Stem-Cells and many more

1. Stem cells
Mala Parab

2. Simposium Stem Cell di
FKUI, ULTAH Kalbe 40
tahun, 2 Sept.2006
Definition of stem cells: stem cells are unspecialized cells that have two
defining properties: the ability to differentiate into other cells and the ability to
self-regenerate/self-renew

3. Classification on Basis on Differentiation
property
⚫Totipotent  all cell types. E.g.: zygote
⚫Pluripotent  all three germ layers. E.g.:
human embryonic stem cells
⚫Multipotent  many cell types. E.g.:
hematopoietic stem cells
⚫Unipotent  can produce only one cell
type, but have the property of self-renewal
which distinguishes them from non-stem
cells

4. Classification based on source/ origin

5. Stem Cell Types Based on The Source of Stem Cells
⚫ Embryonic stem cells obtained from the undifferentiated
inner mass cells of a blastocyst, (an embryo that is between 50
to 150 cells)
⚫ Umbilical cord blood  derived from the blood of the placenta
and umbilical cord after birth
⚫ Adult stem cells  undifferentiated cells found among
differentiated cells of a specific tissue and are mostly Multipotent
cells
◦ Bone marrow
◦ Adipose tissue
◦ CNS stem cells

8. Propertiesof adult stemcells
⚫ Astemcell possessestwoproperties:
⚫ Self-renewal, which is the ability to go through numerous cycles of cell
division whilestill maintainingits undifferentiatedstate.
⚫ multipotency or multidifferentiative potential, which is the ability to
generate progeny of several distinct cell types, (for example glial cells and
neurons) as opposed to unipotency, which is the term for cells that are
restricted to producing a single-cell type. However, some researchers do
not consider multipotency to be essential, and believe that unipotent self-
renewingstemcellscanexist.
⚫ These properties can be illustrated with relative ease in vitro, using
methods such as clonogenic assays, where the progeny of a single cell is
characterized..

9. ⚫ Adult stem cells express transporters of the ATP-binding cassette family
that actively pumpadiversity oforganicmoleculesoutof thecell..
⚫ Adult stem cell research has been focused on uncovering the general
molecular mechanisms that controltheir self-renewal anddifferentiation.
⚫ Notch :The Notch pathway has been known to developmental biologists for
decades. Its role in control of stem cell proliferation has now been
demonstrated for several cell types including haematopoietic, neural, and
mammary, stemcells.
⚫ Wnt :These developmental pathways are also strongly implicated as stem
cell regulators.

10. HowDoesCell TherapyWork?
 Stem cells can be used to
generate healthy and functioning
specialized cells, which can then
replace diseased or dysfunctional
cells.
 It is similar to the process of organ
transplantation only the treatment
consists of transplanting cells
insteadoforgans.

11. HowDoesCellTherapyW
ork?
 Bone marrow transplants are an example
of cell therapy in which the stem cells in a
donor's marrow are used to replace the
blood cells of the victims of leukemia.
 Cell therapy is also being used in
experiments to graft new skin cells to
treat serious burn victims, and to grow
new corneas for the sight-impaired.
 In all of these uses, the goal is for the
healthy cells to become integrated into the
body and begin to function like the patient's
own cells.

12. What DiseasesCanbe
CuredbyStemCellTherapies
Any disease in which there is
tissue degeneration can be a
potential candidate for stem
cell therapies
1. Parkinson's and Alzheimer's diseases
2. Spinal cord injury
3. Stroke
4. Burns
5. Heart disease
6. Diabetes
7. Muscular dystrophy
8. Osteoporosis injuries
9. Cirrhosis hepatic
10.Leukemia
11.Sickle cell anemia
12.Osteoarthritis
13.Rheumatoid arthritis
14.Cancer

13. Stem Cell Characteristics Make Them Good Candidates
for Cell-based Therapies
⚫ Potential to be harvested from patients
⚫ High capacity of cell proliferation in culture to obtain large number
of cells from a limited source
⚫ Ease of manipulation to replace existing non functional genes via
gene transfer methods
⚫ Ability to migrate to host's target tissues, e.g. the brain
⚫ Ability to integrate into host tissue and interact with surrounding
tissue

16. Embryonic Stem Cells for Therapy
⚫ Advantages:
◦ Easilyavailable fromfertilityclinics
◦ Pluripotent haveability todifferentiateintocells derivedfromall 3germ
layers but not theembryonicmembranes
◦ Immortal proliferate inculture&maintainedincultureforseveral
hundreddoublings
⚫ Disadvantages:
◦ Tumorigenic anycontaminatingundifferentiatedcells couldgiveriseto
cancer
◦ Alwaysallogenic immunerejection
◦ Ethicallycontroversial

17. Adult Stem Cells for Therapy
⚫ Advantages:
◦ Canbetakenformpatient’sowncells  notrejectedbytheimmunesystem
◦ Alreadysomewhatspecialized: Inducementmaybesimpler
◦ Lessethical problems
⚫ Disadvantages:
◦ Rarein maturetissues difficulttoobtaininlargequantities
◦ Theydon't liveaslonginacultureasembryonicstemcells
◦ Generallylimited todifferentiating into differentcell typesoftheirtissueoforigin
eventhoughplasticitymayexist

18. SkinReplacement
⚫ Theknowledgeofstemcells hasmadeit possible forscientiststogrowskinfroma
patient'spluckedhair.Skin(keratinocyte)stemcells residein thehairfollicle and
canberemovedwhenahair isplucked.
⚫ Thesecells canbeculturedtoformanepidermalequivalentofthepatientsown
skinandprovidestissueforanautologousgraft,bypassingtheproblemof
rejection.
⚫ It is presentlybeingstudied in clinical trialsasanalternativetosurgicalgraftsused
for venousulcersandburnvictims

19. BrainCell Transplantation
⚫ Theidentification andlocalization ofneuralstemcells,bothembryonicandadult,
hasbeenamajor focusof current research.
⚫ Potentialtargetsofneuralstemcell transplantsincludestroke,spinalcordinjury,
andneurodegenerativediseasessuchasParkinson'sDisease.
⚫ Stemcells canprovidedopamine- achemicallackinginvictims ofParkinson's
Disease
⚫ Over250patientshavealreadybeentransplantedwithhumanfetaltissue

20. Cell-based therapy for Parkinson’s Disease

21. Dopamine-NeuronT
ransplantation

22. TreatmentofDiabetes
⚫ Recently,insulinexpressingcellsfrom
mousestemcellshavebeengenerated. In
addition,thecellsselfassembleto form
structures, whichcloselyresemblenormal
pancreatic isletsandproduceinsulin
⚫ Futureresearchwill needtoinvestigatehow
to optimizeconditionsfor insulinproduction
with theaimof providingastemcell-based
therapytotreat diabetestoreplacethe
constant needfor insulininjections

23. Bone/CartilageRegeneration
⚫ Osteocel
◦ 1st commercial product inUS:bonematrixcontainingallogenichuman
mesenchymal stemcells
◦ Launchedin2005byOsirisTherapeutics
⚫ Otherbrands:Chondrogen,TissueRepairCells (TRCs),Prochymal,and
Mesoblast

24. DrugScreening
⚫ Twooftheworld’smostsuccessfulblockbuster biotechnologyproducts–Epogen&
Neupogen–werediscovered&developedthroughtheuseofin vitro assays
involvingbloodstemcell technologies
⚫ Stemcellswhichcanbeexpandedexvivoanddifferentiatedintospecializedcell
typescouldenablethedevelopmentofhigh-throughputscreensfortestingthe
effectsandpossible toxicity ofarangeofdrugsearlyin thedrugdevelopment
pipeline

25. Stemcells–Blindness
⚫ In clinical trials at Moorfields EyeHospital in London,surgeonsrestored eyesight
for six patients wholost their sight after chemicalaccidents andgenetic diseases.
Thepatientswentundersuccessfulstem-celltransplant.
LimbalstemCell therapy
o Thetreatmentis knownaslimbalstemcell
therapy,andthepatientswhoreceived the
treatmentsufferedfromchemical burnor
geneticdiseaseknowasaniridia
o Byreplacingthelimbal stemcells, the
corneabeginstoclear upasthecellsare
replacedwiththehealthytransparentlayer
again.

27. Neural Stem Cells
⚫ Neural stem cells (NSCs) are self-renewing, multipotent cells that
generate the main phenotypes of the nervous system.
• They are located in: Sub-ventricular zone lining the lateral
ventricles, where they give rise to newly-born neurons that migrate
to the olfactory bulb via the rostral migratory stream and Sub-
granular zone, part of the dentate gyrus of the hippocampus
⚫ NSCs primarily differentiate into neurons, astrocytes,
and oligodendrocyte.
⚫ NSCs are stimulated to begin differentiation via exogenous cues
from the microenvironment, or stem cell niche. This capability of
the NSCs to replace lost or damaged neural cells is
called neurogenesis.
The ependymal cells and astrocytes form glial tubes
are used by migrating neuroblasts. The astrocytes in the tubes
provide support for the migrating cells as well as insulation from
electrical and chemical signals released from surrounding cells. The
astrocytes are the primary precursors for rapid cell amplification.
⚫ Neurons usually show radial migration pattern.

28. Section of the hippocampus,
blue dots are neural stem
cells
Mature neuron (red)

29. ⚫ Functions of NSCs during differentiation:
Radial, astrocytes-like, GFAP-positive cell model:
The niche composed of blood vessels, astrocytes, microglia, ependymal cells,
and extracellular matrix promote the quiescent state (Type B) of NSCs.
Once activated, the Type B cells develop into Type C cells, active proliferating
intermediate cells, which then divide into neuroblasts consisting of Type A
cells.
The undifferentiated neuroblasts form chains that migrate and develop into
mature neurons. In the olfactory bulb, they mature into GABAergic granule
neurons, while in the hippocampus they mature into dentate granule cells.
NSCs play a vital role during development producing the
enormous diversity of neurons, astrocytes and oligodendrocyte
in the developing CNS.

30. ⚫ Function of NSCs during diseases:
⚫ NSCs are involved in migration and replacement of dying
neurons
⚫ Hippocampal stem cells migration elicited by SDF-1a, a
chemokine during stroke helps to repair the injury
⚫ responses during stroke, multiple sclerosis, and Parkinson's
disease in animal models and humans is part of the current
investigation
⚫ Used to treat neurodegenerative disorders and spinal cord injury.

31. Hematopoietic stem cells
⚫ Hematopoietic stem cells (HSCs) are the blood cells that give
rise to all the other blood cells.
⚫ They give rise to all the blood cell types:
• Myeloid
basophils,
(monocytes and macrophages, neutrophils,
eosinophils, erythrocytes, megakaryocytes/
platelets, dendritic cells)
• Lymphoid (T-cells, B-cells, NK-cells)
• HSCs contain cells with long-term and
capacities
regeneration
committed multipotent, oligopotent,
short-term
and
and
unipotent progenitors.
• HSCs are found in the bone marrow of adults, with large
quantities in the pelvis, femur, and sternum and can mobilize
out of the bone marrow into circulating blood
• Markers: CD 34+, CD 59+, Thy+, CD 38, C-Kit

32. ⚫ Sources of HSCs:
1. Bone Marrow: About 1 in every 100,000 cells in the marrow is a
long-term, blood-forming stem cell. Clinically used for bone
marrow transplants.
2. Peripheral blood: It has been known for decades that a small
number of stem and progenitor cells circulate in the bloodstream,
but in the past 10 years, researchers have found that they can coax
the cells to migrate from marrow to blood in greater numbers by
injecting the donor with a cytokine, such as granulocyte-colony
stimulating factor (GCSF). Clinically used for autologous and
allogenic transplants blood cell transplants.
3. Umbilical Cord Blood: There have been suggestions that umbilical
cord blood contains stem cells that have the capability of developing
cells of multiple germ layers (multipotent) or even all germ layers,
e.g., endoderm, ectoderm, and mesoderm (Pluripotent). Have been
used in treatment of Fanconi anemia

33. 4. Fetal Hematopoietic System: Hematopoietic cells appear early in
the development of all vertebrates. the earliest hematopoietic
activity is indicated by the appearance of blood islands in the yolk
sac (day 7-8) and in the AGM—the region where the aorta,
gonads, and fetal kidney (mesonephros) begin to develop. an HSC
in the bone marrow has four actions in its repertoire: 1) it can
renew itself, 2) it can differentiate, 3) it can mobilize out of the
bone marrow into circulation (or the reverse), or 4) it can undergo
programmed cell death, or apoptosis. Understanding the how,
when, where, which, and why of this simple repertoire will allow
researchers to manipulate and use HSCs for tissue and organ
repair.

34. Clinical Uses of HSCs:
1. Leukemia and Lymphoma: Among the first clinical uses of HSCs
were the treatment of cancers of the blood—leukemia and lymphoma,
which result from the uncontrolled proliferation of white blood cells.
In these applications, the patient's own cancerous hematopoietic cells
were destroyed via radiation or chemotherapy, then replaced with a
bone marrow transplant, or, as is done now, with a transplant of HSCs
collected from the peripheral circulation of a matched donor.
2. Inherited Blood disorder: treatment of hereditary blood disorders,
such as different types of inherited anemia (failure to produce blood
cells), and inborn errors of metabolism (genetic disorders
characterized by defects in key enzymes need to produce essential
body components or degrade chemical byproducts). The blood
disorders include aplastic anemia, beta-thalassemia, Blackfan-
Diamond syndrome, globoid cell leukodystrophy, sickle-cell anemia,
severe combined immunodeficiency, X-linked lymphoproliferative
syndrome, and Wiskott-Aldrich syndrome. Inborn errors of
metabolism that are treated with bone marrow transplants include:
Hunter's syndrome, Hurler's syndrome, Lesch Nyhan syndrome, and
osteopetrosis.

35. 3.Rescue for Cancer Chemotherapy: cancer
patients can be given an autologous stem cell
transplant to replace the cells destroyed by
chemotherapy. Once the drugs have washed
out of a patient's body, the patient receives a
transfusion of his or her stored HSCs.
Epithelial Stem cells:
• These give rise to epithelial cells which
constitute 60 percent of the differentiated
cells in the body.
• Responsible for covering the internal (i.e.
intestinal lining) and external surfaces (i.e.
skin) of the body, including the lining of
vessels, glands, and other cavities.
• Epithelial stem cells are also found in the
bulge region of the hair follicle

36. ⚫ In skin epithelial stem cells are usually located in the basal lamina,
near the dermal region. Based on the proliferative and
morphological characteristics, Potten (1974) coined the term
‘‘epidermal proliferative unit’’(EPU) to describe skin stem cells.
⚫ The EPU is a stratified squamous epithelium constituted by
different layers of cells. The innermost layer, called the basal layer,
is strongly attached to its underlying dermis and contains
mitotically active progenitor cells that divide and give rise to the
differentiated suprabasal cells, which eventually differentiate in to
different component cells of skin.
⚫ The IFE continuously self-renews throughout life to replace the
cells that are constantly sloughed off from the skin surface. In mice
and humans, it takes about 3 to 4 weeks to replace all the cells of
the IFE.

37. ⚫ Applications of Skin Stem cells:
1. Skin stem cells for treatment of burns: Human keratinocyte SC
cultured on fibroblast feeder layers, have an enormous
proliferation potential, and only few cells can regenerate sufficient
keratinocytes to cover the all-human skin surface. These cultured
human keratinocyte SC can differentiate and reform a functional
skin barrier that can be transplanted into patients suffering from
severe burnt injuries.

38. 2. Cellular therapy for inherited genetic diseases:
Skin is a highly accessible source of SC, which have an
extraordinary capacity of cellular expansion in culture, and that
can be stably genetically modified. here are many different
genetic diseases affecting the skin epidermis. Among the most
devastating genetic skin diseases are the epidermolysis bullosa,
which is characterized by an extreme fragility of the skin that
results in painful blistering lesionss and require long time to
heal. ue to permanent state of wound healing that stimulates SC
proliferation, these patients are at high risk of developing skin
cancer.
The group of De Luca, have demonstrated that correction of a
severe form of epidermolysis bullosa could be achieved by
transplantation of genetically modified keratinocyte SC.

39. isolated keratinocyte SC from a
⚫ They
patient presenting epidermolysis bullosa
caused by mutation in the gene encoding
laminin 5, a critical component of
hemidesmosomes.
⚫ They expanded these skin SC in vitro and
transduced these cells with a retrovirus
expressing Laminin 5.
⚫ They isolated cells stably expressing
Laminin 5 and made genetically corrected
cultured epidermal grafts that have been
transplanted onto the patient skin lesions..
⚫ The genetically corrected skin tissue
healed perfectly well and biopsies from the
graft demonstrate that the recombinant
skin presents a normal histology and the
expression of the transgene remains stable
even a year after the treatment.

40. ⚫ Intestine Stem cells:
⚫ The gastrointestinal epithelial stem cell niche is believed to be
formed and maintained by the underlying cells of the
mesenchymal lamina propria and their secreted basement
membrane factors, which regulate stem cell behaviour through
the paracrine secretion of various growth factors and cytokines,
the receptors for which are situated on gastrointestinal epithelial
cells
⚫ The epithelial stem cell lineages of gastrointestinal tract undergo
constant turnover, with complete self renewal every 2-7 days
under normal circumstances
damage.
⚫ The Notch signaling pathway
and increases following tissue
and E2F transcription factor
regulates gastrointestinal epithelial cell fate and differentiation of
the four specialized epithelial lineages of the gastrointestinal
tract. Increased levels of Notch protein negatively regulate the
transcription of the Math1 gene, a basic loop-helix transcription
factor, via up regulation of the Hes1 transcriptional repressor.

41. ⚫ Molecular markers for identification of GISCs: LGR5, BMI-
1,CD133 are markers of crypt stem cell population.
⚫ The GISCs are proposed to be used for treatment of colon cancer,
chronic ulcers and transplants for small intestine.
⚫ Mammary stem cells:
⚫ The mammary gland in humans and in other mammals is a
dynamic organ that undergoes significant developmental changes
during pregnancy, lactation, and involution.
⚫ Under the regulation of systemic hormones, as well as local
stromal epithelial interactions, these cells proliferate extensively,
differentiate during each pregnancy and lactation, and undergo
apoptosis during mammary involution the mammary gland is
subjected to major changes in morphology during distinct
developmental windows
⚫

42. ⚫ In humans, the mammary epithelium consists of a network of
ducts that form before birth, by branching and invading the
mammary fat pad. The ducts are formed by a basal layer of
contractile, myoepithelial cells and a luminal layer of specialized
epithelial cells.
⚫ During puberty, ductal outgrowth rapidly increases under
hormonal stimulation, resulting in side branching.
⚫ The final differentiation stage is achieved in the mammary gland
during pregnancy and lactation, when numerous lobulo-acinar
structures containing the milk-secreting alveolar cells are formed
through extensive proliferation, followed by terminal
differentiation. Cessation of lactation following weaning is
accompanied by massive apoptosisand tissue remodelling, and
the gland reverts to a structure resembling that before pregnancy.

43. ⚫ The existence of self-renewing, multipotent mammary stem cells
was first suggested decades ago by the work of Daniel et al. Their
studies in mice and rats demonstrated that an entire mammary
gland can be generated from serially transplanted random
fragments of epithelium.
⚫ Mammary stem cells are usually represented as a population of
small light cells without polarity.
⚫ Cellular Markers: ESA+ , MuC1-, , alpha 6 integrin+, CD10+,

44. Table:
I]Examples of genes with
overlapping
neurospheres,
haematopoietic stem cells
and embryonic stem cells
1. Integrin, beta 1
2. Growth hormone receptor
3. Platelet-derived
factor receptor,
polypeptide
5. Policystic kidney disease
2
II.]
and
Genes upregulated
mammospheres
neurospheres
3. Fibroblast growth factor
receptor 1
expression 4. Tenascin C
patterns in mammospheres, III]Genes upregulated in
mammospheres and
haematopoietic stem cells
1. Notch 3
2. Latent transforming
growth factor beta binding
growth protein 3
beta 3. Apolipoprotein E
4. GATA-binding protein 2
4. p53 target zinc finger
protein 1V]
mammospheres
Genes upregulated in
and
in 2.
embryonic stem cells
1. Nidogen 1 and 2
Glypican 4
3. Insulin-like growth factor
binding protein 4 and 7
1. p59fyn (FYN) oncogene
2. ATP-binding cassette, 4.
subfamily A, member 1
WNT1 inducible signalling
pathway protein 1

47. ⚫ The Notch trans-membrane receptor proteins are part of a
signaling pathway that is critical for the correct developmental
fate of cells and various tissues.
⚫ Activation of the Notch pathway results in changes of cell fate,
such as proliferation of undifferentiated cells, blockage of
differentiation or differentiation along a particular lineage.
⚫ The vertebrate Notch 4 gene has been shown to be involved in
normal mammary development. In vitro, over-expression of
the constitutively active form of Notch 4 inhibits differentiation
of normal breast epithelial cells. In vivo, transgenic mice
expressing a constitutively active form of Notch 4 in the
mammary gland fail to develop secretory lobules during
gestation, and subsequently develop mammary tumours. Thus
alterations in Notch 4 signalling might play a significant role
in breast-cancer development

48. ⚫ Apart from Notch 4, over expression of Wnt signaling pathway,
β- catenin, factors like APC, TCF-1 in mouse mammary glands
increased MMTV promoter increases mammary tumor formation.
LIF and PTEN are two other proteins involved in tumorogenesis
of mammary glands.
⚫ Applications:
1. In-vitro developed mammospheres have been used as models to
study evolution of breast cancers. Such models enables study of
chemotherapeutic agents on receptors like estrogens, BRCA-1
and integrin mediated signal transductions.
2. It allows one to bring about genetic manipulations followed by
transplants as a part of treatment .
3. Recent research demonstrated that mammary stem cells respond
to both hormone withdrawal and stimulation. Thus anti-
hormone therapy, can be used as a treatment aimed for
preventing breast cancer recurrence
4. Mammary stem cells have been shown to repopulate the fat pad
of mammary glands post intensive cancer therapy.

49. In the presence of female hormones, the
mammary stem cell outgrows to generate an
extensive mammary tree (left). Following
hormone deprivation, the mammary stem cell
has reduced outgrowth potential (right).

50. ⚫ In the presence of female hormones, the mammary stem cell
outgrows to generate an extensive mammary tree (left).
Following hormone deprivation, the mammary stem cell has
reduced outgrowth potential (right).
⚫ Prostate Stem cells:
⚫ Prostatic epithelium is composed of multiple differentiated cell
types, including basal, luminal (secretory), and neuroendocrine
cells.
⚫ Luminal secretory cells make up the majority of the epithelial
layer and because they express androgen receptors (ARs), they
can respond directly to androgens by simulating production and
secretion of prostatic proteins, such as prostate-specific antigen
(PSA) and prostatic acid phosphatase.
⚫ In the human prostate, basal cells form a continuous layer,
whereas in other species they are more scattered in appearance.
⚫ Prostatic epithelial cells are identified by their morphological
appearance, location, and also distinct patterns of marker
expression. Basal cells express cytokeratins (CKs) 5 and CK14

52. ⚫ Linear, bi-directional and lineage differentiation models are three
models proposed for in-vivo development of prostrate glands.
⚫ A diagnostic feature of human prostate cancer is the loss of basal
cells.
⚫ Prostate cancer can potentially arise from oncogenic
transformation of CK5C8K basal cells resulting in rapid
differentiation to a luminal phenotype, or alternatively from stem
or multipotent progenitor cells within the CK5C8C intermediate
or CK5K8C luminal populations where stem cells or CARNs are
proposed to reside.
⚫ Multiple lines of evidence demonstrate initiation of prostate
cancer from luminal (and possibly intermediate) cells, based on
targeted gene disruption by Cre-recombinase under the control of
the probasin (either probasin or ARR2/probasin) or PSA
promoters that show luminal-specific expression.

53. ⚫ A body of work by the Witte Laboratory provides evidence to
suggest that basal cells can initiate preneoplastic and cancerous
lesions. For example, loss of PTEN negatively regulates p63C
prostatic basal cell proliferation without
differentiation, resulting in an expansion of a
blocking
prostate
stem/progenitor-like subpopulation, using defined cell lineage
markers including Sca-1C, Bcl-2C, and CD49fhi cells.

54. Bone and Muscle Stem cells
⚫ Skeletal muscle is a dynamic tissue that is capable of responding
to physiological stimuli (i.e., intense exercise training) or a severe
injury by mounting a well orchestrated regenerative response that
restores the cytoarchitecture within a 2-wk period.
⚫ The capacity for this regenerative response is primarily due to a
mononuclear cell population termed satellite cells, usually present
in peripheral (or satellite) position in relation to the adjacent,
larger multinucleated myofiber.
⚫ In response to injury or disruption of the basal lamina, the satellite
cells become activated and have a remarkable proliferative
capacity.
⚫ Ultimately, the satellite cells either fuse to form multinucleated
myotubes or re-establish a residual pool of quiescent satellite cells
that have the capability of supporting additional rounds of
regeneration

55. ⚫ Skeletal Muscle development during embryogenesis
Skeletal muscle formation is derived from the paraxial mesoderm
early during vertebrate embryogenesis.
The paraxial mesoderm coalesces to form segmented, epithelial
spheres referred to as somites that occupy paired structures on
either side of the neural tube.
The somite is further specialized to produce the dermomyotome the
sclerotome.
These progenitors migrate beneath the dermomyotome to form the
myotome and adopt a skeletal muscle fate.
In addition, proliferating cells (Pax3+Pax7+ cells) from the central
dermomyotome migrate directly to the myotome and continue to
proliferate without expression of differentiation markers such as
members of the MyoD family.
MyoD transcriptional factors are expressed in somites and
established skeletal muscle during embyogenesis

56. ⚫ In adults muscle stem/precursor cells including
the myogenic satellite cell population originate
somite and vascular components
principally quail cells, bone marrow cells,
like
endothelium, pericyte, mesoangioblast
⚫ Satellite cells lack expression of the myogenic
regulatory factors, including members of the
Myod family
⚫ A tyrosine kinase receptor (Met, for hepatocyte
growth factor) is expressed broadly during
development in the myotome of the somite and
neural crest derivatives and is localized to the
quiescent satellite cell population in adult
skeletal muscle
⚫ These studies demonstrate that myogenic
stem/progenitors are capable of generating
nonmuscle lineages including blood
derivatives, vascular components, osteoblasts,
and adipocytes in response to specific signals

57. ⚫ For example, satellite cells that are cultured in the presence of
bone morphogenic proteins (i.e., Bmp2) promote the osteogenic
molecular program with an induction of the master transcriptional
regulators Runx2 and Osterix
⚫ Morphological analysis reveals interstitial edema with neutrophils
that release trophic factors to activate the satellite cells within 2 h
of injury
⚫ Following activation of the satellite cell population, they re-enter
the cell cycle and demonstrate a significant proliferative capacity
between 2 and 3 d following injury
⚫ This proliferative period is followed by a differentiation phase
where myoblasts withdraw from the cell cycle and form small
basophilic centronucleated myotubes (the hallmark of skeletal
muscle regeneration)
⚫ Fusion of myoblasts and further growth of the newly regenerated
centronucleated myofibers ultimately results in restoration of the
cellular architecture within an ∼2-wk period

58. ⚫ Role of Stem cells in Disease and aging
Congenital and acquired myopathies are common and deadly. The
most common lethal myopathic disease is DMD, which is an X-
linked recessive disease.
Studies indicate that mutations within Lamin A are associated with
Emery- Dreifuss and Limb-Girdle muscular dystrophy patients.
Age-related myopathies (i.e., sarcopenia) also have impaired
regenerative defects
The impaired proliferative capacity of satellite cells that reside in
aged muscle has been shown to be due, in part, to perturbed
Notch signaling, as forced expression of activated Notch restored
the regeneration capacity of old (or aged) muscle
The study of these cells shall thereby aid an sound understanding of
somatic stem cell biology and serve as a platform for cell-based
therapies directed toward patients with congenital birth defects
and debilitating myopathies