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Zebrafish as a model for studying regeneration
Types of Regeneration

Gets slower and slower as the
animal gets older

The zebrafish can heal his own
heart, which has become the focus
of scientific and media attention in
the past few years …

This involves the formation of a blastema, which is the starting point
of regeneration
...

e
...
is the newt limb
...
(They then proliferate heavily and
redifferentiate into needed structures)
...

This is not true regeneration
...


Repair of one cell type
• Local damage rather than whole organ
• Good example is human skeletal muscle

Why is regenerative medicine so important?
Humans and other mammals have lost the ability to replace lost or damaged body parts
...
This is being outstripped by disease caused by an aging population and lifestyle
...
Analysing animals
that can regenerate naturally will give us insights into how we might provoke regeneration in humans
...
In the vertebrate lineage, only the lower vertebrates can regenerate as
adults
...
Lower
vertebrates are structurally and genetically similar to humans
...

Can respond to infection in similar ways to us
...


natural ability to regenerate
...


Zebrafish as a model for studying regeneration Page 1

natural ability to regenerate
...


Can regenerate a
large number of
organs!

The fin is analogous to limb…

Zebrafish Fin Regeneration
Caudal fin amputation regenerates within 7-21
days post amputation True epimorphic
regeneration – formation of a blastema
...

Possible source of cells is the cut ray
(osteoblasts) and the intra-ray fibroblasts
...
Almost completely
replaceable
...
(Not
essential in fish viability)
...

Can put chemicals in water, e
...
N-ethyl-N-nitrosourea (ENU),
to cause point mutations… can make mutant knockout/down)
...
Lines bred
and positional cloning identifies the specific
mutated genes, therefore genes important
in regeneration
...
Many have been found
...


Chemical screens are beginning to identify compounds that can alter regeneration, by using
commercially available drug libraries
...
(Drugs can be put in water at
specific time points and it is possible to look at the dose-response relationship)
...
g
...

Leads to a problem reorganising tissue and
patterning
...
This drug causes an
irreversible effect, as even when drug is
removed, the fin still does not regenerate
...
By adding AGN192403 at different times during regeneration it was discovered that
the drug disrupts outgrowth of the fin
...
Finding specific pathways in
fin regeneration will allow us to recognize differences between the mechanisms present in the
fin and those in mammals, in order to determine why mammals have lost their ability to
regenerate their limbs
...


Zebrafish as a model for studying regeneration Page 2

Zebrafish Heart Regeneration
Injuring zebrafish
hearts is simple, via
three methods
...


The process is
rapid
...

However the
tissues are the
same as the
human
...

Positioned
close to the
head
...
The process

of regeneration involves these three layers as well:
• A blood clot forms at the apex where the damage is (all
three tissues have been damaged), activating the
endocardium
...

• The activation and proliferation of GATA4+ cardiomyocytes
is promoted by endocardial production of retinoic acid
...


• Within roughly 30 days, you can no longer see the damage
...

Look at the difference between these cells in zebrafish and humans!)
...
Although mammalian epicardium
produces growth factors and the endocardium produces retinoic acid which encourages cardiomyocyte
proliferation, mammals don’t have GATA4+ cardiomyocytes
...

Zebrafish cardiomyocytes are
Studies show that, cardiomyocyte dedifferentiation, which is typically
MONONUCLEATE
...

mononucleate too; however
This dedifferentiation phenotype is reminiscent of those seen in embryonic
mammal adult hearts only
mammalian cardiomyocytes! Revert our cells back?? Find signalling pathways
...

The neonatal mouse heart has been shown to possess robust regenerative capacity during a short
window of time after birth
...

Acute myocardial infarction causes necrotic muscle and cardiac scar which is non -contractile
...
Other therapies are needed for treatment, and
by looking at the mechanisms in species that naturally regenerate this could provide valuable insights for better
treatments
...

Zebrafish as a model for studying regeneration Page 3

Retina Regeneration in Zebrafish
Loss of retinal cells is one of the key problems with aging
...

Structurally very similar to the human eye (histology)
...
Muller cells are a population of glial
cells in the inner nuclear layer (INL), and they dedifferentiate and become stem cell like
...

• Dying photoreceptors signal to Muller cells,
activating them, causing them to become
embryonic-like by expressing a stem
marker, Pax6
...

• Some cells migrate to the outer nuclear
layer (ONL) and differentiate into rods and
cones
...

While mammalian Muller cells can dedifferentiate, they undergo symmetrical division and form a scar
...
By
stopping Pax 6 in Muller cells, this stops the regeneration process
...
Turn on
expression of notch in mammals?? This asymmetric
division causes the clustering which is crucial
...


SUMMARY

Zebrafish display rapid epimorphic regeneration in a
number of important tissues
...
Ongoing
chemical screens will identify drugs that promote
regeneration
...


Multiple cell cycles can be
dangerous to humans, as we have
long lives and don’t want
mutations
...
Within a couple of hours of
amputation the wound begins to heal forming a ‘wound epithelium’ ( Stoick-Cooper et al
...
Like the
limb bud, the blastema is an outgrowth of mesenchymal cells that are already tissue restricted and
covered by an epithelium
...
, 2014)
...
In both cases the ultimate goal is for undifferentiated
Zebrafish as a model for studying regeneration Page 4

involved in regeneration (Nacu and Tanaka, 2011)
...
In embryonic development, the cells migrating from various sections of mesoderm have
the potential to differentiate into multiple tissues, compared with cells originating from adult tissues
...
, 2009)
...
This forms a
heterogeneous collection of undifferentiated mesenchymal cells that are divided into distinct subsections;
however under a microscope they all have the same histology (Kragl et al
...

Since the cells of the blastema have identical histology, it was long thought that all the cells were
dedifferentiated back to a state of pluripotency, with the ability to form any cell of the limb
...
g
...
In fact the mesenchymal cells of the blastema have restricted potential based
upon their adult tissue origin (Kragl et al
...
For instance, blastema cells originating from adult
muscle tissues will only differentiated into muscle in the newly regenerated limb, hence the cells are
progenitors of muscle
...
,
1986)
...
The only progenitors that show a difference from their adult tissue
origin are those from dermis tissues
...
In the embryo
dermis is formed from the lateral plate mesoderm, as are the cartilage cells, hence the cells from dermis
origin in the blastema have reverted back to a lateral plate mesoderm progenitor from the neurula stage of
the embryo
...
, 2009)
...


Zebrafish as a model for studying regeneration Page 5



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