Basic Biology of Stem Cells
Week 2
Defining stem cells
• self-renewal, or the ability to generate at least one daughter cell with
characteristics similar to the initiating cell;
• multi-lineage
differentiation of a single cell;
• in vivo functional
reconstitution of a given
tissue TRENDS in Cell Biology (2002) 12:502
SYMMETRIC DIVISION
THE EMBRYO STEM CELL DIVIDES TO YIELD
TWO IDENTICAL TOTIPOTENT
DAUGHTER CELLS
ASYMMETRIC
DIVISIONTHE TISSUE PROGENITOR CELL GIVES RISE TO ONE DAUGHTER CELL WHICH REMAINS A PROGENITOR CELL AND ONE DAUGHTER CELL WHICH BEGINS THE
PROCESS OF DIFFERENTIATION LEADING TO TERMINATION
EMBRYONIC STEM CELLS
TISSUE STEM CELLS LOGARITHMIC EXPANSION
CONSTANT CELL NUMBER
TRANSIT
AMPLIFYING CELLS
ESC vs Adult SCs
• ES cell from mouse, non-human primates and human blastocysts fulfills all these
principles.
• Most adult stem cells in theory also fulfill this criteria, even though the degree of self renewal and differentiation potential is less than that seen for ES cells.
– Hematopoietic stem cell , neural stem cells,
mesenchymal stem cells, epidermal stem cells
Symmetric and Asymmetric division
• Stem cells can divide by either asymmetric or symmetric modes of division.
• The balance between these two modes is controlled by developmental and
environmental signals to produce appropriate numbers of stem cells and differentiated daughters.
Stem Cell division
Morrison 2006
Stem Cell with plasticity
• self-renewal ability;
• differentiation of a single cell into cells of the tissue of origin and into at least one
cell type different from the tissue of origin;
• Functional differentiation in vivo into cells of the tissue of origin and at least one cell type of a tissue other than the tissue of
origin.
Stem cells and asymmetric cell division
• Intrinsic mechanism
• asymmetric partitioning of cell components (regulators of cell polarity and fate determinants) that determine cell fate
• Extrinsic mechanism
• asymmetric placement of daughter cells relative to external cues
Morrison 2006
Examples ???
Stem cells and symmetric cell division
• Development
• Wound healing and regeneration
• Outcome is “increase in the number of stem cells”
C. elegans germ cells divide symmetrically during larval development
Stem cells
Differentiated cells Niche
(Morrison 2006)
Other
Examples ???
Stem cell plasticity
• It has been described as the unexpected potential of cells from bone marrow, muscle or brain to give rise to cells of
tissues other than the tissue of origin.
• Examples include bone-
marrow-derived cells or even purified hematopoietic stem cells that can give rise to
endothelium, skeletal muscle, cardiac muscle or hepatocytes.
TRENDS in Cell Biology (2002) 12:502
Plasticity of hematopoietic bone marrow cells
• More than 80% of studies reporting adult stem cell
plasticity have been performed using bone marrow (BM), or BM or peripheral blood (PB) enriched for HSC.
• Differentiation not only into hematopoietic cells but also cells with characteristics of skeletal muscle, cardiac
muscle, endothelium, neuroectoderm, skin epithelium and endodermal cells, including hepatocytes,
gastrointestinal epithelium and lung epithelium, has been described.
• most studies also did not fulfill the third criterion of stem cells, namely functional differentiation in vivo.
Stem cell plasticity of mesenchymal bone marrow cells
• MSCs can be cultured ex vivo for several
passages and differentiate at the single-cell level into mesodermal cell types, including
osteoblasts, chondroblasts, adipocytes, fibroblasts and skeletal myoblasts.
• When introduced in vivo, MSCs differentiate into the same array of cell types.
• Several recent reports have shown that MSCs might acquire characteristics of cells outside the mesoderm, including endothelium,
neuroectoderm and endoderm.
Plasticity of skeletal muscle cells
• It has been shown that the transplantation of muscle cells or muscle side population cells, could give rise to hematopoietic cells following lethal irradiation, and could compete with BM- derived HSCs, a criterion commonly used to demonstrate the presence of functional HSCs.
• The same cell that gives rise to hematopoietic cells also gives rise to myoblasts.
Plasticity of neural cells
• Murine and human NSCs cultured ex vivo for several population doublings could
differentiate into hematopoietic cells.
Plasticity of other tissues
• Cells isolated from dermis of mouse and human can be cultured ex vivo for several months and can differentiate into cells with phenotypic
characteristics of neurons and glial cells, as well as adipocytes and smooth muscle, in vitro.
• Human pancreatic cell line as well as fetal pancreatic tissue can be induced to acquire phenotypic characteristics of hepatocytes in vitro.
• However, the differentiated cells were not functionally characterized, and in vivo
repopulation studies were not performed.
J Pathol 197: 441
Possible explanations for the plasticity
• Stem cells for a given tissue might exist in an unrelated organ.
• Plasticity might be caused by the transplanted cells fusing with a host cell of a different lineage, leading to transfer of the genetic information of the
transplanted cell to the host-derived cell.
• Plasticity might occur via de- and re-differentiation, as is seen in cloning or in limb regeneration in amphibians.
• Cells with pluripotent characteristics might persist even after the initial steps of embryological development
Wagers 2004
‘Although stem cell plasticity is not
fully proven, there is sufficient evidence to warrant continued efforts to prove or disprove that
some adult stem cells might be more pluripotent…’
Stem Cell Niches
Niche hypothesis
• In 1978, Schofield proposed the “niche”hypothesis to describe the physiologically limited microenvironment that supports stem cells
• Variety of coculture experiments in vitro and by bone marrow transplantation supported niche hypothesis.
Stem Cell Niche
• Stem cell niche as “a specific location in a tissue where stem cells can reside for an indefinite
period of time and produce progeny cells while self-renewing”
• The likely existence of microenvironmental
factors produced by niche stromal cells has long cautioned that some aspects of stem cell biology may be difficult to deduce from purified stem
cells
The importance of stem cell niches
• The potential of stem cells in regenerative medicine relies upon removing them from their natural habitat, propagating them in culture, and placing them into a foreign tissue environment. To do so, it is
essential to understand how stem cells interact with their micro- environment, to establish and maintain their properties.
Stem Cell Niches
• Exactly when and how most somatic stem cell niches develop is still a
mystery. And there is considerable variation in niche design.
Stem Cell Niches
Cell, 116: 769
Stem Cell Niches
Stem cells and their niches
Xie 2007
Proposed niche types.
• Simple niche. A stem cell (red) is associated with a permanent partner cell (green) via an adherens junction (blue). The stem cells divides
asymmetrically to give rise to another stem cell and a differentiating
daughter cell (orange).
Proposed niche types.
• Complex niche. Two (or more) different stem cells (red and pink) are
supported by one or more partner cells (green).
Their activity is
coordinately regulated to generate multiple product cells (orange and yellow) by niche regulatory
signals.
Proposed niche types.
• Storage niche.
Quiescent stem cells are maintained in a niche until activated by external signals to divide and migrate
(arrows).
Adult stem cell niche model
Walker 2009
Simple Niches
• Bone morphogenetic protein (BMP) in
Drosophila germ-line stem cells (GSCs), directly controls growth and inhibits differentiation
• Wnt signaling may play a similarly direct role in regulating stem cells in the mouse intestinal
crypt
• Niches must ensure that daughter cells
differentiate appropriately as they leave the niche.
• Seemingly simple niches may exhibit complex temporal behavior. For example, it may be possible to support new stem cells without maintaining any special, pre- existing stromal architecture (‘empty niches’).
• Local structures to anchor and maintain new stem cells might simply be induced following stem cell arrival
• Finally, some tissues may have the capacity to support stem cells without any anatomical specializations beyond a large expanse of basement membrane. The basement membrane of mammalian epidermis or seminiferous
tubule may fall in this category.
Complex niches
• Some niches appear to be more complex than the relatively simple ones.
• Subventricular zone (SVZ) neural stem cells closely
associate with and sometimes specifically contact other astrocytes, neuroblasts, ependymal cells, endothelial cells and a factor-rich basal lamina.
• Complexity may also arise when multiple stem cells reside within a niche
• Niches should not be thought of as units of stem cell
maintenance, but rather as units of production of specific cellular outputs — spermatogonial, cysts, ovarian
follicles, intestinal villi,
Storage Niches
• the ‘storage niche’, may contain quiescent stem cells
• Storage niches may simply be normal niches that are located in favorable, damage-resistant regions or they may contain unique mechanisms to facilitate the safe maintenance of quiescent cells.
Programming daughter cells
• Niches with active stem cells must contain routes for progeny cells to exit.
• A cell should leave the niche when it reaches a location that cannot itself support a stem cell because one or more critical adhesive or signaling factors is no longer present.
• Even before it has done so, the daughter cell may begin to differentiate. Thus, niches are likely to contain specific structural features and mechanisms designed to ensure appropriate daughter cell movement and to initiate
differentiation.
Niches in different organisms
• Studies regarding stem cells and their
location/niche in other genetic model systems, including Drosophila and Caenorhabditis elegans
provide clues since locating and identifying stem cell
niches in mammals has
been extremely difficult due to their complex anatomic
structure. Cell, 116: 769
Stem Cell niches in drosophila
• Two or three GSCs are located at the tip of the ovariole in the structure referred to as the
germarium.
• These GSCs are
surrounded by three types of somatic cells:
terminal filament, cap
cells, and inner germarial sheath (IGS) cells.
Annu. Rev. Cell Dev. Biol. 2005
Cap cells are the niche for GSCs.
• GSC divides to generate two daughter cells:one daughter that stays in association with cap cells and another daughter that moves away from the cap cells to form a cystoblast, which eventually becomes, through incomplete cytokinesis, an interconnected 16-cell cyst.
• The anchorage of GSCs to cap cells through E-cadherin-
mediated cell adhesion is
essential for maintaining GSCs.
• Cap cells express genes, such as dpp, gbb, hh, piwi, and Yb, that are known to be important
for maintaining GSCs Annu. Rev. Cell Dev. Biol. 2005
SSCs in the germarium
• Two or three SSCs
located in the middle of the germarium are
responsible for
generating somatic follicle and stalk cells
• The follicle cell
encapsulate 16-cell cysts, whereas the stalk cells connect adjacent egg
chambers. Annu. Rev. Cell Dev. Biol. 2005
The interaction between GSCs and Niches
• The molecular glue that anchors GSCs to their niches is at least in part DE-cadherin, which along with its partner, Armadillo (β-catenin in vertebrates), concentrates at GSC-niche
borders
• A) Apical tip of a wild-type germarium; DE-cadherin (red), Armadillo/β-catenin (green), and nuclei (blue). The yellow band indicates colocalization of DE-cadherin and Arm, prominent at the border between cap cells and GSCs. AJ, adherens junction. (B) Electron micrograph illustrating an adherens junction (arrow) between a cap cell (left) and a GSC (right). Nu, nuclei. Mi, mitochondria.(C) Schematic of the adherens junction in (B), depicting a
homotypic interaction of the extracellular domains of the transmembrane DE-cadherins (red) that are linked via catenins to the actin cytoskeleton (orange).
Cell, 116: 769
GSC Niche in the Drosophila Testis
• In the apical tip of the Drosophila testis, two types of stem cells, GSCs and SSCs are responsible for
producing differentiated germ cells and somatic cyst cells.
• Seven to nine GSCs, each containing a spectrosome, are attached to the hub
Annu. Rev. Cell Dev. Biol. 2005
GSC niche in C.Elegans
• The putative germ line stem cells (GSCs) are directly associated with their distal tip cell (DTC) niche cell, whereas their differentiated progeny move away from the DTC, progressing from the mitotic phase to the meiotic phase.
Annu. Rev. Cell Dev. Biol. 2005
Stem cell niches in mammalians
– the epithelial stem cell location in the bulge area of hair follicles,
– and the intestinal stem cell near the crypt base.
– In 2003, osteoblastic cells, primarily those lining the trabecular bone surface, as the key component of the HSC niche
– In the neural system, the stem cell niche was found in endothelial cells located at the base of the
subventricular zone (SVZ) and subgranular zone (SGZ)
The Hematopoietic Stem Cell Niche
• Bone marrow serves as the pioneer system for studying stem cell
• However, the way in which HSCs interact with their local environment to promote
stem cell maintenance has not been clear
Recent developments in HSC niche
• A subset of osteoblastic cells (N-cadherin+CD45−) to which HSCs physically attach in the bone marrow are identified,
• N-cadherin/β-catenin adherens complex between HSCs and osteoblastic cells have been identified,
• Jagged1, generated from osteoblasts, influences HSCs by signaling through the Notch receptor have been
shown
• A number of Ncadherin+ osteoblastic lining cells control on the number of HSCs have been demonstrated
• GFP-labeled HSCs after transplantation also pointed to the endosteal surface as a possible stem cell niche
• In vitro coculture of HSCs with osteoblasts can expand the HSC population
• depletion of osteoblasts leads to loss of hematopoietic tissue.
• In addition, N-cadherin is a key target of Angiopoietin-1 (Ang-1)/Tie-2 signaling that maintains HSC quiescence
The niche
that regulates the birth and differentiation of blood-forming HSCs
• Depleting osteoblasts of a receptor for bone
morphogenetic protein (BMP) caused a doubling in both the osteoblast population and the stem-cell population.
• expansion of the stem-cell population when they
increased the numbers of osteoblasts by using
parathyroid hormone (PTH).
Nature 425, 836–841 (2003)
HSC niche.
• The HSC niche is located primarily on the surface of trabecular bone, where a small subset of spindle- shaped N-cadherin-positive
osteoblastic cells (indicated as
SNO cells) are the key component of the HSC niche.
• N-cadherin and β-catenin form an adherens complex at the interface between stem cells and niche
cells, assisting stem cells in attaching to the niche.
• Multiple growth factors and cytokines are involved in stem- niche interaction. (SCF/Kit, Jagged/Notch, SDF-1/CXCR4, and Ang1/Tie2)
Annu. Rev. Cell Dev. Biol. 2005
HSC niche (unknowns)
• BMP4 is expressed in osteoblastic cells, but which type of receptor is expressed in HSCs is unknown.
• The Wnt signal is important for stem cell self-renewal, but which Wnts are present in the niche is unknown.
• The same is true for FGF and
hedgehog. In vitro data suggest they affect HSC behavior; however, whether they are present as niche signals is unknown.
• Different types of stromal cells (illustrated as different colors and shapes) may regulate stem cell activation, proliferation, and
differentiation by secreting different microenvironmental signals.
• Finally, maturated blood cells migrate and infiltrate into blood vessel.
Annu. Rev. Cell Dev. Biol. 2005
The Epithelial Stem Cell Niche
• Skin, provides an excellent system for studying the molecular mechanisms that regulate stem cell self-renewal, proliferation, migration, and lineage commitment
• Each hair follicle is composed of a permanent portion, which includes sebaceous glands and the underlying bulge area
• The bulge area functions as a niche, where
epithelial stem cells (are located and maintained
Epidermal stem cells
• Stem cells are located in the bulge region of the hair follicle beneath the sebaceous gland.
• Upon activation, stem cells undergo division; the daughter cells retained in the bulge
remain as stem cells while other daughter cells migrate down to become hair-matrix progenitors responsible for hair regeneration.
• Stem cells can also migrate upward and convert to
epidermal progenitors that replenish lost or damaged epidermis.
Annu. Rev. Cell Dev. Biol. 2005
Epidermal stem cells
• The bulge area is an
environment that restricts cell growth and differentiation by expressing Wnt inhibitors, including DKK,Wif, and sFRP as well as BMPs.
• Wnts from dermal papilla (DP) and Noggin, which is derived from both DP and bulge,
coordinate to overcome the restriction signals imposed by both BMPs and Wnt inhibitors;
this leads to stem cell
activation and subsequent hair regeneration.
Annu. Rev. Cell Dev. Biol. 2005
The Intestinal Stem Cell Niche
• ISCs are located above the Paneth cells. Telomerase, Tcf4, EphB3, P- PTEN, P-Akt, 14-3-3ζ , Noggin, and Musashi-1—are expressed
• Mesenchymal cells adjacent to the ISCs function as the niche.
• BMP4 expressed from the niche influences the ISCs.
• Wnt signaling is present throughout the crypt
• Noggin is proposed to be a molecular switch coordinating with Wnt signaling to fully activate stem cells by
overriding BMP restriction signaling
• Wnt signaling plays a positive role in promoting ISC activation/self-renewal
• in contrast, BMP signaling restricts ISC activation/self- renewal
• in intestine overriding the restriction of BMP activity by Noggin as well as by active Wnt signaling are required to fully activate stem cells and support ongoing
regeneration
The Neural Stem Cell Niche
• NSCs can be isolated from various regions in the adult brain and peripheral nervous
system.
• the subventricular zone (SVZ) and the subgranular zone (SGZ) of the
hippocampus region are the primary and well-characterized regions in which NSCs reside and support neurogenesis in the
adult brain
The subventricular zone (SVZ)
• Astrocytes (B) lining the ependymal cells (E)
function as NSCs; they give rise to transient amplifying cells (C) (green), which further produce neuroblast cells (A) (blue).
• Endothelial cells in the blood vessel/ laminar maintain contact with
astrocytes, which regulate NSC self-renewal and
proliferation by generating different types of signals.
Curr.Opin. Genet. Dev 12:543
The subgranular zone (SGZ)
• Astrocytes (B) directly attach to the blood
vessel and receive signals from the
endothelial cells that direct NSCs to
undergo self-renewal, proliferation (D), and differentiation (G).
Curr.Opin. Genet. Dev 12:543
Signals generated from NSC niche
• BMPs and their antagonist Noggin, FGFs, IGF, VEGF,TGFa.
• BMP favors astrocyte lineage fate
• Noggin favors neurogenesis
• Overexpression of beta catenin leads to expansion of NSC, through the activation of Wnt signalling.
The Germ Line Stem Cell Niche in Mice testes
• The GSCs in mice are single cells that are
located in the periphery of seminiferous tubules and that have the ability to selfrenew and
generate a large
number of differentiated gametes
Cross section of mouse testis.
• A germ line stem cell (GSC) (red) directly contacts a Sertoli cell’s (purple) basement
membrane (gray) secreted by myoid cells (pink), and
specialized region (green), which together may form a
putative GSC niche. Myoid cells (pink) may also participate in niche function, as they are close to GSCs.
• The differentiated
spermatogonial cells (yellow) are germ-line cysts that move through different domains
formed by Sertoli cells toward the lumen, where mature sperm are released.
Adult stem cell niche model
Walker 2009
Common Features, Structures, and Functions of the Stem Cell Niche
• The stem cell niche is composed of a group of cells in a special tissue location for the
maintenance of stem cells.
• The niche’s overall structure is variable, and different cell types can provide the niche
environment.
• The niche functions as a physical anchor for stem cells.
• E-cadherin-mediated cell adhesion events or integrins, may help anchor stem cells to
extracellular matrixes.
Common Features, Structures, and Functions of the Stem Cell Niche
• The niche generates extrinsic factors that control stem cell fate and number.
• Many signal molecules (hh, Wnts, BMPs FGFs, Notch, SCF, Ang-1 , SCF, and LIF) have been shown to be involved in regulation of stem cell behavior, and among these, the BMP and Wnt signal pathways have emerged as common
pathways for controlling stem cell self-renewal and lineage fate from Drosophila to mammals.
Common Features, Structures, and Functions of the Stem Cell Niche
• The presence of signaling components of
multiple conserved developmental regulatory pathways in stem cells supports the ideas that stem cells retain the ability to respond to these embryonic regulatory signals and that
orchestration of these signals is essential for proper regulation of stem cell self-renewal and lineage commitment.
Common Features, Structures, and Functions of the Stem Cell Niche
• The stem cell niche exhibits an asymmetric structure. Upon division, one daughter cell is maintained in the niche as a stem cell (self- renewal);
• the other daughter cell leaves the niche to
proliferate and differentiate, eventually becoming a functionally mature cell.
