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DB3 > Mammalian Morphogenesis > Flashcards

Mammalian Morphogenesis Flashcards

1
Q

first steps

A

cleavage division - mitosis w no growt

cant make feeding structures w/out many specialised cells
so need to use resources at hand
large oocyte divides w/out growth
cell cycle just goes:
S
M
S
M
…

get to about 1000 cells where they start to synthsise proteins

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2
Q

when does zygotic gene transcritption begin

A

4 cell stage
one of first ones is E-cadherin
causes cells to stick together and compact

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3
Q

compaction and trophectoderm formation

A

E-cadherin expression
causes cells to compact
some cells will be entirely in the middle
some will be at free edge

cells at edge differentiate into different type of epithelium - trophectoderm
rest remain as middle mass

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4
Q

blastocyst formation

A

trophectoderm forms
inner cell mass inside
trophectoderm begins letting fluid in - forming a blastocoel at the centre

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5
Q

blastocyst hatching

A

blastocyst forms and then hatches from the Zona Pellucida from the oocyte

the trophoblast of this hatched blastocyst invades the uterine epithelium
-forms interdigitated columns that will develop into placenta

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6
Q

hypoblast/epiblast formation

A

blastocyst has blastocoel cavity inside
some cells of ICM face this cavity
-causes them to differentiate forming layer called the hypoblast
this will then line the trophectoderm on the inside and surround what will be the yolk sac

the remaining layer of ICM cells touching the hypoblast polarises and lets go of the overlying trophectoderm cells
-forms the epiblast

now have a kind of disk inside with hypoblasr and epiblast formed from the ICM

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7
Q

monozygotic twins cause

A

most errors at stage of forming hypo/epiblast are lethal
but some subtle and rare errors are embryo tolerable resulting in identical/monozygotic twins

2 ways of this happening
-cells separate inside the zona pellucida (1/3)
-two ICMs form - almost the rest of MZ twins (2/3)

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8
Q

Cells separating inside the ZP

A

have two trophectoderms form
ICM forms hypoblast and epiblast in each
2 distinct systems with all the structures each

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9
Q

two ICMs form inside same trophectoderm

A

Two ICMs form
so get two sets of epi/hypoblast wihtin same trophectoderm

will share a placenta

at risk of foetal transfusion syndrome where one twin steals supplpy from the other and one twin turns out smaller

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10
Q

axes of embryo after epi/hypoblast formation

A

are currently radial like a jellyfish
need to generate other axes for morphogenesis
have top and bottom but need others
need to mark specific part to distinguish this out of radial symmetry

no way to transform 2 coordinate system of a disc to a 3 coordinate system of 3d object

but if the embryo can somehow mark one part of the edge of the disc different to mark “12 o clock” then it can have 3 coordinates
and these can be transofrmed into the body axes

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11
Q

solution for “marking 12 o clock” and generates the beginnnings of the axes

A

cells at centre of HYPOBLAST express Hex
Hex expressing cells move out to rim - congregating at one point
these cells at one end mark the head end and make inhibitory signals inhibiting progress in the epiblast cells above them at this end

the furthest away cells in the epiblast (so most towards the posterior)
and so are not inhibited
can begin making tail end of the primitive streak
these sites on the discs mark the future trunk and head

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12
Q

3rd and rarest way of making MZ twins

A

the formation of one body axis depends on the Hex-expressing cells being in ONE point on the rim of the hypoblast

if there ends up being two distinct sites -then two heads will form and maybe two primitive streaks

can end up with conjoined twins

can also get a partial axis duplication if the two head organising areas still agree on one site for the tail end
this can form 2 headed andimals
usually prenatal in humans but has seen survivals

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13
Q

germ layer formation

A

GASTRULATION
as primitive streak grows forwards on the epiblast
the advancing end is called the NODE
gastrulation fillows the node - organisation of germ layers of the body

gastrulation:
epiblast cells converge on the midline
-some of these cells push through and move the hypoblast aside and form new cell layer - endoderm (epiblast cells that push in form this)
-other cells that push through stay in the middle and spread out in the space between the epi-/hypoblast discs to form the mesoderm
-other epiblast cells remain in this layer and form ectoderm

the middle part of the new endoderm rises to make notochord plate
this notochord plate then detaches to become the notochord

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14
Q

germ layer cell fates determinant

A

depends on where and when they dived down
-never dive down - ectoderm (CNS, Epidermis)
-dive down firs right through the node - become endoderm and then form the notochord
-cells that dive early - but not directly through node - become endoderm (gut and most abdominal organs)
-cells that dive down later become mesoderm (muscles, connective tissue, urogenital system)

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15
Q

mouse cup peculiarity

A

mice are annoying
they make a cup shape rather than staying flat
this cup is arranges inside-out
hypo and epiblast - epiblast on inside of cup
-ectoderm on inside
mesoderm between
endoderm inside

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16
Q

neurulation basic

A

whole of CNS formed by forming a tube

ectoderm over the back
two edge stripes and one centre stripe forms
cells at these stripes deform their shape
centre stripe folds to form infolded valley
edge stripes form ridges and meet at top of tube

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17
Q

neurulation: when the cells at top meet

A

stick to each other and let go of their neurectodermal neighbours and adhere to each other

pinches off the tube and separates under the sealed ectoderm forming the neural tube

top of the tube “zips” up

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18
Q

failure of neural tube closure

A

failure to zip up properly can leave it open - vertebrae cannot form where this has happened

causes Spina Bifida

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19
Q

Mesoderm development basic

A

each side of neural tube the mesoderm is also developing
embryo is lengthening too
-ends of endoderm pulled out into tubes,
-firther lengthening makes these tube sections linger
-connection to the yolk sac eventually appears as just a minor branch from the tubular gut

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20
Q

components of the mesoderm

A

paired on each side of neural tube under the ectoderm covering
-notochord below neural tube
-somite directly to side of neural tube
-lateral mesoderm next to that
>wolfian duct in it next to somite
>celom inside it
>somatic mesoderm on part above celom
>plachnic mesoderm below celom

endoderm below this

aorta is below somite

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21
Q

formation of structures from somites:

A

somites scatter cells and reform to make vertebrae, ribs, muscles and dermis

somite location kind of prefigures vertebrae location
first half of one somite and back half of another come together to become vertebrae (similar to drosophila parasegments)

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22
Q

other mesoderm component diagram

A

-neural tube
-notochord below it (chorda mesoderm)
-paraxial mesoderm next (head and somites formation)
-intermediate mesoderm next to that
-lateral plate mesoderm next

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23
Q

paraxial mesoderm give rise to

A

head

somites:
sclerotome, syndotome
myotome
dermatome, endothelial cells

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24
Q

intermediate mesoderm

A

kidney
gonads

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25
Q

lateral plate mesoderm

A

splanchnic
somatic
extra-embryonic

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26
Q

the neural crest basic

A

small region of cells formed after neural tube pinches off
some remains neural - others become different

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27
Q

neural crest migration

A

forms:

peripheral nervous system

endocrine and paraendochrine derivatives

pigment cells (melanocytes)

facial cartilage and bone

connective tissue

point of this is that they go all over

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28
Q

E9 twisting of mouse embryo

A

at about E9 the embryo turns
ends up bent around the other way - the “normal way”
from endoderm on outside - “belly out”
to ectoderm on outside - back facing out

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29
Q

phylotypic stage

A

head somawhat distinct
gut tube present
neural tube present
somites present

v early embryogenesis can look different throughout vertebrate phylum (due to environmental adaptation)
BUT it always converges on this v similar (relatively speaking) form
THE PHYLOTYPIC STAGE wit all the same basic architecture

then they go on to diverge again after this

30
Q

circulatory system basic

A

forms from 2 layers of endothelium
in the middle - joins to form a simple 2 chambered heart
-left and right endocardial tubes cometogether
-end up with aortic sac
>right and left aortic arches
>right and left Sinus Venosus
then get folding to form 2 (and then 4 sometimes) chambered heart

in mammals this becomes a 4 chambered heart

31
Q

E12 onwards

A

mostly organigenesis

endoderm makes a tube
forms:
-gut
-lungs
-liver
-pancreas

branched from the gut bud off and form these organs

32
Q

bone formation

A

from mesoderm
commited cartilage cells
compact
form compact nodules
chondrocytes proliferate from this node
as they proliferate hypertrophic chondrocytes eventually form bone

so bone forms in oler parts nearer middle as bone extends

33
Q

intermediate mesoderm forms:

A

reproductive system and kidney

pronephros
nephric duct growing
nephrogenic cord next to duct
mesonephros form there
mesonephros attach to nephric duct
pronephros go away??
metanephrogenic mesenchyme elow these - connected to lower part of nephric duct

cloaca at end of duct

gonad forms next to the mesonephros

34
Q

foetus in fetu

A

where you get twinning

a delayed embryo can become enveloped by the other twin
end up with a delayed foetus growing within another animal

35
Q

generation of in vitro cell types needs understanding:

A

of fate choices
so you can replicate them in culture
understanding developmental decisions is important dor generating in vitro cell types from pluripotent cells

36
Q

understanding genetic defects:

A

the more like a mammal you are
the more relevant to mammalian development genetic studies will be

37
Q

Pax6 mutant similar in mammals

A

Pax6 mouse mutant homozygote has no eyes
heterozygote small eyes

equivalent defect in humans - Aniridia - lack of iris - from Pax6+/-
small eyes was what we could recognise in mice - couldnt ask them how they say - turns out they also lacked an iris

38
Q

public health perspective of development study

A

pregnant people given folic acid supplements to reduce neural tube defects that lead to exposed nerves

curly tail neural tube defect in mice is not rescuable by folic acid - instead inositol treatment protects
-can protect against other defects in humans that folic acid can’t

looking at mouse defects can have impact on human public health issues - like spina bifida and other neural tube defects

39
Q

mouse as good model animal

A

short 9wk generations (shirter than zfish)
large litter size for mammal
easy husbandry
experimental embryology is possible at specific stages
excellent for developmental genetics
-mutagenesis screens
-targeted mutations

40
Q

preimplantation development

A

in the oviduct:
free floating embryo pre-implantation
-fertilisation
-few rounds of cell division
-passes through oviduct
-emerges in uterus as a blastocyst
-then implants in the uterus

it is easy to culture while in this free floating period
can culture from fertilised egg to blastocyst
can replicate oviduct conditions nicely

can study in detail the things that happen to the embryo ready to implant in nucleus

41
Q

4 cell stage polar body

A

extra bump on 4 cell stage
this is polar body
extruded chromosomes not needed (from X inactivation ig??)

42
Q

8 cell to 9 cell stages

A

compaction is occurring
flattening
then after this cavity forms - becomes blastocyst
cavity expands
acellular protective membrane surrounds the blastocyst
then hatches out of this in order to implant

43
Q

mouse preimplantation embryos stages:

A

cell division w/out growth here - cleavage division
gives the 8 cell stage
the sides of the cell flatten into each other - compact to form the MORULA
then these cells secrete fluid into cavity to become blastocyst

maternal RNA used until 2 cell stage
then zygotic transcription begins
(opposed to zebrafish where zygotic genome begins activating after the synchronous divisions enf and the asynchronous ones begin)

44
Q

in compaction to form morula:
and morula polarisation

A

cells flatten
tight junctions and gap junctions form
cells polarise
-inside and outside distinction
-apical outisde w microvilli
-and basal on inside
-cells can wither divide symetrically in circumferential plane to give identical cells
-or divide asymmetriucally in radial plane to give non polarised cells in the middle
gives difference between the inside and outside of embryo
-these outer polarised cells become the trophectoderm

45
Q

trophectoderm vs ICM lineage decision

A

E2.5 - Oct4, Cdx2 in all cells
E3.5 - Oct4 in ICM only, Cdx2 in trophectoderm only

how does Cdx2 become restricted to TE

46
Q

Cdx2 restriction to future trophectoderm cells

A

Hippo signalling pathway regulates Cdx2 expression
-8 cell stage - Yap protein in all cells
-in late morula - big difference in Yap localisation between cells - outer have nuclear Yap, Inner have cytoplasmic Yap
(outer cells w nuclear Yap become TE
inner cells w cytoplasmic Yap become ICM)

This occurs via differential activation of Hippo signalling
-Hippo is active in inner cell:
>activates LATS kinase
>LATS phosphorylates Yap - Yap can no longer co-localise with Tead4 co-TF
>so doesnt go to nucleus - TEAD4 alone leaves Cdx2 OFF
>Cdx2 OFF in inner cells

-Hippo is inactive in outer cell
>hence Yap can co-localise with tead4 - activating Cdx2 in outer cells

47
Q

consolidation of TE and ICM separation

A

mutual repression by Oct4 and Cdx2

consilidates difference between trophectoderm and pluripotent ICM lineage difference once it is set up

48
Q

one more lineage segregation after TE and ICM before implantation:

A

preimplantation cell types:
depends on cells having slightly different FGF signalling levels (similar to the diff levels of notch and delta leading to lineage segregation)

Cells with lower FGF signalling
through Feedback loops secrete more FGF
but have low capability for FGF signalling
-Become epiblast

Some ICM cells have high levels of FGF signalling
-become the primitive endoderm (hypoblast?????????? - looks like that on diagram)

49
Q

ICM vs primitive endoderm lineage segregation markers

A

nanog (ICM) and gata4 (PE) used as lineage markers

segregation of these factors is important for the lineage differentiation
Nanog important for epiblast formation
Gata4/6 for PE

50
Q

Epiblast and primitive endoderm segregation mechanism

A

Future epiblast:
-Oct4+
-secrete FGF (due to Oct4)
-Oct4 promotes Nanog
-Nanog inhibits Gata6

future primitive endoderm:
-receive FGF signalling from future epiblast
-FGF4 signalling inhibits Nanog
-so Gata6 no longer inhibited by nanog so much (gata6 also inhibits Nanog - feedback)
-Gata6 activates Gata4 and FfgR2 (positive feedback by upregulating Fgf sensitivity)

thought that these two layers are different adhesively so separate out

51
Q

pre implantation lineage segregations - mammal and amniote specific

A

ICM-TE segragation - mammal specific

Epiblast-Primitive endoderm - amniote specific

Totipotent blastomeres/ICM/Epiblast:
>totipotent
>give rise to embryo

trophoblast/primitive endoderm:
-make up extra-embryonic structures

trophoblast is mammal specific

52
Q

extra-embryonic lineages importance:

A

for interactions

trophoblast important for interactions (placenta forms from it (in placental mammals) i think as well hence tropho)
Primitive endodewrm will be important for patterning the embryo

53
Q

embryonic layers at implantation

A

has epiblast lineage now
with 2 extraembryonic lineages

polar trophectoderm goes on to differentiate into polar (at the top) trophectoderm
and mural (at the wall[of blastocoel ig]) trophectoderm
atm are just trophectoderm

blastocyst hatches
burrows into uterine lining

54
Q

embryo 1 day after implantation

A

dramatic shape change
in mouse epiblast makes U shaped cup
cavity in the middle of the cup and around the outside

polar trophectoderm has divided a lot and has made a stock known as the egg cylinder

primitive endoderm has extended to cover inside (migrates all around the cavity) and outside of egg cylinder and lines the outside of the polar trophectoderm

cells derived from the mural trophectoderm - trophoblast giant cells on outside

now have eggcylinder - primitive endoderm - and epiblast cup
now to get epiblast ready for gastrulation

55
Q

Mouse AP axis formation in the cylinder shape (as opposed to the disc one earlier)

A

epiblast and polar trophectoderm are lined by PE
2 diff cell types within the PE

formed because Ptrophectoderm signals to the epiblast - with BMP4 signalling it induces Nodal and Wnt3 expression at rim of cup (proximal end)

this causes the distal visceral endoderm (descendent of PE at distal end of egg cup)
this begins expressing nodal and wnt3 inhibitors (lefty1, Cer1, Dkk1)

meanwhile - BMP4 from polar ectoderm causes Nodal and wnt3 expression in the proximal epiblast

the distal visceral endoderm moves to one side - this will become the future anterior of the visceral endoderm
neighbouring visceral endoderm follows the distal visceral endoderm- forming the anterior visceral endoderm

formation of the AVE causes the nodal and wnt3 expression to move to posterior of the epiblast cup

somewhat forming an AP axis

56
Q

anterior visceral endoderm action

A

reaches the embryo/extraembryonic junction
inhibiting nodal and wnt3 at the anterior
-this induces the anterior extoderm (future head and forebrain

57
Q

nodal and wnt3 signalling at posterior of epiblast action

A

-the nodal and wnt3 signalling at the post end causes it to make mesoderm - forming the primitive streak (the structure from whicb mammals make mesoderm)
- cells here also make the definitive endoderm of the gut - which inserts between cells in the primitive endoderm and push them out of the way
-primitive streak initiates gastrulation in the epiblast posterior

58
Q

gastrulation in cup shape mouse

A

cup shape
primitive streak induces at posterior
cells there make mesoderm
mesoderm cells then crawl to the anterior of the embryo - make the mesoderm wings and then line the whole cavity
meanwhile cells have been inserting into the primitive endoerm to make definitive gut endoferm

59
Q

production of the 3 primary germ layers by gastrulation

A

xenopus gastrulates by takinf outer pluripotent cells and bringing them inside the embryo

MOUSE INSTEAD:
takes its inncer cells and pushes them outside to make the mesoderm and endoderm

can envisage the cup as the flat disc from earlier
chicks and humans also do this (flat disc w primitive streak at one end - mesoderm cells migrate forward and eventually make 3 germ layers)

60
Q

fate choice in early embryo

A

after the first 2 lineage segregations from earlier

after these the epiblast makes the mesoderm endoderm and ectoderm
all 3 germ layer lineages will be source of more differentiated lineages

61
Q

germ layer elaboration mouse

A

cup shape of embryo with beginnings of germ layers
-epiblast inside
-mesoderm middle
-endoderm on outer side

one day later see something more reminiscent of typical vertebrate shape
-head fold
-beginning of gut
-still has epiblast w primitve streak at back end of embryo

62
Q

germ layer elaboration - cell movements in the primitive streak region

A

cells move from epiblast layer thru primitive streak where cells do epithelial to mesenchymal transition
then move out of streak as mesoderm

or if they dont move thru streak as mesoderm - will likely move away from primiitve streak and make ectodermal lineage

can have mesoderm and ectoderm lineages coming from same region - just depends whether they did the EMT at mid line

63
Q

similarity of mouse to other vertebrates

A

differences seen as vertebrates approach gastrulation
sizes of embryos
-mouse timy
- chick embryo has v large yolk

then cells divide
and they do diff gastrulation strats:
moving cells inside (fish xenopus)
moving cells outside

all end up at phylotyic stage

64
Q

vertebrate hourglass

A

vertebrates have distinct adult forms
and distinct early embryos

but converge on a phylotypic stage in embryo that looks similar

so hourglass of convergence and divergence to and past Phylotypic stage

65
Q

frog/fish gastrulation strat

A

mouse pulls mesoderm from in to out
frogs pull mesoderm from outside to inside -(along that mesoderm band arround middle of blastula)

mouse is like an inside out frog

66
Q

chick specification of AP axis from external signal

A

used gravity and egg rotation
yolk rotates in shell
causes primitive streak to appear on one side

67
Q

Axis determinants in mouse

A

evidence against:
-absence of yolk, early genome activation
-huge capacity for regulation

68
Q

regulation in preimplantation embryos - evidence against axial determinants

A

can take 2 morula embryos and sit next to each other in well
morulas stick together readilt and form chimeric embryo
survive just as efficientpy as non chimeric embryos and implant in foster mother

-suggests lack of obligate AP axis defines it yet as smashing two diff morula together is feasible
can also sort out inner or outer cells and make mouse out of each - also same survival rates as non chimera
also suggest lack of AP axis determines

69
Q

evidence for cytoplasmic determinants in mouse zygote

A

biasing towards animal or vegetal cytoplasm by removing part of zygote - gives exact same embryos

so no evidence for determinants here

or could there be
as there was a low frequenct of liveborn mice (not jsut looking at embryi formation but survival post birth)
could just be due to a detrimental effect of general loss of cytoplasn - not necessarily due to axis determinants

70
Q
A

DB3 (11 decks)

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