with 1/3 of anterior piece of posterior sclerotome
● Posterior part of the sclerotome from the anterior somite combines with the anterior part of the sclerotome from the following (posterior) somite to form vertebral rudiment, and eventually vertebra – resegmentation
● Resegmentation enables innervation of muscles (developed from myotome) to coordinate movement - allows compression and bending between vertebrae.
- What are two major factors involved in determination of the myotome cell to become a myoblast? How do they influence gene expression?
Two Factors = Myf5 and MyoD
● Act as transcription factors and change gene expression
HOW THEY WORK
● Paracrine factors induce MyoD which induce myotome cells
○ Lateral area of dermamyotome – Pax3 induced which induces MyoD (abaxial muscles)
○ Medial area of dermamyotome – Myf5 induced which induces MyoD (primaxial muscles)
● MyoD activates other genes
- What are the three distinct cell lineages involved in formation of skeleton? Which germ layer(s) they originate from?
Cranial Neural Crest – from ectoderm (creates branchial arch, craniofacial bones, cartilage)
Somites – from paraxial mesoderm (creates axial skeleton in vertebrae)
Somatic Part of – lateral plate mesoderm (creates limb skeleton)
- What are the two types of bone formation? What are the main differences between them (think about tissue origin and type of ossification)?
Intramembranous (Perichondral) Ossication or direct ossification of neural crest cells
Endochondral Ossification or indirect ossification uses hyaline cartilage model for bone construction
● Requires breakdown of hyaline cartilage before ossification
- What are the steps in endochondral ossification (we roughly divided endochondral ossification into 7 steps)?
- Mesenchymal cells commit to become cartilage (shh induces pax1)
- Cells condense into compact nodules, differentiate into chondrocytes (BMP induces N-cadherin, N-CAM and Sox9)
- Chondrocytes proliferate and lay down collagen 2 matrix (cartilage specific ECM)
- The chondrocytes in the middle stop dividing, increase in size and become hypertrophic chondrocytes (Runx2). They secrete factors that lead to either differentiation into osteoblasts or their own death by apoptosis. After death and disintegration they leave cavity within cartilage (ECM shifts; increase in mineralization and secretion of vascular endothelial growth factor VEGF- will cause angiogenesis).
- Replacement of chondrocytes by osteoblasts dependent on mineralization (Ca++).
- Blood vessels invade, bringing in osteoprogenitor cells. They will develop into octeoclasts.
- Remodeling releases more VEGF, and increases the number of blood vessels in the dying cartilage.
- Osteoblasts begin forming bone matrix on the partially degraded cartilage; bone matrix forms a bone collar around dying cartilage cells.
- Perichondrium is replaced by periosteum
- At the same time osteoclasts are (re)forming the bone marrow cavity by hollowing out (degrading) the internal region of the bone
- Continued development through a balance of osteoclasts and osteoblasts.
- Secondary ossification centers form in epiphyses (blood vessels migrate into epiphyses carrying osteoblasts….)
