Module - V

Stem Cell
&
Tissue Engineering

R. K. Gothwal, PhD
Department of Bioengineering and Biotechnology
BIT Mesra, Ranchi
Jaipur-Campus

Stem Cells
 A stem cell is a cell with the unique ability to develop into specialised cell
types in the body. In the future they may be used to replace cells and
tissues that have been damaged or lost due to disease.
 Stem cells are unique cells present in the body that have the potential to
differentiate into various cell types or divide indefinitely to produce other
stem cells.
• Stem cells are essential cells that replace
damaged cells or cells lost due to diseases.
• These cells are the earliest cells of the cell
lineage in all tissues and are found in both
embryonic and adult organisms.
• These cells provide a continuous supply of
new cells that make up the tissues and organs
of animals and plants.
• Stem cells have been of great interest as a
therapeutic method for various diseases and
conditions.
• The stem cells in the embryonic organism are
present in the inner cell mass of the
blastocyte, which then differentiates into all
other cells in the body.
• The stem cells in adults, however, are
localized to specific areas within the body like
in the bone marrow and the gonads.

1
Stem Cell Renewal and Differentiation

Properties of Stem Cells

 The two fundamental characteristics of stem cells are the ability of
unlimited self-regeneration and the production of at least one type of
differentiated progeny cells.
 All the stem cells found throughout all living systems have three important
properties. These properties can be visualized in vitro by a process called
clonogenic assays, where a single cell is assessed for its ability to
differentiate. The following are some properties of stem cells-
1. Stem cells, of all origins, are capable
of dividing and renewing themselves
for long periods of time. These cells
undergo a period of cell proliferation
while preserving the undifferentiated
state.
2. All stem cells are unspecialized or
undifferentiated. These are present
as a mass of cells that differentiate
later during their period of division.
3. Another essential property of stem
cells is their ability to differentiate into
specialized cells that together make
up different tissue types. These cells
can be either pluripotent or
multipotent.
yourgenome.org/facts/what-is-a-stem-cell

2
Totipotent, Pluripotent, & Multipotent Cells
 Totipotent stem cells: Cells able to give rise to all embryonic somatic
cells and germ cells. In other words, they can build a whole animal. The
zygote and a few early cells of the morula are totipotent.

 Pluripotent stem cells: These cells are descendants of totipotent stem

cells and can give rise to cells of the three germ layers: endoderm,
mesoderm, and ectoderm. They have no contribution to extraembryonic
membranes or the placenta.

 Multipotent stem cells:

These produce cells of a
particular lineage or
closely related family.

Tsonis P. A. (2007). Molecular Interventions, 7: 249-250.

Types of Stem Cells

 Embryonic stem cells- Embryonic stem cells supply new cells for an
embryo as it grows and develops into a baby. These stem cells are said to
be pluripotent, which means they can change into any cell in the body.

 Adult stem cells- Adult stem cells supply new cells as an organism grows
and to replace cells that get damaged. Adult stem cells are said to be
multipotent, which means they can only change into some cells in the body,
not any cell, for example: Blood (or 'haematopoietic') stem cells can only
replace the various types of cells in the blood. Skin (or 'epithelial') stem
cells provide the different types of cells that make up our skin and hair.

 Induced pluripotent stem cells- Induced pluripotent stem cells, or ‘iPS

cells’, are stem cells that scientists make in the laboratory. ‘Induced’
means that they are made in the lab by taking normal adult cells, like skin
or blood cells, and reprogramming them to become stem cells. Just like
embryonic stem cells, they are pluripotent so they can develop into any
cell type.

3
Applications of Stem Cells
 Research – to help us understand the basic biology of how living things
work and what happens in different types of cell during disease.
 Therapy – to replace lost or damaged cells that our bodies can’t replace
naturally.

 We can use stem cells to study how cells become specialised for specific functions
in the body, and what happens when this process goes wrong in disease. If we
understand stem cell development, we may be able to replicate this process to
create new cells, tissues and organs.
 We can grow tissue and organ structures from stem cells, which can then be
studied to find out how they function and how they are affected by different drugs.
 Cells, tissues and organs can sometimes be permanently damaged or lost by
disease, injury and genetic conditions.
 Blood stem cells are used to provide a source of healthy blood cells for people with
some blood conditions, such as thalassaemia, and cancer patients who have lost
their own blood stem cells during treatment.
 Skin stem cells can be used to generate new skin for people with severe burns.
 Age-related macular degeneration (AMD) is an example of a disease where stem
cells could be used as a new form of treatment in the future.
 Currently, damaged organs can be replaced by obtaining healthy organs from a
donor, however donated organs may be 'rejected' by the body as the immune
system sees it as something that is foreign.
 Induced pluripotent stem cells generated from the patient themselves could be used
to grow new organs that would have a lower risk of being rejected.
yourgenome.org/facts/what-is-a-stem-cell

Applications of Stem Cells

 Stem cell research has been used in the field of regenerative medicine,
which deals with the restoration of tissues or organs in the patient suffering
from severe injuries or some chronic disease.
 The progress made in the field of stem cell research has laid the
foundation for other cell-based therapies of disease that cannot be cured
with conventional medicines.
 Studies related to the human stem cell research has enormous potential
for contributing to our understanding of fundamental human biology.
 Many years of research on stem cells has made it possible to transplant
hematopoietic stem cells to the patients after the cancer treatments.
 Stem cell research has also been used for the testing of new drugs before
they can be tested in animals or humans.
 Cultured stem cells are used for the transplant of cells in the case of
various diseases like bone marrow for leukemia, nerve cells for
Parkinson’s and Alzheimer’s disease, heart muscle for heart disease, and
pancreatic islets for diabetes.

https://microbenotes.com/stem-cells

4
Limitations/ Challenges of Stem cell Research
 The most crucial challenge to stem cell research is the ethical issue
related to the use of embryonic stem cells. There are even political and
religious obstructions to stem cell research due to these issues.
 The source of some stem cell lines might have mutations which increase
the chances of mutations in the transplants.
 It is also difficult to transplant the stem cells produced in the laboratories to
the target cells.
 Embryonic stem cells also do not permanently renew themselves in vivo,
but instead, differentiate soon into different lineage progenitor cells of the
three embryonic germ layers.
 Self-renewal of these cells can be achieved in vitro under artificial
conditions, which inhibit their differentiation.
 It is also challenging to obtain a sufficient amount of stem cells with the
ability to differentiate into the desired cell type.
 The differentiation of embryonic, as well as adult stem cells, even if guided
by the addition of differentiation factors, inevitably involves a certain
amount of spontaneous differentiation into various cell types
 Additionally, the differentiation is not synchronizable yet, leading to a
mixture of cells in various stages of development.
https://microbenotes.com/stem-cells/#video-what-are-stem-cells-craig-a-kohn-by-ted-ed

Tissue Engineering

5
Tissue Engineering
 Tissue engineering was defined by Langer and Vacanti in early 90s as “an
interdisciplinary field which applies the principles of engineering and life sciences
toward the development of biological substitutes that restore, maintain, or improve
tissue function”
 Tissue engineering is a biomedical engineering discipline that uses a combination
of cells, engineering, materials methods, and suitable biochemical and
physicochemical factors to restore, maintain, improve, or replace different types of
biological tissues. Tissue engineering often involves the use of cells placed on tissue
scaffolds in the formation of new viable tissue for a medical purpose but is not limited
to applications involving cells and tissue scaffolds. While it was once categorized as
a sub-field of biomaterials, having grown in scope and importance it can be
considered as a field in its own.

Genetic Education Inc.

Overview of tissue engineering strategies

 Two main approaches are utilized in this area to produce engineered tissue.
 Scaffolding- It can be used as a cell support device upon which cells are
seeded in vitro; cells are then encouraged to lay down matrix to produce
the foundations of a tissue for transplantation.
 Using the Scaffold as a growth factor/drug delivery device- This
strategy involves the scaffold being combined with growth factors, so upon
implantation cells from the body are recruited to the scaffold site and form
tissue upon and throughout the matrices. These two approaches are not
mutually exclusive and can be easily combined.

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2475566/

6
Scaffolds
 Scaffold is a support that allows the seeded cells to interact and establish into a new
organ.
 The complexity of in vivo tissue organization allows cells to interact with each other
and with the surrounding extracellular matrix (ECM). In an engineered in vitro model,
the scaffold must be designed to finely replicate in vitro the architecture of the native
tissue, i.e., its ECM framework to let cells to adhere, spread, proliferate, differentiate,
maturate, and produce ECM, similarly to what they do in vivo.
 The choice of the most suitable biomaterial for scaffold fabrication is a key element for
the model design, since it strongly influences cellular functions. Due to their high
stiffness and load-bearing properties, ceramics and their composites are generally
used in TE of hard tissues, while polymers are mainly employed in the engineering of
soft tissues.

Scaffold

Scaffolds requirements
 Biocompatibility- The very first criterion of any scaffold for tissue engineering is that
it must be biocompatible; cells must adhere, function normally, and migrate onto the
surface and eventually through the scaffold and begin to proliferate before laying down
new matrix.
 Biodegradability- The objective of tissue engineering is to allow the body's own cells,
over time, to eventually replace the implanted scaffold or tissue engineered construct.
Scaffolds and constructs, are not intended as permanent implants. The scaffold must
therefore be biodegradable so as to allow cells to produce their own extracellular
matrix.
 Mechanical properties- Ideally, the scaffold should have mechanical properties
consistent with the anatomical site into which it is to be implanted and, from a practical
perspective, it must be strong enough to allow surgical handling during implantation.
 Scaffold architecture- Scaffolds should have an interconnected pore structure and
high porosity to ensure cellular penetration and adequate diffusion of nutrients to cells
within the construct and to the extra-cellular matrix formed by these cells.
 Manufacturing technology- In order for a particular scaffold or tissue engineered
construct to become clinically and commercially viable, it should be cost effective and
it should be possible to scale-up from making one at a time in a research laboratory to
small batch production.
 The final criterion for scaffolds in tissue engineering, and the one which all of the
criteria listed above are dependent upon, is the choice of biomaterial from which the
scaffold should be fabricated.

7
 Selection of cells for tissue engineering strategies
 The production of an engineered tissue in vitro requires the use of cells to populate
matrices and produce matrix resembling that of the native tissue. The main
successes in this field have come from the use of primary cells, taken from the
patient, and used in conjunction with scaffolds to produce tissue for re-implantation.
However, this strategy has limitations, because of the invasive nature of cell
collection and the potential for cells to be in a diseased state. Therefore, attention
has become focused upon the use of stem cells, including embryonic stem (ES) cells,
bone marrow mesenchymal stem cells (BM-MSCs) and umbilical cord-derived
mesenchymal stem cells (UC-MSCs).
1. Embryonic stem cells-
 Allow production of type-matched tissues for each patient, either
through stem cell banking or by the use of therapeutic cloning.
 Have the ability to be maintained for long (theoretically indefinite)
culture periods, therefore potentially providing large amounts of
cells for tissues that could not be derived directly from a tissue
source.
 Proof of the true pluripotent nature of ES cells is teratoma
formation. This property demonstrates the ability of stem cells to
tissue-engineer multiple tissue types but also highlights the
importance of using a terminally differentiated cell stock without
latent stem cell-like properties.
 The ability to control the differentiation of the cells to the desired
tissue lineages

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2475566/

Characteristics of
Embryonic Stem
Cells

8
 Selection of cells for tissue engineering strategies
2. Bone marrow-derived mesenchymal stem cells-
 Able to differentiate from a generic marrow cell population to an osteogenic lineage and have
been used to augment repair of bone.
 The MSC cell population can be isolated as a fraction of the adherent bone marrow colony
forming units – fibroblastic and can be differentiated to the osteogenic and other lineages.
 As marrow is a complex mixture of cells a more defined starting cell population can be
isolated from the mixture on the basis of epitope expression
 These cells can be removed from marrow and used to enhance materials such as the filler
used for stabilizing artificial hip joints or for joining critical sized defects in bone that would
not otherwise heal.

Selection of cells for tissue engineering strategies

3. Umbilical cord-derived mesenchymal stem cells-
 Umbilical cord blood contains MSCs that can undergo multi-lineage differentiation.
 The analysis of their gene expression profile reveals similarities to BM-MSCs with an ability to
differentiate into adipocytes, osteoblasts, hepatocytes and neuronal-like cells.
 If this type of stem cell does function as a BM-MSC it would greatly improve the availability of
matched tissues for treatments.
 This source of stem cells would provide a large pool of material, which could be purified using
non-invasive techniques and could be recipient matched. In addition, as BM-MSCs
differentiating potential may decrease with age, an alternative such as UC-MSCs would be of
huge benefit.

Differentiation capacities of
hUC-MSCs and their clinical
application prospects

9
Selection of cells for tissue engineering strategies
 Autologous cells- Cells are harvested from the individual intself. Under
some circumstances like genetic diseases and burns the cells are not
available.
 Recently there has been a trend towards the use of Mesenchymal Stem
Cells from bne marrow and fat tissues. These cells can differentiate into a
variety of tissue types, including bone, cartilage, fat and nerve.
 Allogeneic Cells- come from the boby of a donar of the same species.
 Xenogenic Cells- isolated from individuals of another species particularly
animal cells are used for construction of cardio vscular implants.
 Syngenic or Isogenic Cells- are isolated from genetically identical
organisms, such as twins, clones, or highly inbred research animal models.

Physicochemical Stimuli
 The in vivo environment guarantees the presence of fundamental molecular cues
that direct cell behavior, while the vascularization provides nutrient supply and waste
removal. Thus, the presence of molecular factors influencing cellular division, shape,
spreading, proliferation, death, and secretion of ECM components is necessary to
successfully model morphogenetic events.

 Microscale technologies, such as novel platforms based on microfabrication and

microfluidics, have shown to be another important tool, allowing real-time monitoring
and high-throughput results, with the possibility to test a single parameter in an
independent way.

 Cells are subjected to extracellular and

intracellular mechanical forces in vivo
that determine their fate. In particular,
cells respond to dynamic cues, such as
electric fields, osmotic and hydrostatic
pressure, stress, strain, fluid flow, and
streaming potential, by modifying the
surrounding ECM. Mechanical stimuli
are usually provided to tissue-
engineered constructs by bioreactors
specifically designed to reproduce the in
vivo conditions.

Caddeo et al. (2017), Front. Bioeng. Biotechnol.

10
Classic tissue engineering (TE) paradigm

Caddeo et al. (2017), Front. Bioeng. Biotechnol.

Classic tissue engineering (TE) paradigm

Caddeo et al. (2017), Front. Bioeng. Biotechnol.

11
Applications of Tissue Engineering
 Therapeutic cloning for tissue repair.
 Cardiovascular system
 Cardiac-tissue engineering
 Blood vessels
 Heart valves
 Muscoskeletal system
 Bone regeneration
 Bone and cartilage
reconstruction
 Artificial skin regeneration
 Urological repairs
 Oral mucosa tissue engineering
 Artificial pancreas
 Artificial liver

Limitations of Tissue Engineering

 Complexity using human tissue implants

 Variability in cell growth
 Standardization of process
 Irreversible process
 Risk assessment in clinical trials & commercialization
 Ethical issues

12
Challanges in Tissue Engineering
Selecting most effective-
- Cell type
- Scaffold
Fundamental Challanges - Growth factors (which?, combination)
Achieving proper vascularization
Achieving seamless host integration
Donor versus host cell contribution
Appropriate immunomodulatory biomaterials/agents
Knowledge limitation
Possible side effects/complications of donor cells
Most appropriate animal models
Regenerated bone quality
Evaluation challenges Regenerated bone functionality
Long-term tracking of the regenerated bone
Food & Drug Administration (FDA) Approval
Multi- vs. Single-component
Clinical Challenges
Expensive
Patient specific

Topics
 Central dogma of molecular biology
 Methods in genetic engineering and applications
 PCR and its application
 ELISA and its application
 Stem cell and tissue engineering
Artificial intelligence in biology
Plant factory

13
 BE-101

Biological Science for Engineers

M1: Basic Cell Biology
M2: Bioenergetics and Metabolism
M3: Enzymes and its Applications
M4: Biological Signal Generation & Propagation
M5: Engineering Biological Systems and its
Applications

THANK YOU

14