An assignment

On

Biomaterials and other factors in stem cell differentiation, and tissue engineering
for limbal stem cell transplantation
Course Title:

Course Code: PHR5107

Section: 1

Submitted By,

Anika Tabassum Shama, ID: 2016325672

Submitted To,

Dr. Hasan Mahmud Reza

Professor

North South University

Submission Date: 4th June 2020

Biomaterials
Introduction

To work out stem cell proliferation efficiently extracellular microenvironment is a very

important factor that can be provided by biomaterials (1). Biomaterials have a very important
role in tissue growth. Is has the ability to control the growth of a tissue and also the ability to
control the behavior of a cell (2). These are made of synthetic or natural polymer. By processing
the biomaterials into scaffolds they can be used in tissue differentiation and tissue proliferation
that will be able to produce new tissues (2). Stem cells can work hand in hand with biomaterials
to treat fault tissue or organ (2). Along with cell proliferation, biomaterials also support lineage-
specific differentiation, cellular attachment (3). The basic requirements for a biomaterial to be
used as scaffolds are surface properties and biocompatibility (3). Without the need of any kind of
chemical factors it is possible to control cells using biomaterials (4). A variety of biomaterials
are being developed now. New development of a brilliant biomaterial can act like a backbone for
the clinical therapeutics (5).

Discussion

Depending on the microenvironment, different types of biomaterials might be needed (5). For
this reason, different types of ceramic-based, polymer-based and metal based biomaterials are
being developed to drive the microenvironment of the stem-cell differentiation. To mimic the
ECM (5) , biomaterials are composed of different types and can also be transformed in fibrous,
hydrogel or porus scaffold (5). Biomaterials can be of different type. For example it can be a
natural polymer or synthetic polymer or ceramics or metals. Different types of biomaterials are
described in the following.

Natural polymer

Several biocompatible and biodegradable polymers are used as biomaterials. These natural
polymers enzimatically or hydrolytically degrades into wastes or nutrients (5). These materials
are highly used in stem cell differentiation. Two natural polymers: proteins and polysaccharides
are described below.

 Proteins
 Collagen: Collagen is highly conserved and 30% of the total mammalian cells are of this
substance (5). Collagen is derived from bovine cartilage. Collagen can be used to
maintain chondogenesis (6). Collagen microbeads can expand hUCBCs and also can
improve cell viability (6).
 Fibrin: Fibrin is used in creating 3D scaffold materials. Fibrins are derived from tissues.
It has been found out that using fibrin along with growth factor can significantly increase
the oligodendrocyte and neuron production (7).
 Silk: Silk is a versatile polymer that is derived from silkworm cocoons. Silk can be
transformed into porous scaffolds and can be used in salt leaching or gas foaming method
(8).
 Matrigel: Matrigel is composed of various ECM components which includes collagen IV,
laminin and heparin sulphate proteoglycans. It is extensively used in cell culture (9). Pre-
cultured cell, when injected with matrigel can produce vascular structure (tube-like) (10).

Fig: A flow diagram of stem cell differentiation by using biomaterials

 Polysaccharide
 Chitosan: Chitosan is derived from crustacean shells.It is biocompatible and
biodegradable. Chitosan along with coralline is used to study MSC-osteogenesis (10).
  Hyaluronic acid: It is one of the very important biomaterial which is used in cell
signaling and cell behavior. It is a gel like substance and can be easily modified into
membranes, fibres and microspheres (11). It can be used to maintain the undifferentiated
and pluripotency of hESCs (12) .
 Chondroitin Sulfate
 Agarose
 Alginate: It is derived from algae cell walls. It is used for the differentiation and
encapsulation of ESC (13).

Synthetic polymer

 Polyethylene Gycol: Polyethylene glycol helps in regulating the expression of the

chondrogenic markers (10).
 L-lactic acid, glycolic acid, lactic-co-glycolic acid: These polymers contain ester and
they are hydrolytically degradable. These polymers are used in cell based therapeutics
(5).
 Polycaprolactone : In physiological condition this important biomaterial can be degraded
via the hydrolysis of ester bond (5). Scaffold porus PCL electrosporum fibre has been
used in the rat’s bMSCs to form tissues alike bones (5).
 Micellaneous polymers: Micellaneous polymers such as polyphosphoesters can be used
in tissue engineering as drug or gene carriers or as scaffold by degrading them into non-
toxic products (5).

Ceramic

This includes bioactive glasses, calcium phosphates and bioceramics, which are used to study
stem cell differentiation as well as osteoporosis (10). They can integrate with the bone easily can
drive matrix formation and mineralization. Ceramics are used in tissue engineering extensively
(14).

Metals

Metal for example titanium is widely used as biomaterial. It is used in dental and orthopedic
surgery (10).

Other factors in stem cell differentiation

Introduction

Stem cells are very important for our body as they have the capability to generate into any type
of cell according to the need of our body (15). It is very important to keep control over the tissue
homeostasis and for organogenesis (15). These cells are considered as a promising too for
treating different diseases and organ damage (16). Stems cells, depending on their differentiation
capability can be classified into pluripotent or adult or embryonic stem cells (16). On the other
hand, based on the potency of a stem cell it can be classified into totipotent (it can generate all
type of cells) , pluripotent (it can generate all type of body cells which includes germ cells as
well), multipotent ( it can generate all tissue cells) and unipotent cells ( it has the ability to
generate a single cell type) (17).

Fig: Stem cell differentiation

Discussion

Other than biomaterials there are different types of factors that work behind stem cell
differentiation. Such as: small molecules, nano particles, growth factors, morphogenic factors.
These factors are described below:

Small molecule

Stem cell differentiations can be effectively regulated by these small molecules. . Retinoic acid
(RA) has the ability to enhance the expression of the neural crest. It also has the ability to reduce
the mesodermal differentiation, which is an alternative of the mesenchymal origin. Use of
sodium butyrate results in producing hepatocyte type cells by expressing the glycolytic
phenotype. The steroid derivatives of cholesterol metabolism, thyroid hormones are also been
proved as an important differentiation factors. The small molecules are cell permeable. Some
examples of these small molecules are: vitamin C, dexamethasone, vitamin C, thyroid hormones,
sodium pyruvate, prostaglandin E2, concavalin A, dibutryl cAMP, vanadate as well as retinoic
acids.

Growth Factor

In stem cell differentiation different types of growth factors and cytokines are responsible. For
example it has been found out in a study that, the epidermal growth factor (EGF) stimulates the
differentiation of human mesenchymal stem cells into cells that can form a bone (18). Different
factors for example Wnt proteins, hedgehog proteins (Hhgs), the members of TGF-β super
family of the growth factors Notch ligands or FGFs have a very significant role in controlling
and maintaining the regulation of transcription factors. This may cause differentiation to a certain
lineage. Both BMP4 and TGF-β1 plays a very important role in Stem cell differentiation. It
causes bone morphogenesis by initiating chondro-progenitor cell determination and
differentiation. In the regenerative application, the control release of the growth factors through
diffusion plays a very important role. Enhanced cell differentiation and formation are some
achievements done by the controlled release of the growth factors. On the surface of the
biomaterials, growth factors are modified which provides some essential factors that can ease the
stem cell differentiation and the survival of in vivo.

Nano Particles

Nano particles have played an important role in scaffold modification (16).It is used to offer a
suitable and efficient microenvironment for cell differentiation, proliferation and adherence (16).
Nano particles are also involved into iPSC differentiation and reprogramming (16). Nano
particles include particles which are 1-100 nm in size (16). As they have a very small size and a
big surface area they are widely applied in the stem cell research (16).

Morphogenic Factor

Morphogenic factor is another important factor behind stem cell differentiation. It can lead to
tissue formation and commitment by modulating differentiation signaling pathways.
Chondrocyte secretes some morphogenic factors, which has the ability to regulate MSC
differentiation. It also has the ability to enhance osteogenic and promotes the chondrogenic
potential of the MSCs. The morphogenetic factors secreted by chondrocytes can regulate MSC
differentiation and also promote osteogenic as well as chondrogenic potential of MSCs. The
autocrine and paracrine factors that are secreted by differentiated cell also have their
significance. These factors, in proximity with the stem cells have the ability to lead a efficient
and direct transduction of the molecular signals. This might help in the expression of markers
that are organ specific. Conditioned medium of neural stem cell has also been proven as an
important reagent in can of promoting the differentiation of the embryonic stem cells. The
secreted factors in the conditioned media might be suitable for the prolonged expansion of
embryonic stem cells for tissue specific differentiation right before the transplantation. To
regenerate a fully functional tissue the co-transplantation and co-culture of different type of cells
can act an important therapeutic approach.

Tissue engineering for limbal stem cell transplantation

Introduction

Limbal stem cells are located in basal layer of limbus. They are responsible for the healing of the
epithelial of the cornea after any type of injury (19) . The cells that are originated from limbus
are found to be responsible to heal the damaged or impaired cornea. After the damage of the
cornea, limbal stem cell first proliferates, and then differentiates and finally it migrates to the
located injury (19). Tissue engineering is a process though which new tissues are formed in the
replacement of the old or damaged tissues (16). This method is usually used to restore the usual
function of the body tissues which medications and therapies doesn’t work (16). In this part we
will discuss the use of tissue engineering in the limbal stem cell transplantation.

Fig: Tissue Engineering

Discussion

Davanger and Evensen first observed in a study that limbal epithelial cells have the capability to
regenerate into corneal epithelial cells. According to their study, in the pigmented eyes while the
healing of the defected corneal epithelium, the pigmented line of the epithelium moves from
limbal region to central cornea. The basal epithelial cells of the limbus consists the lowest
number of corneal epithelium differentiated cells. It is found in a study that, k3 the 64kDA
keratin is present in the corneal epithelial cells. In suprabasal layer of limbus along with the
corneal epithelium, the k3 is expressed. Here the k3 was found to be absent in the limbal basal
cells. This proves that the limbal basal cells represent a non differentiated population, more
primitive cells that do not express cytokeratin. According to another study, it was found that, a
cornea specific keratin names K12 is expressed in the superbasal cell (limbus). It was found
absent in the basal limbal cell.

Cell turn over constantly occurs under the corneal epithelium. Here the loss terminally
differentiated cells are replaced by the basal epithelial cells. According to a study, the progenitor
cells are the originator of the corneal epithelial cells. These progenitor cells are located at the
limbus (in the basal cell layer). These progenitor cells are responsible for the regeneration and
for the renewal of the corneal epithelium. Transient amplifying cells are formed by the division
of the limbal stem cells. To gain superbasal location of limbus the transient amplifying cells
migrates superficially. They migrate centrally in the aim to form basal layer of corneal
epithelium. These transient amplifying cells have the capability to differentiate and forms post
mitotic cells. This superficial migration of the cells results in taking up the final characteristic
(phenotypic). The cells that are terminally differentiated and the post mitotic cells don’t have the
capability to differentiate.

There are some special structure related features that are important for the microenvironment of
the stem cells. The structural feature of limbus is perfect for protecting corneal stem cells. In a
human body, stem cells are located in the deep tissue layer. The limbal epithelium consists of 8-
10 thick layers of cells. On the other hand the corneal epithelium consists of 5 layers of cell. The
limbus is pigmented heavily and for this reason it has the capability to protect basal cells from
UV (Ultraviolet) ray’s carcinogenic effects. The stroma like component of limbus consists rich
network of vesicles, which allows the regulations and regulation of the growth of the limbal stem
cell. It also allows the proliferation by various pathways (cytokine mediated and neural
mediated).

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