Stem cell Biology

Embryonic development and origin of

stem cells.

General Education Program

Biology

Presented by:Dr. Shaimaa Nasr Amin

Lecturer of Medical Physiology

INTRODUCTION

Historical Perspectives

• Major changes in regenerative medicine

(replacement of damaged or diseased cells

and tissues with new cells and tissues) due

to advances in stem cell technologies

Historical Perspectives

• Some stem cell therapies in existence for over

50 years. First successful bone marrow

transplant done in 1956 on leukemia patient.

Bone marrow contains adult‐derived

hematopoietic stem cells (able to regenerate

tissues similar to the specialized tissues in

which they are found.

Historical Perspectives

• Embryonic stem cells believed to have greater

potential. This line of stem cell research has

been the most controversial.

1998 - Researchers first extract stem cells from human

embryos

1999 - First Successful human transplant of insulin-making

cells from cadavers

2001 - President Bush restricts federal funding for embryonic

stem-cell research

2002 - Juvenile Diabetes Research Foundation International

creates $20 million fund-raising effort to support stem-

cell research

2002 - California ok stem cell research

2004 - Harvard researchers grow stem cells from embryos

using private funding

2004 - Ballot measure for $3 Billion bond for stem cells

Historical Perspectives

Historical Perspectives

• 2007 Nobel Prize in Medicine

• Mario R. Capecchi, Martin J. Evansand Oliver

Smithies for their discoveries of the principles

for introducing specific gene modifications in

mice by the use of embryonic stem cells.

STEM CELL BASICS

Stem Cell – Definition

A cell that has the ability to continuously divide and differentiate (develop) into various other kind(s) of cells/tissues

Stem Cell Characteristics

• „Blank cells‟ (unspecialized)

• Capable of dividing and renewing

themselves for long periods of time

(proliferation and renewal)

• Have the potential to give rise to

specialized cell types (differentiation)

Self renewal is needed because if the stem

cells didn‟t copy themselves, you would quickly

run out. It is important for the body to maintain a

pool of stem cells to use throughout your life.

• Differentiation is important because

specialized cells are used up, damaged or die

all the time during your life. Specialized cells

cannot divide and make copies of themselves,

but they need to be replaced for your body to

carry on working.

Kinds of Stem Cells

Stem cell

type Description Examples

TotipotentEach cell can develop

into a new individual

Cells from early

(1-3 days)

embryos

PluripotentCells can form any (over

200) cell types

Some cells of

blastocyst (5 to 14

days)

Multipotent

Cells differentiated, but

can form a number of

other tissues

Fetal tissue, cord

blood, and adult

stem cells

Kinds of Stem CellsEmbryonic stem cells

• five to six-day-old embryo

• Tabula rasa

Embryonic germ cells

• derived from the part of a human embryo or fetus

that will ultimately produce eggs or sperm

(gametes).

Adult stem cells

• undifferentiated cells found among specialized or

differentiated cells in a tissue or organ after birth

• appear to have a more restricted ability to produce

different cell types and to self-renew.

• Unlike gametes (egg and sperm cells), all

other cells (somatic cells) have the same

DNA content and the same genes.

Multipotent stem cells

•Multipotent stem cells – limited in what the cells can become

EMBRYONIC STEM CELLS

• Embryonic stem cells: found in the blastocyst,

a very early stage embryo that has about 50

to 100 cells

Embryonic stem cells: Challenges

• Scientists around the world are trying to

understand how and why embryonic stem

cells produce skin, blood, nerve or any other

particular kind of specialized cell.

• What controls the process so that the stem

cells make the right amount of each cell

type, at the right time?

ADULT STEM CELLS

What are Adult Stem Cells?

• An adult stem cell is an undifferentiated

(or partially-differentiated) cell found in

tissues and organs

• They can self-renew and differentiate to

become most or all of the specialized cell

types within their specific tissue lineage.

What are Adult Stem Cells?

• Adult stem cells

• Maintain cell populations

• Help you heal

• Play a role in aging

Homeostasis

• The ability to regulate internal conditions,

usually by a system of feedback controls

Stabilize health and functioning, regardless

of the outside changing conditions.

Homeostasis

• One piece of homeostasis is the constant

or periodic generation of new cells to

replace old, damaged, and dying cells

• Adult stem cells fulfill this role through the

process of regeneration

How Regeneration Works

• Adult stem cells normally remain quiescent (non-dividing)

for relatively long periods of time until they are activated

by signals to maintain tissues

• When activated they divide through a process called

asymmetric cell division

• Through this process they are able to maintain their

populations and differentiate into the desired cell types by

the creation of a progenitor cell

• A progenitor cell, in contrast to stem cells, is already far

more specific: they are pushed to differentiate into their

"target" cell.

Asymmetric Cell Division1. Proliferates

2. Maintains pop.

3. Creates Progenitor Cell

Progenitor cell

Stem cell Stem cell

Location of Adult Stem Cells

• Adult stem cells and progenitor cells reside

through out your body

• These stem cells reside in a specific area

of each tissue called the “stem cell niche”

Location of Adult Stem Cells

• This niche is a particular microenvironment

that fosters the growth of resident stem

cells

• Mutations in cells, signals they receive, and

changes in the microenvironment can

activate a stem cell

Types of Adult Stem Cells

1. Hematopoietic stem cells: blood and

immune system

2. Mesenchymal stem cells: bone, cartilage,

fat, muscle, tendon/ligament

3. Neural stem cells: neurons, glial cells

Epithelial stem cells: skin, linings

Hematopoietic stem cells

Give rise to all the blood cell types:

• Myeloid (monocytes and macrophages,

neutrophils, basophils, eosinophils,

erythrocytes, megakaryocytes/platelets,

dendritic cells)

• Lymphoid (T-cells, B-cells, NK-cells)

Hematopoietic stem cells

Found in the bone marrow from very early

on in development, as well as in umbilical

cord blood and placental tissue

Mesenchymal stem cells

• These stem cells will

differentiate into:

• cartilage cells

(chondrocytes)

• muscle cells (myocytes)

• fat cells (adipocytes)

• tendons, ligaments, and

connective tissue

(epithelial cells including

osteoblasts)Smooth muscle cells (red)

© CIRM

Mesenchymal stem cells

• These cells are located

throughout the body

• Bone marrow, fat, and cord

blood are easiest to isolate

Smooth muscle cells (red)

© CIRM

Neural stem cells

• They are located in:

• Subventricular zonelining the lateral

ventricles, where they

give rise to newly-born

neurons that migrate to

the olfactory bulb via the

rostral migratory stream

• Subgranular zone, part

of the dentate gyrus of

the hippocampus

Top: Section of the

hippocampus, blue

dots are neural stem

cells

Left: Mature neuron

(red)

© CIRM

Neural stem cells

• Neural stem cells (also

called Neural

precursor cells) give

rise to neurons,

oligodendrocytes, and

astrocytes Top: Section of the

hippocampus, blue

dots are neural stem

cells

Left: Mature neuron

(red)

© CIRM

Epithelial stem cells

• Give rise to epithelial

cells which constitute

60 percent of the

differentiated cells in

the body.

Retinal pigment epithelial cells

© CIRM

Epithelial stem cells

• 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.

Retinal pigment epithelial cells

© CIRM

Epithelial stem cells

• Epithelial stem cells

are also found in the

bulge region of the hair

follicle

Retinal pigment epithelial cells

© CIRM

Adult Stem Cell Therapies

Bone Marrow Transplant

Tissue Specific Organs

• In November 2008,

scientists in Spain

carried out a trachea

transplant for a woman

whose windpipe had

been damaged by

tuberculosis.

Tissue Specific Organs

• The doctors took adult stem cells and some other cells from the healthy right airway of the woman needing the trachea transplant, grafted those cells onto the stripped-down donated (cadaver) trachea, and marinated the trachea in chemicals in a lab to coax the trachea into rebuilding itself.

Adult Stem Cell Mobilization from

Bone Marrow

Clinical Trials

Clinical trials are conducted in phases.

The trials at each phase have a different

purpose and help scientists answer

different questions:

Clinical Trials

Phase I trials: researchers test an

experimental drug or treatment in a small

group of people (20-80) for the first time to

evaluate its safety, determine a safe

dosage range, and identify side effects.

Clinical Trials

Phase II trials: the experimental study drug

or treatment is given to a larger group of

people (100-300) to see if it is effective and

to further evaluate its safety.

Clinical Trials

Phase III trials: the experimental study drug or

treatment is given to large groups of people

(1,000-3,000) to confirm its effectiveness,

monitor side effects, compare it to commonly

used treatments, and collect information that will

allow the experimental drug or treatment to be

used safely.

Clinical Trials

Phase IV trials: post marketing studies

delineate additional information including

the drug's risks, benefits, and optimal use.

Risk vs. Benefits of Participating in a

Clinical Trial

Risk

• The patient must

stop taking other

treatments before

the trial

Benefit

• Play an active role

in their own health

care.

Risk vs. Benefits of Participating in a

Clinical Trial

Risk

• There may be

unpleasant, serious

or even life-

threatening side

effects to

experimental

treatment.

Benefit

• Gain access to new

research treatments

before they are

widely available.

Risk vs. Benefits of Participating in a

Clinical Trial

Risk

• The experimental

treatment may not

be effective for the

participant, or given

a placebo

Benefit

• Obtain expert

medical care at

leading health care

facilities during the

trial.

Risk vs. Benefits of Participating in a

Clinical Trial

Risk

• The protocol may

require more of their

time and attention than

would a non-protocol

treatment, including

trips to the study site,

more treatments,

hospital stays or

complex dosage

requirements.

Benefit

• Help others by

contributing to medical

research.

• The patient may get

better as a result of the

experimental

treatment.

Stem Cell Tourism

• In what is called “stem cell tourism” patients

travel to other countries with less

restrictions to receive stem cell therapies.

• Sometimes experimental and can be dangerous

• There are many legitimate therapies going

through national regulatory processes in these

countries.

Stem Cell Tourism

• December 2008 study of stem cell clinic

web sites

• Sites claimed to treat a range of diseases that

go beyond the scope of the early evidence on

stem cells' efficacy

• Played up the benefits and talked little about

risks

• Each treatment costs around $21,500

PLEURIPOTENT STEM CELLS

Embryonic Stem cells

Totipotent

Pluripotent

Multipotent

Unipotent

iPSCs, are a type of pluripotent stem cell

artificially derived from a non-pluripotent

cell, typically an adult somatic cell, by

inducing a "forced" expression of certain

genes.

first produced in 2006 from mouse cells

and in 2007 from human cells

WHAT ARE iPSCs ?

Genes responsible for pluripotency

• Group 1

ES cell-Specific transcription factors

Essential for pluripotency in ES cell & early embryos

Oct¾, Sox2, Nanog…

• Group 2

(Proto-oncogene's)

Important for proliferation of ES cells, but not in early

embryos

TCL1, Stat3, c-Myc, ERas, Klf4

Genes responsible for pluripotency

• Group 3

Less famous

Specifically expressed in ES cell

But less defined function

ECAT1, Esg1,Fbx15, …

Oct3/4:

• Involve in the maintenance of self renewal of

pluripotent cells.

• Repression in ES cells leads to the formation

of trophoectoderm.

• Overexpression leads to the formation of

various lineages including primitive

endoderm.

Sox2:

• Essential for embryonic development

• Downregulation by siRNA silencing leads to

the differentiation of cell in murine ES cells.

• Nanog: In embryonic stem cells, Nanog,

along with Oct-3/4 and Sox2, is necessary

in promoting pluripotency.

• LIN28: LIN28 is an mRNA binding protein

expressed in embryonic stem cells and

embryonic carcinoma cells associated with

differentiation and proliferation.

Nuclear reprogramming to a pluripotent state by three approaches

Shinya Yamanaka & Helen M. Blau

NATURE|Vol465|10 June 2010|doi:10.1038/nature09229

Nuclear reprogramming to a pluripotent state by three approaches

Shinya Yamanaka & Helen M. Blau NATURE|Vol 465|10 June

2010|doi:10.1038/nature09229

Production of iPSCs

• Typically derived by transfection of certain

stem cell-associated genes into non-

pluripotent cells, such as adult fibroblasts.

Production of iPSCs

• Transfection is typically achieved through

viral vectors, such as retroviruses.

Production of iPSCs

• After 3–4 weeks, small numbers of transfected

cells begin to become morphologically and

biochemically similar to pluripotent stem cells,

and are typically isolated through morphological

selection, doubling time, or through a reporter

gene and antibiotic selection.

(1)Isolate and culture donor

cells. (2)Transfect stem cell-

associated genes into the

cells by viral vectors.

(3)Harvest and culture the

cells according to ES cell

culture, (4)A small subset of

the transfected cells become

iPS cells and generate ES-

like colonies.

Human induced pluripotent stem cells

• Produced in November 2007.

• With the same principle used earlier in

mouse models, Yamanaka had successfully

transformed human fibroblasts into

pluripotent stem cells using the same four

pivotal genes: Oct3/4, Sox2, Klf4, and c-

Myc with a retroviral system.

Pluripotency induction is a slow and gradual

process

• After transfecting the cells with all of the four

factors (Sox2,OCct3/4,Klf4 and c-Myc) initially

only eight colonies were picked at 11th day and

ten colonies on 16th day.

(Alexender Meissner et al.;

Pluripotency induction is a slow and gradual

process

• Only one out of eight colonies from 11 day and

four colonies from ten colonies from 16 day old

colonies gave rise the ES like cells.

(Alexender Meissner et al.;

Complications

• Because of viral transfection systems, the

created cells might be prone to form

tumors.

Complications

• However Konrad Hochedlinger and his

Harvard University research team

successfully used an adenovirus to

transport the requisite four genes into the

DNA of skin and liver cells of mice.

Complications

• Since the adenovirus does not combine

any of its own genes with the targeted host,

the danger of creating tumors is eliminated.

Identity

The generated iPSCs were remarkably similar to naturally-isolated pluripotent stem cells

Cellular biological properties:

Morphology, Growth properties, Stem Cell Markers, Stem Cell Genes, Telomerase Activity:

Pluripotency of iPSCs :

Neural Differentiation, Cardiac Differentiation, Teratoma Formation, Embryoid Body,

Cells :iPSApplications of

• iPSC regarded as holy grail stem cell research

• Studying disease models in vitro

• Drug screening

• Toxicological testing of new drugs

Cells :iPSApplications of

• Generating patient specific & disease specific

pleuripotent stem cells

• Allow unprecedented access to all stages of

human biology

• Studying development & function of human

tissue

• Regenerative medicine

iPS CELLS TO CURE SC ANEAMIA

Cell replacement therapy seems particularly

suitable for Parkinson’s disease,

can be used for treatment of iPS

Parkinson’s disease

A common neurodegenerative disease caused by loss

of midbrain dopamine neurons . Transplantation of

fetal midbrain cells has been shown to restore

dopamine function in animal models and in human

patients .

can be used for treatment of iPS

Parkinson’s disease

cellsiPSMouse kidneys created using

A team of scientists has successfully used

induced pluripotent stem (iPS) cells to create

kidneys inside a mouse whose parents were

genetically engineered so their offspring would

not be born with the organ.

• iPS cells can be differentiated into electrically

active motor neurons, a discovery that may aid

in studying and treating neurological disorders.

iPSc can be used to create Electrically

Active Neuron

• The motor neurons derived from the iPS cells

appeared to be similar in function and

efficiency to those derived from human

embryonic stem cells, although further testing

needs to be done to confirm that.

iPSc can be used to create Electrically

Active Neuron

Somatic cells can be used to generate

the β-cells

Pancreatic Exocrine cells can be converted

into β-cells closely related to the islet β-

cells.

Can be used to cure diabetes.