Lecture 5

Stem cell Technology

Dr. Imran Riaz Malik

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Stem Cell

Can differentiate into other types of cells and can

also divide in self renewal to produce more of the same
type of stem cells.

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Dr. Imran Riaz Malik
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Significance of Stem Cell

The main function of stem cells is

- To serve as internal repair system
- To provide new cells to damaged organs as long as
person or animal is alive.
- When stem cells multiplies, each new cell has potential
to either continue as a stem cell(reservior) or become
another type of cell of the body such as neural cell, or
red blood and muscle cell.
Interestingly, stem cells are different from other cell types
in two ways:
- First, stem cells are unspecialized cells that are
capable of renewing themselves through cell division
- second, under special physiological or experimental
conditions, they can be differentiated to become tissue
or specific type of cells with
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cardiomyocytes and neurons.
Significance of Stem Cell

- In some organs, such as gut and bone marrow, stem

cells regularly divide to repair and replace damaged
tissues in body, whereas in other organs such as
pancreas and heart , stem cells only divide under
special conditions or requirements.
- In principle, there are two types of stem ells in human
body, they are ESCs and nonembryonic or somatic or
adult stem cells.

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Dr. Imran Riaz Malik
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Significance of Stem Cell

- In 1981, ESCs were isolated from early mouse embryos,

which later led to development of a method to derive
stem cells from human embryos which can grow in lab.
- Stem cells that are isolated from human embryo are
called human ESCs.
- Human embryos used in these studies are created for
reproduction by using in vitro fertilization procedures.
- When these embryos are no longer needed by
patients, they can be donated for research with the
informed consent of donor.
-

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Significance of Stem Cell

- Moreover, stem cells are important to living organisms

form many reasons
- The 3-5 day old embryo that is called a blastocyst
contains cells normally referred to as inner cells.
- The inner cell mass contains stems cells that give rise
to entire body of organism, including many specialized
cell types and organs such as skin, sperm, eggs, heart,
lung and other tissues.

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Significance of Stem Cell

- Interestingly, all stem cells, regardless of their source,

normally have three general properties:
i. They are capable of multiplying and renewing
themselves for long periods of time.
ii. They are unspecialized
iii. They can give rise to specialized cells types upon
differentiation.

- Stem cells are unspecialized cells and one of

fundamental properties of stem cells is that it does not
have any tissue specific structures or morphology that
allows it to perform specialized functions.
- For instance, a stem cell can not work like adult or
somatic cells to pump blood through body and stem
cells cannot carry oxygen
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stream.
Significance of Stem Cell

- Nevertheless, these unspecialized stem can give rise to

specialized cells through a process called
differentiated.
- During this process, stem cells usually become
specialized somatic cells in several stages.

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Dr. Imran Riaz Malik
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Classification of Stem Cells

- ESCs that isolated from developing embryo known as

blastocyst

- Adult stem cells that are found in adult tissues or

organs.
1. Embryonic stem cells

- In humans, ESCs are pluripotent stem cells derived

from the inner cell mass of blastocyst(an early stage
embryo).
- Moreover, human embryos reach the blastocyst stage
4-5 days post fertilization. At that time, they consist of
50-150 cells; these cells are called ESCs.
- ESCs are pluripotent stem cells and have the
capability to differentiate into
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layers
1. Embryonic stem cells

- Three germ layers later on give rise to more than 220

different cell types in adult body.
- ESCs can generate all cell types in the body, whereas
adult stem cells are multipotent and can produce an
inadequate number of cell types
- Furthermore, under well defined conditions, ESCs are
capable of propagating themselves for long periods of
time without losing their pluripotency.
- ESCs can be employed as useful models both for
conducting research and as regenerative medicine
because ESCs can produce unlimited numbers of cells
and can be transplanted into patients.
- with the help of ESCs, it is possible to treat various
diseases such as genetic diseases, diabetes,
Parkinson's disease, blindness,
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cord injuries.
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 Historical perspective of ES cells

- It is started in 1964 with the research work carried out

to isolate a single type of cell from a teratocarcinoma, a
tumour now known to be derived from a germ cell.
- These specialized cells were isolated from
teratocarcinoma, replicated, and grown in cell culture
media;
- These specialized cells were later named embryonic
carcinoma cells.
- In 1981, ESCs were derived for the first time from
mouse embryos by two research groups.. Martin Evans
and his coresearchers and Mathew Kaufman and his
co-researchers in their pioneering research, showed
for first time a technique to derive ESCs from mouse
embryo.
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 Historical perspective of ES cells

- Also, G.R. Martin, showed that embryos could be

cultured in lab using in vitro cell culture method.
- In 1998, a breakthrough occurred when researchers led
by James Thomson first developed a technique to isolate
and grow human ESCs under lab conditions.
- Isolation and culture of ESCs
- ESCs can be isolated from inner cell mass of early
embryo that is donated by mother.
- The method of isolating ESCs involves the delay in
embryo implantation, which allows the inner cell mass
to increase in size.
- The isolation process includes removing donor mother
ovaries and treating her with hormone progesterone,
which cause the embryos to remain free in uterus.
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- Isolation and culture of ESCs
- Furthermore, after 4-6 days of culture, the embryo are
harvested and grown in petri plates until the inner cell
mass forms an egg or cylinder like structures.
- These egg or cylinder shaped inner cell mass is then
detached into a single cell population, and plated on
fibroblast cells that previously treated with mitomycin-
c.
- The single cells are then processed for isolating clonal
cells. Moreover, clonal cells grown from these in vitro
cell cultures can form three germ layers.
- After successful isolation and culture of mouse ESCs,
efforts have also been made to isolate ESCs form
humans.
- For that purpose, a method to derive human ESCs and
cultured embryonic cells was established.
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- Isolation and culture of ESCs
- In the first step, embryos are removed from donor
mother at approximately 76h after fertilization and
cultured overnight in media containing serum and
nutrients.
- After inner cell mass is removed from late blastocyst
using microsurgery, the extracted inner cell masses are
cultured on fibroblasts treated with mitomycin-c in
media that contains serums.
- One week later, colonies of cells grow out and these
cells demonstrates pluripotent. The colonies of cells
have ability to form three germ layers, differentiate in
vitro, and form embryoid bodies.
- These embryoid bodies are commonly known as ESCs.
-

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- Isolation and culture of ESCs.
- Additionally, to maintain the pluripotentcy of stem
cells, the culture media is delivered with leukemic
inhibitory factor and bone morphogenetic proteins that
are necessary to prevent ES cells from differentiating.
- Moreover, these factors are extremely important for
efficiency of deriving ES cells.
-

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Dr. Imran Riaz Malik
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- Identification of ESCs
- While isolating and deriving ESCs, it is very important
to know the identity of stem cells and their cellular,
molecular and functional characteristics.
- The various characteristics of ESCs
-

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Dr. Imran Riaz Malik
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- Differentiation of ESCs
- One of the main characteristics of ESCs is to give rise
to many cell types, which have been demonstrated in a
number of experiments conducted using in vitro cell
method
- Under specific culture conditions and balanced
nutrients presence, these ESCs can be differentiated to
form various body cell types such as heart, skin,
kidney and nerve and many other cell types.
- Over the last few years, efforts have been made to
develop basic methods to differentiate ESCs into some
specific cell types.
- These differentiated cells such as neural cells or
cardiomyocytes can be used to treat Parkinson disease
and heart infarction.

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Types of Stem cells

1.Embryonic Stem cells

-- Isolated from the inner cell mass of blastocytes

2. Adult Stem cells

They are found in various tissues.

-- In adult organisms, stem cells and progenitor cells act

as a repair system for body, replenishing adult tissues.

-- In a developing embryo, stem cells can differentiate into

all the specialized cells- ectoderm, endoderm and
mesoderm but also maintain the normal turnover of
regenerative organs such as blood, skin or intestinal
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Sources of autologous adult stem cells in human

--bone marrow

--adipose tissue

-- blood

Stem cells can also be taken from umbilical cord blood

just after birth.

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--Adult stem cells are frequently used in various medical
therapies as bone marrow transplantation.

--Stem cells can now be artificially grown and

transformed or differentiated into specialized cell types
with characteristics consistent with cells of various
tissues such as muscles or nerves.

-- Embryonic cells lines and autologous embryonic stem

cells generated through somatic cell nuclear transfer or
dedifferentiation have also been proposed as promising
candidates for future therapies.

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Properties

Self renewal
the ability to go through numerous cycles of cell division
while maintaining the undifferentiated state.

Potency
-- the capacity to differentiate into specialized cell types.

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Self renewal

1. Obligatory asymmetric replication

--A stem cell divides into one mother that is identical to
original stem cell, and another daughter cell that is
differentiated.

--when stem cell self renews it divides and does not

disrupt the undifferentiated state. This self renewal stem
demands control of cell cycle as well as upkeep of
mutlipotency or pluripotency, which all depends on stem
cell.

2. Stochastic differentiation
-- when one stem develop into two differentiated daughter
cells, another stem cell undergoes mitosis and produce
two stem cells identical Dr.
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original.
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Potency

--potency specifies the differentiation potential of stem

cell.

1.Totipotent
-Stem cells can differentiate into embryonic and
extraembryonic cell types. Such cell can construct a
complete, viable organisms.

- Produced from fusion of egg and sperm cell. Cell

produced by first few divisions of fertilized egg are also
totipotent.
2. Pluripotent
- Descendants of totipotent cells can differentiate into
nearly all cells. I.e. Cells derived from any of three germ
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3.Multipotent
Stem cells can differentiate into a number of cell types,
but only those of closely related family of cells.

4. Oligopotent
Stem cells can differentiate into only a few cell types such
as lymphoid or myeloid stem cells.

5. Unipotent
Cells can produce only one cell type, their own, but have
the property of self renewal, which distinguishes them
from non stem cells e.g progenitor cells which can not
self renews

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Dr. Imran Riaz Malik
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