18) limb development notes PDF

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This is a compilation of notes I made for the embryology topic which is assessed in the anatomy MCQ and essay papers....

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Limb development - Embryology notes

Limb development is initiated in the 4th week of development. Limbs receive information about their correct positioning and use this in combination with their history when determining where to be placed and what structures to form connections with. Atypical limb development is the 2nd most common cause of congenital abnormality. There may be some environmental influence on this e.g. mothers taking thalidomide. Limbs develop from limb buds which consists of a core of loose mesenchyme surrounded by a layer of ectodermal epithelial cells. Surrounding this is a thickened ectodermal layer called the AER. Limbs grow from proximal to distal i.e. proximal layer differentiates first and distal layers differentiate sequentially. It has been suggested that the AER plays a role in this pattern of proximal-distal growth patterning. Replacing the leg AER with a wing AER on a lower limb bud (in a chick experiment) caused the leg to still grow. Removal of AER causes truncation of limb developemtn. The earlier the AER is removed, the less the limbs develop. It is thought that AER secretes a signal called FGF8 protein. Replacing the AER with a bead of FGF8 still causes normal development. The same effect was found when replacing AER with a bead of FGF4 which is found more posteriorly.

Older mesenchyme forms more distal structures. This is because the structures that form depend on the time that cells are exposed to FGF8 signals. Structures lying closest to the AER remain undifferentiated (progress zone). Proximal cells exit first therefore they are exposed to signal for shortest period of time and form proximal structures. Whereas distal cells exit last and exposed to signal for longest duration thus form distal structures. SHFM (split hand foot malformation) is a clinical condition caused by failure of the AER which causes failure of the progress zone and hence the formation of the autopod. Phocolemia is a limb deformity induced by thalidomide caused by lack of proliferation of cells in the progress zone. There are two types: 1st type - hand is connected to trunk, 2nd type - short forearm is connected to trunk.

Dorsal-palmar patterning is determined by the ectoderm. If the ectoderm is removed and reversed, then the dorsal-palmar pattern switches to palmar-dorsal i.e. pattern of

muscles/tendons reverses to correspond to transplanted ectoderm. Wnt7a is a secreted protein that diffuses into the mesoderm where it co-ordinates information to determine dorsal fates. Eliminating Wnt7a results in double ventral limbs i.e. mirror image ventral structures.

Cranial caudal patterning was thought to be determined by the ZPA (zone of polarising activity). Removing graft from the posterior aspect of a limb bud and inserting it onto the anterior aspect caused a double ventral duplication of the limb. The growth of the bud determined by position and amount of graft. It is thought that these grafted cells produce some kind of diffusible signal - the area emitting this diffusible signal/morphogen is known as the zone of polarising activity. This signal was later identified to be Shh, the gradient of which establishes limb patterning. Polyadectyly is a congenital condition caused by changes in Shh activity of which there are sporadic/inherited cases.

Morphogenesis Different parts of the limb will express different transcription factors and different Hox genes. There are different types of Hox genes: a/b/c/d. Hox a/b/c is involved in cranial/caudal patterning whilst Hox d is involved in limb development. Ectopic Shh will up regulate ectopic Hox d expression - there are other signalling centres involved in this too. Cartilage formation requires the condensation of mesenchyme - a pathway which requires BMP type proteins e.g. BMP4/GDF5. Mutation in GDF5 can cause CGT/CHTT. Another congenital abnormality is synadactyly which is the 2nd most common hand abnormality caused by fusion of digits/webbing of skin. It requires surgical correction by performing zig zag incisions and closing these bits of skin inwards as well. Cartilage then becomes bone. GH/IGF-1/thyroid hormones act on germinal zone stem cells. FGF3 inhibits bone formation by binding to FGF3R - a dominant mutation in which will cause achondrplasia (dwarfism).

Limb positioning Placing a bead of FGF8 between upper and lower limb can result in the development of a new limb with its own AER and ZPA. The type of limb that develops depends on where the FGF8 is expressed and whether Tbx5 expression is in the right place. Tbx5 defects caused by 5;12 chromosomal translocation can result in Holt-Oran syndrome which is characterised by severe skeletal malformation. Postioning of limb also depends on Hox genes. Knocking out Hoxb5 in mice results in limbs developing in a more anterior position. The correct combination of Hox proteins can result in localised formation of Tbx5 and FGF which is needed for early stages of limb development.

VOILA :)...