STEM CELLS

Undifferentiated (unspecialized)
cells

(i) that can renew themselves

(ii) can give rise to one or more

specialized cell types

(iii) each new cell type has

specific functions
Embryonic Stem Cells
 HUMAN STEM CELLS

Embryonic Fetal Infant Adult

Blastocyst
(5-7 days) Gonadal ridgeAbortus Umbilical Whartons GermlineSomatic
(6 weeks) (fetal tissues) cord blood Jelly
Embryonic
stem cellsEmbryonic Fetal Umbilical cordUmbilical cord Oogonia
Spermatogonia
(HES) germ cellsstem cells blood stem cells
matrix stem cells
(HEG) (FSC) (UCB) (UCM)

HemopoieticMesenchymalLiver Epidermal Neuronal Eye Gut Pancreas?

(skin,hair)

Bone PeripheralBone marrow

marrow blood stroma
 ICM
Human
blastocyst
TE

ICM (hES
cell)

Hypoblast
Epiblast

Yolk sac
Ectoderm Mesoderm Endoderm Germ
cells

Skin NS CT Eyes RS GI Liver

Sperm
Ears Pancreas
Eggs

Muscle UGS VS Bone Cartilage BM

Cardiac Skeletal Smooth

 Cultured BlastocystIsolated Inner Cell Mass
First Plating

Cultured inner Mouse fibroblast

cell mass feeder cells

inner cell mass

Cultured inner 9 - 15 days

cell mass clusters
trophectoderm

7 - 10 days Replatedonto
new feeder cells
hES cells

Established Cultures Second Plating to Establish Colonies

A B

C D
Human Embryonic Mouse Embryonic
Stem Cells Stem Cells
Collagen I HEM Matrigel
Laminin

M. Richards, C-Y Fong, W-K Chan, P C Wong, A.

Bongso, Nature Biotech, (2002) 20: 933-936
 Produce ES cells
ES cells on
feeder
cells

Heart Nerv Blood

e
Blastocyst
c
c t ri
Ele lse
pu
Tranplantatio
Donor n to correct
Recipient cell disease or
enucleated injury
egg
Cell
(Human/animal) Fusion Donor cell for
Autotransplantatio
n
CHARACTERIZATION AND CONFIRMATION OF PLURIPOTENCY

Surface marker SSEA-1 -ve

antigens: SSEA-3 +ve
SSEA-4 +ve
GCTM +ve
TRA-1- +ve
60
TRA-1- +ve
81

Molecular genetic Endoder AFP

markers: m:
Mesoder BMP4
m:
Ectoderm NEURO
:OCT 3/4; TDGF1;
D1 Sox2; Rex1;
LeftyA
SCID mice teratoma
formation
Alkaline phosphatase +ve
Cytogenetics
Embryoid body formation
Chimaeras (blastocyst)
 hES CELL LINES: NIH REGISTRY
78 hES cell lines (All on MEF-Xenoproteins)

12 hES cell lines available for shipping (All on MEF-

Xenoproteins)

* HES1, HES2, HES3, HES4, HES6 (ESI Singapore)

Genetic diversity limited

Gender
Ethnicity
HLA typing

Behavioural differences between cell lines

Characterization
Early differentiation markers
In vitro
In vivo
 XENOSUPPORT AND XENOPROTEINS
hES CELL LINES
EMBRYOS
Culture media + HS
ICSI (Bovine/Ovine hyaluronidase)
Freeze-thawing: HS
DERIVATION
Guinea pig complement
Rabbit anti-human antibodies
Murine embryonic fibroblasts (MEF)
DMEM + FCS
Bovine insulin-porcine transferrin-selenium
(ITS)
Feeder-free supports: animal origin
PROPAGATION
Murine embryonic fibroblasts (MEF)
DMEM + FCS
Feeder cell attachments: Gelatine
Passaging: Trypsin
Bovine insulin-porcine transferrin-selenium
(ITS)
Feeder-free supports: animal origin
 USES OF ES CELLS

Transplantation therapy
ES cell directed tissue transplantation
ES cell-scaffold organ transplantation

Understanding normal human development

What goes wrong in birth defects,
cancer

Testing drugs for safety and potential

efficacy

Gene therapy
Terada N et al, Nature
Ying QL et al, Nature
(2002)
Dental advance will let adults grow n
Stem cell plan to grow human teet

Explores the possibility of growing new teeth

in the lab or inside the mouth as a
replacement. "Five years from now, we will
know if in principle we can produce human
teeth from cultured cells,

Milk teeth are an "attractive source" of stem

cells, he said, though he added it could take
many more years to commercialise, even if
the experiments are successful.
 Recent Progress of Teeth
Stem Cells
Scientists successfully grown a human tooth in a
mouse entirely from cultured cells - they were of the
right size, surrounded by new bone and connective
tissue, and showed evidence of root formation.

There is already a milk tooth stem cell bank operated

by BioEden's labs in Daresbury, Cheshire.

The stem cells are extracted and then frozen until

they are needed for general use: seen as a 'repair kit'
for the body, stem cells are able to turn into more
than 200 different cell types, from muscle to nerve
cells, replacing the diseased, dead and worn out cells
behind a range of conditions.
The figure demonstrates a
scaffold where cells are seeded.
A normal mouse tooth on the left, where ameloblast cells that produce enamel
are glowing in red. On the right is a tooth with the Ctip2 gene deleted, and little
enamel has been able to form. (Credit: Image courtesy of Oregon State
University)
A normal mature mouse incisor is shown on the top, where ameloblasts (am)
that produce enamel (e) is clearly visible. On the bottom is a mature incisor with
the Tbx1 gene deleted, and no enamel and ameloblasts were able to form.
(Credit: Image courtesy of University of Zurich)
What are tooth stem cells?
Tooth stem cells are stem cells found in the deciduous or baby
teeth of children. Recent studies show that baby teeth stem cells
appear to have the ability to develop into more types of body
tissue than other types of stem cells. This difference opens the
door to more therapeutic applications.
What can tooth stem cells do for my child?
Stem cell therapy is emerging as a revolutionary new way to treat
disease and injury, with wide-ranging medical benefits. It works by
introducing stem cells into an area where the normal cells have
lost their function due to disease or damage. The stem cells then
replace or repair the damaged cells and restore normal function.
Is there a difference between baby teeth and wisdom
teeth?
Although it has been known for several years that wisdom teeth
will yield valuable cell populations, research has found that those
cell populations have less therapeutic potential than the cell
populations found in primary, or "baby" teeth. For example, baby
teeth cells more readily form neural tissue, and have the
demonstrated ability to form bone and fatty tissue as well, which
has not been achieved with wisdom teeth cells.
Implanting Stem Cells From Teeth In Humans
Dr. Robey outlined three approaches presently being investigated by
different laboratories to implant stem cells from teeth in humans:

Place the stem cells into a mold of a tooth crown made of an

enamel-like substance along with "scaffold" material. Loop
blood vessels through this "construct" and implant it elsewhere in the
body until it matures. Then, extract it and implant it in the mouth. For
instance, researchers have already been able to use bone marrow to
grow the jaw bone of a man in his shoulder blade until it was mature,
then they extracted it and attached it to his jaw.

Harvest a wisdom tooth, release the stem cells from the

dental pulp then implant the cells directly into a severely
injured tooth. For example, if teeth are seriously injured in a car
accident, usually root canals are needed and the teeth die. Instead,
using stem cells, the goal is to regenerate the pulp inside the teeth.
"It would be like bringing the teeth back to life, sparing them root
canals," Dr. Robey said.

Lastly, if no tooth is present (whether by accident, severe dental

disease, or a genetic condition such as cleft palate) researchers can
take stem cells from unerupted wisdom teeth, organize them into